Evolutionary Ecology of Marine Invertebrate Larvae

Evolutionary
Ecology of Marine
Invertebrate Larvae

Ed i t e d by

Tyler J. Carrier
University of North Carolina at Charlotte, USA

Adam M. Reitzel
University of North Carolina at Charlotte, USA

Andreas Heyland
University of Guelph, Canada

1
3
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We dedicate this edited volume to the “Larval Marvel,” Larry McEdward. Larry edited
Ecology of Marine Invertebrate Larvae (1995), the first comprehensive edited volume with a
marine larval ecology focus. His keen interest and knowledge in the field started many
years before, and was nurtured under the guidance of Dr. John Lawrence and Dr. Richard
Strathmann. In the late 1980s, Larry organized a group of biologists whose expertise
included a diversity of topics centered around larval life histories such as fertilization,
feeding, development, dispersal, and settlement, which resulted in the publication
of Ecology of Marine Invertebrate Larvae. In the past two decades, this text has inspired
new and experienced researchers alike and brought a new generation of students into
the field. Larry’s passing in 2001 was a great loss to the field of marine larval biology.
We hope that the current volume, Evolutionary Ecology of Marine Invertebrate Larvae, will
continue inspiring scholars of the field to investigate the many marvels of larvae. We miss
you, Larry.

Benjamin G. Miner
Adam M. Reitzel
Andreas Heyland
Many of the authors of this edited volume cut our teeth on Ecology of Marine Invertebrate
Larvae (1995). The work of Larry McEdward as well as others of his generation and
their mentors built a foundation for understanding a diverse set of topics about marine
invertebrate larvae. With this formative groundwork laid, we have been able to pose
and actively pursue answers to difficult questions about mechanisms and evolution of
larval development and life histories; to apply our knowledge to systems beyond our
easily accessible nearshore environment; and to seek answers to pressing questions about
marine life in the context of our rapidly changing climate.
Diane K. Adams was a rising star in developmental and larval ecology who dove
headfirst into these difficult questions. In her short career, she significantly advanced our
understanding of the mechanisms that underlie phenotypic plasticity of larval feeding
structures and contributed groundbreaking work on dispersal of deep-sea hydrothermal
vent larvae. We will miss our friend, colleague, and collaborator, and dedicate Chapters
8 and 16 of this volume to the memory of Diane. We promise to keep tackling the hard
questions, Di.

Shawn Arellano
Justin McAlister
Abstract

For more than a century, evolutionary biologists, ecologists, and oceanographers alike have been intellectually
stimulated by marine invertebrate larval forms. In 1995, Ecology of Marine Invertebrate Larvae, edited by the
late Dr. Larry McEdward, captured the fundamental phenomenon and tremendous diversity of reproductive,
biological, and oceanographic aspects of larval ecology. Now, more than twenty years later, this edited vol-
ume provides an update to many of the original 13 chapters, while also reviewing several branches of larval
ecology and evolution that have developed since. In Evolutionary Ecology of Marine Invertebrate Larvae,
authors review the origins of marine invertebrate larvae and the developmental mechanisms and ecological
factors that may generate this great diversity, and how these microscopic organisms feed, develop, and be-
have in the pelagic environment. Whether in coastal areas or the deep sea, larvae are exposed to a diversity
of environmental factors, which affect their physiology and development as well as subsequent stages of
their life history. Now, in an age of climate change, larvae face a warmer, more acidic, and more toxic ocean
than ever before. Responses to these plus many other stressors and facets of larval biology can be broadly
profiled, with current technological advances. This edited volume provides a major synthesis of an important
interdisciplinary field, as it aims to foster stimulating discussions centered on the evolution and ecology of
marine invertebrate larvae.

Keywords
Marine invertebrate larvae, ecology, evolution, oceanography, life history
 Preface
In the early twentieth century, the field of larval ecol-
ogy materialized into an independent discipline of
study. The following decades brought tremendous
advances in the appreciation and understanding of
larvae, including the sheer morphological diversity,
mechanisms of feeding, and processes of metamor-
phosis. The conceptual framework of larval ecology
as a field was not summarized until the end of that
century, when the late Larry McEdward edited a
13-chapter volume entitled Ecology of Marine Inver-
tebrate Larvae (1995). Now, following more than two
decades of development, we have decided to organ-
ize an update for many of the chapters featured in
Ecology of Marine Invertebrate Larvae as well as re-
views of several new directions the field has taken.
Because many of the chapters in this volume are
highly integrative and cover many different species
as well as approaches, we grouped them conceptu-
ally into four sections: (1) Evolutionary origins and
transitions in developmental mode; (2) Functional
morphology and ecology of larval forms; (3) Larval
transport, settlement and metamorphosis; and (4)
Larval ecology at the extremes.
Section 1, Evolutionary origins and transitions in
developmental mode: In Chapter 1, Claus Nielsen pro-
vides an overview of the diversity of larval types
from groups of marine invertebrates as diverse
as  the echinoderms, crustacean, and annelids, as
well as emphasizes some of the central hypoth-
eses regarding the evolution of larval types and
how they evolved in the first place. In Chapter  2,
Heather Marlow provides a detailed overview of
the evolutionary development of marine larvae.
She revisits the question, “What is a larva?” and
guides her discussion by comparing specific sen-
sory structures of the planktonic life form and how
it provides unique insight into how gene regulatory
networks may have diversified over evolutionary
time in patterning the larval body. In Chapter 3,
Dustin Marshall and colleagues revisit questions
of parental investment and life history trade-offs,
as originally addressed by Vance, McEdward, and
others. These important questions, as they have
for decades, continue to have a fundamental influ-
ence on the life history theory of marine larvae. In
­Chapter 4, Rachel Collin and Amy Moran primar-
ily discuss transitions in developmental mode.
Namely, while planktotrophy (feeding) and lecitho-
trophy (nonfeeding) have largely dominated this
discussion in the past, a thorough phylogenetic ap-
proach, using broad taxon sampling provides excit-
ing new evidence for how these primary modes of
development and feeding may have changed over
evolutionary time. Lastly, in Chapter 5, Jonathan
Allen and colleagues summarize the current under-
standing of asexual reproduction by larvae, includ-
ing the diversity of developmental strategies and
environmental factors eliciting a cloning response.
Section 2, Functional morphology and ecology of
larval forms: In Chapter 7, Bruno Pernet reviews
fundamental principles for mechanisms of larval
feeding, as well as detailing the specifics for more
than a dozen phyla of marine invertebrates. He
then compares rates of feeding and performance
for many larval types in nature. In Chapter 8, Justin
McAlister and Benjamin Miner outline the pheno-
typic plasticity of feeding structures in marine in-
vertebrate larvae. They discuss the evolution and
molecular underpinnings of this trait, as well as
proper experimental design and statistical analy-
ses. Finally, in Chapter 9, William Jaeckle focuses
his chapter on the physiology of larval feeding: he
x   P r e fa c e
begins with the ingestion of a food particle or dis-
solved organic material and ends with the delivery
and assimilation of the associated biomolecules.
Section 3, Larval transport, settlement, and meta-
morphosis: In Chapter 11, Jesús Pineda and Nathalie
Reyns define and discuss larval transport, both as
a concept and as a measurement. In doing so, they
provide an overview of components of larval trans-
port, from behavior to advective and diffusive pro-
cesses, as well as present challenges the field faces
and emerging approaches to combat these chal-
lenges. In Chapter 12, Peter Marko and Michael
Hart provide a lucid discussion of approaches for
genetic analysis of larval dispersal and population
connectivity. They discuss the use of genetic data
in characterizing connectivity, the differentiation
of individual- vs. population-based methods, and
the utility of isolation-by-distance patterns and dis-
persal kernels. In Chapter 13, Jason Hodin and col-
leagues emphasize the integration of fluid dynamics
and sensory ecology, across spatial and temporal
bounds, in the process of settlement and metamor-
phosis to holistically assess how larvae use diverse
cues to find settling grounds. Finally, in Chapter 14,
Jan Pechenik discusses latent effects, the embryonic
and larval experience and its influence on life after
metamorphosis. He outlines how abiotic and biotic
stressors that larvae face shape this experience, as
well as begins to detail how many mechanisms me-
diating these consequences remain unknown.
Section 4, Larval ecology at the extremes: In Chap-
ter  16, Craig Young and colleagues assess larval
transport patterns in the deep-sea and the ecologi-
cal and evolutionary constraints on such patterns.
They specifically hypothesize how these environ-
ments may select for specific reproductive and lar-
val characteristics in these vast habitats. In Chapter
17, Maria Byrne and colleagues shift our perspective
of larval evolutionary ecology toward the predicted
future ocean, by outlining how marine invertebrate
larvae are affected by ocean warming and acidifica-
tion, as well as the interaction between these stress-
ors. They discuss how a changing oceanic climate
favors poleward movement, and why certain larval
types may be more resilient to these stressors than
others. In Chapter 18, Ilaria Corsi and Luis Fer-
nando Marques-Santos discuss how toxicants affect
embryonic and larval functionality with a focus on
the ABC transporters of echinoids. They provide a
history of ABC transporters, discuss the utility of
echinoids in ecotoxicology research, and emphasize
the need to find complementary models. In Chapter
19, Elizabeth Williams and Tyler Carrier provide an
-omics-focused perspective on marine invertebrate
larvae. Specifically, they outline the use of a multi-
omics approach in deciphering the molecular mech-
anisms of life history evolution and the resiliency
to environmental stress, as well as new fronts in
the field, including characterizing larval-associated
microbiota. In Chapter 21, we conclude the volume
with a summary of its contents from Richard Strath-
mann and Alan Love, who propose how marine in-
vertebrate larvae continue to collectively serve as a
model for studying life history evolution.
The editors and many of the chapter authors
who contributed to this edited volume would like
to acknowledge the impact that Ecology of Marine
Invertebrate Larvae (McEdward, 1995) had on our re-
search interests and, indeed, even on our decisions
to pursue science as a profession. We anticipate that
research utilizing marine invertebrate larvae will
continue to grow, and just as the previous two dec-
ades have opened up new avenues in research since
McEdward (1995), the coming years will continue
to bring new ways to understand marine inverte-
brate larvae. If this book can have even a small im-
pact on promoting the growth of the field, then we
will feel this project will have been successful. As
a field, we still know surprisingly little about vast
numbers of marine invertebrates, including whole
phyla in some cases, and parts of the globe remain
virtually unexplored. Lastly, marine invertebrate
larvae have at different times in history held par-
ticular sway in the understanding of central ques-
tions in biology, ranging from macro-evolution of
life histories to cell differentiation, developmental
pathways, and gene regulatory networks; the time
may be ripe, again, for larvae to take center stage.
There are many people who contributed substan-
tially to the planning and preparation of this volume.
We first thank Ian Sherman, Senior Commissioning
Editor for Biology at Oxford University Press, for
agreeing to undertake this project and especially for
guiding us through the publishing process. Thanks
to Lucy Nash and Bethany Kershaw, Senior Assis-
tant Commissioning Editors at Oxford University

Press, for their immense assistance in organizing
this volume. Lastly, we thank the external reviewers
commissioned by Oxford University Press for this
project, as well as the external reviewers as selected
by the editors, for providing invaluable remarks on
improving our original drafts of our proposal and
chapters submitted by the authors, respectively.
The editorial work of Tyler Carrier was done in
the Department of Biological Sciences at the Uni-
versity of North Carolina at Charlotte and at both
the Friday Harbor Laboratories of the University of
Washington and the Darling Marine Center of the
University of Maine. The editorial work of Adam
P r e fa c e    xi
Reitzel was done in the Department of Biologi-
cal Sciences at the University of North Carolina at
Charlotte and supported by a Junior Faculty Devel-
opment Award from UNC Charlotte. The work of
Andreas Heyland was done in the Department of
Integrative Biology at the University of Guelph and
at the Friday Harbor Laboratories of the University
of Washington. We greatly appreciate the facilities
and assistance from the staff at these institutions.
Tyler J. Carrier
Adam M. Reitzel
Andreas Heyland
 The Editors
Tyler J. Carrier, BS, is an NSF Graduate Research
Fellow, an NSF Graduate Research Opportunity
Worldwide awardee, and a PhD student in the De-
partment of Biological Sciences at the University of
North Carolina at Charlotte.
He received his BS from the University of Maine
in 2015, was a visiting research scholar at Brown
University that summer, and began his PhD that
fall. As part of his doctoral work, he has been a
visiting graduate researcher at the Friday Harbor
Laboratories of the University of Washington, the
Darling Marine Center of the University of Maine,
the Duke Marine Lab of Duke University, and the
University of Sydney (Australia).
Tyler has been the recipient of research grants
from the National Science Foundation, the Maine as
well as North Carolina Sea Grants, and Sigma Xi. To
date he has published six peer-reviewed papers in
international journals.
His research interests are in how oceanographic
phenomena shape evolution in the sea with an
emphasis on marine invertebrate larvae, as well as
host-microbiota partnerships and how these rela-
tionships promote evolutionary innovation.
Adam M. Reitzel, PhD, is an Assistant Professor in
the Department of Biological Sciences at the Uni-
versity of North Carolina at Charlotte.
Dr. Reitzel obtained his MSc in zoology from the
University of Florida in 2002, a PhD in biology from
Boston University in 2008, and was a postdoctoral
scholar and fellow with Ann Tarrant and Mark
Hahn at Woods Hole Oceanographic Institute from
2008 to 2012.
Dr. Reitzel has published more than 60 peer-
reviewed publications in journals such as Molecu­
lar Ecology, BioEssays, and Marine Ecology Progress
Series, and has organized various meetings and
symposia. Dr. Reitzel has received funding from
National Science Foundation, the National Insti-
tutes of Health, the Human Frontiers Science Pro-
gram, and other organizations in support of his
research program.
He is interested in the ecology and evolution of
coastal invertebrates, particularly echinoderms and
cnidarians. His research combines comparative de-
velopment, phylogenomics, and functional biology
to test hypotheses concerning the origin of signal-
ing pathways and mechanisms mediating organ-
ism-environment interactions.
Andreas Heyland, PhD, is an Associate Professor
in the Department of Integrative Biology at the Uni-
versity of Guelph.
Dr. Heyland obtained his MSc in zoology from
the University of Zurich in 1999, a PhD in zoology
from the University of Florida in 2004, and was
trained as a postdoctoral fellow with Leonid Mo-
roz from 2004 to 2007 at the Whitney Laboratory for
Marine Biosciences.
He has published more than 60 peer-reviewed
scientific articles, published abstracts and book
chapters, many in international journals such as Bio­
Essays, Evolution, Evolution & Development, Nature,
and Cell. He co-edited the book Mechanisms of Life
History Evolution with Thomas Flatt. He is regularly
invited to speak at universities and conferences and
to review journal articles and grant proposals.
Dr. Heyland is interested in understanding the
physiological and molecular mechanisms under-
lying marine invertebrate life histories. Using a
combination of evolutionary-developmental biol-
ogy, functional genomics, and physiology, his work
examines the role of hormones in development,
metamorphosis, and life history plasticity in marine
invertebrates, such as echinoderms.
 List of Contributors
Jonathan D. Allen  College of William & Mary, USA
Shawn M. Arellano  Shannon Point Marine Center,
Western Washington University, USA
Maria Byrne  University of Sydney, Australia
Tyler J. Carrier  University of North Carolina at
Charlotte, USA
Rachel Collin  Smithsonian Tropical Research Institute,
Republic of Panama
Ilaria Corsi  University of Siena, Italy
Symon A. Dworjanyn  Southern Cross University,
Australia
Matthew C. Ferner  San Francisco Bay National
Estuarine Research Reserve and San Francisco State
University, USA
Brian Gaylord  Bodega Marine Laboratory, University
of California at Davis, USA
Michael W. Hart  Simon Fraser University, Canada
Jean-François Hamel  Society for the Exploration and
Valuing of the Environment, Canada
Andreas Heyland  University of Guelph, Canada
Jason Hodin  Friday Harbor Laboratories, University of
Washington, USA
William B. Jaeckle  Illinois Wesleyan University, USA
Alan C. Love  University of Minnesota, USA
Heather Marlow  Pasteur Institute, France
Luis F. Marques-Santos  University of Paraiba, Brazil
Dustin J. Marshall  Monash University, Australia
Justin S. McAlister  College of the Holy Cross, USA
Annie Mercier  Memorial University, Canada
Peter B. Marko  University of Hawaii at Mānoa, USA
Benjamin G. Miner  Western Washington University,
USA
Amy Moran  University of Hawaii at Mānoa, USA
Claus Nielsen  Natural History Museum of Denmark,
University of Copenhagen, Denmark
Laura M. Parker  University of Sydney, Australia
Jan A. Pechenik  Tufts University, USA
Bruno Pernet  California State University Long Beach,
USA
Jesús Pineda  Woods Hole Oceanographic Institution,
USA
Adam M. Reitzel  University of North Carolina at
Charlotte, USA
Nathalie B. Reyns  University of San Diego, USA
Pauline M. Ross  University of Sydney, Australia
Richard R. Strathmann  Friday Harbor Laboratories,
University of Washington, USA
Elizabeth A. Williams  Max Planck Institute for
Developmental Biology, Germany
Craig M. Young  University of Oregon, USA
 C h a pt er 1
Origin and Diversity of Marine Larvae
Claus Nielsen
1.1  Introduction—Defining a Larva
Marine larvae have fascinated zoologists for centu-
ries, not only for their beauty, but also for their in-
formation about animal evolution, about food webs
in the plankton, and, more recently, about the gen-
etic regulation of development (Davidson, 2001).
But what is a larva? (For an additional discussion of
this issue see Marlow chapter in this volume.)
Intuitively it seems easy to define a larva, based
on observations of life cycles. Two main types are
generally recognized: indirect (biphasic) life cycle
with a larval stage (morphologically and ecologic-
ally different from the adult) and direct without
a larva. However, even this simple distinction is
problematic. There are numerous definitions both of
the term larva (e.g., Hickman, 1999; Young, 2002b)
and of the term metamorphosis (e.g., Bishop et al.,
2006). The typical indirect development of marine
invertebrates involves an embryo (within the fertil-
ization membrane), hatching from the fertilization
membrane, a larva (with special larval structures),
metamorphosis (with loss of larval structures), and
a juvenile/adult (with special adult structures).
This type of life cycle is shown, for example, in
many molluscs, annelids, and echinoderms.
Hatching intuitively appears as a well-defined
process, but the egg membrane becomes incorpo-
rated into the larval cuticle, for example, in annelids
such as Phragmatopoma, Polygordius, and Phascolo-
soma (Rice, 1967; Eckelbarger and Chia, 1978; Smart
and Von Dassow, 2009) and in gastropods such as
Ifremeria (Reynolds et al., 2010). Most often, meta-
morphosis marks the transition from one lifestyle
to another, for example, from pelagic to benthic or
Nielsen, C., Origin and diversity of marine larvae. In. Evolutionary Ecology of Marine Invertebrate Larvae.
Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018).
© Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0001
from planktotrophy to benthic feeding. This transi-
tion involves loss of larval feeding structures, such
as larval ciliary bands, the velum in gastropods and
bivalves, or long spines or chaetae in crustaceans
and polychaetes.
Metamorphosis may be very gradual, as, for ex-
ample, in the Müller’s larvae of polyclad turbellar-
ians, where the larval tentacles are lost gradually
(Rupper­ t, 1978), or “catastrophic,” where larger
larval structures are cast off or devoured, as, for ex-
ample, in the annelid Polygordius lacteus ­(Figure 1.2F)
and the phoronid Phoronis muelleri ­(Herrmann, 1979).
A change of habitat/lifestyle, such as planktonic
to  benthic (free-living or sessile) or parasitic, is
observed in a majority of species. Only a few hol-
oplanktonic species, such as some heteropod gastro-
pods, have a life cycle with a planktotrophic veliger
larva and a carnivorous adult. One complication
is seen, for example, in many marine gastropods
which go through a nonfeedin­g larval stage with a
velum within an egg capsule and then hatch as ju-
veniles without the velum (e.g., Littorin­a obtusata
and Buccinu­m undatum; Frette­r and Graha­m, 1994).
In Galeodea (Cassidaria), the intracapsular veliger
is even “filtering” yolk particles with the ciliary
bands (Fioroni, 1966). Another complication is the
“metagenesis” of the medusozoans, where the larva
settles and metamorphoses into a juvenile (polyp),
which then buds off ephyra stages rather differ-
ent from the sexually mature medusae; these small
ephyras (Figure 1.1D) are often called larvae.
As should be expected from a continuously
evolving group such as the metazoans, develop-
mental types do not fall into well-defined “boxes,”
so a pragmatic definition will be used here: a larva
4   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v e rt e b r at e L a r va e

(A) (B) (C) (D)

(E) (F) (G)

(H) (I) (J)

(K) (L) (M)

Figure 1.1  (A) Larva of the sponge Spongia ceylonensis (courtesy of Liu Li-Lian, National SunYat-sen Univ.). (B–D) Cnidarian larvae: (B) Larva of the
anthozoan Nematostella vectensis (courtesy of Gáspár Jékely, Max Planck Institute for Developmental Biology, Tübingen). (C) Larva of the anthozoan
Isarachnanthus nocturnus manii (courtesy of Alvaro Esteves Migotto, Univ. São Paulo). (D) Ephyra “larva” of Aurelia sp. (courtesy of Peter Bryant, Univ.
California, Irvine). (E) Cydippid larva of a ctenophore (courtesy of Otto Larink, TU-Braunschweig). (F) Müller’s larva of a polyclad turbellarian (courtesy
of Kate Rawlinson, Cambridge Univ.). (G,H) Nemertine larvae: (G) Pilidium larva of Maculaura cerebrosa and (H) lecithotrophic larva of Pantinonemertes
californiensis (both courtesy of Svetlana Maslakova, Univ. Oregon). (I,J) Brachiopod larvae: (I) Terebratalia transversa (© Nina Furchheim), (J) Lingula
anatina (courtesy of Karen Chan, STRI). (K,L) Bryozoa: (K) Cyphonautes larva of Membranipora sp. (courtesy of Peter Bryant, Univ. California, Irvine). (L)
Drawing of the lecithotrophic larva of Scrupocellaria scruposa (after Calvet, 1900). (M) Actinotrocha larva of a phoronid (courtesy of George van Dassow,
Univ. Oregon) (see Plate 1).
 O r i g i n a nd D i v e r s i t y o f M a r i ne L a r va e    5

(A) (B) (C)

(D) (E) (F)

(G) (H) (I)

(J) (K) (L)

Figure 1.2 Annelid (A–F) and mollusc (G–L) larvae. (A) Metatrochophora larva of Hydroides sp. (courtesy of Brian Nedved, Univ. Hawai´i at
Mānoa). (B) Large larva of a chaetopterid (courtesy of Karen Osborn, Smithsonian Institution). (C) Pelagosphaera larva of a sipunculan (courtesy
of Mary Rice, Smithsonian Marine Station at Fort Pierce). (D,E) Sabellaria sp. (D) undisturbed larva, (E) protective pose with spread chaetae (both
courtesy of Otto Larink, TU-Braunschweig). (F) Drawings of the pericalymma larva and its metamorphosis of Polygordius sp. (after Woltereck, 1902).
(G) Larva of a polyplacophoran (courtesy of Jenna R. Valley, Univ. Oregon). (H) Unidentified gastropod veliger (courtesy of Peter Bryant, Univ.
California, Irvine). (I) Veliger of the heteropod Carinaria japonica (courtesy of Jackie Sones, Univ. California, Davis). (J) Unidentified gastropod
veliger (courtesy of Peter Bryant, Univ. California, Irvine). (K) Veliger of Mytilus californianus (courtesy of Eric Sanford, Univ. California, Davis).
(L) Unidentified bivalve veliger (courtesy of George von Dassow, Univ. Oregon) (see Plate 2).
6   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v e rt e b r at e L a r va e

is the part of the life cycle stage between hatch-
ing and “metamorphosis,” where metamorphosis
marks more-or-less radical changes in both mor-
phology and lifestyle.
1.2  Origin of Larvae
It seems generally accepted that the ancestral meta-
zoan was a spherical, ciliated organism with direct
development (Richter and King, 2013; see Marlow
chapter in this volume for additional hypothesis),
probably consisting of choanoflagellate-like cells,
a choanoblastaea (Nielsen, 2008). The biphasic life
cycle with a larva and an adult can in principle have
developed in two ways; either by intercalation of a
larval stage into the life cycle between the zygote
and the adult—the intercalation theory, or by add-
ing a new adult stage to the life cycle and retaining
the “old” adult as a larva—the terminal addition
theory (Figure 1.3). The terminal addition theory
can be explained by adaptive evolution from small,
probably holopelagic, planktotrophic ancestors to
forms with planktonic, planktotrophic larvae and
larger benthic adults, which could exploit a variety
of new feeding modes (Nielsen, 2012b). The inter-
calation theory cannot explain the evolution of the
several specialized structures of the larval feeding
structures, which can only be of adaptive value
when fully formed (Nielsen, 2009). A very illus-
trative example is the downstream-collecting cili-
ary system of the trochophora larvae (Figure 1.4),
which can only function when fully differentiated

Terminal addition theory

zygote

planktotrophic
adult gastraea

zygote

planktotrophic benthic deposit-feeding adult

larva

Intercalation theory

zygote

benthic deposit-feeding adult

zygote
Figure 1.3 Two theories for the “origin of larvae.”
planktotrophic benthic deposit-feeding adult The added stage of the life cycle is dark gray (modified
larva from Nielsen, 2013).

stomach
mouth
anus
(A)
Figure 1.4 The “typical” planktotrophic trochophora larva exemplified by serpulid polychaetes. (A) Light microscopy of Hydroides sp. (courtesy
of Bruno Pernet, California State Univ., Long Beach). (B) SEM of Serpula columbiana (Claus Nielsen).
(Nielsen, 2013). Further, an intercalation of the feed-
ing trochophora larva would involve convergent
evolution of the complex downstream-collecting
ciliary system hundreds of times in a number of
phyla, which appears highly unlikely. Moreover,
it appears that the eumetazoans may have evolved
from larvae of a poriferan or a poriferan ancestor,
because the choanoflagellates and the adult sponges
have undulatory cilia, whereas the sponge larvae
and most of the eumetazoans, both larvae and
adults, have “effective stroke cilia” (Nielsen, 2012a).
The new hypothesis proposing that the ctenophores
are the sister group of all other metazoans (Ryan et
al., 2013; Moroz et al., 2014) is still much discussed
(Pisani et al., 2015). It is not important for the pre-
sent discussion.
Direct development is found in many metazoan
lineages, and is prevalent in small, interstitial spe-
cies. This lifestyle and developmental type, perhaps
originating in anaerobic environments, has been
interpreted as ancestral (Boaden, 1989) and is still
discussed (Laumer et al., 2015), but this would im-
ply that the planktotrophic larval stages with com-
plicated feeding structures characteristic of many
phyla have evolved convergently numerous times.
It should be mentioned that evolution of non-
planktotroph­y in clades with mainly planktotrophic
O r i g i n a nd D i v e r s i t y o f M a r i ne L a r va e    7
apical tuft
prototroch
metatroch gastrotroch
(B)
development is well documented, whereas evolu-
tion in the opposite direction is poorly documented
(see the following and chapter by Collin and Mora­n,
this volume).
1.3  Variation in Larval Types
As mentioned earlier, invertebrate larvae show an
immense diversity (Levin and Bridges, 1995). This
brief discussion will center around the variation in
developmental types: planktotrophic-lecithotrophic-
direct, but special larval structures which protect
the larvae from predation is amply demonstrated
through the illustrations.
The developmental type varies in several of the
larger clades. Ciliated “primary” larvae are found
in many clades, but, for example, all ecdysozoan
larvae are characterized as secondary larvae be-
cause they lack ciliated epithelia (Hadfield et al.,
2001). Direct development is found in most free-
living platyhelminth groups, but planktotrophic
larvae are found in some polyclads (Figure 1.1F).
Cephalopods are almost exclusively direct develop-
ers, but species of the family Chirotheuthidae have
a “paralarva” stage with a very special morphol-
ogy, which was originally described as a separate
genus, Doratopsis (Young, 1991). The ascidians are
8   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v e rt e b r at e L a r va e
especially interesting, because most species have a
life cycle with a tadpole larva, but a number of spe-
cies of the families Molgulidae and Styelidae have
“anuran” larvae without the tail (Hadfield et al.,
1995). Experimental work has demonstrated that
the silencing of just one gene changes the develop-
mental type from the “tailed” to the “tail-less” type
(Swalla and Jeffery, 1996), and this, together with
the scattered occurrence of the anuran larval type in
the two families, shows that the direct-developing
type can easily change from the indirect, whereas
evolution in the opposite direction is not known.
Many marine larvae are planktotrophic, that is,
they depend on feeding in the plankton, whereas
others are lecithotrophic, that is, they contain suf-
ficient yolk for the whole planktonic development
until metamorphosis. However, a few species are
facultative planktotrophs—they may develop
through metamorphosis to juveniles without feed-
ing, but if food is available they feed and grow to a
larger size before metamorphosis, for example, the
echinoid Brisaster latifrons (Hart, 1996).
It appears that lecithotrophy has evolved in many
smaller or larger lineages, for example, many times
within the gastropod family Littorinidae (Reid,
1989), and larger groups as cephalopods (although
this does of course depend on the interpretation of
mollusc phylogeny). There is no well-documented
example of an evolution in the opposite direction.
The re-evolution of planktotrophy in some littorinid
lineages and in Crepidula is a simple “reactivation”
of the ciliated velum present in species with intraca-
psular development (Collin et al., 2007; Nielsen,
2013). Independent evolution of lecithotrophy has
been directly demonstrated by the rich fossil record
of spatangoid echinoids, where lecithotrophy has
evolved independently in five lineages (Cunning-
ham and Abt, 2009).
Another illuminating example is the rare type
of development called poecilogony (Knott and
McHugh, 2012), where a species shows two types of
development, either in different periods of the year
or even within the same egg capsule. The annelid Py-
gospio elegans deposits egg masses in their tube, and
each egg cocoon may contain embryos of the usual
feeding trochophora type and almost globular, yolky
embryos (Rasmussen, 1973) (Figure 1.4). Also, the
egg capsules of the gastropod Calyptraea lichen may
contain embryos with completely different develop-
mental types, direct-developing embryos which feed
on nurse eggs/embryos and embryos which hatch
as planktotrophic veligers (McDonald et al., 2014).
There seems to be no intermediate embryo types, so
this must be interpreted as a genetic shift similar to
that observed in some ascidians, where tailed, plank-
tonic and anuran (tail-less), benthic larvae are found
in closely related species of ascidians (see earlier). It
must be assumed that in the poecilogenous species,
all the fertilized eggs contain the genetic informa-
tion for making a trochophora larva, but that this
is somehow switched off in the direct-developing
embryos. This is a further indication that the indi-
rect development with planktotrophic larvae is the
ancestral developmental type at least in the bilate-
rians. The gastropod Alderia willowi shows seasonal
changes in developmental type, with smaller eggs
producing planktotrophic larvae being most com-
mon in the summer months and larger eggs pro-
ducing lecithotrophic larvae more common in the
winter; intermediate development occurs but is rare,
and experiments show that the developmental type
is environmentally induced (Krug et al., 2012).
1.4  An Overview of the Diversity
of Marine Larvae
Many animal phyla show examples of both indirect
and direct development, and many groups have
characteristic larvae, which have been given spe-
cial names, in some cases because the larvae could
not be linked with the adults. An example is the
actinotroch, which was described long before the
adult Phoronis was discovered. A large selection of
larvae is pictured in the Atlas of Marine Invertebrate
Larvae (Young, 2002a).
Porifera: The compact, almost completely cili-
ated, lecithotrophic larvae can be difficult to distin-
guish from other small ciliated larvae (Ereskovsky,
2010). Some demosponge larvae show a ring of
longer, photosensitive cilia around the posterior
pole (Figure 1.1A).
Cnidaria: Almost all cnidarians have spherical
to ellipsoidal, ciliated planula larvae, which are
difficult to identify; some of the anthozoan lar-
vae are planktotrophic (Figure 1.1B). Among the

anthozoans some cerianthids have juvenile pelagic
polyps (Figure 1.1C) and some zoanthids have
zoanthina or zoanthella larvae with special cili-
ary bands (Martin and Koss, 2002). In the hydroid
Tubularia, the embryos grow to a polypide-like,
free-swimming stage called actinula (Martin and
Koss, 2002). The scyphozoan polyps bud off stacks
of ephyra “larvae” (Figure 1.1D), which during
growth gradually transform into the adult medusa.
Ctenophora: Adult ctenophores show a wide mor-
phological variation, from spherical types (e.g.,
Pleurobrachia) to blown-up types with large oral lap-
pets, found in both planktonic types (e.g., Bolinop-
sis) and creeping types (e.g., Coeloplana), and even
leathery, sessile types such as Tjalfiella (Mortensen,
1912). However, all types show a characteristic “cy-
dippid” larval stage when hatching (Martindale,
2002) (Figure 1.1E), and all feed on smaller or larger
plankton organisms.
1.4.1  Spiralia (Lophotrochozoa)
Many spiralians have indirect development with
characteristic, ciliated larval types, but direct devel-
opment is characteristic of the phyla Gastrotricha,
Gnathostomulida, Micrognathozoa, and Rotifera
(Nielsen, 2012a).
Annelida: Annelids show a wide variety of devel-
opmental types. Some have filter-feeding, almost
schematic trochophora larvae (Figure 1.5), feeding
with a downstream-collecting ciliary system of pro-
totroch, metatroch, and adoral ciliary zone (Riisgård
et al., 2000), but many are lecithotrophic with only
one or two prominent ciliary bands, prototroch or
telotroch, and some capture larger organisms with
palps or other structures, for example, the larva of
Magelona (Lebour, 2009). A special type is the perica-
lymma larva, in which the “adult” body develops in
a deep invagination. In Polygordius and Eulalia, where
the body below the metatroch develops in the invagi-
nation, the main part of the adult body is covered by
a “serosa.” At metamorphosis, the larval structures,
including the large prototroch, are shed and may be
devoured (Figure 1.2F). In the planktotrophic larva of
the annelid Owenia, the part of the body behind the
first setigerous segment develops in an invagination
(Smart and Von Dassow, 2009). Most of the sipuncu-
lans have pelagosphaera larvae (Figure 1.2C).
O r i g i n a nd D i v e r s i t y o f M a r i ne L a r va e    9
E.R
Figure 1.5 Poecilogeny. Egg capsule of the polychaete Polydora
ciliata containing a fully differentiated trochophora larva and three
bumpy lecithotrophic embryos (from Rasmussen, 1973).
Mollusca: Solenogasters, caudofoveates, poly-
placophorans, and schaphopods have lecitho-
trophic larvae with only the prototroch, for
example, in polyplacophorans, (Figure 1.2G)
or prototroch and telotroch, for example, in so-
lenogasters (Okusu, 2002). Cephalopods have
direct development. Gastropods have plank-
totrophic or lecithotrophic veliger larvae, but
some of the lecithotrophic veligers lose the ve-
lum before hatching from the egg capsule. Most
of the planktotrophic veligers show a velum
with small rounded or oval lobes, but a number
of species have the velum extended into long
lappets (Figure 1.2H–J). The veligers are ciliary
feeders using a downstream-collecting system
(Riisgård et al., 2000). Protobranch bivalves have
pericalymma larvae, where the huge prototroch
cells cover the remaining parts of the larval body
(Zardus and Morse, 1998), whereas almost all
autobranchs have veliger larvae, usually with a
small roundish or bilobed velum (Figure 1.2K,L).
Nemertea: Many nemertines have lecithotrophic
development (Figure 1.1H), but a number of the
pilidiophorans have indirect development with
the characteristic planktotrophic pilidium larvae
(Figure 1.1G). Feeding by the pilidia of the species
Micrura alaskensis was described by von Dassow
et al., (2013). Their observations revealed a unique
feeding strategy consisting of a multi-stage process
composed of ciliary-driven water movement and
behavioral mechanisms.
10   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v e rt e b r at e L a r va e
Platyhelminthes: Most of the species have direct de-
velopment, but some polyclads have planktotrophic
larvae with a number of ciliated lappets, called ei-
ther Müller’s or Goette’s larvae (Figure 1.1F). The
parasitic groups show various types of ciliated, lec-
ithotrophic larvae, for example, the oncomiracidium
of monogenes; the cestodes have complicated life
cycles with ciliated miracidia and unciliated cercaria
(Smith et al., 2002).
Bryozoa: The planktotrophic cyphonautes larva
(Figure 1.1K) is found in basal eurystome groups,
and lecithotrophic larvae of various types are found
in the remaining groups (Figure 1.1L). The cypho-
nautes larvae are ciliary filter feeders with the cilia
situated on a “ciliated ridge” with ciliary bands like
one side of an adult bryozoan tentacle (Nielsen,
2002). The filtering mechanism is of the ciliary-
sievin­g type (Strathmann, 2006).
Entoprocta: All entoprocts develop through a tro-
chophora stage, which may be planktotrophic or
lecithotrophic (Nielsen, 1971).
Phoronida: All phoronids but one develop through
the characteristic planktotrophic actinotrocha stage
(Figure 1.1M). Its ciliated tentacles are similar to
those of the adults; their feeding mechanism is cili-
ary sieving (Temereva and Malakhov, 2010).
Brachiopoda: All brachiopods have ciliated
larvae, which are planktotrophic in the lingu-
liforms (Figure 1.1J) and lecithotrophic in rhyn-
chonelliforms and craniiforms (Figure 1.1I). The
planktotrophic larvae are filter feeding, using a
ciliary-sieving system (Strathmann, 2005).
1.4.2 Ecdysozoa
All ecdysozoans larvae are characterized as second-
ary larvae because they lack ciliated epithelia (Had-
field et al., 2001).
Arthropoda: Only two of the chelicerate groups have
remarkable marine larvae: the pycnogonids have
brood protection where the males carry the brood un-
til a fairly advanced juvenile stage. The early stages
resemble crustacean nauplii, but the first pair of ap-
pendages are cheliceres (Figure 1.6A). The xipho-
surans deposit their eggs in the intertidal and the
larvae hatch as so-called trilobite larvae (Figure 1.6B).
Crustacea: Some crustacean groups, such as am-
phipods and isopods, have direct development, but
most groups have larvae, and the nauplius larva
(Figure 1.6C,D) is now generally regarded as the
ancestral larval type of the crustaceans (Martin et
al., 2015b). Nauplii of some copepods create small
water currents and intercept food particles indi-
vidually (Bruno et al., 2012). Larger larvae show a
wide variety of feeding modes, from filter feeding
to carnivory (Anger, 2001). Most free-living cope-
pods go through a gradual development with ad-
dition of segments, but many other groups show
characteristic larval forms, which have been given
special names (Figure 1.6E–J). A series of larval
stages is seen, for example, in cirripedes (Figure
1.6D,E) and in many decapods (Figure 1.6G,H).
Many types show spines on various parts of the
body, which must protect against predation (Figure
1.6G–J). Also the extremely expanded, thin body
of the phyllosoma larvae (Figure 1.6F) must be of
protective value. A detailed treatment of crustacean
larvae is found in Martin et al. (2015a).
Loricifera: The tiny loriciferans have a meioben-
thic larval type called the Higgins larva; its feeding
biology is unknown (Kristensen, 1991).
Priapula: The benthic priapulan larvae resemble
the adults, but have a ring of longitudinal plates at
the body; it is probably carnivorous like the adults
(Higgins et al., 1993).
1.4.3 Ambulacraria
Enteropneusta: Indirect development with the char-
acteristic planktotrophic tornaria larva with an an-
terior convoluted ciliary band used in feeding and
a large posterior band of compound cilia used in lo-
comotion is found in genera such as Ptychodera and
Schizocardium (Figure 1.7A,B). The anterior ciliary
band is used in particle collection, probably using a
ciliary reversal method like that of the echinoderm
larvae. Lecithotrophic larvae with only the poste-
rior band are found, for example, in Saccoglossus.
The planktotrophic larvae go through a series of
stages, which have been given special names, and
the last stage before settling resembles that of the
lecithotrophic larvae (Figure 1.7C).
Echinodermata: All described crinoid larvae are
lecithotrophic, but the four other classes show
much variation, from planktotrophic larvae to com-
pletely featureless lecithotrophic “shmoo” larvae
 O r i g i n a nd D i v e r s i t y o f M a r i ne L a r va e    11

(A) (B) (C)

(D) (E) (F)

(G) (H) (I)

(J)

Figure 1.6  Arthropod larvae: (A,B) Chelicerates, (C–K) Crustaceans. (A) First instar larva of the pycnogonid Achelia assimilis (courtesy of Tobias
Lehman, Bavarian State Collection of Zoology, Munich). (B) Trilobite larva of the xiphosuran Tachypleus tridentatus (courtesy of Arild Hagen,
Slaastad, Norway). (C) Nauplius larva of the copepod Macrocyclops (courtesy of Anne Bruce, Lancaster, UK). (D,E) Nauplius larva of the barnacle
Elminius modestus and cypris larva of an unidentified barnacle (courtesy of Otto Larink, TU-Braunschweig). (F) Phyllosoma larva of the rock lobster
Sagmariasus verreauxi (courtesy of Alan Blacklock, NIWA). (G,H) Zoea and megalopa larvae of a crab (courtesy of Guillermo Guerao, Barcelona,
Spain). (I) Pseudozoea larva of the stomatopod Squilla mantis (© Jean Lecomte). (J) Zoea larva of a porcellanid crab (courtesy of Christian Sardet,
Observatoire Océanologique, Villefranche-sur-Mer) (see Plate 3).

(Figure 1.7F). The different larval types are clearly larvae (Figur­e 1.7H). The early bipinnaria type as-
variations over the supposedly ancestral larval stage, teroid larva develops shorter or longer arms and
the dipleurula, which can be recognized in early de- becomes a brachiolaria (Figure 1.7D), and a juvenile
velopmental stages in most of the planktotrophic rudiment differentiates on the left side of the body
12   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v e rt e b r at e L a r va e

(A) (B) (C)

(D) (E)

(F)

(G) (H)

(I) (J)

Figure 1.7 Ambulacrarian larvae. (A–C) Enteropneusts, (D–J) Echinoderms. (A) Tornaria larva of Schizocardium sp. (courtesy of Paul Gonzalez, Stanford
Univ.). (B) Tornaria larva with a most complicated ciliary band extended onto small tentacles (courtesy of Russell R. Hopcroft, Univ. Alaska, Fairbanks).
(C) Metamorphosing larva of Schizocardium (courtesy of Poul Gonzales, Stanford Univ.). (D) Brachiolaria larva of the asteroid Pisaster ochraceus (Univ.
Saskatchewan Archives & Special Collections, TC Lacalli fonds, MG 210). (E) Echinopluteus larva of the echinoid Strongylocentrotus franciscanus (Univ.
Saskatchewan Archives & Special Collections, TC Lacalli fonds, MG 210). (F) Lecithotrophic larva of the asteroid Solaster stimpsonii (courtesy of Jochanan
Aronowicz, Stanford Univ.). (G) Metamorphosing larva of the asteroid Asterias rubens; the larval body is being resorbed (© Dr Richard Kirby, reproduced
with permission). (H) Early bipinnaria larva of an asteroid (© Wim von Egmond). (I) Ophiopluteus larva of the ophiuroid Ophiura albida (© Wim von
Egmond). (J) The ophiuroid Ophiopholis aculeata: fully differentiated ophiopluteus (above) and late metamorphosis stage showing the small juvenile at the
center (below) (courtesy of Richard R. Emlet, Univ. Oregon) (see Plate 4).

(Figure 1.7G); the larval body becomes resorbed.
The planktotrophic pluteus larvae of echinoids
(Figur­e 1.7E) and ophiuroids (Figure 1.7I,J) have
long ciliated arms with delicate calcareous skele-
tons. Most holothurians have lecithotrophic larvae,
but a few groups have planktotrophic larvae, which
first resemble asteroid bipinnaria, but their ciliary
band breaks up and becomes reorganized into five
rings like those of the crinoid larvae, a larval type
called doliolaria (Sewell and McEuen, 2002). The cil-
iary bands are used in locomotion and feeding. The
feeding mechanism is an upstream-collecting sys-
tem that involves reversal of individual cilia “push-
ing” particles to the oral field (Strathmann, 2007).
Pterobranchia: Only lecithotrophic larvae have
been reported (Lester, 1988).
1.4.4 Chordata
Urochordata: Most urochordates have the character-
istic lecithotrophic “tadpole larva” with the noto-
chord and neural tube in the tail, which becomes
resorbed after settling (Figure 1.8A,B).
Cephalochordata: Amphioxus has pelagic larvae,
which gradually attain the adult morphology
(Figure 1.8C).
(A)
(B)
(C)
Figure 1.8  Chordate larvae: (A) larva of the solitary ascidian Ciona
intestinalis (courtesy of Yasunori Sasakura, Tsukuba Univ.). (B) Larva of
the compound ascidian Botrylloides violaceus (© Lin Baldock). (C) Late
larva of Branchiostoma japonicum (courtesy of Kinya Yasui, Hiroshima
Univ.) (see Plate 5).
O r i g i n a nd D i v e r s i t y o f M a r i ne L a r va e    13
1.5 Summary
The myriad of planktonic larvae shows all sorts
of adaptations not least to a life in the free water
masses. Some clades, such as protobranch bivalves
and linguliform brachiopods, show very little vari-
ation in developmental type. Other clades show
enormous variations, even between closely related
species, such as the echinoids Heliocidaris tuberculata
with a planktotrophic echinopluteus larva and H.
erythrogramma with a lecithotrophic “shmoo” larva
without gut and larval arms (Laegdsgaard et al.,
1991), and the occurrence of tailed and anuran (tail-
less) larvae in closely related species of the ascidian
genus Molgula (Jeffery et al., 1999). In these cases,
the adults resemble each other very much, but their
larvae are profoundly different.
The study of marine larvae, their shapes, feeding,
reactions to biological and non-biological stimuli,
adaptations to various types/availability of food
and to predators, etc., is fascinating in itself, but it
also throws light on many evolutionary, ecological,
and physiological principles. It should be remem-
bered that although marine larvae have been stud-
ied for centuries, there are still important gaps in
our knowledge of almost all aspects. Many species
are still undescribed, and the development of many
of the known species is still unknown.
So, there should be plenty of interesting study
subjects for any type of scientific inclination.
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 Ch a pt er 2
Evolutionary Development
of Marine Larvae
Heather Marlow
2.1 Introduction
Embryology and larval biology have been linked
from the early years. Pioneering embryologists
were among the first to establish and populate
marine biology field stations with the aim to ac-
cess a wide diversity of early animal stages. Their
descriptions of developmental programs came at a
critical juncture in our understanding of selective
mechanism and evolutionary theory, leading to
some of first theories of body plan evolution (Hall,
2000; Horder, 2006). Organisms with distinct larval
life history stages such as molluscs as well as non-
vertebrate deuterostomes served as popular study
animals (Conklin, 1897; Goodrich, 1903), with some
models, such as the sea urchin and tunicate, contin-
uing to serve as valuable models for gene regulation
(Davidso­n et al., 2002; Bertrand et al., 2003). A focus
on the establishment of systems appropriate for
functional studies in the early days of genetic tech-
niques later pushed investigators inland to research
centers and led to the establishment of laboratory
models for early development such as Drosophila
melanogaster and Caenorhabditis elegans, thus shifting
focus away from marine species and their charac-
teristic larval forms. New breakthroughs in high-
throughput sequencing technology and genetic
interference have removed many of the technical
barriers to the study of marine organisms (Cong
et al., 2013; Doudna and Charpentier, 2014) and are
pushing new generations of researchers to examine
the diversity of development from a molecular and
mechanistic perspective (see Williams and Carrier,
this volume). These emerging studies inform not
Marlow, H., Evolutionary development of marine larvae. In. Evolutionary Ecology of Marine Invertebrate Larvae.
Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018).
© Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0002
only our understanding of developmental diversity
but also help us tackle classical questions regarding
the origin and diversification of larval forms.
Among the earliest pursuits of marine embryolo-
gists was to uncover the origins of marine larvae, to
determine how often such forms have arisen (once
or at multiple points independently on the animal
tree), and what relationship they share with adult
body plans. Such endeavors to assign homology to
entire life history stages are challenging, as discrete
features, including ciliary or sensory structures, ex-
cretory cells, and even the gut, can evolve in one
stage and shift by heterochrony to another—in large
part driven by environmental and perhaps genetic
factors (Raff et al., 1990; McDougall et al., 2006).
Genomic studies have revealed that most animal­s in
the clade encompassing the Cnidaria and Bilateria
use very similar gene repertoires to build this vast
morphological array of structures, necessitating that
individual genes are used and reused in seemingly
unrelated structures (Glassford et al., 2015). Adding
further to the problem, the expression of batteries
of genes is often linked, and the expression of any
single gene in such a battery cannot be treated as an
independent character without prior knowledge of
its function in development. The challenge for evo-
lutionary-developmental biology in the molecular
era lies in determining if these disparate approaches
to the study of larval developmental features and
cell types, modular genetic landscapes, and charac-
ter co-option can be integrated and if a signal for the
origin of life history transitions will emerge. Were
the first animals direct developers (either holope-
lagic or holobenthic) or were they benthic animals

that dispersed their developmental stages via the
plankton from the very start?
The marine larva represents an interesting devel-
opmental paradox. It is clearly an embryonic stage
which will continue to form new cell types and struc-
tures and organs, and in many cases, an entirely new
body axis. However, it is also a functional organism
which locomotes, may feed, and is capable of coordi-
nating a sensory response. While this juxtaposition
of functional sensory, feeding, and locomotory ma-
chinery on top of an early embryonic form is rarely
seen in terrestrial systems, it is commonplace in the
marine ecosystem, and as life evolved in the sea, it is
likely that such a dichotomy of life history was pre-
sent from the very beginnings of animal evolution.
2.2  Homologous Larval Features
A larva can be most broadly defined as an interme-
diate developmental stage between the embryonic
and adult stages. For evolutionary studies, several
more specific definitions have been proposed to ac-
count for the wide diversity of larvae and the per-
spective of the researcher in considering this stage
Box 2.1  Terminology utilized in the discussion of life history strategy evolution. Such terms
are often nonexclusive and many larval forms or life history strategies can be assigned to
multiple categories.
Term Definition
Primary Larva Ciliated, swimming larva common to
marine organisms
Secondary Larva Independently acquired larval form
through intercalation (often of a
modified adult form)
Planktotrophic A feeding larva
Lecithotrophic A non-feeding larva
Maximally Indirect Bi-phasic life history in which little
Development of the larval body plan is re-used in
formation of the
Minimally Indirect A larva which undergoes a minimum
Development amount of tissue remodeling during
Catastrophic Laraval form that undergoes dramatic
Metamorphosis body plan modification during
Direct A life history in which no larval form is
Development present
E v o l u t i o n a ry D e v e l o p me n t o f M a r i n e L a r va e    17
(Box 2.1). One of the most formidable challenges
has been the comparison and study of “primary
larvae” in Bilateria, which are also the focus of this
chapter and the discussions which follow (also see
Nielsen, this volume). Primary larvae here will be
considered those with a ciliated apical (anterior)
epithelium that often possesses an apical organ of
long tufted sensory cilia. The primary body axis of
the larva is that of an apical (anterior)-oral (poste-
rior) poles, and the body plan in this context refers
to the position of the blastopore, mouth, and apical
organ in relation to the primary axis of symmetry—
in many lineages additional features such as ciliary
swimming bands, eyes, kidneys, chaetae, or muscu-
lature may be present but are not essential features
of the primary larval body plan.
Due to the widespread distribution of primary
larvae among bilaterians, it has been proposed
that primary larvae are an ancient metazoan fea-
ture to facilitate the widespread distribution of
benthic adult forms via a planktonic intermediary
stage. It has been argued that this dispersal stage
can be traced to the root of the bilaterian animals.
By contrast, a novel, lineage-specific larval form
Origin Examples (Non-Exhaustive)
Jägersten, 1972 cnidarians, sea urchins, annelids, hemichordates,
Mollusks, nemerteans
Peterson et.al, 1997 insects, vertebrates (aneurans)
sea urchins, molluscs
annelids
Davidson et.al, 1995 phoronids, sea urchins, nemerteans, ascidians
Marlow, et.al, 2014 cnidarians, amphioxus, hemichordates
Emig, 2001 nemerteans, sea urchins
sea urchins,vertebrates (mammals)
18   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
is referred to as a “secondary larva” and can be
clearly recognized in ecdysozoan groups such as
crustaceans. It is often the case, however, that the
origin of a particular larval form may be more am-
biguous. Diversity of larval forms and the gradi-
ent, which often exists in defining the larval vs.
adult stage, as is the case for animals which un-
dergo a gradual metamorphosis, pose a challenge
to determining if modern larvae can indeed be
traced to a single emergence.
2.3  History of the Debate
for Ancestrality of Larval and Adult
Forms
Two opposing viewpoints, and several intermedi-
ate variations, dominate the debate regarding the
appearance of metazoan larval forms (Figure 2.1).
A larva-centric theory proposed by Nielsen (see
Nielsen chapter in this volume), and anchored in
morphological affinity of diverse larval forms,
holds that the first metazoans were holopelagic
larva-like forms that fed and reproduced in the
plankton (Nielsen, 1985). This theory expands upon
the initial observation made by Ernst Haeckel that
larval forms likely originated from an ancestral
swimming gastrula-like form and that adult stages
arose from this gastrea (Haeckel, 1874). Importantly,
the trochaea theory makes specific homology state-
ments regarding the apical organ and blastopore of
extant metazoans as well as larval feeding systems,
and sets the stage for recent molecular work com-
paring these structures (Nielsen, 2013). This theory
is consistent with geochemical data which suggests
that low oxygen conditions limited animal body
size, complexity (Sperling et al., 2013 and Raff and
Raff, 1970). A second, diametrically opposed the-
ory recognizing the similarity of adult body plans,
suggests that the first animals were adult benthic
forms. This theory, proposed by Raff, suggests that
the larval forms are convergent stages added to the
life cycle via intercalation (Raff, 2008). Under this
scenario, adult bilaterian body plans are homolo-
gous and larvae have been derived independently
in echinoderms, hemichordates, lophotrochozoans,
and ecdysozoans. No distinction is made between
primary and secondary larvae, as all forms would
be considered secondary to the evolution of the an-
cestral, shared bilaterian body plan.
A third scenario in which the larval and adult
forms existed at the base of metazoans, serves as an
intermediate between the larval and adult-centric
views of body plan and life history evolution. In this
scenario, first proposed by Jagerston (1972) and later
supported by Rieger (1994), both larval and adult
body plans existed at the base of bilaterian animals.
In this view, the ancestral larval form would have
been a nonfeeding form that could have been elabo-
rated subsequently in different bilaterian lineages
(Haszprunar et al., 1995). If similarities which exist
in both larval and adult forms are considered ho-
mologous, it can be considered that these two body
plans and, by extension, the biphasic life cycle, are
ancestral metazoan or bilaterian features. For the
ancestrality of the biphasic life cycle to be examined,
distinct homologies for adult and larval body plans
must be identified and the process of metamorpho-
sis among lineages should be compared (see later).
2.4  Molecular Approaches to Comparing
Larval Features
As the number of genomes and transcriptomes
available for study has rapidly increased in the
last decade (Dunn and Ryan, 2015), evolutionary-
developmenta­ l biologists have sought to utilize
these new “molecular characters” as characters
when comparing larval tissues and cell types
­(Santagata et al., 2012; Marlow et al., 2014; Hiebert
and M­ aslakova, 2015). Often this involves attempts
to define a particular tissue by the expression of a
single molecular marker, such as a transcription
factor or effector molecule. An analogous approach
has been utilized with great success to determine
the relationship between species and solve the
phylogenetic relationships among animal groups
(Dunn et al., 2008; Rouse et al., 2016).
The ever-growing number of sequenced genomes
available for analysis now roughly covers the main
branches of the animal phylogenetic tree: we now
have draft assemblies from both vertebrate and
invertebrate deuterostomes, protostome models
including lophotrochozoans (annelids, molluscs,
and rotifers included), non-insect ecdysozoans (pri-
apulids, crustaceans, and chelicerates) and all of the
early branching non-bilaterian lineages (cnidarians,
ctenophores, sponges, and placozoans) (Dunn and
Ryan, 2015). While many additional genome studies
 E v o l u t i o n a ry D e v e l o p me n t o f M a r i n e L a r va e    19

Cnidarian Lophotrochozoan Deuterostome Cnidarian Lophotrochozoan Deuterostome

pelagic benthic pelagic benthic pelagic benthic pelagic benthic pelagic benthic pelagic benthic

ancestor pelagic benthic ancestor

pelagic

(A) (B)

Cnidarian Lophotrochozoan Deuterostome

pelagic benthic pelagic benthic pelagic benthic

ancestor
benthic

(C)

Figure 2.1 The three scenarios show the origin of the pelagic larval body plan. The presence of pelagic forms is indicated by black lines and that
of benthic forms by gray lines. A single-colored line indicates a monophasic life cycle that would be pelagic in scenario B and benthic in scenario C.
Double lines (black and gray) indicate a biphasic, pelago-benthic life cycle (with pelagic larval and benthic adult forms). Note that the biphasic life cycle
is assumed to have evolved multiple times independently in scenario C. (A) The classical view implies homology of both ciliated larvae and benthic
adults that, once evolved, have remained part of the eumetazoan life cycle (Jagersten, 1972). (B) Nielsen (2012) modified this view to propose that
the holopelagic neuralian ancestors persisted beyond the initial divergence of the major neuralian clades, and that the biphasic life cycle with benthic
adults arose independently in the cnidarians and once or twice in the bilaterians. (C) In stark contrast, other authors assume that today’s ciliated larvae
arose convergently many times by the repeated intercalation of a pelagic dispersal larva into primarily monophasic, holobenthic life cycles and are thus
evolutionarily unrelated. This view implies that the characteristics of today’s swimming larvae such as apical organs and equatorial ciliary bands evolved
convergently (Raff, 2008). Figure and legend taken from Marlow et al. (2014).

will likely fill in the remaining gaps, these data have and allowed us to plot broad patterns of gene gain
served as the basis for a number of phylogenetic and loss, genome reorganization, duplications and
studies which have convincingly structured the core conservation of synteny (gene order) (Dunn et al.,
branching order for the divergence of key lineages 2008; 2014). Clear patterns of “ancestral linkage
20   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
groups” indicate that there has been conservation
of gene order within the linear genome sequence,
but that this order is only retained in some lineages
while it has been lost in others (Irimia et al., 2012;
Simakov et al., 2013; 2015). A similar pattern can be
observed for the internal structure of genes. While
the scale of gene loci can vary drastically between
lineages, the structure of key features such as exon-
intron boundaries have been stable in some lineages
and can be traced to the last common ancestor of
Cnidaria and Bilateria (Raible et al., 2005; Putnam
et al., 2007). Despite the significant variations in ge-
nome structure and size, the protein-coding gene
content is surprisingly similar among bilaterians,
and, to a large extent, this gene content is shared
with the cnidarians, while other basal metazoan lin-
eages such as sponges and ctenophores show con-
siderably less similarity to bilaterian protein-coding
genes (Putnam et al., 2007; Ryan et al., 2013). Ad-
ditional genome sequencing projects will be needed
to determine whether this represents a secondary
loss of ancient genome features or whether these
lineages predate the origin of the genomic features
shared among Cnidaria and Bilateria.
While the construction of species trees has pro-
vided great insight into the evolution of animal
body plans and life histories, considerably more
caution must be taken when using molecular char-
acters to assess the origin of new phenotypic traits.
Limitations to the molecular character approach
are highlighted by key differences in the analysis
of species trees and phenotypic character trees (e.g.
cell type trees). Specifically, genes are rarely passed
between lineages in a species tree (later gene trans-
fer), but are often co-opted between cell types in a
cell-type tree (co-option of genetic modules).
2.5  Distinguishing Larval Features
and Adult Features: Body Plan
and Innovations in Gene Regulation
Three defining features characterize what is gener-
ally accepted to represent the bilaterian body plan,
and by extension, the body plan of the last common
bilaterian ancestor (LCBA): (1) a neuralized anterior
region—the degree of centralization and structure
of an anterior brain of this ancestral form remains
debated, (2) an extended hox-patterned trunk re-
gion, and (3) a tripartite through-gut. Determining
how these hallmarks are distributed in extant adult
and larval forms has helped distinguish whether
the LCBA body plan was more similar to a pelagic
larva, a benthic adult, or perhaps was represented
by an ancestral biphasic life cycle. Molecular stud-
ies over the last 30 years have contributed signifi-
cantly to our understanding of body plan evolution
by providing a vastly expanded set of genetic
characters which can be used to test hypotheses re-
garding the homology of features of the bilaterian
body plan. While many open questions remain, it is
now rather clear that the ciliated swimming larvae
and benthic muscular body plans originate from a
biphasic ancestral life history that occurred at the
base of the Bilateria.
Among the essential functions of the LCBA body
plan were to feed and locomote, but tracing the ori-
gin of feeding stages has been particularly difficult
when considering the evolution of primary larval
forms considering the number of feeding transitions
and reversals which have occurred within phyla—
the echinoderms are a particularly illustrative ex-
ample of this challenge (Strathmann, 1985; Rouse,
2000). There is, however, a stronger phylogenetic
signal for the origins of locomotory mechanisms
among adults and larvae. The advent of locomo-
tory mechanisms likely drove the evolution of the
three key bilaterian features, through-gut, trunk,
and anterior neuralization. In general, primary
larval forms use ciliary beating in response to en-
vironmental sensory stimuli and passive transport
as their primary mode of locomotion (Jékely et al.,
2008; Jékely, 2011). In contrast, despite the tremen-
dous variation in locomotory modes (e.g., burrow-
ing vs. undulatory movement) in benthic bilaterian
animals, nearly all of these modes rely primarily
upon muscle-based mechanisms (Clark, 1981).
In larval forms, the anterior neurogenic ecto-
derm often termed the “apical plate” or “larval epi-
sphere” coordinates the locomotory behavior and
is characterized by the expression of what has been
termed the “head patterning network” (Lemon­ s
et al., 2010; Sinigaglia et al., 2013; Arendt et al.,
2016b). Considerable recent progress has begun to
uncover the critical role of the larval apical plate in
ciliary swimming, settlement, and modulation of
 E v o l u t i o n a ry D e v e l o p me n t o f M a r i n e L a r va e    21
behavior via neurohormones. Members of the genetic
network that patterns the larval apical plate have been
described in conserved, nested expression domains in
both protostome and deuterostome larvae, strongly
supporting that the anterior ectoderm and its most
prominent sensory structure, the apical tuft, were
present in the LCBA (Marlow et al., 2014; Sinigagli­a
et al., 2015). However, this network and the transcrip-
tion factors and signaling molecules that comprise it
are not restricted solely to larvae and a multitude of
direct-developing groups—those with secondary lar-
vae also employ this network for sensory organ and
brain development (Lemons et al., 2010, 10.1016/j.
cub.2016.10.047), with a study in direct-developing
centipedes highlighting the role of these factors in
the development of an ancient discrete brain neuro-
hormone center that was likely lost in many insects
(Hunnekuh­l and Akam, 2014). The widespread dis-
tribution of this network in marine larvae argues in
favor of an ancestral larval body plan (Marlow et al.,
2014); its integration into protostome and deuteros-
tome adult brains presents the intriguing possibility
that the anterior brain may have had its roots in larval
sensory cell types (Tosches and Arendt, 2013).
While larval-specific characters are exemplified
by the sensory epithelium and apical organ, adult
features are associated with the development of the
trunk, its musculature and innervating neural com-
ponents (Shankland and Bruce, 1998; Shankland
and Seaver, 2000; Arendt et al., 2016b). Develop-
ment of the trunk is linked to the development of the
mesoderm, the formation of the ventral nerve cords
of protostomes, when present, and the formation of
the coelomic cavities, structures which may form
in larvae but persist into the adult stage (Arendt et
al., 2015). Its formation is concurrent with the settle-
ment process in marine organisms in which a bipha-
sic life cycle is present (Gonzalez et al., 2016; Hejnol
and Vellutini, 2016). The developing trunk tissues
across adult bilaterian animals express the hox fac-
tors, homeodomain transcription factors famous for
their role in body plan patterning, with numerous
functional roles in development (Kulakova et al.,
2007). A unifying theme in their expression is the
nested anterior-posterior progression of cluster ex-
pression along the main body (Kmita and Duboule,
2003). It has been strongly argued that the elabora-
tion of the hox genes has driven the elaboration of
animal body plans by facilitating the development
of the musculature and neurons (hallmarks of the
bilaterian trunk) (Holland, 2015). A second clus-
ter of colinear antp-class homeodomain genes, the
parahox genes, arose through duplication of the
ancestral hox-like genes and, in association with
the winged-helix foxA and GATA transcription
factors, has evolved to pattern both the larval and
adult tripartite gut (Fröbius and Seaver, 2006; Boyle
and Seaver, 2008; Annunziata and Arnone, 2014;
Martín-Durán and Hejnol, 2015). Hox and parahox
gene expression, when present in marine larvae,
is most commonly confined to tissues which give
rise to the adult gut and trunk, the larval rudiments
(Arenas-Mena et al., 2000; Hiebert and Maslakova,
2015). While larval metamorphoses can be vastly
different and range from gradual to “catastrophic,”
the result is nearly universal, the reduction of the
anterior ciliated epithelium and the generation via
rapid development of the mesoderm, the adult
musculature. So, in short, a diffusely neuralized
anterior epithelium or episphere which coordinates
ciliated locomotion is a larval feature, while an ex-
tended mesodermally patterned trunk which gives
rise to locomotory musculature represents an adult
feature. It is therefore feasible that hox and parahox
gene diversification accompanied the evolution of
the trunk and through-gut prior to the bilaterian
radiation.
While features of this biphasic life cycle—most
prominently, a neuralized apical plate and apical
organ patterned by the “head patterning network”
(Lemons et al., 2010)—can be recognized in cnidar-
ians and perhaps even earlier branches of the ani-
mal tree, significant work needs to be conducted to
understand the relationship of the adult cnidarian
polyp body plan to that in bilaterians. As the hox
cluster organization in Cnidaria comprises a mini-
mal cluster and clear orthologs of posterior parahox
genes are absent, it is possible that the adult ben-
thic polyp body plan arose prior to the advent of
the benthic body plan shared by bilaterian adult
stages (Chourrout et al., 2006; Ryan et al., 2007).
Conversely, it is possible that the cnidarian adult
polyp body plan has been significantly modified
from the ancestral state, however, such an asser-
tion will remain purely speculative until further
studies can be conducted in additional cnidar-
ian species. Therefore, it can be concluded that a
ciliated swimming dispersal stage was present in
22   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
the last common ancestor of cnidarians and bilate-
rians, and that a biphasic life cycle with a ciliated
larvae bearing significant similarity to that found
in Cnidaria was present at the base of the Bilateria.
2.6  Comparison of Cell Types
The use of molecular characters in assessing homol-
ogy has significantly informed our understanding
of the evolution of the body plan, and these same
molecular characters in conjunction with new and
highly sensitive sequencing technologies are set to
re-enliven the classical pursuit of tracing cell-type
homologies. Traditionally, photoreceptors, osmoreg-
ulatory structures, ciliated cells, muscle cells, and
neurons have been identified and compared based
upon subcellular structures via differential stain-
ings, later via EM microscopy, and now through
the generation of spatial in situ hybridization data,
wherein labeled riboprobes are used to detect gene
expression through hybridization to endogenous
RNA molecules (Arendt et al., 2004; Martindale
et al., 2004), and increasingly through single-cell
RNAseq technologies. This approach follows the
same logic wherein cell morphologies (e.g., micro-
tubule organization, vesicle presence or structure.
etc.) have been used as characters (Bartolomaeus
and Ax, 1992; Purschke et al., 2006). The result is a
gene expression signature which is then taken as a
phenotypic character and compared among species
in an effort to assert or refute homology and used
to build “cell-type trees” in which the origin of cell
types and their diversification in distinct lineages
can be traced within animals (Arendt, 2008).
The number of cells which can be recognized
microscopically in marine organisms, particularly
marine larvae, is extremely limited, leading some
to term the diversity of morphologically nonde-
script, but molecularly distinct, cell types as “cryp-
tic complexity” (Miller and Ball, 2008). RNA in situ
hybridization techniques have provided a valuable
tool in allowing these cells to be identified, but are
still greatly limited by their low-throughput and re-
liance upon the candidate gene approach. Despite
these limitations, this characterization has allowed
for the identification of a complex repertoire of
larval sensory cells, including distinct photorecep-
tor cell types utilizing opsins (Tosches et al., 2014;
Gühmann et al., 2015), the same molecules re-
sponsible for photoreception in the vertebrate eye.
Additional studies have uncovered ancient neuro-
secretory cells of the larval apical plate as well as
neuropeptide-expressing cells involved in settle-
ment and metamorphosis (Tessmar-Raible et al.,
2007; Conzelmann et al., 2013b).
The reliance on the candidate gene approach in
particular has likely led to an overestimation of the
number of homologous cell types shared among
bilaterians, and, as genome-wide sequencing tech-
nologies have advanced, a clearer picture of the
startling diversity of animal cell types is beginning
to emerge. Strikingly, of the sequenced marine in-
vertebrate genomes characterized, most show in-
credible diversity in neural molecule complexity,
much of which is independently derived in these
lineages—hinting that much of the cell-type diver-
sity of marine organisms may be independently
evolved (Moroz et al., 2014). Examples include
heavily diversified ion channels (Li et al., 2015;
Mora­n et al., 2015) and neuropeptides (Shapiro et
al., 2013), indicating that while many neural fea-
tures can be traced to origins that predate Bilateria,
an incredible amount of subsequent diversification
has occurred in animal lineages. The independent
elaboration of metazoan nervous systems, while
striking, is not entirely unexpected given the highly
specific sensory stimuli and environmental con-
ditions which pose strong selective pressure for
adaptive change. Such a diversity in sensory cells
explains how larval forms navigate the vast pelagic
environment and ultimately identify a minuscule,
and highly specific micro-niche that is appropriate
for settlement and initiation of the benthic stage of
their life cycle.
While technical barriers have limited the number
of cells which can be investigated using molecular
data, this field is poised to advance dramatically
in the coming years with the advent of single-cell
sequencing technologies which allow for genome-
wide profiling of single cells in high throughput
and without reliance upon the candidate gene ap-
proach (Shapiro et al., 2013). This field is currently
uncovering previously unrecognized cellular diver-
sity even in well-studied organ systems such as the
retina and spleen (Jaitin et al., 2014; Shekhar et al.,
2016), and will be instrumental in describing the
 E v o l u t i o n a ry D e v e l o p me n t o f M a r i n e L a r va e    23
full complement of cells in marine larvae. Highly
specialized ciliary swimming band cells in proto-
stomes and deuterostomes as well as osmoregu-
latory structures and the tufted cells of the apical
organ await detailed molecular characterization.
2.7  Comparison of Global Transcriptional
Signals—Assessing Homology
of Ontogenetic Process
Examination of molecular data associated with
distinct cell types has driven evolutionary-
developmenta­l studies to a reductionist level, and
allowed for the identification of cell types or func-
tions in larval forms and the comparison of these
cell types between species. A complementary ap-
proach in which global transcriptional signatures
are compared at the phylum or species level have
also been used to generate evolutionary hypoth-
eses regarding the evolution of animal body plans
(Kalink­a et al., 2010; Levin et al., 2016).
Comparison of global developmental programs,
like the examination of morphological cell types, is a
classical pursuit most famously popularized by the
German embryologist Ernst Haeckel, in which he
ascertained that, within vertebrates, a stage in which
a common “phylotypic stage”—a developmental
stage in which intermediate developmental stages
most closely resembled the presumed ancestral form
for that lineage—could be identified (Haeckel, 1866).
This concept was later revisited by Duboule, who ar-
gued that embryogenesis resembled an hourglass in
which distinct developmental processes may be con-
strained in order to give rise to critical morphologi-
cal phenotypes characteristic of particular phyla, in
that particular case, the role of hox genes in anterior-
posterior patterning and growth (Duboule, 1994). In
both instances, the aim of the studies was to charac-
terize and compare developmental programs in or-
der to identify which features of a group of animals
are “ancient” and which developmental programs
are constrained in defining these ancient features.
To test these early comparative frameworks in
relation to animal life histories, contemporary stud-
ies have focused on transcriptomics to pinpoint or
molecularly characterize the “phylotypic stage”
and to argue for the ancestrality of organ systems
and tissues as well as the developmental programs
required for the generation of these phenotypes.
Studies which characterize the age of genes ex-
pressed at a given ontogenetic time point and the
similarity of expression between species within
a phylum appear to support the developmental
hourglass model (Domazet-Lošo and Tautz, 2010;
Kalinka et al., 2010). Put simply, conserved develop-
mental stages express genes which originated early
in metazoan evolution and lineage-specific innova-
tions are characterized by developmental programs
utilizing young genes. Evolution and diversifica-
tion of the minicollagen genes characteristic of
the lineage-specific cnidarian cnidocytes (stinging
cells) is an example of “young genes” in novel cell
types (Beckmann and Özbek, 2012). In vertebrates
and fruit flies, these studies pinpoint the segmen-
tation period and the germ band extension stage
as hotspots of conservation (10.1038/nature09632
and 10.1038/nature09634), heavily arguing for the
ancestrality of adult patterning mechanisms. How-
ever, both insects and vertebrates have lost primary
larvae, making it impossible to assess the conserva-
tion of the larval body plan from this dataset.
The utility of these approaches remain to be as-
sessed for marine larvae, as no study has yet fo-
cused on bentho-pelagic life history transitions
characteristic of marine invertebrates. However, a
recent study assessing ten different phyla did find
evidence for a conserved intaphyletic stage cor-
responding to what the authors interpret as the
phylotypic stage. Interestingly, conservation be-
tween phyla was observed at early stages and late
stages, but diverged significantly at the phylotypic
stage (Levin et al., 2016). Subsequently, it has been
pointed out that such approaches are sensitive to
noise from highly divergent lineages or insuffi-
ciency of sampling (Hejnol and Dunn, 2016; Levin
et al., 2016), and it is conceivable that heterochro-
nic shifts in the appearances of structures or organ
systems, or the elaboration or lengthening of life
history stages would likely pose significant chal-
lenges in the interpretation of such datasets. Such
approaches serve as a strong counterpoint to the
cell-type specific analyses discussed previously as
they attempt to integrate the total developmental
signal across a complete ontogeny rather than dis-
crete cellular units. An understanding of cell-type
24   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e

Minimally Indirect Development: A hypothesis for the

evolution of the Metazoan Biphasic Life Cycle

Conserved Larval Features Patterns of Developmental Development of the Adult

Conservation in Biphasic Life Trunk
Histories
Extension and Growth of the
Patterning of the Larval Axis Trunk Following Settlement
embryogenesis
and Apical Organ Establishment of the
Episphere-Blastopore
six3/6 Axis
(Larval Epithelium Acquisition of
Planktotrophy
Patterning) (Heterochrony)
functional
larval form
Wnt
Precocious
Settlement Development
Neurohormone-Mediated of trunk Settlement via Eversion
Settlement rudiment of Pre-formed Rudiment
(Heterochrony)

metamorphosis
Extension and
Patterning of the
Trunk
(Adult Mesoderm)
reproductive
adult
(A) (B) Feeding (C)

Figure 2.2  Conservation of life history transitions in minimally indirect development. A. Patterning of the larval episphere-blastoporal axis is
dependent on Wnt signaling and the conserved expression of the transcription factor Six3/6 is a common feature in primary marine larvae (Marlow et
al., 2014). The transition from the larval-to-adult transition is mediated by conserved signaling via a GLWamide-like peptide (MIP) in multiple species
of primary marine larvae (Conzelmann et al., 2013b). B. Assuming homology of the biphasic life cycle among marine invertebrates, the larval form,
the adult form, and the metamorphic transition between these forms are homologous. Placing this hypothesis within the hourglass framework would
generate a trimeric pattern of conservation. Heterochronic shifts in the development of adult feeding structures and the adult trunk frequently result
in the precocious development of these structures in larval stages. C. Two examples of the formation of the adult trunk. Extension of the post-ciliary
band region of a hemichordate larva in which gradual extension gives rise to the adult trunk; reproduced from Burdon-Jones (1952). Rapid extrusion
of a pre-formed trunk rudiment in a polychaete larva gives rise to the adult trunk; reproduced from Wilson (1932).

specific gene expression, as well as global develop-
mental programs, is necessary to understand the
evolution of animal developmental programs and
how the output (morphologies generated) of these
programs may be compared—one such hypothesis
is presented here (Figure 2.2).
2.8  Developmental Networks
and the Modularity of Gene Expression:
Integrating Global and Cell-Type Specific
Transcriptional Programs
Utilizing molecular characters to compare mor-
phological features is significantly more chal-
lenging than the construction of species trees for
several reasons: (1) changes in protein-coding gene
complements do not fully account for phenotypic
changes in animals, meaning that new features may
arise without drastically new gene complements;
(2) genes, particularly transcription factors, can be
reused to build incredibly diverse features of the
body plan; (3) genes do not act independently and
equivalently and therefore may face distinct selec-
tive pressures.
To understand how a relatively stable protein-
coding gene repertoire among the extant Bilateria
gives rise to the great diversity of animal body
plans, organ systems, and cell types found therein,
one must have an understanding how of how genes
give rise to specific developmental programs and
resultant phenotypes. Our understanding of the
mechanisms of gene regulation in an increasing
number of model and non-model animal species
has advanced dramatically, providing critical in-
sight into how regulatory programs function, and,
 E v o l u t i o n a ry D e v e l o p me n t o f M a r i n e L a r va e    25
relatedly, how they are then subject to selective
pressure (Schwaiger et al., 2014; Cook et al., 2016).
One framework for understanding the hierarchical
nature of gene regulation during development is
the concept of the “gene regulatory network,” most
notably employed in modeling the development of
sea urchin endomesoderm specification (Davidson
et al., 2002; Davidson, 2010a; 2010b).
In such a framework, genes can belong to early
patterning and specification cassettes and down-
stream effector target genes. Of particular note is
the modular nature of such interactions (Davidson,
2010a). Genes which act in concert to produce a
particular phenotype are often wired “recursively”
to allow for robust maintenance of expression
(McCaule­ y et al., 2010). One observation arising
from such a framework is that individual network
components (e.g., genes) are not independent, and
that the redeployment of a single gene (i.e., tran-
scription factor) can influence and in some cases di-
rect the expression of other interdependent genes.
Therefore, in order to compare molecular features
such as gene expression patterns, one must have
a prior knowledge of the way in which such fac-
tors are linked to other genes, a measure of non-­
independence, and a solid understanding of the
developmental networks in each species compared.
Gene networks are not static entities; significant
modifications occur rapidly, even among closely
related lineages. It has been demonstrated that
such changes in gene regulation are responsible
for change at micro- and macroevolutionary scales
and can even be linked to changes at the level of the
body plan (Davidson and Erwin, 2006; Peter and
Davidson, 2011). Once a useful genetic module has
appeared, such a module may be reused or “rede-
ployed” under different conditions to give rise to
additional structures (Hinman et al., 2003). Clear
examples can be found from the head patterning
network, the retinal determination network, and
that employed in muscle striation (Lemons et al.,
2010; Martik and McClay, 2015; Brunet et al., 2016).
Just as early developmental network compo-
nents can be co-opted to give rise to new struc-
tures, downstream terminal selector transcription
factors and the effector genes they regulate can be
deployed in novel contexts, thus giving rise to new
cell types. Work in C. elegans has demonstrated that
neuron identity is tied to the combinatorial expres-
sion of selector transcription factors and that fate
conversion can occur simply through the dysregu-
lation of a single transcription factor (Gordon and
Hobert, 2015), indicating that effector modules can
be redeployed in new contexts, thus conferring new
functional properties and thereby generating new
cell types. While cell-type identity concepts have
matured to take into account the modularity of gene
expression and the non-independence of the activ-
ity of individual genes, many of these frameworks
still focus on the cell as the unit of selection and con-
sider that cells can be traced back over evolutionary
time by tracing their divergence of function (Achim
and Arendt, 2014; Arendt, 2015).
A recent framework for understanding cell-type
evolution argues that emphasis for cell identity
should be placed on terminal selectors and effec-
tor genes and that less emphasis should be placed
on the upstream regulatory cascade. However,
this framework still specifies that cells evolve via
a bifurcation process much akin to speciation in
which “sister cell types” are produced (Arendt et
al., 2016a); even so, this concept risks potentially ob-
scuring cases in which genetic modules can be as-
sembled entirely de novo. Nevertheless, it must still
be taken into account that cell state is not equivalent
to cell identity and that genetic modules can be co-
opted into new functions, both within the lifespan
of a single animal and across divergent species.
A view more in line with modern network biol-
ogy and consistent with previous network evo-
lution models, such as the modular networks
proposed by Davidson and Erwin (2009), suggests
that the co-regulation of gene expression is tied to
functional protein modules which can be deployed
in a variety of developmental or cellular contexts
and whose physical interaction gives rise to emer-
gent functional properties (Wan et al., 2015), but
whose deployment can be rapidly changed over
short evolutionary timescales.
Despite the many ways in which networks can
be modified, there are still clear patterns of conser-
vation for the linkage between some genetic mod-
ules and the developmental processes they govern.
These observations necessitate that networks—not
just their terminal branching points and resultant
cell-specific transcriptomes—must be considered
26   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
when comparing tissue- and cell-level homology.
These master transcription factors and their associ-
ated genetic modules have been termed “kernels”
due to the centrality of their function in establish-
ing animal body plans (Davidson and Erwin, 2006;
Erwin and Davidson, 2009). Such an ability to es-
tablish and maintain a tissue-level competency may
therefore represent a selectable molecular trait that
has been subsequently preserved in tissues, which
are shared across many species. Some insight into
how particular developmental processes and their
“master transcription factors” may be linked comes
from an improved understanding of the function
of these transcription factors in regulating gene
activity.
To understand how transcription factor expres-
sion may be tied to a specific developmental process,
we can consider two analogous cases: the anteri-
orly expressed homeodomain transcription factor
Six3/6 and FoxA, a winged-helix transcription fac-
tor expressed in the foregut. Anterior expression of
Six3/6 is conserved from cnidarians and across bi-
laterians in the anterior epidermis (­ Sinigaglia et al.,
2013; Marlow et al., 2014), while FoxA is expressed
in the foregut in both protostomes and deuteros-
tomes, as well as the pharynx of cnidarians (Mar-
tindale et al., 2004; Boyle and Seaver, 2008; de-Leon
and Davidson, 2010). In both cases, these factors
are expressed early in the developmental process
in a large population of spatially contiguous cells
that are fated to give rise to a heterogeneous cell
population within a wide diversity of animals, and
while they operate via different mechanisms, they
share a common capacity to establish and main-
tain the state of a progenitor field. Recent work in
liver cells has identified FoxA as a “pioneer tran-
scription factor” capable of opening chromatin and
facilitating the binding of additional, and perhaps
more cell-type specific, transcription factors (Zaret
and Carroll, 2011; Iwafuchi-Doi et al., 2016). Six3/6
has been identified as a co-factor of the GROUCHO
co-repressor (Kobayashi et al., 2001), and has been
shown to act in establishing anterior neural pro-
genitor fields (Gestri et al., 2005). While Six3/6 and
FoxA have clearly distinct roles in regulating gene
expression, both share the ability to establish and
maintain a heterogeneous field of lineage and fate-
restricted progenitors; such factors could therefore
be considered as “competency factors” akin to the
kernels of GRNs hypothesized to establish prolif-
erative zones (Erwin and Davidson, 2009).
2.9  A Case Study in Heterochrony:
Modularity Gives Rise to Heterochronic
Shifts in Feeding Structures
While the larval anterior epithelium and the adult
locomotory trunk can be generally described as
stage-specific features, many organ systems and
tissues may emerge in one stage and later become
selectively advantageous in another, resulting in a
heterochronic shift of a specific morphological fea-
ture or behavior. Shifts in the development of mor-
phological features occur in both directions, with
larval features retained in adult organisms and
adult structures deployed in the planktonic larval
stage (Seuss et al., 2012).
Larval and adult feeding organs are conten-
tious features, particularly as relates to the evolu-
tion of the lophophore (byrozoans, brachiopods,
and phoronids) and ambulacrarian feeding bands.
While some of these controversies have been satis-
factorily “solved” by modern phylogenetic meth-
ods—as is the case for the convergent evolution
of lophophore-like structures (Halanych, 1996;
Philippe et al., 2005)—others are much less clear. It
has been demonstrated that the presence of larval
feeding is a non-stable character which evolves rap-
idly in the presence of environmental pressures (see
Collin and Moran, this volume), in part driven by
the duration of the larval period.
Much effort has been devoted to the ciliary feed-
ing bands of deuterostome larvae and their com-
parison to feeding structures of adult and larval
lophophorate phyla (bryozoans, brachiopods, and
phoronids). While these structures are superfi-
cially similar, structural differences in their form
and function indicate that they appeared conver-
gently. Similarity in adult feeding organs in hemi-
chordates and amphioxus and the tornaria larvae
of hemichordates has also raised the possibility of
a conserved evolutionary relationship between all
three structures, and supports that a heterochronic
shift from larvae to adults (or vice versa) has oc-
curred somewhere along the deuterostome lineage
 E v o l u t i o n a ry D e v e l o p me n t o f M a r i n e L a r va e    27
(Brambell and Cole, 1939). In phoronids, the ac-
tinotroch larva bears a complex feeding apparatus
consisting of many ciliated tentacles and large oral
hood. While this structure is shed and replaced by
a new lophophore in some species upon settlement,
this feeding apparatus is clearly derived from the
adult structure. Similar studies have focused on
the feeding bands of molluscan veliger larvae and
annelid trochophores, and conclude that while
the prototroch (swimming band) is ancestral, the
specialized feeding bands of various lophotrocho-
zoa are likely convergently acquired traits (Rouse,
1999). Although the emergence and homology of
these bands is contentious, it is clear that they dis-
play a higher level of lineage restriction than more
ubiquitous structures such as the apical organ—
for example, while apical organs share homology
across Bilateria, feeding bands have likely evolved
through convergent evolution in echinoderms,
molluscs, and annelids secondarily. Furthermore,
in cases where larval feeding organs are present, a
fully functional through-gut is also present, and like
the adult gut, shows regional expression of parahox
genes (Brooke et al., 1998; Annunziata and Arnone,
2014). These points provide compelling evidence
that larval feeding and digestion represent adult
features which have heterochronically shifted to
earlier developmental stages to allow for the pro-
longation of the larval period. It remains to be de-
termined how often and with what frequency these
shifts have occurred.
2.10  Pelago-benthic Metamorphosis
Is an Ancestral Metazoan Feature
While the morphological features linked to larval-
and adult-specific behaviors, ciliary swimming,
and adult locomotion are useful starting points for
examining homology relationships among lineages,
it can be difficult to assign these features exclusively
to one or the other stage. One feature, however,
stands out as particularly suitable to a core ques-
tion in the evolutionary development of marine lar-
vae: the larval-to-adult transition—metamorphosis.
Metamorphosis itself occurs in many forms among
marine larvae, which, at first inspection, would
make this process an unlikely starting point in a
search for homologous features. However, many
commonalities exist in this transition among spe-
cies. This process has been examined extensively,
but can be generally defined by two phases:
1. Induction of the metamorphic process via the
apical organ (likely through G-protein-coupled
receptors)
2. Rapid generation of the adult form via extension
of the trunk (selective pressure for speed—con-
vergent extension)
In order to transition from the pelagic, free-­
swimming form, larvae need to successfully
identify a suitable substrate and in many cases ir-
reversibly initiate a transformation to the adult
form. Functional evidence has identified the apical
organ as the mediator of the larval metamorphic
transition (Hadfield et al., 2000; Rentzsch et al.,
2008). Biotic and abiotic factors which trigger this
metamorphic process are incredibly diverse and
range from specific bacterial, fungal, or algal sub-
strates that, by way of the co-occurrence with suit-
able habitats, have become biotic homing beacons
for the larvae they attract (Hadfield and Paul, 2001).
Similarly, abiotic factors such as oxygen, tempera-
ture, and dissolved substances factor into the set-
tlement choice. While there is astounding diversity
in the cues larvae utilize in finding a suitable habi-
tat, it is likely that these cues are detected and re-
layed to the nervous system via a familiar route, the
G-protein-coupled family of receptors (GPCRs).
These molecules can be traced to the very root of the
animal tree and appear in great diversity from nearly
the beginning of their appearance (Churcher and
Taylor, 2011). GPCRs can change function rapidly
via change of a small number of residues (Bockaert
and Pin, 1999). This makes them a highly adaptable
system for the detection of settlement cues (much as
they are for odorants and many other environmen-
tal stimuli). While it is currently unknown which
receptors function in the detection of environmental
stimuli, exciting new work has begun to pinpoint
the receptor necessary to relay this settlement signal
to the rest of the organism (Conzelmann 2013b).
Once an initial settlement cue is detected, this
signal must be transmitted via the nervous system
or through paracrine transmission to other tissues
so that a global metamorphic process requiring
28   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
the remodeling of the body architecture can be in-
itiated. Several neurohormones have been inves-
tigated and found to play a role in the settlement
process in marine larvae, although the way in
which these hormones act and the developmental
processes they regulate are varied (Heyland et al.,
2005). Recent work has highlighted the key role
of GLWamide-like neuropeptides in mediating
the larval-to-adult transition in cnidarians, an-
nelids, and potentially sponges (Iwao et al., 2002;
Conzelmann et al., 2013b), and the widespread oc-
currence of this hormone across animals suggests
that its role in the pelagic-to-benthic transition
may have arisen early in the metazoan lineage—a
hypothesis that awaits investigation in additional
lineages. Functional evidence suggests that this
hormone acts via a conserved receptor to mediate
the larval metamorphic transition (Conzelmann
et al., 2013b).
How a neuropeptide acting as a neurohormone
came to govern the larval settlement process is an
interesting question. A diffusible neurohormone
can propagate the settlement signal via synaptic
and non-synaptic transmission and lead to changes
via the nervous system as well as at a global level.
This ability to globally alter the physiology and be-
havior of nearly every cell within the animal allows
the coordination of an organismal-level response.
Interestingly, this same class of neuropeptides has
been shown to mediate the activity of musculature
in both insects and cnidarians (Takahashi et al.,
2003; Lange et al., 2012) as well as feeding behavior
in polychaetes (Williams et al., 2015). It is conceiv-
able that metamorphosin/GLWamide first acted to
coordinate the contraction of musculature for ho-
meostatic processes such as respiration, peristaltic
digestion, or perhaps locomotion but was later co-
opted to mediate the settlement process. It is also
conceivable, however, that its first role was in the
mediation of settlement and that it was later co-
opted into additional roles in the adult organism. In
one such scenario, GL Wamide may in and of itself
been the first intrinsic settlement cue to signal to the
animal nervous system that the adult musculature
and feeding system were mature and functional and
that the initiation of settlement was appropriate.
Metamorphosis, once triggered, leads to an of-
ten rapid and irreversible transition to the adult
form. In some cases, the transition is moderate, in-
volving only the elaboration of pre-formed adult
structures. In other cases, the transition is dra-
matic and involves the de novo growth of entirely
new structures. In both cases, a common outcome
is the rapid extension of adult features through
the process of convergent extension. Convergent
extension is the reshuffling of cellular neighbors
in such a way as to allow for the elongation of the
primary body axis. Such processes are common
in animals and can be found in the germ band
extension of Drosophila (Irvine and Wieschaus,
1994) and the posterior growth of vertebrates
(Wallingfor­ d et al., 2002). During settlement, a
larva is virtually immobilized and unable to es-
cape potential predators, which provides a major
selective pressure for this transition to occur rap-
idly. As a result, most metamorphoses are driven
by rapid cell rearrangements, and occasionally
cell death, rather than de novo growth or prolif-
eration of new structures. In numerous cases, the
primary mechanism driving this transition has
been identified as convergent extension of the epi-
thelium. This process has been described during
the metamorphosis of ascidians and polychaetes
and is likely an underlying mechanism in many
other species (Munro and Odell, 2002; Jiang et al.,
2005; Steinmetz et al., 2007).
While convergent extension takes place in many
contexts—and it is not currently possible to rule out
that this module represents a homoplastic character
redeployed in the face of strong selective pressure—
the molecular underpinnings of this process repre-
sent a conserved genetic activity of the Wnt/PCP
pathway (Heisenberg et al., 2000; Ninomiya et al.,
2004; Roszko et al., 2009). In cnidarians, PCP signal-
ing has been implicated in epithelial polarity and
cnidarian noncanonical Wnt ligands can regulate
convergent extension behaviors in Xenopus (Rigo-
Watermeier et al., 2011; Momose et al., 2012). Both
canonical and noncanonical Wnt ligands have been
identified to specifically localize to the blastopore
(posterior pole of the embryo) in numerous species.
Here, they have been demonstrated to play a role in
both ectodermal patterning and endoderm specifi-
cation. It is likely that ligands also mediate the con-
vergent extension of tissues during axis elongation
during settlement.
 E v o l u t i o n a ry D e v e l o p me n t o f M a r i n e L a r va e    29
2.11 Summary
1. A strictly limited definition of the larval form as
“minimally indirect” can encompass the likely
ancestral mode of development for metazoan
lineages.
2. The larval sensory epithelium (including the api-
cal organ) and the adult locomotory trunk repre-
sent life history stage-specific features for which
hypotheses of homology can be tested.
3. Network studies hold particular promise for
testing the homology of larval features as they
integrate both ontogenetic developmental pro-
grams as well as cell-type specific data.
4. Metamorphosis is a widespread process among
marine invertebrates and employs similar
mechanisms across phyla. Homology of the
metamorphic process, by extension, supports the
ancestrality of a biphasic life cycle with an adult
and larval form.
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 Ch a pt er 3
Evolutionary Ecology of Parental
Investment and Larval Diversity
Dustin Marshall, Justin S. McAlister, and Adam M. Reitzel
3.1 Introduction
Marine larvae vary enormously in the amount of
care—be it in the form of energy or other costly
caregiving that increases offspring fitness—they re-
ceive from their parents. At one end of the spectrum
of parental investment, the offspring of some spe-
cies receive very little from their parents: they are
released as tiny eggs (< 40 μm diameter), and must
complete every element of their development in-
dependently, from fertilization all the way through
to metamorphosis. At the other extreme, some spe-
cies are released as fully formed juveniles ~2000
μm in length, with their entire development com-
plete. Between these two extremes lie every possi-
ble level of parental investment, and most species
lie somewhere along this continuum. In contrast
to terrestrial taxa, parental investment is less cou-
pled to phylogeny in marine taxa, such that closely
related species may have wildly different parental
investment strategies (Marshall et al., 2012). For
example, the congeneric sea urchins Heliocidaris
erythrogramma and H. tuberculata live in the same
habitat but produce offspring that differ in volume
by 35-fold (Marshall et al., 2012). Such diversity
demands explanation, and marine biologists have
been fascinated by variation in parental investment
for over 100 years. In this chapter, we will review
patterns in parental investment in space, review the
theory of parental investment in life history theory,
explore the key assumptions of life history theory
as it pertains to parental investment, and then ex-
amine the evolutionary causes and ecological con-
sequences of variation in parental investment for
Marshall, D., McAlister, J., and Retizel, A., Evolutionary ecology of parental investment and larval diversity. In. Evolutionary
Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland:
Oxford University Press (2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0003
marine organisms. Finally, we will provide some
suggestions as to future work in this field.
Before we consider parental investment in detail,
it is first important to define it. General life history
theoreticians, such as Clutton-Brock and Stearns,
have provided elegant descriptions and justifica-
tions of parental investment, and we use a similar
definition here: the best measure of parental invest-
ment is the proportion of surplus power devoted
to reproduction for each offspring (Clutton-Brock,
1991; Stearns, 1992). Whether that surplus power is
direct energy investments into offspring in the form
of provisioning eggs, embryos, or juveniles; the con-
struction of protective capsules or egg masses; or
the energy expended ventilating a clutch of eggs—
all these acts involve the use of energy specifically
for reproduction. One key element of parental
investment we will focus on in this chapter is off-
spring size. We have that focus for several reasons:
first, offspring is an axis of parental investment that
shows enormous variation among species; second,
it maps to major differences in life history; third, it
is likely that this axis captures much (but certainly
not all; see following) of the variation in parental
investment in marine organisms; and, finally, off-
spring size is a component of parental investment
for which we have the most data both in terms of
its variation and in terms of its consequences for the
ecology marine organisms.
As we begin the chapter, we would first like to ac-
knowledge and recommend the work of the giants
in this field, including but not limited to the early
works of Mortensen, Thorson, Strathmann, Emlet,
and McEdward (e.g., Mortensen, 1922; Thorson, 1950;

Strathmann, 1985; Emlet et al., 1987; McEdwar­d and
Miner; 2001). Furthermore, this chapter hopefully
builds on work by Will Jaeckle and Jon Havenhand
(see Havenhand, 1995; Jaeckle, 1995), and we strongly
recommend these as an excellent starting point.
3.2  The Biogeography of Parental
Investment in the Sea
Biologists have noticed patterns in parental invest-
ment in marine systems for at least 100 years. The
great larval biologist Gunnar Thorson reviewed
the literature in the early 1900s and concluded
that there were latitudinal gradients both in larval
development types and offspring size in marine
invertebrates (Thorson, 1936). Specifically, he sug-
gested that species that lacked a larval stage were
more common in high latitudes and that offspring
sizes were generally larger at the poles relative to
the tropics (Thorson, 1950). Though Thorson was
always troubled by the fact that some species with
small eggs and extended larval phases still thrived
in the poles, he came to believe that the poles “sup-
pressed” such species and favored species that
lacked a larval phase and produced larger offspring
(Thorson, 1950). Thorson believed that the colder
temperatures of the poles and the lack of planktonic
food were the drivers of the life history patterns he
observed. These ideas remained highly influential
for the next 30 years, with extensions of the pattern
beyond marine invertebrates to marine fishes. This
eventually came to be known as “Thorson’s rule”
(Laptikhovsky, 2006).
In the 1990s, marine biologists started to chal-
lenge the orthodoxy of Thorson’s rule for several
reasons (Pearse, 1994). Thorson’s ideas were not
without problems; his understanding was limited
by the data at the time. He thought that species with
nonfeeding larvae were rare and did not occu­r at
the poles. As studies accumulated, it became clear
that many species have nonfeeding larvae and these
were particularly common at the poles (Pearse,
1994). Thorson assumed that the poles were food-
less deserts, but later studies showed that phyto-
plankton blooms, while brief, were significant in
both the Arctic and Antarctic (Marshall et al., 2012).
By the end of the 1990s there was a growing con-
sensus that Thorson’s ideas were outdated and that
Par e n ta l I n v e s t m e n t a n d Lar va l D i v e rsi t y    35
latitudinal patterns in marine organisms were less
clear than once thought (Pearse and Lockhart, 2004).
Over the next decade, life history data continued
to accumulate and explorations of biogeographi-
cal patterns in marine life histories became more
sophisticated (Fernandez et al., 2009). More recent
studies have taken advantage of technological
achievements that were unavailable in Thorson’s
day. These include, for example, more formal sta-
tistical analyses of the association between life his-
tory and latitude, an exploration of the biophysical
drivers underlying latitudinal gradients, and the
introduction of phylogenetic techniques to control
for common ancestry in constraining biogeographi-
cal patterns (Marshall et al., 2012). Today, a modi-
fied version of Thorson’s rule has been revived
(Marshal­l et al., 2012). Generally, higher latitudes
are associated with larger offspring sizes, regardless
of clade or hemisphere. This pattern appears most
associated with latitudinal gradients in mean tem-
perature. The tropics tend to have a higher propor-
tion of species with feeding larvae while the poles
are dominated by species with larvae that do not
feed. Higher latitudes in the southern hemisphere
tend to have much higher proportions of species
that lack a larval phase, but this pattern does not
occur in the northern hemisphere. While the poles
and tropics are extremes and most different with
respect to marine life history patterns, it is worth
noting that they don’t drive these patterns exclu-
sively—the gradients exist within just the temper-
ate latitudes as well (Marshall et al., 2012).
Interestingly, some life history patterns were not
correlated with differences in mean environmental
conditions. For example, the proportion of species
with planktonic-feeding larvae and the size of off-
spring in this group are both unrelated to the mean
abundance of planktonic food. This finding is sur-
prising given that feeding larvae depend on plank-
tonic food to complete their development. Instead,
it seems that the predictability and seasonality of
planktonic food at any one latitude is more impor-
tant than the simple mean abundance for determin-
ing the prevalence and offspring size of species
with feeding larvae (Marshall and Burgess, 2015).
Accordingly, future studies should move beyond
using simple means as a descriptor of environmen-
tal conditions and formally incorporate variability
36   E v o l u t i o n ary E c o l o g y o f M ari n e I n v e rt e b rat e Lar va e
and predictability, as these clearly also act to shape
marine life histories (Marshall and Burgess, 2015).
Given the strong biogeographical relationship
we observe between temperature and develop-
mental mode, it seems reasonable to predict that
future global change will alter the global distri-
bution of developmental modes. Specifically, we
would predict a poleward extension of species with
feeding larvae associated with global temperature
increases. Initial evidence supports this prediction,
with some species with feeding larvae greatly ex-
tending their range to higher latitudes (Marshall et
al., 2012). There is some support for this prediction
already (Ling et al., 2009). Whether species with
nonfeeding or aplanktonic development are made
more rare by global climate change remains unclear
but is certainly a concerning possibility.
3.3  Theory of Parental Investment
in Marine Organisms
One of the starkest dichotomies in marine inver-
tebrate life histories is the grouping of species with
feeding vs. nonfeeding larvae (Strathmann, 1985).
There are a very few species, known as facultative
planktotrophs, which can feed but can also complete
development without feeding (Herrera et al., 1996),
Egg size distributions (n = 1167)
150
100
Count
50
0 0.00
100 1,000
Egg size (μm)
but the overwhelming majority of marine inver-
tebrate larvae either do not feed at all or must feed
to complete development. Offspring size covaries
with developmental mode strongly (Figure 3.1): eggs
smaller than around 120 μm in diameter tend to pro-
duce feeding larvae, while eggs larger 300 μm tend
to only produce nonfeeding larvae (with the notable
exception of Conus, in which feeding larvae can be up
to 400 μm in diameter; Kohn and Perron, 1994). For
a detailed consideration of the differences between
planktotrophy, lecithotrophy, and feeding and non-
feeding larvae, see Strathmann (1985). The covariance
between egg size and developmental mode has long
fascinated biologists, and there have been repeated
theoretical attempts to explain the evolution of egg
size and development starting almost 50 years ago.
Richard Vance developed a theoretical model for
predicting offspring size in 1973, and he developed
it specifically for marine invertebrates (Vance, 1973).
Smith and Fretwell published a more general model
the following year (Smith and Fretwell, 1974), and
this has received more attention from a broader au-
dience, but it is worth noting that the first optimal-
ity model of offspring size was Vance’s—a fact not
often appreciated among life history theoreticians
more generally. Vance’s model has two key com-
ponents, an offspring size-number trade-off and
0.12
0.10
0.08
Proportion
0.06
0.04
Figure 3.1  Distribution of offspring sizes (data taken
0.02
from Marshall et al., 2012) across three developmental
modes. The darkest bars are for species with feeding
larvae, the intermediate shade for species with
nonfeeding larvae, and the lightest bars are for species
that lack a planktonic larval phase (see Plate 6).

an offspring size-fitness function (Vance, 1973). As
far as we are aware, these two basic components
have been retained in all models of offspring size
since then. In this section we will consider the ba-
sic model of Vance, and outline some modifications
that have been added in order to better reflect our
understanding of how offspring size affects ma-
rine invertebrate life histories. We will later deal
with empirical evidence for each component (the
size-number trade-off and the offspring size-fitness
function) separately. We will now briefly consider
Vance’s model (for a more detailed review see
Marshal­l and Keough, 2008a).
In six relatively simple equations, Vance laid out
his assumptions about how egg size affected fitness
in marine invertebrates (Vance, 1973). He assumed
that larger eggs took longer to develop through the
embryonic stage before becoming larvae that could
either feed or not feed. Thus, in species with non-
feeding larvae, the length of the larval phase was
positively correlated with egg size. This was a par-
ticularly prescient assumption given there was lit-
tle supporting data in 1973, but since then several
studies have confirmed this relationship (Marshall
and Keough, 2008a). Vance further assumed that in
species with feeding larvae, larger eggs developed
into larvae that spent less time in the plankton be-
fore accumulating sufficient resources to metamor-
phose—relative to the pre-feeding phase, Vance
assumed that the length of the feeding period was
much greater. Vance made no assumptions about
size-specific mortality rates, though later models
did (Christiansen and Fenchel, 1979). Vance also as-
sumed that larvae would only ever be large enough
to complete metamorphosis, a size he called s—
there was no fitness advantage to producing eggs
that are larger than s. Later studies by Herrera et
al. (1996) and McEdward and Miner (2006) further
discussed and quantified s in relation to maternal
investment and nutritional mode of larvae. Vance
also assumed that mortality rates were constant in
the plankton, such that any extension of the larval
period incurred more mortality. Finally, Vance as-
sumed a direct energy trade-off between the size
and number of offspring that mothers could pro-
duce: any increase in size yields a concomitant re-
duction in the number of offspring that mothers
could make due to energy constraints.
Par e n ta l I n v e s t m e n t a n d Lar va l D i v e rsi t y    37
Vance’s model predicted that mothers should ei-
ther produce the smallest possible offspring or the
largest possible offspring (to a maximum size s = 1),
and that local conditions would alter the balance be-
tween these extremes (Vance, 1973). The model pre-
dicted that food availability should affect the relative
advantages of producing feeding vs. nonfeeding lar-
vae: higher food abundances should favor the pro-
duction of feeding larvae. However, as we saw in the
previous section, this intuition is not supported in
a straightforward way. Vance’s model had the prob-
lem of “lost souls”: infinitely (and therefore infinitely
numerous) small offspring still had some fitness
because the model always predicts fitness is maxi-
mized by producing infinitely small eggs—Vance
assumed an arbitrary minimum viable offspring size
to work around this problem.
The theoretical framework that Vance provided
was an invaluable contribution to the field, and the
paper should still be considered required reading
by today’s marine evolutionary ecologists. Never-
theless, the model contains assumptions that today
have less support, and is limited to only one section
of the life history, whereas it now appears that egg
size has pervasive consequences across the entire
life cycle. In what follows, we consider the key com-
ponents of Vance’s model and revise these in light
of our current understanding.
3.4  Parental Investment, Egg Size,
and the Size-Number Trade-Off
Vance’s original suggestion that mothers must bal-
ance the size and number offspring that they can
produce due to energy constraints has reverber-
ated through the literature ever since. Upon initial
consideration, such a constraint has intuitive ap-
peal—mothers have a limited amount of energy
they can devote to reproduction and so increases in
the energy contained in each offspring must come
at a cost to the number of offspring that can be pro-
duced. Thus, it could be argued that the natural unit
of parental investment is the energy content of off-
spring rather than the size of offspring, and so stud-
ies should seek to measure energy content wherever
possible. Such an approach raises the practical dif-
ficulty of measuring energy content of very small
individuals, often in destructive ways. If offspring
38   E v o l u t i o n ary E c o l o g y o f M ari n e I n v e rt e b rat e Lar va e
size is well correlated with energy content, then off-
spring size would be a convenient proxy for energy
content, with the additional advantage of being a
nondestructive measure. Offspring size also has
the convenient property of capturing other poten-
tial trade-offs and can directly affect performance
(more about this later). Thus, the crucial question
remains: how well are offspring size and energy
content correlated?
3.4.1  Does Offspring Size Reflect Energy
Content?
Interspecific studies show strong relationships be-
tween egg size and energy content, but these are
largely irrelevant to our considerations here given
selection acts within species (Jaeckle, 1995). Like-
wise, while Vance’s model has often been invoked
to explain differences among species, the processes
that operate to increase fitness of one pheno-
type over another operate within species. Indeed,
Vance’s model is explicitly intraspecific in its ap-
proach, though it is used to make inferences about
interspecific patterns. Thus, while macroevolution-
ary patterns among species may provide clues as
to the microevolutionary processes that produced
them, the key relationships to be considered are the
covariances between offspring size, energy content,
and offspring performance within species.
Two problems with estimating the relationship
between offspring size and energy content within
species are that we’re often working with relatively
small amounts of variation in offspring size, such
that the “signal” of covariation is likely to be small,
and relatively imprecise methods for estimating
content—though technological advances are im-
proving methods all the time—such that the “noise”
is likely to be substantial. This low signal-to-noise
ratio makes tests of the relationship between off-
spring size and content highly susceptible to Type II
statistical errors (false negatives). One might errone-
ously conclude that there is no relationship between
offspring size and content simply because the range
of sizes tested is too small and the imprecision of
the measurement of content is too great. A good
illustration of this problem is provided by Solaster
stimpsoni, where an influential study using only a
small number of replicates found no relationship
between egg size and content (McEdward and
Coulter, 1987), but a better replicated study later
found that such a relationship did exist (McEdward
and Chia, 1991), suggesting the earlier analysis had
suffered a Type II error. There are a few different
ways of dealing with this issue that we recommend.
One approach would be to simply use a different
α for concluding that a relationship is significantly
different from zero (for a detailed consideration
of adjusting α, see Quinn and Keough, 2002). We
would argue, however, that even this approach is
probably not the most informative.
Imagine that we find a significant relationship be-
tween offspring size and content. Under a standard
frequentist approach, this tells us that the relation-
ship is significantly different from zero, but it tells
us nothing about how a single unit increase in size
results in a corresponding increase in content. Yet
for the strict energy-based trade-off that is often
modeled to apply, there needs to be a one-to-one
relationship between offspring size and content. We
recommend that future studies focus on this one-
to-one relationship; specifically, a more appropriate
statistical approach would be to test whether the
relationship between offspring size (volume) and
content scale significantly differently from 1.
There are several ways of doing this. The simplest
might be to determine whether the confidence in-
tervals on the coefficient linking size and content
overlap 1. Alternatively, a Wald test—where the dif-
ference between the coefficient and 1 is divided by
the standard error of the coefficient and then com-
pared against a t distribution (as in a standard Wald
test for whether a slope is sufficiently different from
0, but here 0 is replaced by 1)—could be used to
determine whether the relationship is significantly
different from 1. The equivalent approach under a
Bayesian framework would also work well. Regard-
less of the specific approach, we suggest that simply
testing whether the relationship between offspring
content and size is different from 0 is relatively
uninformative, and future studies should instead
formally test whether offspring size (volume) and
content scale differently to a one-to-one ratio (Mora­n
et al., 2013). As always, a power analysis should also
be considered to determine whether such an effect
can even be detected. This approach also reduces
the likelihood of committing a Type II error.

3.4.2  Does Energy Content or Size Reflect Total
Per-Offspring Investment?
Vance’s original focus on the energy content of off-
spring means that subsequent studies have typi-
cally estimated the energy content of offspring once
they are released from the parent, or have com-
pleted development. For example, there have been
several subsequent reviews that cover the differ-
ences in composition of eggs from species with vari-
ous developmental modes (Moran and McAlister,
2009). An issue with such an approach is that it only
captures a proportion of the energy that the parent
expended on each offspring. Similarly, offspring
size only estimates the physical dimensions of an
offspring upon release, and may not estimate all of
the investment made to produce that offspring.
Ideally, any measure of parental investment
should include the total costs of producing each
offspring; this would include the costs of creating
the reproductive structure that produced the egg,
the reproductive tract for either receiving the sperm
or spawning the egg, any accessory costs associated
with the egg (e.g., follicle cells, egg coats, mucilage,
and thickeners), and costs of moving to a spawning
site. Gametes stored in the gonad are unlikely to be
completely metabolically inert, and will consume
some resources until release. In species that brood
developing offspring, some require costly ventila-
tion (e.g., crustacean egg broods) and others draw
resources from the mother while developing (e.g.,
bryozoans, echinoderms; McClary and Mladenov,
1990). For mothers that release offspring into egg
masses or capsules, the costs of the protective struc-
tures, as well as nutritive eggs or yolk material, must
also be factored into the energy costs of producing
the offspring. These costs matter because they will
influence the per capita energy costs of producing
offspring and alter the optimal balance between the
size and number of offspring that mothers should
produce. For example, Sakai and Harada (2001) pre-
dict that, because brooded offspring (in their case
seeds, but the theory equally applies to brooded
offspring) consume resources while increasing in
size, the rate at which offspring consume resources
relative to their supply while being brooded alters
the predicted optimal offspring size that mothers
should produce in order to maximize fitness. While
Par e n ta l I n v e s t m e n t a n d Lar va l D i v e rsi t y    39
a few studies have estimated the costs of brooding,
and ventilation in marine invertebrates (McClary
and Mladenov, 1990), the energy content of acces-
sory structures (Bolton et al., 2000), and the provi-
sioning of nurse egg structures (Collin and Moran,
this volume), there are too few to generalize, and
existing theory is yet to incorporate these overhead
costs into offspring size-number models.
3.4.3  Does Energy Content Reflect the Proximal
Constraints on Maternal Investment?
One problem with assuming that mothers are lim-
ited by energy in determining how they should
balance the size and number of offspring that they
produce is that it assumes that energy is the proxi-
mal limiting factor. Instead, in any one reproduc-
tive bout, the reproductive capacity of a female may
simply be limited by size. For species that broad-
cast spawn, total investment in reproduction may
be limited by maximum gonad size, and for species
that brood, total investment might be limited by the
maximum size of a brood that can be maintained
or oxygenated (Strathmann and Strathmann, 1982).
From this perspective, offspring size may repre-
sent a more natural currency to consider as a direct
trade-off for number. However, too few studies
have examined how space limited brood or gonad
capacity is in marine invertebrates (though Strath-
mann and others have argued that it does represent
an important limit).
3.5  Offspring Size-Fitness Functions
Regardless of what costs (space, energy) are associ-
ated with increasing offspring size, it seems reason-
able to assume that if offspring size does not affect
subsequent offspring fitness, then mothers should
produce the smallest, cheapest (in terms of energy)
offspring possible because this will maximize ma-
ternal fitness. If, on the other hand, offspring size
does positively affect fitness, then mothers must bal-
ance the costs of producing larger, more expensive
offspring with the benefits of each offspring having
higher fitness. Ever since Vance’s assumption that
offspring size affects planktonic duration alone, we
now have a wealth of data that suggests that off-
spring size affects every aspect of offspring fitness,
40   E v o l u t i o n ary E c o l o g y o f M ari n e I n v e rt e b rat e Lar va e
from initial fertilization through metamorphosis,
and can affect adult reproduction and even lifespan.
Throughout the following, we will consider only
the effects of offspring size on subsequent perfor-
mance within species, as this is an appropriate scale
for considering the eco-evolutionary dynamics of
parental investment. For an exploration of among-
species covariation between offspring size and per-
formance, see Marshall and Keough (2008a).
3.5.1  Offspring Size and Fertilization Success
By recent estimates, around 50% of all marine inver-
tebrates have external fertilization, where eggs and
sperm must meet in water (Monro and Marshal­l,
2015). External fertilization is fraught, with the
probability of being fertilized strongly dependent
on sperm concentration and thus the density of
spawning males. When sperm concentrations (and
male densities) are too low, the probability of an egg
being contacted by a sperm is low and so fertiliza-
tion success is sperm-limited. When sperm concen-
tration (and male densities) are too high, then the
probability of eggs being contacted by multiple
sperm simultaneously is high, and if multiple sperm
enter the egg before the egg has a chance to create
a block, then the egg suffers a condition known as
polyspermy and usually dies. While there has been
debate regarding the prevalence of both sperm limi-
tation and polyspermy in natural populations, it is
reasonable to assume that both happen and their
likelihood depends mostly on the density of spawn-
ing males and local hydrodynamic conditions. It
now seems that egg size also affects fertilization
with consequences for selection on offspring size.
Vogel et al. (1982) predicted and Levitan (1996)
showed that egg size affects the kinetics of fertiliza-
tion: larger eggs are more likely to be contacted by
sperm simply because they present a larger target
for sperm to hit. Levitan showed that under sperm-
limiting conditions, larger eggs were favored;
Levitan incorporated the influence of egg size on
fertilization into a modified version of the Vance
model (Levitan, 2000). Later, Marshall et al. (2002)
showed that not only were larger eggs more likely
to be fertilized under sperm-limiting conditions,
but they are also more likely to suffer polyspermy
under sperm-saturating conditions. Thus, all else
being equal, high densities of sperm/males should
favor smaller eggs and low densities should favor
larger eggs, a prediction that has some empirical
support (e.g., Crean and Marshall, 2008).
3.5.2  Offspring Size and the Planktonic Period
Looking across species and developmental modes,
species with small eggs and feeding larvae tend to
have longer planktonic periods than species with
larger eggs and nonfeeding larvae. A study of echi-
noderm life histories found that planktonic duration
did not correlate well with developmental mode
(Mercier et al., 2013); in only two classes out of four
(echinoids and ophiuroids) did species with feed-
ing larvae have longer durations than species with
nonfeeding larvae. The failure to find an effect of
developmental mode for holothuroids and asteroids
is surprising, and we decided to reanalyze those
data. Importantly, we included the effect of rear-
ing temperature in the analysis for which data were
available because planktotrophic species are more
likely to occur in warmer waters and nonfeeding
species are more likely to occur in cooler waters; in
other words, temperature could confound the effect
of developmental mode. Given that temperature is
likely to drive developmental times strongly and
that the key comparison of interest is the effect of
developmental mode on developmental period for
a given temperature, we therefore statistically con-
trolled for temperature by including it as a covariate.
Usin­g an ANCOVA approach, we found a strong ef-
fect of developmental mode in both holothuroids
(F1,24 = 15.15, P <0.001) and asteroids (F1,50 = 22.59,
P <0.001). For asteroids, we find that for a given tem-
perature, feeding larvae have a developmental pe-
riod that is twice as long as nonfeeding larvae, and
for holothuroids, feeding larval periods are almost
eight times longer than nonfeeding larval periods.
Thus, we would agree with Mercier et al. (2013) that
larval developmental mode alone is a poor predictor
of developmental period across wide temperature
ranges. Nevertheless, once the confounding influ-
ence of temperature is taken into account, we would
instead conclude that larval mode is an excellent
predictor of developmental period across all echino-
derm classes. Thus, Vance’s intuition that plankto-
trophs are cheaper to make but spend longer in the

plankton and therefore accumulate more mortality
holds true for Echinodermata in any one thermal
environment. We will now examine patterns within
each developmental mode and within species.
Vance’s predictions about how offspring size af-
fects planktonic period were remarkably prescient.
In species with nonfeeding larvae, larger eggs gen-
erally take longer to develop and hatch than smaller
eggs (though there are exceptions). Larger nonfeed-
ing larvae also tend to be more selective with respect
to settlement, taking longer to reach competence to
metamorphose, but are also more likely to reject
low quality settlement sites for longer (Marshal­l
and Steinberg, 2014).
In species that are released as competent non-
feeding larvae (e.g., colonial ascidians, bryozoans,
and sponges), larger larvae tend to have longer
planktonic periods than smaller larvae. Though
larger larvae are capable of ending the larval period
as soon as smaller larvae, it is thought that larger
larvae are more likely to delay settlement in the ab-
sence of cues for high quality habitat. A recent study
(Pettersen et al., 2015) suggests that larger larvae
not only begin life with more energy, but because
of the allometric relationship between offspring
size and offspring metabolism, larger larvae also
utilize a lower proportion of their energy reserves,
such that delaying metamorphosis is less costly in
larger larvae relative to smaller larvae. Regardless
of the mechanism, it seems that larger larvae have a
higher chance of settling into higher quality.
In species with feeding larvae, limited evidence
suggests that larger eggs take longer to hatch into
feeding larvae than smaller eggs. A series of el-
egant manipulations of egg size by Sinervo and
McEdwar­d (1988) as well as Hart (1995) showed
that egg size also affects the duration of the larval
feeding period in sea urchins in the manner pre-
dicted by Vance. Later studies also show that gener-
ally, larger eggs require less time as feeding larvae
in order to complete development.
While the effects of egg size on the overall plank-
tonic period are increasingly well understood, more
detailed explorations of the mechanisms by which
offspring size affects fitness in species with feeding
larvae are lacking. For example, it is easy to imagine
Par e n ta l I n v e s t m e n t a n d Lar va l D i v e rsi t y    41
how offspring size may affect the ability of larvae
to feed, exploit different food resources, or resist
starvation. Furthermore, offspring size has long
been predicted to affect susceptibility to predation
(Christiansen and Fenchel, 1979), a prediction that
has some initial support (Allen 2008).
3.5.3  Offspring Size Effects on Post-metamorphic
Performance
Offspring size affects the survival, growth, repro-
duction, and longevity of marine invertebrates
(Table 3.1). Within a range of species and devel-
opmental modes, larger offspring tend to survive
better than smaller offspring. The mechanisms by
which larger offspring have a survival advantage
are unclear. Larger offspring can be better buffered
against starvation, more resistant to predation, and
more competitive than their smaller conspecifics
(Marshall and Keough, 2008a).
Larger offspring also tend to show higher rates
of post-metamorphic growth than smaller offspring
(Table 3.1). In contrast, one study (Jacobs and
Sherrard, 2010) on seven ascidians failed to find a
relationship between initial offspring size and sub-
sequent size though this study had limited replica-
tion (an average of nine replicates per species). In
some colonial marine invertebrates, offspring size
affects initial asexual budding rates with larger
offspring producing more daughter zooids than
smaller offspring. In other species, the mechanism
by which offspring size affects growth remains
unclear, though viable explanations include size-
based differences in feeding structures or foraging
behavior such that larger offspring can gain re-
sources more efficiently (Kosman and Perne­t, 2011).
A recent study (Pettersen et al., 2015) also shows
that metamorphosis is relatively less costly for
larger offspring than smaller offspring. Petterse­n
et al. (2015) show that, because metabolism scales
allometrically with size, smaller offspring burn
~47% of their energy reserves in order to complete
metamorphosis while larger offspring will burn
~22%. Thus, as well as possibly beginning life with
a higher research of energy, larger offspring are
also more efficient in passing through the costly
42   E v o l u t i o n ary E c o l o g y o f M ari n e I n v e rt e b rat e Lar va e

Table 3.1  Studies Examining Effect of Offspring Size on Post-Metamorphic Performance in Marine Invertebrates.

Study Location Species Survival Growth Reproduction

Emlet and Sadro, 2006 Field Balanus glandula ✗ ✓

Kosman and Pernet, 2011 Lab Bugula californica ✓
Burgess et al., 2013 Field Bugula neritina ✓ ✓ ✓
Monro et al., 2010 Field Bugula neritina Mixed ✗
Monro et al., 2010 Lab Bugula neritina ✗ ✓
Kosman and Pernet, 2011 Lab Bugula neritina ✓
Dias and Marshall, 2010 Field Celleporaria sp. ✓ ✓
Kosman and Pernet, 2011 Lab Cryptosula pallasiana ✓
Allen and Marshall, 2014 Field Hydroides diramphus ✗ ✓
Allen and Marshall, 2013 Field Hydroides diramphus ✓ ✓
Kesselring et al., 2012 Field Janua pagenstecheri ✓(negative effect) ✓ ✓
Gehman and Bingham, 2009 Lab Leptasterias aequalis ✓ ✓
Rius et al., 2010 Field Microcosmus squamiger ✓ ✓
Sun et al., 2015 Lab Urticina felina ✗
Davis and Marshall, 2014 Field Watersipora subtorquata ✓
Marshall and Monro, 2012 Field Watersipora subtorquata ✓ ✓
Marshall and Keough, 2008 Field Watersipora subtorquata ✓ ✓
Lange and Marshall, 2016 Field Watersipora subtorquata ✓
Crean et al., 2011 Field Styela plicata ✓ ✓
Jacobs and Sherrard, 2010 Field Boltenia villosa ✗
Jacobs and Sherrard, 2010 Field Styela gibbsii ✗
Jacobs and Sherrard, 2010 Field Corella inflate ✗
Jacobs and Sherrard, 2010 Field Diplosoma macdonaldi ✗
Jacobs and Sherrard, 2010 Field Distaplia occidentalis ✗
Jacobs and Sherrard, 2010 Field Botrylloides violaceus ✗
Jacobs and Sherrard, 2010 Lab Boltenia villosa ✗
Jacobs and Sherrard, 2010 Lab Styela gibbsii ✗
Jacobs and Sherrard, 2010 Lab Corella inflate ✗
Jacobs and Sherrard, 2010 Lab Ciona savigni ✗
Jacobs and Sherrard, 2010 Lab Diplosoma macdonaldi ✗
Jacobs and Sherrard, 2010 Lab Distaplia occidentalis ✗
Jacobs and Sherrard, 2010 Lab Botrylloides violaceus ✗
Carrasco et al., 2012 Lab Cominella virgata ✗ ✓
Carrasco et al., 2012 Lab Cominella maculosa Mixed ✓
Pernet et al., 2012 Lab Capitella teleta ✓ ✓

Note. Unless otherwise stated, crosses indicate no significant effect, and check marks indicate a significant, positive effect of offspring size on performance. Where
mixed effects were recorded, the effects of offspring size were inconsistent among different experiments.

metamorphic stage. Though there are fewer stud-
ies, offspring size effects on post-metamorphic per-
formance also occur in species with feeding larvae
(Allen and Marshall, 2010). It seems that even small
differences in initial investment can still manifest in
the adult stage despite an extended larval feeding
stage. This persistent difference may be due to the
differences in efficiency between smaller and larger
offspring discussed above, but tests of metabolic
scaling in species with feeding larvae suggest oth-
erwise (Moran and Allen, 2007).
Experimental studies within species that differ in
relative investment suggest that offspring size may
have complex relationships with reproduction. For
example, both Marshall et al. (2003) and Dias and
Marshall (2010) find that larger offspring tend pro-
duce more offspring themselves relative to smaller
offspring. These effects are not always straightfor-
ward, however: Kesselring et al. (2012) showed that
while larger offspring tended to reproduce more in
any one reproductive bout, they also died sooner
than smaller offspring such that they took part in
fewer reproductive events. These experimental re-
sults are essential in order to provide empirical tests
for hypotheses developed in the context of models
or comparative datasets. Given that reproductive
output is one of the best estimates of true fitness,
we would argue that more studies that follow the
entire life history, including life span and total re-
productive output, are needed.
3.6  Eco-evolutionary Dynamics
of Parental Investment
3.6.1 Ecological Importance of Offspring Size
Given the effects of offspring size on subsequent
performance, it seems reasonable to assume that
variation in offspring size is likely to be an impor-
tant driver of ecological dynamics. While marine
ecologists have long focused on the role of variation
in the quantity of larvae entering the population as
a driver of population dynamics, offspring size ef-
fects suggest that we should also consider the qual-
ity of larvae entering the population. Indeed, larval
quality effects more generally can overwhelm the
effects of larval quantity effects in marine popula-
tions (Burgess and Marshall, 2011a). Examination
Par e n ta l I n v e s t m e n t a n d Lar va l D i v e rsi t y    43
of the ecological importance of offspring size
for populations are exceedingly rare but the one
study that has explored this formally found that
offspring size plays only a limited role: offspring
size only accounted for between 0.1% and 15% of
total variation in post-metamorphic performance
in the bryozoan Watersipora subtorquata (Lange and
Marshall, 2016). Importantly, offspring size may
not only affect the dynamics of the focal species in
which offspring size varies, rather it can also affect
the assembly of the surrounding community. Davis
and Marshall (2014) found that offspring size in a
resident species can be more important in driving
community assembly in the field than the density
of the resident species. Paradoxically, when the resi-
dent species originated from larger offspring (and
therefore grow faster), the community that subse-
quently assembled was denser than when the resi-
dent species originated from smaller offspring (and
therefore grew slower). These results suggest that
offspring size affects not only the performance of
the focal species but also affects its niche usage and
the capacity for other species to coexist. Whether
offspring size has community-level effects in other
systems or species remains completely unexplored,
but as Davis and Marshall (2014) argue, such effects
are likely to be widespread.
3.6.2  Drivers of Among-Environment Variation
in Offspring Size
Previous reviews (e.g., Marshall et al., 2008a; 2012)
have identified the drivers of broadscale temporal
(e.g., seasonal) and spatial (e.g., latitudinal) patterns
in offspring size, and so here we focus on local-scale
source of variation in offspring size. Similarly, the
role of maternal phenotypes such as size in altering
the offspring size relationship has also been dealt
with in a separate review.
Life history theory predicts that changes in the
offspring size-performance relationship should al-
ter the offspring size that mothers should produce
(Parker and Begon, 1986; Kindsvater and Otto,
2014). Formal theory predicts that optimal offspring
size is dependent on the steepness of the relation-
ship between offspring size and performance. Gen-
erally, if the relationship is steep, then mothers will
be better off producing larger offspring because the
44   E v o l u t i o n ary E c o l o g y o f M ari n e I n v e rt e b rat e Lar va e
per capita fitness return of high performing, larger
offspring exceeds the per capita cost of producing
larger offspring. Conversely, when the relationship
is shallow, mothers should produce smaller off-
spring because they get much of a less fitness return
for a given increase in per capita investment and are
better off maximizing the number of offspring that
they can produce. Thus, any changes in the envi-
ronment that alter the offspring size-performance
relationship should alter the fitness returns of a
given offspring size.
If the maternal environment changes, and it is a
good predictor of the offspring environment, then
selection should favor mothers that alter the size of
their offspring accordingly (Burgess and Marshal­l,
2014). There is increasing evidence that such
transgenerational plasticity is widespread in marine
invertebrates both among and within populations.
The environmental cues to which mothers modify
the investment in eggs include diverse factors: biotic
and abiotic, as well as natural and anthropogenic.
In the bryozoan Bugula neritina, colonies reared in
high-competition environments produce larger off-
spring that are better competitors and more likely
to disperse away from local competitive conditions
(Allen et al., 2008). Similarly, colonies exposed to a
heavy metal stress will also produce larger, more
resistant offspring that are more likely to disperse
away, potentially to avoid the physiological stressor
in the next generation (Marshall, 2008). Mothers
exposed to higher temperature tend to produce
smaller offspring that perform better in higher tem-
peratures themselves (Burgess and Marshall, 2011b).
Mothers can also alter the phenotype of their off-
spring in relatively subtle ways. For example, in the
ascidian Styela clava, individuals reared at high den-
sities produce smaller eggs than individuals reared
at low densities (Crean and Marshall, 2008). The
change in overall egg size comes from a decrease
in the size of egg accessory structures (follicle cells)
such that the overall target of the egg is smaller and
less susceptible to polyspermy. Even though the
overall egg size is smaller, the ovicell, the portion of
the egg that provides nutrition for the offspring, ac-
tually increases in size in individuals reared at high
densities—presumably to increase the dispersal po-
tential and competitive ability of those individuals
(Crean and Marshall, 2008).
Such size-based changes in offspring size are
not universal. For example, in tubeworms, moth-
ers exposed to lower salinities or higher tempera-
tures do not change the size of their offspring, even
though the performance of their offspring changes
dramatically, suggesting that mothers can manipu-
late the phenotype of their offspring in numerous
ways (Jensen et al., 2014; Guillaume et al., 2015),
which may differ between populations (Collin and
Salaza­r, 2010).
3.6.3 Environmental Dependent Variation
in Offspring Quality Other than Size
Mothers may be able to manipulate the phenotype
of the eggs and offspring in ways that are unrelated
to size. Maternal effects beyond offspring size are
common in diverse organisms (e.g., oviparous ver-
tebrates, insects, vascular plants; Rossiter, 1996) and
are surely present in marine invertebrates as well.
Thus, while much of the theoretical and empirical
work on ecologically and evolutionary related vari-
ation in marine invertebrate offspring has focused
on size as a proxy for investment, additional ele-
ments of the egg phenotype affect offspring fitness.
Vitellogenesis broadly refers to the period of oo-
cyte development in the ovary where protein, lipids,
and carbohydrates are synthesized or assimilated
into the maturing egg. These biochemical compo-
nents can either be synthesized in a non-ovarian tis-
sue (liver in mammals, fat body in insects) or in the
ovary and transported into the oocyte via endocytosis
(termed, heterosynthetic), or they can be synthesized
by the egg itself after small molecule uptake (termed,
autosynthetic). Research by Eckelbarge­r (2005) sug-
gests that species with heterosynthetic mechanisms
of yolk synthesis can produce mature eggs rapidly
because large molecules like vitellogenin are deliv-
ered and endocytosed in mature form. Furthermore,
Eckelbarger (2005) hypothesized these mechanisms
and the ovary may exert significant effects on egg in-
vestment, among other factors, important for larval
development. Thus, these vitellogenic mechanisms
could promote different forms of plasticity in egg
composition. For species with heterosynthetic yolk
production, the quantity of yolk deposited into each
egg could be altered by environment-dependen­ t
shifts in vitellogenin and other large molecules

available during oocyte maturation or the modifica-
tion of lipoproteins and vitellogenin-specific cell re-
ceptors on the oocyte membrane. For autosynthetic
species, the environment may shift the proportion
of amino acids, monosaccharides, and fatty acids
circulating in the female during egg maturation and
thus the small molecules available from which larger
nutritive molecules are synthesized. In both cases,
the mature egg may more closely match the mater-
nal nutritive profile and thus may better match the
current environment. Mature oocytes provide the
initial nutrition to the embryo in the form of yolk
protein. In many oviparous organisms, vitellogenin
serves as the precursor yolk protein that is enzymati-
cally cleaved into lipoproteins and phosphoproteins.
Biochemical measurements of mature oocytes have
determined the quantity of lipids, proteins, and car-
bohydrates and, in some cases have treated these as
species’ characteristics that represent maternal in-
vestment. However, females may shift not only the
total caloric investment but potentially the represen-
tation of each category of molecule in functionally
important ways, particularly in response to envi-
ronmental variation (Moran and McAlister, 2009;
Marshall and Keough, 2008b). For example, females
experiencing lower food environments at time of re-
production may produce eggs with higher relative
protein in place of lipid for their size than conspecif-
ics in more food rich environments. Thus, while the
size and total caloric content may remain the same,
the molecules may shift.
Furthermore, mature eggs are incredibly complex
cells with subcellular localization of proteins to the
membrane and other regions of the cell, diverse RNA
types, and organelles. For example, proteomic ap-
proaches applied to oocytes have shown that mature
eggs are composed of hundreds to thousands of pro-
teins that are not explicit sources of energy but instead
involved in the development of the embryo (Lotan et
al., 2014). These proteins have diverse roles including
cell cycle regulation, RNA silencing, membrane sign-
aling, and metabolism. Thus, females may provision
their eggs with proteins to better suit their offspring to
the current environment. For example, the aforemen-
tioned tubeworms exposed to low salinity may have
increased relative amounts of mRNA or proteins into
their eggs favoring performance in these more stress-
ful environments. Potential candidates could include
Par e n ta l I n v e s t m e n t a n d Lar va l D i v e rsi t y    45
general chaperon proteins like heat-shock proteins or
process-specific proteins like aquaporins.
We should also recognize that the evolution of
environmental manipulations to eggs is limited by
the costs and limits common to any plasticity re-
sponse. The environment may not represent reliable
cues to mothers such that there is no selection for
mothers to alter their offspring size—formal esti-
mates of environmental predictability are necessary
if tests of transgenerational plasticity are to be reli-
able (Burges­s and Marshall, 2014).
3.6.4  Within-Brood Variation
Offspring size varies not only among broods and
mothers, but it also varies within broods. Whether
this variation is adaptive is hard to determine. It
has been suggested that within-brood variation
represents a form of bet-hedging, mothers may
make a range of offspring sizes when their future
environment is unpredictable (Crean and Marshall,
2009). Among species patterns in within-brood
variation suggest that species with more disper-
sive larvae (and less predictable experience) tend
to have higher within-brood variation in offspring
but any number of factors could drive these effects
(Marshall et al., 2008b). Within species tests of this
idea are rare and inconclusive (Crean and Marshall,
2009). Empirically distinguishing between adaptive
bet-hedging strategies and simple developmen-
tal instability may prove difficult in most systems
and so for now, the adaptive significance of within-
brood variation in offspring size remains unclear.
3.7  Future Directions
Marshall et al. (2012) reviewed the biogeography of
marine invertebrate life histories, and their Figure 1
provides a striking visual indication that there are
still vast swaths of the global marine environment
that remain unexplored (or at the least, undocu-
mented). With respect to marine invertebrate life
histories, the possibility exists that some of these
unexplored areas have in fact been studied and
that difficulties in obtaining published work in lan-
guages utilizing a non-Latin alphabet may bias our
understanding. However, the majority of global lo-
cations are represented by only one to two published
46   E v o l u t i o n ary E c o l o g y o f M ari n e I n v e rt e b rat e Lar va e
studies, and in only three areas (the Puget Sound
region of North America, the English Channel, and
Eastern Australia) do the number of studies fall
into the article’s highest category (55–104 published
studies). These results suggest that well-studied ar-
eas are outliers to the overwhelming paucity of data
on marine invertebrate life histories, globally.
Further study of this image reveals broader-
scale patterns of our lack of information. There is a
bias toward more information from northern than
southern hemisphere locations, toward temperate
over tropical regions, and data from polar loca-
tions is largely absent, aside from two moderately
well studied locations in Antarctica adjacent to
polar research stations. Other largely unexplored
coastal areas include the majority of the coasts of
Africa, South America, and Antarctica, the Middle
East and India, Indonesia, northern Russia and the
Kamchatka Peninsula, northern Canada, Alaska
and the Aleutian Islands, as well as many isolated
island archipelagos worldwide. The life histories of
marine invertebrates inhabiting the vast majority of
the deep-sea benthos (the largest habitat on Earth)
are also unknown to science.
Going forward, how may exploration into these
biogeographically understudied areas be important
for refining a general theory of offspring investment?
Temperature and food availability are recognized as
the dominant variables contributing to latitudinal
patterns in offspring size and maternal investment
(Thorson, 1950; Vance, 1973; Laptikhovsky, 2006;
O’Connor, 2007). Additional latitudinal and longitu-
dinal sampling will allow us to ascertain if these vari-
ables function in similar ways at similar geographical
positions, or if other organism or ecosystem specific
variables play significant roles. For example, marine
habitats off the western coasts of North and South
America, and Northwestern and Southwestern
Afric­a all experience wind-driven coastal upwelling.
Comparing patterns of offspring size and maternal
investment within and among taxa from these four
areas would provide a strong test of whether pat-
terns of offspring size and maternal investment hold
across species found in similar marine ecosystems
at similar latitudes or if they differ longitudinally
or across taxa. Increasing depth may also yield pat-
terns of offspring size and maternal investment that
are similar to those found by latitude. Although the
available data to test this hypothesis are limited and
results appear to vary by taxa and location, there is
a trend toward the production of larger eggs with
increasing depth (Gage and Tyler, 1991; Auel, 2004).
Refinement of this hypothesis would require addi-
tional deep-sea research efforts to specifically exam-
ine reproductive biology. If these types of studies are
conducted within a comparative framework, taking
advantage of unique replicated environments, they
may provide for powerful tests of the biogeographi-
cal ideas presented here. Lastly, modern technologi-
cal advances in computing and social connectivity
make it feasible to establish online databases that
compile offspring size data that is collected and
uploaded directly by local researchers. Large-scale
projects of empirical data collection based on a
crowd-source model may be possible and may yield
unique insights into how reproductive patterns are
changing globally in our rapidly warmin­g world.
3.8 Summary
1. Parental investment varies dramatically in ma-
rine invertebrates and is largely unrelated to
phylogeny. Instead, strong biogeographical pat-
terns in parental investment exist with the larg-
est offspring being produced at the poles and the
smallest in the tropics.
2. Offspring size is a convenient proxy for parental
investment but uncertainty remains regarding
how completely offspring size captures total pa-
rental investment.
3. Some of the first life history models of offspring
size were developed for marine invertebrates,
and for over 40 years marine biologists have
sought to parameterize these models but key
gaps our understanding regarding the offspring
size-number trade-off remain.
4. Offspring size not only affects the larval period,
as was initially thought, but instead affects the
entire life history, from fertilization through to
post-metamorphic performance.
5. There is increasing evidence that mothers manip-
ulate the size and phenotype of their offspring in
response to local environmental variation in or-
der to maximize offspring fitness.
6. Despite decades of study, much of the world’s
life history variation remains unknown; instead,

we know a lot about only a few places. We also
have little understanding of the mechanisms by
which mothers manipulate the phenotype of off-
spring in order to maximize fitness.
Acknowledgments
JSM would like to thank Amy Moran, Bob Podolsky,
Jon Allen, Steve Stancyk and Richard Strathmann
for many insightful conversations on the ecological
and evolutionary implications of maternal provi-
sioning. AMR would like to thank the University
of North Carolina at Charlotte for support through
a Junior Faculty Development Award.
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 Ch a pt er 4
Evolutionary Transitions in Mode
of Development
Rachel Collin and Amy Moran
4.1 Introduction
Since the field of larval biology was first framed by
Thorson (1946; 1950), much attention has focused
on the strategies that marine invertebrates employ
to transform an egg into a juvenile. Because these
strategies can generally be placed into one of two
more-or-less discrete categories (planktotrophic and
lecithotrophic; see Table 4.1), a main line of inquiry
has been to understand the ecological and evolu-
tionary mechanisms underlying transitions be-
tween these strategies. A large body of literature has
focused on studies of the evolutionary origin, loss,
and potential regain of feeding larvae. In this chap-
ter, we argue that in the past, most of the focus has
typically been on the long view; that is, evolution of
complex larval traits is generally discussed in the
context of phylogenetic patterns that have formed
over long evolutionary timescales, and that are de-
tectable when making comparisons within families,
classes, or phyla. We increasingly use an analytical
approach incorporating comparative phylogenetics
to address these long-view questions. Such an ap-
proach is necessary for studies of the origin of larval
types and broad macroevolutionary patterns, but
will not be as effective for understanding the select-
ive forces causing evolutionary transitions in larval
feeding modes or the evolutionary processes that
facilitate or limit change. Here we discuss what has
been learned from taking a comparative phylogen-
etic approach and the limitations of this approach.
We propose that approaches based on a closer
view—analyses based on genetic, morphological,
and functional variation as well as selective forces
Collin, R. and Moran, A., Evolutionary transitions in mode of development. In. Evolutionary Ecology of Marine Invertebrate Larvae.
Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018).
© Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0004
that act on that variation among individuals, popu-
lations, or closely related congeners—are more ap-
propriate to answer questions about mechanisms
underlying the loss and regain of major complex
characters such as feeding larvae.
Many groups of marine invertebrates include
both species with planktotrophic larvae and species
that lack a feeding larval stage, and a few groups
also include unusual intermediates like faculta-
tive feeding larvae and polymorphic development
(Allen and Pernet, 2007; Collin, 2012; Table  4.1).
Nonplanktotrophic development has been vari-
ously subdivided in the literature, depending on
whether larvae are pelagic or spend part or all of
development in capsules, gel masses, or brooded;
however, a commonly listed dichotomy is between
planktotrophic and lecithotrophic (yolk-feeding)
development, where lecithotrophy encompasses
all nonplanktotrophic modes as well as nonfeeding
planktonic larvae. Planktonic larvae are free-living
animals with complex morphologies that are gen-
erally characteristic of a phylum, class, or family.
Feeding larvae may live for days, weeks, months,
or even years in the water column, whereas pe-
lagic lecithotrophic larvae generally spend hours to
days in the plankton (Olson, 1985; Strathmann and
­Strathmann, 2007; Neilson, this volume; Marshall
et al., this volume). While the evolutionary loss of
feeding is often the main focus of comparative stud-
ies (Strathmann, 1978; 1985; Wray, 1995; 1996; Rouse,
2000; Collin, 2004), the evolutionary loss of plankto-
trophic larvae entails not just the loss of feeding per
se, but also loss or modification of other characteris-
tics that are associated with planktotrophy such as
 E v o l u t i o n ary Tra n s i t i o n s i n M o d e o f De v e l o p me n t    51
Table 4.1  Summary of Terms as Used in This Contribution.
Term Explanation
Planktotrophic development Including a larval stage that swims
and feeds on plankton
Lecithotrophic development Development without feeding on
= nonfeeding development plankton
Lecithotrophic larva A free-living larva that does not
feed on plankton
Protected development Development that is maternally
protected or encapsulated and does
not include a planktonic larval stage
Embryo Any developmental stage before
hatching
swimming structures, anti-predatory defenses, ma-
ternal protection, maternal investment, and many
others (Strathmann, 1978; 1985), as well as likely
modification of the genetic architecture underlying
these traits (see Israel et al., 2016).
4.2  The Analytical Approach
Historical recognition of the gross morphological
similarity between feeding larvae in each phylum
or class, along with reduction in morphological
complexity in those species that lack a feeding lar-
val stage, has led to two main inferences from the
long view: (1) the evolutionary transition from spe-
cies with feeding to species with nonfeeding larval
stages has happened multiple times in diverse lin-
eages (Strathmann, 1978; 1985; Hart, 1996; Smith
et  al., 2007; Krug et  al., 2015), and (2) the evolu-
tionary reversal of this transition, or the regain of
feeding larvae, is either very rare or impossible. The
asymmetry in (2) has been attributed to Dollo’s Law,
which posits that complex characters that are not
used are quickly lost and cannot be regained (Hart,
2000; Smith et al., 2007; Collin and Miglietta, 2008).
It is indeed difficult to imagine how the profound
losses of characteristic larval structures in the most
derived types of lecithotrophic development could
be reversed to give rise again to the typical plank-
totrophic larval form for the group; in many taxa,
nonfeeding larvae or embryos with nonfeeding de-
velopment entirely lack the obvious structures used
for feeding by planktotrophic larvae of related spe-
cies. Examples of this include the loss of the long
ciliated arms that are key to particle capture in the
pluteus larva of planktotrophic echinoids and ophi-
uroids (Emlet, 1994; Hart, 2000; Selvakumaraswam­y
and Byrne, 2000; McEdward and Miner, 2001), loss
of the shell and feeding apparatus of the bryozoan
cyphonautes larva (Nielsen and Worsaae, 2010), and
loss of the pilidium body form and ciliated lateral
lobes necessary for particle capture in some lecitho-
trophic nemerteans (Maslakova and von Dassow,
2012; Martín-Durán et  al., 2015; Maslakova and
Hiebert, 2015). Many lecithotrophic larvae also lack
a complete digestive system or functional gut, and
in taxa that lack a free-living larval stage, the ability
to swim has also frequently been lost (Moran, 1999;
Chaparro et  al., 2002; Hofstee and Pernet, 2011).
Along with such dramatic morphological changes,
other obvious but equally essential feeding char-
acters are likely to be lost in many lecithotrophic
taxa, including sensory, behavioral, or physiological
pathways that facilitate feeding, though these types
of characters have received less attention.
Although it is indisputably difficult to imagine
evolutionary pathways that could lead to the reversal
of the loss of complex suites of larval feeding struc-
tures, this failure of our imaginations is likely a func-
tion of taking the long rather than the closer view
of the evolutionary dynamics of mode of develop-
ment. Here we discuss results of recent phylogenetic
studies that test the three major predictions derived
from the traditional long view that feeding larvae are
lost but not regained. We describe the strengths and
weaknesses of this approach for testing the general
theory of irreversibility of the loss of larval feeding
and other transitions in mode of development.
Prediction #1: Species lacking feeding larvae should
be nested within clades of species with feeding larvae;
species with nonfeeding development should not be re-
constructed as the ancestors of species with feeding
development.
Most types of feeding larvae (e.g., echinopluteus,
veliger, pilidium, actinotroch) are widespread in
their phylum or class and appear to have evolved
only once, and long ago (Strathmann, 1978; 1993).
However, phylogenetically widespread does not
necessarily equate to ancestral, and formal phyloge-
netic analyses followed by ancestral character-state
reconstructions are necessary to determine whether
52   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae

lecithotrophic species are nested within clades of
species with feeding larvae. Analytical methods of
ancestral trait reconstruction must then be applied
to assess the direction of character-state change.
Such approaches have been used in efforts to un-
ravel the evolution of mode of development within
a number of gastropod taxa; surprisingly, how-
ever, these analyses have been carried out in only
a handful of other taxonomic groups (Table  4.2).
Overall, these analyses show that ancestral states
can be reconstructed with high confidence if evolu-
tionary transitions are infrequent. For example, in a
molecular phylogeny of temnopleurid sea urchins,
the ancestral mode of development was recon-
structed as planktotrophic with complete certainty
(Jeffery et al., 2003). The results of this analysis were

Table 4.2  A Summary of Phylogenetic Studies Used to Document Evolution of Mode of Development.

Group Number of species Phylogenetic Conclusions of comparative Reference

included / included with coverage (total analysis
developmental data species in group)

Gastropods
Conus s.l. 70/60 8% (~750) Planktotrophy lost often and recently. Palumbi and
Duda 1999
Calyptraeids 72/72 51% (~140) Change too common to reconstruct Collin 2004
patterns with confidence.
Crepipatella 6/5 62% (8) Planktotrophy regained once. Collin et al. 2007
Sacoglossans 202/113 39% (~290) Planktotrophy lost; often not regained. Krug et al. 2015
Muricids 45/45 3% (~1600) Planktotrophy evolved three times. Pappalardo et al. 2014
Nucella 9/8 80% (10) Transitions between lecithotrophy with Marko et al. 2014
large eggs and with nurse eggs are
common.
Phestilla 6/4 100% (4) No formal analysis; plantktotrophy Faucci et al. 2007
regained twice.
Bivalves
Lasaea 27/27# 100% (23) Three independent losses of Li and Ó Foighil 2016
planktotrophy; no regains.
Ostreidae 21/21 38% (~55) Maternal care evolved once and is Ó Foighil and
ancestral for the only lecithotrophic Taylor 2000
species.
Echinoderms
Asterinids 28/150 19% (~150) Results vary with model parameters. Keever and Hart 2008
Macrophiothrix 15/9 20% (45) No formal analysis; change is frequent. Hart and
Podolsky 2005
Temnopleurids 24/24 39% (61) Planktotrophy is lost once; change is rare. Jeffery et al. 2003
Bryozoans
Gymnolaemate 48/43 <1% (>5000) Ancestral state is ambiguous with Waeschenbach
bryozoans parsimony reconstruction. et al. 2012
Ascidians
Styelidae & 45/45 10% (~400) Swimming has been lost multiple times, Maliska et al. 2013
Molgulidae but cannot be regained.

*Phylogenetic coverage is rough estimate, calculated as the number of species with developmental data in the phylogeny divided by the total number of species in the
group (obtained from the cited reference or from WoRMS).
#Number of asexual lineages. Since lineages may not coincide with named species, this could result in an upward bias in taxonomic coverage.
 E v o l u t i o n ary Tra n s i t i o n s i n M o d e o f De v e l o p me n t    53
clear because there was only a single loss of feeding
in the phylogeny, and all descendants of that node
were also nonfeeding (Jeffery et  al., 2003). Ances-
tral states can also be reconstructed with high con-
fidence when one state substantially outnumbers
the other, as occurs in plakobranchacean sea slugs
(Krug et  al., 2015). In this clade, 21 of 62 taxa for
which developmental information is available are
lecithotrophic. An evolutionary quantitative genet-
ics model reconstructed ancestral character states
as planktotrophic at many internal nodes with pos-
terior probabilities greater than 95% (Krug et  al.,
2015). In contrast, ancestral states in the sister group,
Oxynoace­ a, which includes seven lecithotrophic
species and nine planktotrophic species, could not
be reconstructed with any certainty; both states
were equally likely at many nodes (Krug et  al.,
2015). Analyses of other groups in which mode of
development is highly variable have also generally
failed to assign ancestral character states with con-
fidence, including calyptraeid gastropods (Colli­n,
2004), muricid gastropods (Pappalardo et al., 2014;
in their Figure  4.2), bryozoans (Waeschenbach
et  al., 2012), and asterinid sea stars (Keever and
Hart, 2008). Recent analysis of continuous charac-
ters (e.g., egg size and number) suggests that use
of continuous characters and the inclusion of other
life history traits that might promote or constrain
evolutionary transitions in the focal character may
be powerful aids to reaching a fuller understanding
of the subtleties of the evolution of development
(Marko et al., 2014; Krug et al., 2015).
Prediction #2: Reconstructed rates of transitions between
modes of development should be highly asymmetrical.
If (as seems likely) evolutionary transitions from
planktotrophy to lecithotrophy are more probable
than reversals, or if reversals are impossible, spe-
cific patterns should be recovered by analyses of
character-state evolution. Support for irreversibility
could be found in two aspects of a given analysis.
First, likelihood and Bayesian models of character
evolution provide a platform to determine if the
data are best fit by a single-rate or a two-rate model.
A single-rate model is favored when the data do not
support different rates of transitions and their re-
versals. When the data favor unequal rates of tran-
sitions and reversals, a two-rate model is favored.
Irreversibility would be supported if the estimated
rate of transitions far exceeds the rate or reversals or
if the rate of reversals is estimated as zero. Second,
given a particular distribution of traits on a phylog-
eny, likelihood ratio tests can be used to compare
the fit of different models of evolution between an
irreversible model and an unconstrained model that
allows both transitions and reversals. Phylogenetic
analyses that have taken these approaches have gen-
erated a variety of estimates of rates of transitions
and reversals and may also produce results that
seem contradictory. In temnopleurid sea urchins,
the group in which a clade of lecithotrophs is nested
within a clade of pure planktotrophs, the one-rate
model fits significantly better than a two-rate model,
showing that the null hypothesis that losses and re-
gains are equally likely cannot be rejected (­Jeffery
et  al., 2003). In contrast, the irreversible model of
character change fit the data significantly better
than the unconstrained model (as only one transi-
tion and no reversals were evident), providing sup-
port for the idea that reversals do not occur (Jeffery
et  al., 2003). These different results may be due to
the small number of changes, limiting the power of
the test to reject the single-rate model. However, in
a larger phylogeny of calyptraeid gastropods, the
single-rate model was also the best fit for both gains
and losses of planktotrophy and gains and losses of
the ability to swim (Collin, 2004).
Mode of development may show patterns of phy-
logenetic correlation with other features like extra-
cellular yolk (ECY) (Krug et al., 2015) or soft vs. hard
substrate (Pappalardo et al., 2014). For ­example, in
muricid gastropods the estimated rates of change
between pelagic and benthic development were
virtually zero in both directions for species living
on hard bottoms, but estimated transitions from
benthic to pelagic development were significantly
greater than transition to benthic development in
species living on soft bottoms ­(Pappalardo et  al.,
2014). Unfortunately, scant taxonomic sampling
makes these results difficult to interpret. A study
of sacoglossan sea slugs with substantially more
thorough taxonomic sampling showed that mod-
els allowing covariance of lecithotrophy and ECY
were highly preferred over those with independ-
ent evolution of the two traits: in the presence of
ECY, lecithotrophy was twice as likely to evolve as
54   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
it was in lineages lacking ECY (Krug et  al., 2015).
Likewise, in Nucella whelks, egg size and relative
number of nurse eggs are significantly evolutionar-
ily correlated (Marko et  al., 2014). Nurse eggs are
estimated to have been lost twice and significantly
reduced in two other lineages; however, no esti-
mates of the confidence in these reconstructions are
reported (Marko et al., 2014).
Surprisingly, several phylogenies provide some
support for a reversal: the evolution of planktotro-
phy from lecithotrophic ancestors. In two genera of
calyptraeid gastropods there is clear evidence that
planktotrophic species are nested deeply within
clades of lecithotrophic species, and a likelihood
reconstruction of the character states at specific
nodes supports the re-evolution of feeding larvae
in one genus (Collin et  al., 2007), while a formal
analysis has not been conducted on the other ge-
nus ­(Collin, 2005). A similar result was obtained by
Faucci et al. (2007) who observed that two species of
planktotrophic Phestilla were nested within a clade
of lecithotrophic taxa, although the small num-
ber of species in this genus suggests that a formal
analysis may not resolve the ancestral states with
any confidence. Notably, in both of these analyses,
lecithotrophic species were more numerous than
planktotrophs. In the sacoglossan sea slugs and Co-
nus (Palumbi and Duda, 1999), where planktotrophs
predominate, no patterns of reversals are recovered.
It is not clear if this represents an analytical artifact
or a difference between the mechanisms driving life
history evolution in these taxa.
Prediction #3: Species with lecithotrophic development
should lose the structures used for feeding, but retain
vestigial features that indicate descent from plankto-
trophic ancestors.
Overall, the literature shows strong support for
this prediction. Numerous studies of comparative
embryology and larval biology show that charac-
teristic features of feeding larvae are reduced or
(incompletely) lost in species with lecithotrophic
development. Features used for swimming are also
lost in species with benthic development. However,
groups vary considerably in the degree to which
larval features are lost, and this may in turn influ-
ence the probability of regaining feeding in lecitho-
trophic lineages (Collin and Miglietta, 2008).
The most well-studied examples of feeding-­
structure loss come from sea urchins. The character-
istic larval form of sea urchins, the echinopluteus,
is conserved across all echinoid families, suggest-
ing this is a plesiomorphic character for the group.
In lecithotrophic species, larvae often lack arms or
guts (­Emlet, 1994; McEdward and Miner, 2001).
However, on close examination, vestiges of feed-
ing structures such as arm spicules are present even
in the most derived lecithotrophic echinoid larvae
(Emlet, 1995). This is also the case in other classes
of echinoderms such as asteroids and ophiuroids,
where some lecithotrophic larvae have retained
many features of planktotrophs but others are so
altered as to be almost unrecognizable (McEdward
and Miner, 2001; Selvakumaraswamy and Byrne,
2004; Byrne, 2006). In a larger but less thoroughly
studied group, the gastropods, the characteristic
feeding structure is the velum. The velum is gen-
erally present in lecithotrophic larvae, probably
because it retains a swimming function, although it
tends to be smaller and less elaborate than in plank-
totrophs (Moran, 1999). The velum is also retained
in many species with encapsulated lecithotrophic
development (Moran, 1999; Chaparro et  al., 2002),
but in other species, all outwardly visible vestiges
of the velum are lost (e.g., Collin, 2000; Figure 4.1).
Similarly, in lecithotrophic larvae of bryozoans and
nemerteans, the characteristic feeding structures of
the planktotrophic larva are absent, and in many of
these lecithotrophic taxa, the larva bears little re-
semblance to the planktotrophic form (Zimmer and
Woollacott, 1977; Maslakova and Hiebert, 2015).
4.3  Limitations of the Long View
There are a number of challenges to taking the broad
approach to studying evolutionary transitions in
mode of development. For transitions that likely hap-
pened many tens or hundreds of million years ago,
such as the presumed loss of feeding larvae by an
entire class of echinoderms (Crinoidea; McEdward
and Miner, 2001), these challenges are perhaps in-
surmountable due to the difficulty in reconstructing
ancient events. However, for more recent (but still
long-view) transitions, impediments to inferring an-
cestral states fall into two main groups: (1) the fact that
phylogenies of many (most) marine invertebrate taxa
 E v o l u t i o n ary Tra n s i t i o n s i n M o d e o f De v e l o p me n t    55
(A) (B)
s s
(C) (D)
n v
are woefully incomplete, and (2) shortcomings inher-
ent to most methods of character-state reconstruction.
Comparative data from a growing number of gen-
era or families indicate that in at least some cases,
transitions in mode of development occur often and
rapidly. This means that the results of comparative
phylogenetic approaches could be significantly im-
pacted by taxa that are missing from the phylogeny,
or for which mode of development is not known.
Only a small number of published analyses include
50% or more of the currently recognized species in
the genus or family under study (Table 4.2). Under-
standably, studies with dense taxonomic coverage
of extant species are those focused on small taxa
(e.g., a genus with only a handful of species; Marko
et al., 2014; Faucci et al., 2007), and this may limit
both the power of the analysis and the opportunity
to capture rare evolutionary events. Even if all liv-
ing species in a taxon were included in a phylogeny
and the developmental mode was known for all
the species, comparative phylogenetic studies may
still be compromised by incomplete taxon sampling
due to loss of species through extinction. In some
n
Figure  4.1 Lecithotrophic modifications of
embryos in three adelphophagic calyptraeid
species. A. Mid-stage embryo of Crepidula
coquimbensis lacks a velum entirely.
(E) n The velum would be clearly visible in the
same stage of an embryo of species with
planktotrophic development. B. Late-stage
embryo of Bostrycapulus odites, which retains
a small but distinct velum. C. Early stage of
C. coquimbensis at which the embryo has
already begun to ingest siblings. D. Early-
stage C. coquimbensis embryo ingesting a
sibling nurse embryo. E. A mid-stage embryo
of Crepipatella dilatata using the velum to
s position a nurse egg for ingestion. v = velum;
s = shell, n = nurse egg/embryo.
cases (e.g., bivalves, shelled gastropods, some echi-
noids, brachiopods, and bryozoans) it is possible to
determine developmental mode from fossil mor-
phology, but this is generally not the case. It is also
difficult to include fossil taxa with any confidence
in phylogenies of extant taxa generated from DNA
sequence data, and it is not generally possible to
generate well-estimate­d branch lengths with fossil
data, as is required for most methods of ancestral
state reconstruction. Overall, because sampling
needs to be dense relative to the events of interest
(transitions in mode of development), the compara-
tive phylogenetic approach may be useful in groups
where evolution in mode of development is slow
and changes occur infrequently. To be effective in
taxa where changes in mode of development are
rampant, extremely dense taxon sampling of recent
clades where extinction is thought to be minimal is
likely the best approach.
Methods for reconstructing character-state evo-
lution have come a long way in the last 20 years.
In particular, current methods have made sub-
stantial progress toward addressing the circularity
56   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
inherent in early attempts at using parsimony to
describe asymmetry in frequencies of transitions
and reversals (Strathmann and Eernisse, 1994;
­Cunningham et al., 1998; Cunningham, 1999; Collin
and Migliett­a, 2008; Keever and Hart, 2008). How-
ever, modern methods and the ways they are used
to examine character evolution still make assump-
tions about the pattern of character-state evolution
and are subject to bias from phylogenetic uncer-
tainty (Duchêne and Lanfear, 2015). A clear example
of the importance of model selection: Keever and
Hart (2008) used asterinid sea star phylogenies to
illustrate the importance of differentiating between
(1) the use of phylogenetic comparative analyses to
determine which model of character evolution is
supported by the data and (2) their use to recon-
struct ancestral states at specific nodes. They clearly
demonstrated that using a model based on our heu-
ristic understanding of larval biology (incorporat-
ing the idea that characteristic larval morphologies
may be difficult for lecithotrophs to regain) pro-
duces different results from application of a model
with unconstrained rates. Reconstructions of char-
acter evolution in sacoglossans also depended on
the model selected; the unconstrained model esti-
mated the rate for loss of planktotrophy as lower
than the rate of reversals, a result considered to be
biologically unreasonable by Krug et  al. (2015). In
addition, both rates were very high. A model that
prohibited reversals estimated the rates of losses as
about 1/10 of those estimated by the unconstrained
model, clearly showing the impact of model choice
on both the estimated rates and directions of change
(Figure 5 in Krug et al., 2015).
Researchers agree that accurate reconstruction
of ancestral developmental mode requires a model
of evolution that is corroborated with independent
traits, but anything more than the simplest models
have yet to be developed (Collin, 2007; Keever and
Hart, 2008; Krug et al., 2015; see following). Newly
developed evolutionary quantitative genetic models
may be more appropriate than discrete-state mod-
els for traits like mode of development because the
longer a lineage drifts away from the character-state
transition, the less likely reversals become (Revell,
2014; Krug et  al., 2015). This matches our under-
standing that the likelihood of reversals decreases
following the loss of feeding larval development,
although it remains to be seen how accurately this
approach models life history evolution.
Unfortunately, when more sophisticated models
are developed, more sophisticated inputs are often
necessary and more sophisticated problems arise.
For example, a recent phylogenetic analysis of sa-
coglossans provided support for state-specific spe-
ciation and extinction rates. Such differences in rate
could challenge estimates of character-state transi-
tions. If speciation and extinction rates depend on
the mode of development such that the derived state
(lecithotrophy) has a lower diversification rate than
the ancestral state, the phylogeny is likely to lack old
lecithotrophic lineages. This pattern could bias esti-
mated reversal rates upwards (Krug et al., 2015). Be-
cause differences in life history characters are thought
to alter dispersal and gene flow and therefore impact
speciation and extinction probabilities, state-specific
extinction and speciation rates could potentially oc-
cur in many or most clades of marine invertebrates.
Summing up the long-view evidence, both the
majority of the literature and common sense dictate
that lecithotrophic taxa are generally derived from
planktotrophic ancestors, and reversals are rare
but perhaps not impossible. Supporting evidence
comes largely from homologous feeding struc-
tures that are found across classes and from phy-
logenies where transitions are relatively rare and
lecithotrophic development is uncommon. In these
latter cases results are likely to be robust to the
choice of model selection. However, comparative
analyses of deeply intriguing taxa such as Patiriella,
Macrophiothrix, and calyptraeids—where no one
feeding mode dominates and transitions may be
frequent—fail to provide support for the accepted
view. Unfortunately, comparative methods are not
yet likely to provide satisfying answers about evo-
lutionary transitions in what are arguably the most
interesting taxa: groups where life history charac-
ters evolve rapidly and transitions are common.
4.4  Future Directions and Unanswered
Questions for Analytical Approaches
The development of more sophisticated meth-
ods for comparative analyses now allows some
longstanding questions in larval biology to be
 E v o l u t i o n ary Tra n s i t i o n s i n M o d e o f De v e l o p me n t    57
addressed. First, it has been widely accepted that
(1) species with lecithotrophic development have
lower levels of dispersal and gene flow than do
planktotrophic species (Collin, 2001; Selkoe and
Toonen, 2011; Ellingso­n and Krug, 2015), and that
(2) this results in higher rates of speciation and/
or extinction in lecithotrophic species than in
planktotrophic species (Hansen, 1980; Jablonski
and Lutz, 1983; Marko and Moran, 2009). How-
ever, two recent comparative analyses testing for
state-specific rates of diversification (speciation-
extinction), have shown that diversification rates
are lower in species with development resulting
in lower dispersal. In sacoglossan­s, lecithotrophic
slugs have lower diversification rates compared
to planktotrophs (Krug et al., 2015). In fact, higher
extinction rates lower the net diversity of lecitho-
trophs despite the fact that lecithotrophy originates
frequently. Krug et al. (2015) suggest that this pat-
tern could explain the long-term maintenance of
planktotrophy in clades despite the fact that loss of
planktotrophy is a frequent event. Species selection
against a trait that reduces dispersal has also been
demonstrated in ascidians, where speciation rate
is higher and range size is larger in species with
tailed tadpoles than in those whose development
lacks a tailed swimming larva (Maliska et al., 2013).
The apparent discordance between results from
fossils, population genetics, and comparative phy-
logenetics needs to be investigated in more detail.
It is possible that this discordance is due to specific
features of the biology of sacoglossans (many are
obligate specialist herbivores) or tunicates (many
are fouling organisms subject to rafting). Clearly,
a concerted effort must be made to generate large,
densely sampled phylogenies, especially for taxa
with extensive background data on population ge-
netics and larval biology, so that similar tests on
other taxa can be performed.
Second, in order for phylogenetic models to per-
form well, it is necessary to have well-­characterized
traits. It may be both appealing and ecologically
relevant to dichotomously classify development
as planktotrophic vs. lecithotrophic, pelagic vs.
benthic, or brooded vs. broadcasted, but a poten-
tial downfall of using discrete classification is that
it can become implicit in the phylogenetic method
that the states are homologous. For  example,
gastropods that develop without a feeding larval
stage from large eggs and those that develop from
small eggs with embryos that consume each other
would both be coded as “nonplanktotrophic.”
Therefore, use of continuous measures like egg
size or nurse egg number (Marko et  al., 2014;
Krug, 2015), and more nuanced descriptions of
development that separate modes of development
into multiple discrete states (Collin, 2004; Keever
and Hart, 2008) are likely to produce more robust
results. In addition, simultaneous analyses of cor-
related characters or characters that may influence
or constrain the trait of interest may significantly
improve results (Ó Foighil and Taylor, 2000; Krug
et al., 2015). Therefore, as much care should be put
into coding characters—­including the most appro-
priate traits as correlates—as is put into the analy-
ses themselves.
Finally, questions about whether shifts in mode
of development are associated with the process
of speciation, and if shifts occur during gradual
change in a species or divergence in isolated pe-
ripheral populations, are also understudied. Since
closely related sister species often differ in mode
of development in speciose clades, it is possible
that changes in mode of development contribute
to speciation. The largest study to explicitly ad-
dress this question across a deep phylogeny found
that models allowing divergence in mode of de-
velopment at nodes as well as along branches did
not perform any better than a model that allowed
change only along branches (Krug et  al., 2015),
though a model where change occurred only at
nodes was not tested. Further detailed study of
populations of slugs in the process of diverging
suggested that reduction or loss of dispersal asso-
ciated with a shift toward aplanktic development
could increase local recruitment and inbreeding,
leading to the evolution of reproductive isolation
(Ellingson and Krug, 2015). More detailed studies
of this kind are necessary to assess this possibil-
ity. Detailed genome scans of closely related spe-
cies are now possible for non-model taxa, and are
likely to provide exciting new insights into what
genes are under selection in diverging or recently
diverged taxa, potentially clarifying links between
shifts in developmental mode and speciation
(Zaka­s and Rockman, 2015).
58   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
4.5  A Closer View
We argue that unlike the deep questions surround-
ing the origin of larval feeding modes in large
clades such as echinoids, gastropods, bryozoans,
etc., which have the potential to be answered by
combining data from phylogenies, the fossil record,
and morphological evidence from modern taxa,
many questions about life history transitions are
essentially shallow ones that hinge on microevo-
lutionary processes. If we are really to understand
the causes and processes involved in transitions
between modes of development, it is important to
use what we have learned from broad compara-
tive analyses to look closely at divergence in action.
Ideally, the broadly comparative analytical and the
more detailed descriptive views can be reciprocally
illuminating: phylogenetic analyses can help iden-
tify species or small sub-clades that are of particu-
lar interest, and detailed analyses at the population
or species level can provide a close-up view of the
underlying genetic, morphological, and functional
variation, and selective forces that act on that vari-
ation. In the rest of this chapter we list some major
unanswered questions that we feel would benefit
from greater research emphasis on closer detailed
descriptions and microevolutionary approaches.
Question #1: Does the order of transitional steps in the
evolution of direct development differ among taxa, and
what selective forces act along the way?
When using modern comparative phylogenetic
methods to understand character-state evolution,
two key factors are coding the characters and choos-
ing a model of character-state transformation. At
the most basic this includes the null model of losses
and reversals being equally likely vs. the hypothesis
that losses are irreversible. If there is a continuum of
character states (e.g., egg size) along which plank-
totrophic species must move in order to eventually
make the switch to lecithotrophy (e.g., brittle stars
which may include an ophiopluteus which may
or may not feed and/or a distinct vitellaria stage;
McEdward and Miner, 2001; Selvakumaraswamy
and Byrne, 2004), or if species find alternative solu-
tions to the same problem (e.g., gastropods where
lecithotrophic development can proceed from large
eggs or from small eggs with adelphophagy), the
way the characters are scored can significantly im-
pact the conclusions of the study (e.g., Collin, 2004;
Keever and Hart, 2008). Once the characters are
coded, comparative methods can be used to deter-
mine the rates of c­haracter-state transformations
that are best supported by the data (phylogeny and
the character states of the operational taxonomic
unit (OTUs)). However, this approach ignores any
independent knowledge we have about the likeli-
hood of the transitions, and when this information
is included in a crude way, the results of the compar-
ative analyses often differ (e.g., Krug et al., 2015).
Computational biologists are constantly working
to refine the way independent data can be incor-
porated into such analyses, but larval and devel-
opmental biologists need to put effort into refining
evidence and developing formal models of how
evolutionary transitions among modes of develop-
ment work. This includes articulating explicit mod-
els of how different developmental morphologies
are related to each other, postulating the selective
forces that are acting most strongly on different
kinds of development, and developing hypotheses
about the selective factors that could drive evolu-
tionary change. At this point heuristic models have
been developed for sea urchins (Smith et al., 2007)
and sea stars (Keever and Hart, 2008). The overall
paradigm based on these echinoderms roughly
posits that changes occur in this order: (1) energy
content of eggs increases; (2) development to meta-
morphosis accelerates; (3) the requirement/ability
to feed is lost; (4) time to metamorphosis is short-
ened by additional reduction of larval structures
(Smith et al., 2007). In order to adapt this heuristic
model of transitions between modes of develop-
ment to other taxa, features relevant to those taxa
need to be considered. For many groups these in-
clude encapsulation or maternal protection and
extraembryonic nutrition (extracellular yolk; nurse
eggs; matrotrophy). Some differences in the factors
that are most likely under selection during differ-
ent stages and kinds of development are presented
in Figure 4.2. It is important to emphasize that few
detailed datasets have been developed to describe
these steps and that heuristic schemes describing
evolutionary trajectories are generally based on log-
ical interpretations of known larval morphologies
(McEdward and Janies, 1993; McEdward and Miner,
 E v o l u t i o n ary Tra n s i t i o n s i n M o d e o f De v e l o p me n t    59

An integrative model of transitions Planktotrophy Lecithotrophy

EEN = Extraembryonic nutrition Defenses/stress tolerance Defenses/stress tolerance
Rapid development Rapid development of Rapid development to
to larva juvenile juvenile
Stress tolerance Settlement Settlement
Swimming Swimming
Feeding

Free-living
embryo

Free-living
larva

Protected
Mother embryo, Juvenile
no EEn

Gas exchange
Stress tolerance
Hatching

Egg number Protetcted

Egg size embryo,
Egg content EEN
Maternal care
Fertilization

Gas exchange
Stress tolerance
Consuming EEN
Outcompeting siblings
Hatching

Figure 4.2  Alternative pathways of larval development from egg to juvenile and the major features under selection at each stage. Defining or unique
selective forces for each stage are indicated in bold.

2001; Smith et  al., 2007). Larry McEdward was a
major proponent of this approach (McEdwar­d and
Janies, 1993; McEdward and Miner, 2001) and his
heuristic models still form an important framework
for explicit hypothesis testing. Similar detailed ef-
forts are needed to provide clear models for non-
echinoderm taxa that could help to inform more
sophisticated comparative analyses.
Question #2: What features other than gross larval
morphology change during transitions in mode of
development?
The major focus of marine invertebrate biolo-
gists working on the evolutionary transitions be-
tween modes of development has been the gross
morphology of the embryos and larvae, and in par-
ticular the structures used for feeding and swim-
ming. However, changes in numerous other features
may occur either via genetic drift or through active
selection for alternate function. Understanding
these may help further refine our view of how se-
lection acts on small details of development and
ultimately produces transitions between modes of
development. While egg size has long been a focus
of life history studies, one related character that has
received considerable attention in recent years is
the biochemical composition of the egg.
The major constituents of eggs are protein, lipid,
and carbohydrate, and the proportion and type of
these constituents is correlated with developmental
60   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
mode in what is generally interpreted as an adap-
tive way. As one broad example, the eggs of lecitho-
trophic species generally contain proportionally
more lipid, which is thought to provide energy
for the metamorph as well as the nonfeeding larva
(Emlet and Hoegh-Guldberg, 1997; Mora­ n and
Manahan, 2003; Prowse et  al., 2008; Moran et  al.,
2013; Falkner et  al., 2015) and has also been im-
plicated in buoyancy (Arai et  al., 1993). Feeding
larvae, in contrast, contain proportionally more
protein and build lipid reserves through feeding
(Jaeckle, 1995; Moran and Manahan, 2004; Sewell,
2005; Moran et al., 2013). Lipid profiles also differ
by taxon and mode of development; for example,
in asterinid sea stars, triglycerides are relatively
more abundant in eggs of most lecithotrophic spe-
cies than in planktotrophic species (Prowse et  al.,
2008), while some lecithotrophic ophiuroids sup-
ply eggs with wax esters, a lipid class not found
in planktotrophs (Falkner et  al., 2015). As another
example, thyroid hormones are required for meta-
morphosis in many echinoderms (Heyland et  al.,
2004). For planktotrophic species, these hormones
are acquired through feeding; in lecithotrophs, ex-
ogenous acquisition has been replaced by endog-
enous production (Heyland et al., 2004; Armstrong
and Lessios, 2015).
Other changes we might expect with the switch
to lecithotrophy are the loss of larval sensory struc-
tures and threat responses in species with protected
development. Species lacking free-living larval
stages may also show loss or reduction of the path-
ways associated with triggers for settlement and
metamorphosis (see Section  3 of this volume), as
well as changes in the genetic architecture, maternal
provisioning of mRNA, cell fates, and other compo-
nents of development. For example, in Heliocidaris
the animal-vegetal axis is maternally specified in
the planktotrophic species, while in the lecitho-
trophic species the dorso-ventral axis is also mater-
nally specified (Henry et al., 1990). Clearly, the loss
of feeding larvae is accompanied not just by mor-
phological changes to larvae, but also by more sub-
tle physiological changes, as well as alterations to
gene expression, maternal oogenetic pathways, and
maternal reproductive morphology. The evolution-
ary lability of these less visible features has rarely
been explored, but they may play a major role in
determining the probability and direction of transi-
tions in life history characters.
Question #3: If planktotrophy were to re-evolve, how
would it happen and how could we recognize it?
Thought exercises exploring possible routes to the
re-evolution of feeding larvae have been performed
over deep phylogenies (Strathmann, 1978). Two
primary routes have been considered: larvae of a
non-planktotrophic species could regain function of
vestigial feeding structures, or larvae could regain
feeding through co-option of juvenile or adult feed-
ing structures. For many taxa, the striking reduction
or loss of larval feeding structures suggests that co-
option of juvenile features may be the only option
available for re-evolving feeding larvae. This may
have occurred in the ancestors of modern clades of
inarticulate brachiopods, cerianthid cnidarians, and
cephalopods, all of which have feeding larvae which
are strikingly similar to their juveniles. This option,
however, is not equally open to all taxa; for exam-
ple, in echinoderms the juvenile feeding structures
appear to be poorly suited for planktonic feeding
(Strathmann, 1978). It may be noteworthy that no
cases of the recent regain of feeding via “larvaliza-
tion” of the juvenile has been reported in individual
species or genera; however, one related example has
been described in Pteraster tessulatus, in which non-
feeding larvalization has occurred through the re-
cent evolution of a swimming juvenile (McEdwar­d
and Janies, 1993). Closer examination of these rare
cases could provide important insight into the pro-
cesses involved in transitions of early life history
stages between the benthos and the plankton.
Re-evolution of planktotrophy may also happen
through regain of function of vestigial feeding struc-
tures (Strathmann, 1978). It is intuitively apparent
that the probability of regaining function would be
negatively correlated with the degree of loss; thus,
a facultatively feeding larva that is functionally lec-
ithotrophic but can still feed would much more eas-
ily return to the planktotrophic state than a species
whose lecithotrophic larvae have lost feeding struc-
tures and a functional gut (Strathmann, 1978; 1985).
Similarly, encapsulated embryos that retain the lar-
val structures and their function may facilitate re-
evolution of larval feeding (Collin, 2004; Collin and
Miglietta, 2008). In this case, however, it would be
 E v o l u t i o n ary Tra n s i t i o n s i n M o d e o f De v e l o p me n t    61
very difficult to distinguish a species that had tran-
sitioned back to planktotrophy based on morphol-
ogy alone (McEdward and Janies, 1997).
A number of lecithotrophic species do seem to
retain the unused potential for larval feeding and
swimming (Hookham and Page, 2016). For exam-
ple, embryos of many calyptraeid gastropods retain
the ability to capture and ingest particles (Chaparro
et al., 2002; Collin, 2004; Collin, unpublished data).
Likewise, a number of taxa have recently been
demonstrated to possess facultatively feeding lar-
vae (Allen and Pernet, 2007). What, then, is needed
to identify species that may have reverted back to
planktotrophy from such a state?
We know of no example from the marine inverte-
brate larval literature, but one example from deep-sea
worms illuminates how phylogenetic and morpho-
logical analyses can be used in concert to identify re-
versals of past changes to complex life history traits.
Working with the siboglinid genus Osedax, a group
of saprotrophic worms in which males of most spe-
cies show extreme dwarfism as parasites on females,
Rouse et al. (2015) demonstrated an evolutionary re-
version from dwarf parasitic males (ancestral in this
clade) to free-living full-sized males. The evidence
came from phylogenetic analyses, in which a spe-
cies with free-living non-dwarf males was nested
within a clade of species with parasitic dwarf males,
and from retention of the dwarf-style testicular
structure on the males of the species that re-evolved
large body size. A search for similar relic markers
for direct development or lecithotrophy could help
identify potentially re-evolved larvae that could be
the focus of further study. For example, neutral li-
pid classes that are not present in eggs of plankto-
trophic ophiuroids occur in eggs of lecithotrophic
species (Falkner et  al., 2015); if a planktotroph’s
eggs were found to contain these “lecithotrophic”
lipids, this could provide independent evidence for
a phylogenetically based hypothesis for reversal
(Keever and Hart, 2008), or could indicate a taxon
in the early stages of transition. Similarly, in gastro-
pods, the retention of globose protoconch shapes, or
protoconchs lacking characteristic planktotrophic
sculptures, may be evidence of reversions to plank-
totrophy (Reid, 1989). Finally, it would be interest-
ing to know if it is possible to detect transitions in
mode of development in taxa with morphologically
simple larvae. For example, while many cnidarian
larvae are morphologically similar, they vary sig-
nificantly in their duration in the plankton as well as
their reliance on yolk reserves, autotrophy, parasit-
ism, and larval feeding (Baird et  al., 2009). Due to
their simple forms, these larvae have been largely
overlooked in discussions of evolutionary transi-
tions in mode of development, but they may be a
taxon ripe for study.
Question #4: How do mode of metamorphosis and ma-
ternal protection influence the evolution of mode of
development?
Three different factors may influence the modifi-
cation of lecithotrophic developers compared to
their planktotrophic ancestors: (1) natural selec-
tion for better performance in larval functions (e.g.,
swimming, settlement site selection) results in con-
vergence on ball-like morphology and specific ar-
rangements of ciliary bands; (2) natural selection for
faster development to the juvenile stage selects for
earlier allocation to juvenile structures and reduc-
tion of specific larval features; and (3) genetic drift
in the genes underlying unused structures and func-
tions. For most species with large population sizes,
selection would be expected to act more quickly
than genetic drift. The relative importance of these
factors has been discussed for echinoid models, but
they may be quite different in other taxa.
Echinoids are the model on which much of our
current understanding has been built, as plankto-
trophs have unprotected, maximally indirect devel-
opment. In species with feeding larvae the juvenile
grows in a small pocket inside the larva. At meta-
morphosis the juvenile emerges from the pocket
and the vast majority of the larval body is discarded.
This suggests that the larval and juvenile bodies are
largely independent and should be free to evolve
independently. Therefore, if the larva no longer
needs to feed, the developing juvenile is probably
not strongly impacted by a reduction in the larval
arms or rearrangement of the ciliary bands, and
the gross morphology of the larva could be altered
drastically. In fact, selection may act on a number
of facets of development to speed the loss of larval
features once the transition is underway, including
selection for better swimming performance (Emlet,
1991; 1994) and selection for faster development
62   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
of the juvenile (Raff and Byrne, 2006; Smith et  al.,
2007). Other larvae with similar development of the
juvenile within but largely distinct from the larval
body include the pilidium larva of nemerteans and
mitraria of oweniid polychaetes. In nemerteans, lec-
ithotrophic pilidia tend to converge on ciliated balls
similar to those predicted to perform well at swim-
ming (Emlet, 1991; Maslakova and Hiebert, 2014).
Likewise, in lecithotrophic larvae of bryozoan­ s structures—or whether these features are simply not
and hemichordates, two other groups in which
metamorphosis of a planktotrophic larva involves a
drastic change in body plan, the lecithotrophic lar-
vae appear to be optimized for swimming.
In contrast, larvae of a number of other groups
have a body plan very similar to that of the juvenile.
These include most polychaetes, gastropods, and
polyclad flatworms in which the larval body is al-
most entirely the same as the juvenile body, but with
the addition of ciliated bands and projections used
for feeding and swimming. In these groups lecitho-
trophic developers do not evolve into yolky ciliated
balls, and they generally retain the body form that is
shared between the larva and the juvenile. In these
cases natural selection for rapid development of the
juvenile would not be expected to modify the body
plan other than to perhaps reduce the size or com-
plexity of the specific larval structures. In addition, in
many of these groups the embryos are encapsulated,
and it appears unlikely that natural selection would
act to modify them for swimming. However, in
many cases aspects of the encapsulated environment
may act to retain larval structures (Figure 4.1); for ex-
ample, the velum of gastropod larvae has a feeding
function in many species where nutrition is extraem-
bryonic (e.g., adelphophagy) and species may be
selected for better and faster consumption of nurse
eggs, extraembryonic yolk, or albumen (reviewe­d in
Moran, 1999). The velum (or ciliary bands in other
taxa) may also serve encapsulated embryos by stir-
ring capsule fluid and providing a large ciliated
surface for enhancing gas exchange (Hunter and
Vogel, 1986; Moran, 1999; Hofstee and Pernet, 2011).
The intracapsular environment may in some cases
have selected for dramatic modification to the ve-
liger form; for example, inter-sibling competition for
early nurse egg consumption in adelophophagic em-
bryos of Searlesia dira, and Buccinum undatum results
in embryos that are bags of yolky eggs and hardly
resemble early development of other gastropods at
all (Rivest, 1983; Smith and Thatje, 2013; Figure 4.2).
In general, however, in groups where the larval body
plan is the same as the juvenile body plan, the re-
tention of ancestral larval feeding and swimming
structures appears to be common. Whether this is
due to recent evolutionary origins of lecithotrophic
development or unidentified functions of these
visible to natural selection and take a long time to be
lost by the slow accumulation of mutations via ge-
netic drift—is an outstanding question.
Underlying all of these close-view questions is
the issue of how microevolutionary processes pro-
duce evolutionary change in mode of development.
McEdward (2000) explained the importance of
combining quantitative models that analyze the se-
lective factors that drive evolutionary change with
understanding of the genetic architecture underly-
ing the mechanisms of development. This is as true
today as it was nearly 20 years ago. While the field
of evo-devo has made rapid progress in under-
standing the genetic mechanisms of development,
little progress has been made in understanding
how selection acts on natural variation to generate
divergent life histories. In the last 20 years, research
on evolutionary transitions in mode of develop-
ment has been focused fruitfully on comparative
phylogenetic approaches. We hope that the next 20
years will see an equally fruitful focus on microev-
olutionary models to understand the evolutionary
mechanisms of transitions in mode of development
(e.g., Garfield et al., 2013; Arendt, 2015; Zakas and
Rockman, 2015).
4.6 Summary
1. Despite the call for more phylogenetic analyses
with dense sampling to understand evolution of
mode of development by McEdward and Miner
(2001), few such analyses have been published in
the intervening 15 years.
2. Comparative analyses show well-supported
patterns when transitions are rare but are not
always able to reconstruct evolutionary patterns
of mode of development with confidence when
evolutionary changes are rapid and common.
 E v o l u t i o n ary Tra n s i t i o n s i n M o d e o f De v e l o p me n t    63
3. Improvements in phylogenetic comparative
methods and wider sampling are likely to im-
prove the utility of this approach, but significant
weaknesses remain.
4. Overview of the published analyses shows that
the “what makes sense” view of macroevolution-
ary patterns of the evolution of mode of devel-
opment are not always supported. For example,
contrary to what makes sense, planktotrophic
species showed a higher diversification rate than
lecithotrophs in sacoglossans (Krug et al., 2015).
5. Increased effort to examine closely the microevo-
lutionary processes involved in transitions and
the mechanisms like pleiotropy, plasticity, and
balancing selection, which potentially play a role
in the retention of reversal ability—another area of
study advocated by McEdward (2000)—are likely
to contribute significantly to our understanding.
Acknowledgments
We thank the editors for inviting us to contribute
to this volume, the United States National Science
Foundation (IOS-1019727 to RC and OCE-1505158
and PLR-1341476 to AM) for supporting our re-
search on invertebrate development, and C. Zakas
for helpful comments on the manuscript.
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 C h a pt er 5
Asexual Reproduction of Marine
Invertebrate Embryos and Larvae
Jonathan D. Allen, Adam M. Reitzel, and William Jaeckle
5.1 Introduction
The life histories of marine invertebrates are incred-
ibly diverse and provide a wealth of opportunities
to develop and test hypotheses about how and why
modes of reproduction, development, and behav-
ior evolve within and among lineages. With respect
to the evolution of reproductive and developmen-
tal mode, phylogenetic, adaptive, and functional
hypotheses presented over the past century have
predominantly focused on the evolution of repro-
ductive traits (e.g., free spawning, brooding, en-
capsulation; Rouse and Fitzhugh, 1994), nutritional
mode of larvae (e.g., planktotrophy and lecithotro-
phy; Strathmann, 1985; Hart et al., 1997), and devel-
opmental form (e.g., larval morphology; Jeffery and
Swalla, 1992; direct and indirect development; Wray,
1995; McEdward and Janies, 1997). Frequently, but
not exclusively, these hypotheses have been tied to
changes in per-offspring investment (Emlet et  al.,
1987; Sinervo and McEdward, 1988; Emlet and
Hoegh-Guldberg, 1997), and influential models of
per-offspring investment often serve as a frame-
work for studies of the evolution of developmen-
tal modes (Vance, 1973; Christiansen and Fenchel,
1979; Levita­ n, 2000). Phylogenetic assessment of
the evolution of character states within lineages has
revealed frequent shifts among life histories traits
(McHugh and Rouse, 1998, and references therein).
In addition to these macroevolutionary changes
among species, the life history exhibited by indi-
viduals may change in response to one or more
environmental features (life history plasticity). Plas-
ticity in the evolution of life histories has also been
Allen, J. D., Reitzel, A. M., and Jaeckle, W., Asexual reproduction of marine invertebrate embryos and larvae. In. Evolutionary Ecology
of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018).
© Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0005
addressed with respect to such aspects of sexual re-
production as facultative progenesis, where larval
stages develop gametes (Poulin, 2001), and poecil-
ogony, where one individual or species exhibits dif-
ferent reproductive modes (Krug, 1998). One area of
life history plasticity that is far more common, but
has received less attention, is asexual reproduction.
Asexual reproduction during development oc-
curs in most marine phyla (reviewed by Alvarado,
2000; Blackstone and Jasker, 2003) and may occur
at multiple stages during the indirect development
of some species. Despite, or perhaps because of, its
broad distribution among phyla, diverse explana-
tions for the adaptive value or evolutionary sig-
nificance of asexual reproduction as a life history
strategy have been offered. Adaptive explanations
include the following: as a method to weather
harsh environments (e.g., gemmules in sponges,
statoblasts in bryozoans); as a short-term solution
for escaping mutation load (Hurst and Peck, 1996);
as a means to colonize and/or adapt to chang-
ing or new habitats (Maynard Smith, 1971); or as
a means to persist at low population sizes (Gerber
and Kokko, 2016). Several authors have addressed
adult asexual propagation and regeneration across
the animal kingdom (e.g., Ferretti and Geraudie
2001), but asexual propagation is also employed by
pre-adult stages. In this chapter we aim to synthe-
size the literature describing asexual reproduction
by embryonic and larval stages of marine inver-
tebrates, with a particular focus on echinoderms,
where the ecological and evolutionary costs and
benefits of diverse modes of clonal replication have
been most studied. We conclude with a list of open
68   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
questions that may provide fruitful future research
avenues to better understand this understudied life
history trait of marine invertebrates.
5.2  Types of Asexual Reproduction
of and by Embryos and Larvae
The general life history of a marine invertebrate
has two potential stages where asexual propaga-
tion could occur after fertilization, but prior to
the development of the adult stage: the develop-
ing embryo and, when present, the larva. Asexual
reproduction in various marine invertebrates, as
well as throughout all animal phyla, also occurs
by an elimination of meiosis during reproduction
(e.g., parthenogenesis, apomixis, automixis; Judson
and Normark, 1996) and during the juvenile/adult
stage (Blackstone and Jasker, 2003).
5.2.1 Embryo
Asexual propagation at the embryo stage is referred
to as polyembryony. Polyembryony is defined as
the splitting of one sexually produced embryo into
many independent individuals. Approximately 20
years ago, Craig et  al. (1997) reviewed the occur-
rence of polyembryony in animals. They included
both fragmentation of embryonic stages and larval
budding within the term polyembryony, whereas
here we will limit our definition of polyembryony to
asexual propagation prior to the formation of a de-
finitive larva. Embryos and larvae commonly have
different levels or degrees of tissue and structural
organization, which allow for different mechanisms
of asexual reproduction. For some terrestrial species
(e.g., nine-banded armadillos and various species
of parasitoid wasps), polyembryony is an obligate
part of the life history where a single fertilized oo-
cyte fragments to result in more than one develop-
ing embryo. Polyembryony has also been reported
in a handful of species from the Bryozoa, Cnidaria,
Echinodermata, and Platyhelminthes. Some spe-
cies of cyclostome bryozoans (e.g., Crisia denticulata;
Hughes et al., 2005) exhibit polyembryony via frag-
mentation of the cleaving embryo. Adults produce
one or two primary embryos in the gonozooid that in
turn divide to produce up to 100 secondary embryos
(reviewed in Reed, 1991). These secondary embryos
may also undergo facultative asexual propagation
to yield many hundreds of embryos that complete
development and that are released as lecithotrophic
larvae. Polyembryony has been described for one
species of cnidarian, the coral Acropora millepora,
where environmental turbulence fragments early
embryos and the isolated blastomeres can develop
into complete larvae (Heyward and Negri, 2012).
Among echinoderms, polyembryony has been ob-
served in five species from the class Echinoidea: the
sea urchins Prionocidaris baculosa (Mortensen, 1938),
Eucidaris tribuloides, Lytechinus variegatus, and Stron-
gylocentrotus droebachiensis, and the sand dollar Echi-
narachnius parma (Allen et al., 2015). The incidence of
polyembryony in these species is highly variable and
significantly impacted by changes in environmental
conditions (e.g., salinity, discussed later). Finally, a
species of ectoparasitic flatworm (phylum Platyhel-
minthes), Gyrodactylus elegans, also exhibits polyem-
bryony but through a different mechanism involving
unequal cleavage, instead of embryo fragmentation.
For this species, a single fertilized egg undergoes
unequal cleavage that produces a second embryo
within the initial embryo, a third embryo within the
second, and a fourth embryo within the third (Kath-
eriner, 1904).
5.2.2 Larva
Asexual propagation at the larval stage has been re-
ported in at least four phyla: Arthropoda, Cnidaria,
Neodermata (within the traditional phylum Platy-
helminthes), and Echinodermata (addressed in a
separate section later). For those species where it
has been described, asexual propagation occurs
by budding, although the method of budding can
be quite variable. Within the phylum Arthropoda,
the parasitic rhizocephalan barnacles display a bi-
zarre method of larval budding, where they use
totipotent cells for host invasion (Glenner and
Høeg, 1995). The cyprid larva settles on a decapod
host and transforms into a kentrogon, which then
injects a number of de-differentiated cells into the
host. Each cell forms a vermiform stage that splits
into individual cells that will form independ-
ent adults in different parts of the host. A similar
“infective” single-cell stage may occur via amoe-
boid cells in narcomedusozoans (Russell, 1953),
 A s e x ua l R e p r o d u c t i o n o f M ar i n e I n v ert e b rat e E m b ryo s a n d Lar vae    69
although the specific details are limited. For other
species of narcomedusan hydrozoans (Osborn,
2000), scyphozoan­s (Berrill, 1949), one species of sea
anemone (Edwardsiella lineata; Reitzel et  al., 2009),
and trematodes and cestodes (Katheriner, 1904;
Poulin, 2007), a larva asexually propagates by bud-
ding from the existing larval tissue.
5.3  Asexual Reproduction by Feeding
Larvae of Echinoderms
5.3.1  Class-level Distribution of Larval Cloning
in Echinoderms
Asexual propagation by larvae has been most
thoroughly described for planktotrophic larvae
of echinoderms. A review by Mladenov (1996)
summarized the occurrence of asexual reproduc-
tion in echinoderm classes and life history stages,
where 1.3% of species undergo asexual reproduc-
tion as adults (0% of crinoids and echinoids, up
to 2.2% of ophiuroids). Asexual reproduction by
feeding larvae of echinoderms is known from four
of five taxonomic classes (Asteroidea, Echinoidea,
Holothuroidea, and Ophiuroidea), and is not
­ ing larval body contains only short esophageal and
known for larvae from the Crinoidea. Larval clon-
ing is more broadly distributed among echinoderm
classes than adult asexual reproduction despite the
many fewer larval forms that have been studied,
some exclusively from field samples.
Of all reports, evidence of cloning by larvae in the
field is known only for some members of the Aster-
oidea and Ophiuroidea (Bosch et  al., 1989; Bosch,
1992; Rao et  al., 1993; Jaeckle, 1994; Balser, 1998;
Knott et al., 2003; Galac et al., 2016). In the subtrop-
ical-tropical western Atlantic Ocean, asteroid larvae
showing evidence of current and past asexual re-
production by paratomy are seasonally abundant
(Jaeckle, 1994; Knott et  al., 2003). Rao et  al. (1993)
also reported cloning larvae from the Bay of Ben-
gal (northeast Indian Ocean), and there is an image
of a cloning bipinnaria larva collected from waters
surrounding the Great Barrier Reef (Image Quest
Marine, personal communication). Two other forms
of asexual reproduction are exhibited by field-
collected asteroid larvae from the tropical west-
ern Atlantic Ocean: the release of the tips of larval
arms (budding) by newly collected larvae has been
infrequently observed and there is one larval form
(tentatively assigned to the taxonomic order Paxil-
losida, family Astropectinidae) that asexually re-
produces by autotomy of the preoral lobe (Jaeckl­e,
1994).
Cloning by ophiuroid larvae was originally sug-
gested by Mortensen (1921) and described by Balser
(1998) involving similar morphological changes to
those observed in asteroids, but the clone is formed
after the separation of the larval body from the fully
formed juvenile. The remnants of the “primary”
larva migrate to the vertex of the posterolateral
arms and reiterate gastrulation (by invagination)
and form the digestive system of the secondary
larva. Balser (1998) reported that from a single
fertilized egg of Ophiopholis aculeata two juveniles
could be formed and that the development of second
clonal generation for one individual was initiated.
Among asteroids, a cloning sequence similar to
that exhibited by ophioplutei is hypothetically pos-
sible. Cloning by larvae could occur after a larva
and the fully formed juvenile separate as the larval
remnant may reenter the water column (e.g., Luidia
sarsi). Although the digestive system of the persist-
intestinal segments, these individuals can survive
for some period of time (Tattersall and Sheppard,
1934). However, in culture they appear to be inca-
pable of feeding on particulate foods and “as far
as Luidia sarsi is concerned, regeneration followed
by a second metamorphosis most probably never
occur­s” (Wilson, 1978, p. 475).
5.4  Modes of Asexual Reproduction
in Echinoderms
5.4.1 Asexual Reproduction by Budding
Larval budding has been reported for members of the
echinoderm classes Asteroidea (Jaeckle, 1994; Vick-
ery and McClintock, 2000), Holothuroidea (Eaves
and Palmer, 2003), and Echinoidea (Vaughn and
Strathmann, 2008; Vaughn, 2009; 2010; McDonal­d
and Vaughn, 2010). The details of this form of asex-
ual reproduction among groups are not well de-
scribed and the developmental sequence of events is,
at best, superficially known. For asteroid larvae, the
released apices of the larval arms appear to reiterate
70   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
the sequence of development from a fertilized egg.
Gastrulation by the blastula-stage secondary em-
bryos reportedly occurs through unipolar invagina-
tion and the digestive system of the clone is derived
from the ectoderm of the parent larva (Jaeckle, 1994).
Although Jaeckle (1994) hypothesized that coelomo-
genesis in these embryos occurs through schizocoely,
there are no new data that support this proposal.
5.4.2 Asexual Reproduction by Paratomy
Paratomous asexual reproduction by asteroid larvae
(Figure  5.1) is known from individuals collected in
plankton samples taken throughout the subtropical-
tropical western Atlantic Ocean. To our knowledge,
in all examined clones formed by paratomy of the
posterolateral arms, the anterior-posterior axis of the
developing clone is reversed to the anterior–posterior
axis of the parent arm; the proximal (anterior) portion
Cb
M
uM
cA
Figure  5.1 Scanning electron micrograph (ventral view) of a field-
collected early brachiolaria larva showing an unmodified posterolateral
arm (uM, left side) and a clonal arm (cA, right side) where the
posterolateral arm is transforming into a new “secondary” larva
through the process of paratomy. Symbols: Cb = ciliary band, M =
mouth. Scale bar = 200 μm.
of the parent arm corresponds to the posterior side of
the offspring. How this axial inversion or any other
attribute of asexual reproduction is genetically speci-
fied or regulated has not been identified.
Transformation of the posterolateral arms into
new (secondary) individuals is morphologically
signaled by the dissipation of the ciliary band that
runs along the lateral sides of the posterolateral
arm and the transformation of a simple epithelium
into a stratified epithelium (Bosch et al., 1989). All
known forms of cloning exhibited by asteroid lar-
vae involve an apparent re-differentiation of cell
fates. During the paratomous transformation, the
presumptive ectoderm of the parent larva forms a
new “secondary archenteron,” which forms the di-
gestive system of the clone.
The archenteron develops along the length of the
transforming arm (Bosch et  al., 1989; Figure  5.2).
PL
S
SL
A
Figure  5.2 Light micrograph of a field-collected sea star larva with a
gastrula-stage secondary larva (SL) forming from the posterolateral arm.
Symbols: A = archenteron of clone, E = esophagus of the primary larva,
PL = primary larva, S = stomach of the primary larva. Scale bar = 200 μm.
 A s e x ua l R e p r o d u c t i o n o f M ar i n e I n v ert e b rat e E m b ryo s a n d Lar vae    71
However, it is unclear if the archenteron invaginates
as a longitudinal furrow that closes along its length
to leave a circular posterior opening or if it forms
from a single invagination that extends distally
within the blastocoel of the transforming arm. After
the archenteron achieves its full length, the tubular
gut differentiates to become the esophagus, stomach,
and intestine, and the mouth is established as a new
opening on the distal side of the transforming arm.
Coelom formation during the development of
the secondary (clonal) larva remains incompletely
described, but examination of physical and optical
sections of cloning larvae indicates that secondary
coelomogenesis occurs by enterocoely (Figure 5.3).
Ultimately, a single presumptive coelom may lie
lateral to the developing archenteron and extend
beyond the archenteron in the distal side of the
transforming larval arm.
Clonally produced individuals may separate
from the parent as an early stage or fully formed
S-p PL
I-p
LS
B
A A
? SL
Figure 5.3 Combined optical sections (N=3) of a ventral view of a
primary larva (PL) with an attached secondary larva (SL) forming through
the process of paratomy. The secondary archenteron (A) is continuous
with a blastopore (B) and a thin-walled cavity whose developmental
fate is unresolved. Symbols: I-p = intestine of primary larva, LS = left
somatocoel of the primary larva, S-p = stomach of primary larva. The
specimen was stained with Nile Red and examined using a Leica model
TCS SP2 scanning laser confocal microscope. Scale bar = 150 μm.
bipinnaria larva (Figure  5.4). Fully formed sec-
ondary bipinnariae may ingest particles prior to
separation from the parent larva. The only known
observation of the separation of the clone from the
parent larva (of Luidia senegalensis) occurred as the
clone rotated about the long axis of the transform-
ing arm of the parent arm. This rotation tightened
the stricture between the parent and clone and re-
sulted in a breakage of the epithelial connection.
5.4.3 Asexual Reproduction by Autotomy
Mortensen (1898) illustrated a larva (“Auricularia
paradoxa”) collected from the tropical western At-
lantic Ocean (0° 24’ N; 46° 40’ W) that resembles an
asteroid larva with long posterolateral arms minus
the preoral lobe. He described this larva by writing,
“The opening of the mouth seems to lie very close
to the anterior end, but everything which should
be above the anterior transverse margin is want-
ing.” Whether this individual was an auricularia
larva (Holothuroidea) that was damaged during
Cb
E
S
Cb
Figure 5.4 Combined optical sections (N=3) of a dorsal view of an
independent secondary larva formed by the process of paratomy. The
left axohydrocoel (A) extends a pore canal that fuses with the outer
epithelium (not shown), but there is no evidence of a right axohydrocoel.
Symbols: Cb = ciliary band, E = esophagus, S = stomach. The specimen
was stained with Nile Red and examined using a Leica model TCS SP2
scanning laser confocal microscope. Scale bar = 100 μm.
72   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae

collection or an asteroid larva that had previously
autotomized its preoral lobe is not known. Nearly
100 years later, Jaeckle (1994) described larvae col-
lected from the subtropical western Atlantic Ocean
with a common phenotype—but different from the
larva described by Mortensen—that reproduced by
autotomy of the preoral lobe. Autotomization of
the preoral lobe was then reported for cultured lar-
vae of several forcipulatid asteroids: Pisaster ochra-
ceus (Vickery and McClintock, 2000), Distolasterias
nipon (Kitazawa and Komatsu 2001), and Asterias
forbesi (Blackburn and Allen, 2013, Figure 5.5). Au-
totomy by cultured pluteus larvae of the sand dol-
lar Dendraster excentricus (order Clypeasteroida)
was seen after larvae were exposed to fish mucus
(Vaughn and Strathmann, 2008; Vaughn 2009; 2010)
or acute increases in food abundance (MacDonald
and Vaughn, 2010). Despite multiple observations
of this form of cloning, the mechanism of asexual
reproduction by autotomy is not known.
Examination of the “new” bipinnaria larvae
formed by autotomy of the preoral lobe of the
parent larva reveals apparent differences in the
coelomic cavities. Examination of these small bi-
pinnariae revealed a precocious development of
the coelomic system, compared to a larva of similar
size formed from a fertilized egg. In one individual
at the time of collection, there was no evidence of a
pore canal extending from the left axocoelic coelom
or a dorsal pore (“hydropore”).
5.5  Induction of Asexual Reproduction
Little is known about the factors that induce asex-
ual reproduction in nature, but induction of poly-
embryony and larval cloning have been achieved
using a number of different cues in laboratory
settings. Understanding the forces responsible for
inducing asexual reproduction, especially if they
are rather common, may help us understand the
frequency with which it occurs in nature. Studies
to date suggest both abiotic and biotic factors can
result in larval cloning in different species.
5.5.1 Abiotic
Abiotic cues of asexual reproduction include
changes in environmental parameters, including
pH, salinity, temperature, and even the turbulence
of the water. Few studies have investigated the
degree to which these parameters (many of which
are projected to covary in models of future climate
change) may interact with one another to induce
asexual reproduction. Independently, each condi-
tion has been demonstrated to play at least some
role in inducing asexual reproduction and thus all
are of great interest to both larval biologists and ma-
rine ecologists concerned about how recruitment
patterns vary in space and time.
There is recent interest in how ocean acidifica-
tion will affect the small, often partially calcified

Day 7 Day 8 Day 9 Day 10 Day 11 Day 12 Day 19

Figure 5.5  Development/regeneration of the anterior portion of a bisected larva over 12 days. The preoral lobe was first observed floating in
culture on Day 7. Arrowhead indicates larval mouth. Arrow indicates the reappearance of coelomic cavities. The dark coloration of the stomach on
Day 19 indicates the presence of algal food following resumption of larval feeding. All images are at the same scale. Scale bar = 100 µm. All photos
courtesy of Holly Blackburn.
 A s e x ua l R e p r o d u c t i o n o f M ar i n e I n v ert e b rat e E m b ryo s a n d Lar vae    73
developing stages of marine invertebrates (re-
viewed by Byrne, 2011), but relatively little is known
about the effect that acidification might have on the
propensity of embryos or larvae to clone. Chan et al.
(2013) demonstrated that pCO2 levels predicted for
2100 and 2300 induced high frequencies (>50%) of
budding for early larval stages of the purple sea ur-
chin Strongylocentrotus purpuratus. However, none
of the resultant buds survived for more than two
days, calling into question the viability of asexually
produced propagules and also raising the possibil-
ity that methods for rearing buds may differ from
those sufficient to culture primary embryos and lar-
vae. Low pH has also been reported to increase the
frequency of conjoined twins in larvae of the gastro-
pod Crepidula fornicata (Eyster, 1995), but there is as
yet little evidence that this occurs in nature or that
unjoined twins are ever produced.
Temperature has also been shown to affect the
frequency of larval cloning in echinoderm larvae. In
asteroids, Vickery and McClintock (2000) reported
that temperature exerted a strong effect on the like-
lihood of larval cloning in the bipinnariae of the sea
star Pisaster ochraceus. At low temperatures (7–10 ºC)
no larval cloning was observed, but at moderately
increased temperatures (12–15 ºC) cloning was ob-
served in about 6% of larvae in culture. At more ex-
treme levels of temperature (17–20 ºC) no cloning
was observed, but this result may have been con-
founded with high levels of mortality among pri-
mary larvae (Vickery and McClintock, 2000).
Similarly, few studies have assessed the role of
salinity as a factor that induces asexual reproduc-
tion by echinoderm embryos. In 2015, Allen et  al.
described the role of salinity as a cue inducing
polyembryony in early embryos of two sea urchins,
the cidaroid Eucidaris tribuloides and the echinoid
Echinarachnius parma. Reduced salinity resulted in
higher frequencies of polyembryony in both spe-
cies, but also interacted significantly with increas-
ing temperature to yield even higher frequencies
of polyembryony. Similar results have been found
in a third echinoid species, the sand dollar Den-
draster excentricus (Abdel Raheem and Allen, un-
published data). The interaction between salinity
and temperature suggests that multiple factors may
act in synergistic ways to influence frequencies of
asexual reproduction (see later). Prior to this work,
Mortensen (1938) suggested that polyembryony
may occur as part of the normal developmental
pathway in another cidaroid, Prionocidaris baculosa,
but provided no information on the environmen-
tal conditions (temperature, salinity, etc.) at which
embryos were raised. Interestingly, embryos of two
species of regular urchins (Strongylocentrotus droe-
bachiensis and Lytechinus variegatus) have also been
tested for similar responses to salinity, but very
rarely exhibited polyembryony.
Environmental induction of polyembryony in ir-
regular echinoids but not in regular echinoids is con-
sistent with chemical induction of polyembryony
as reported in a series of papers by Mazia and Vac-
quier (Mazia, 1958; Vacquier and Mazia, 1968a; b).
Vacquier and Mazia (1968b) showed that the close
association of blastomeres with the hyaline layer in
regular echinoids kept cells pressed together even
when cell-cell adhesion was disrupted by applica-
tion of dithiothreitol (DTT). In contrast, applica-
tion of DTT to Dendraster excentricus embryos with
a looser association with the hyaline layer resulted
in dissociation of cells from one another and ulti-
mately polyembryony. Since the presence of Ca2+
ions is also required for normal interactions be-
tween blastomeres (Vacquier and Mazia, 1968a; b),
it has been proposed that environmental induction
of polyembryony at low salinities may result from
lower concentrations of Ca2 + in low-salinity seawa-
ter (Allen et al., 2015), but this has yet to be formally
tested. Regardless, the diversity of responses of
regular and irregular echinoids to similar environ-
mental conditions suggests these lineages may be
promising models for future comparative studies of
the mechanisms of asexual propagation of embryos.
The combination of multiple stressors, either
taken from the previous list or unique ones not
yet considered, may be a fruitful area for future re-
search on the induction of asexual reproduction in
embryos and larvae.
5.5.2 Biotic
In addition to the abiotic stimuli described earlier,
there is increasing evidence that asexual reproduc-
tion may also be induced by interactions between
larvae and other organisms in their environ-
ment. Biotic stimuli suggested to induce asexual
74   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
reproduction include the amount of phytoplankton
in culture (Vickery and McClintock, 2000; Eaves and
Palmer, 2003), a sudden change in food abundance
(McDonald and Vaughn, 2010), and the presence of
chemical cues from a possible predator (Vaughn and
Strathmann, 2008; Vaughn, 2009; 2010). The ability
of larval stages to perceive and respond to biologi-
cally produced chemical cues in their environment
is well known, and can be manifested in a number of
ways, including plasticity in arm length in response
to phytoplankton food (e.g., Hart and Strathmann,
1994; Miner, 2005), plasticity in shell morphology
in response to predators (Vaughn, 2007), delays in
hatching time in response to predators (Miner et al.,
2010), and plasticity in settlement time in response
to prey (Hadfield, 1977). Thus, it may not be sur-
prising that to the degree planktonic larvae are able
to reproduce asexually, they do so in response to
biological cues in their environment. However, the
frequency of biologically induced cloning events in
nature is unknown. Only in a handful of controlled
laboratory studies do we have any evidence for bio-
logical induction of asexual reproduction.
The most detailed examples of biotic stimuli in-
ducing asexual reproduction come from studies of
the effects of larval food supply (i.e., phytoplankton
abundance) on clone production in echinoderms.
In the ochre sea star, Pisaster ochraceus, Vickery
and McClintock (2000) showed that cloning rates
were highest when phytoplankton food was most
abundant in culture and that a mixed diet (multi-
ple species of phytoplankton) yielded higher rates
of cloning than did a single-species diet. Similarly,
phytoplankton abundance is also a cue for cloning
in the pluteus larvae of the sand dollar, Dendraster
excentricus (McDonald and Vaughn, 2010). In the
case of D. excentricus, however, it is not the level
of phytoplankton abundance that induces cloning,
but rather a sudden shift in phytoplankton abun-
dance (from low to high) that correlates with high
frequencies (50–100% of larvae) of clone production
(McDonald and Vaughn, 2010).
Perhaps the most surprising example of biologi-
cal stimuli inducing clone production also comes
from D. excentricus, where pluteus larvae can be
induced to produce clones via budding after ex-
posure to mucus cues from planktivorous fish
(Vaughn and Strathmann, 2008). The production of
clones in response to fish mucus is highly variable
and appears to be influenced by genetic (maternal)
background (Vaughn, 2009). The reduced size of
larval clones relative to primary larvae has been
suggested to be an adaptive response (Vaughn and
Strathmann, 2008; see further discussion later) to
visual predators that are known to be size-specifi­c
in their feeding on larval echinoids, preferring
larger larvae to smaller ones (Allen, 2008; Vaughn,
2010). However, as with phytoplankton cues, the
prevalence of predator-induced cloning in nature is
unknown and the mucus inducer of larval cloning
is isolated from a benthic fish (the dover sole, Micro-
stomus pacificus) that feeds preferentially on benthic
polychaetes and ophiuroids as an adult (Gabriel
and Pearcy, 1981), although the lengthy pelagic
period of its larvae may provide an opportunity
for planktivory on echinoids (Pearcy et  al., 1977),
though that has yet to be demonstrated.
5.6  Other Taxa
To date, only one study has examined the role of
environmental turbulence in asexual reproduc-
tion, occurring in the scleractinian coral Acropora
millepora (Heyward and Negri, 2012). Cloned coral
embryos resulting from turbulence-induced blasto-
mere separations at the two-, four-, and eight-cell
stages were able to successfully complete develop-
ment in the lab, albeit at smaller sizes. For embryos
of A. millepora the levels of turbulence sufficient to
induce cloning in the laboratory are equivalent to
or less than those found in nature during the major-
ity (52%) of spawning events on the Great Barrier
Reef. It is therefore likely that natural turbulence
generates cloned coral embryos, although the fate
of these embryos and the potential adaptive value
of this response remains unknown.
5.7  Is Larval Cloning Adaptive?
As noted earlier, it is tempting to view asexual re-
production by embryos and larvae through the
lens of adaptation, but the adaptive value of poorly
understood phenotypes is difficult to demonstrate.
The hypothesized benefits of asexual reproduc-
tion by embryos and larvae are intuitive, related
 A s e x ua l R e p r o d u c t i o n o f M ar i n e I n v ert e b rat e E m b ryo s a n d Lar vae    75
to increasing the number of genotype copies. They
could contribute to reproductive success in one or
more ways. Cloning:
1. increases female fecundity without an apparent
increase in resource allocation to reproduction;
2. may increase in the likelihood that a member of a
genet survives;
3. may increase the probability that a member of
a genet will locate a suitable settlement site by
sampling a greater geographic area;
4. may reduce the genet’s susceptibility to loss (i.e.,
predation) by increasing propagule number; and
5. may reduce the genet’s susceptibility to loss
(i.e., predation) by decreasing propagule size
(Vaughn, 2010).
Nevertheless, the formation of each new individual
(the clone) is a result of the loss of cell(s) from the
“parent” embryo or larva, and this reduction in bio-
mass may negatively influence the larva, the result-
ing juvenile, or both. The impact of this tissue loss
on the survivorship of the parent is unknown, but
it seems reasonable to hypothesize that the cost of
asexual reproduction to the parent embryo or larva
could include:
1. a decrease in larval feeding rate;
2. a decrease in larval grow rate;
3. an increase in the time to metamorphosis (with
potentially increased exposure to planktonic
predators); and
4. a decrease in juvenile size.
The extent to which any fitness trade-off varies
among different forms of asexual reproduction by
larvae is unknown. However, it seems reasonable
to assume that any cost is correlated with two fac-
tors: (1) when asexual reproduction occurs dur-
ing a species’ life history and (2) the proportion of
the primary individual’s body that is allocated to
cloning. Cloning by budding removes the smallest
volume of material from the parent larva, and we
hypothesize that this form of asexual reproduction
has the smallest cost to the parent larva. Autotomy
of the preoral lobe or paratomy of the posterolateral
arm(s), in contrast, involves loss of a greater volume
of tissue and potentially results in a greater cost to
the parent larva through a reduction of feeding ca-
pacity, loss of biomass, or both. Yet the elements of
both lists of “consequences” are plausible, but re-
main largely untested. The net effect of cloning is
realized as the sum of these positive and negative
consequences, and there is no a priori reason to ex-
pect that the result of this summation is constant
from year to year.
To our knowledge, there is only one formal at-
tempt to model the consequences of larval cloning.
Rogers-Bennett and Rogers (2008) evaluated the
influence of an unspecified form of larval cloning
on dispersal distance. Their simulations predicted
(1) that cloning increases the dispersal potential
of a cohort of larvae, and (2) that the age of par-
ent larvae when asexually produced offspring are
released was positively related to the dispersal po-
tential of the cohort. Their model, however, did not
incorporate any estimates of the cost(s) of asexual
reproduction to the parent larva or to the subse-
quent juvenile.
Although the costs and benefits that may select
for evolution of larval cloning in marine inverte-
brates remain understudied in free-living marine
species, asexual reproduction of developmental
stages in parasitic species are better characterized.
Endoparasitic larval stages of digeneans, cestodes,
and other parasitic Platyhelminthes (Neodermata)
commonly exhibit asexual propagation in inter-
mediate hosts. For example, the miracidium stage
that infects a snail host can asexually replicate itself
to generate thousands of cercariae, the next stage
in the life history. The extent of asexual reproduc-
tion in these larval stages can vary widely due to
taxonomic group (Rohde, 1982, p.  54) and a num-
ber of interrelated life history factors summarized
by Poulin (2007), including host size and infec-
tion longevity (Keeney et  al., 2008), competition
(Hendrickso­n and Curtis, 2002), and adult size or
longevity (Moore, 1981).
In schistosome species, the extent of asexual re-
production in the miracidium stage is an appar-
ent trade-off with oocyte production, where larger
embryos have higher asexual reproduction rates
(Loker, 1983). The increased number of asexually
produced individuals may represent one adaptive
mechanism for increasing successful transmission
to a second intermediate or terminal host, a step in
the life history which may occur at a low frequency
due to death of the current host or low encounter
76   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
rates with the next host. Asexual reproduction of
endoparasitic larvae in hosts may also be a com-
mon strategy in other parasitic species, including
the sea anemone Edwardsiella lineata, where larval
cloning occurs and parasite number correlates with
host size (Reitzel et al., 2009). We can hypothesize
that larvae of nonparasitic species may be selected
for asexual reproduction due to similar trade-offs
in the plankton, where different selection pressures
(e.g., likelihood of predation, feeding environment,
probability of successful settlement) may result
in positive selection for asexual reproduction in a
species-dependent manner. Discerning between po-
tential selective factors will require a combination
of laboratory experimentation and resolved phylo-
genetic relationships for closely related species with
and without larval asexual propagation.
5.8  Open Questions for Future Research
(1) Does asexual reproduction by cultured larvae offer a
window to natural phenomena or represent an induced
developmental response to unnatural environmental
conditions?
Production of asexual propagules in response to
environmental conditions or changes in conditions
(food abundance: Vickery and McClintock, 2000;
Eaves and Palmer, 2003; McDonald and Vaughn,
2010; possible predators: Vaughn and Strathmann,
2008; Vaughn, 2009; 2010) are known from in vitro
experiments. Consequently, it is not certain whether
examples of asexual reproduction by embryos or
larvae in culture represent natural phenomena or
are developmental events induced by culture con-
ditions. There is evidence provided by early work-
ers (e.g., Gemmill, 1914; 1915; MacBride, 1918) that
deviations from normal development (e.g., larvae
with multiple or no pore canals) could be induced
by culture conditions.
Echinoderm embryos and larvae exhibit a tremen-
dous capacity for regeneration. Research has dem-
onstrated that bisected echinoderm larvae of sea
stars (Pisaster ochraceus, Luidia foliolata, and Patiria
miniata) and sea urchins (Dendraster excentricus and
Lytechinus variegatus) are capable of regrowing the
excised portions of their body (Vickery and McClin-
tock, 1998; Vickery et al., 2002; Oulhen et al., 2016).
In most cases, the regenerated larvae successfully
completed metamorphosis to produce a benthic ju-
venile. A more extreme example of the regenerative
potential of developmental stages of echinoderms
was described by Dan-Sohkawa et al. (1986). They
dissociated gastrula-stage embryos of the sea star
Asterina pectinifera using osmotic, ionic, and physi-
cal forces, and then followed the re-­ aggregation
of the isolated cells (or groups of cells). From
the initial suspension of cells, a free-swimmin­ g,
­blastula-like embryo was established after ≈30 h
post-­dissociation. After 80–100 h post-dissociatio­n,
bipinnaria-stage larvae were reconstructed. Tamur­a
et  al. (1998) examined the morphological events
of secondary coelomogenesis and described two
different processes that created the “coelomic
pouches.” In some larvae (37 of 62 individuals) a
coelomic pouch was established through processes
reminiscent of enterocoelly (“­enterocoel-like” coe-
lom formation), while in other individuals (22 of
62 individuals) coeloms were established by an ag-
gregation of mesenchyme-like cells (“schizocoelic-
like” coelom formation); three reconstructed larvae
formed coeloms through both processes. Not all
reconstructed larvae were developmentally nor-
mal as some individuals developed a multiple or a
branched digestive system.
(2) Does asexual reproduction occur in all groups of
echinoderms, but the frequency of occurrence in some
groups is too low to detect in samples from the field?
Among echinoderm species that produce devel-
opmental stages that undergo asexual reproduc-
tion, only cloning larvae of some species of the
Asteroidea and the Ophiuroidea have been identi-
fied from plankton samples. However, species of
the Echinoidea and Holothuroidea are known to re-
produce asexually in laboratory cultures. If asexual
reproduction occurs in all groups, but at a variable
and, in some cases, low frequency, then detection of
cloning by embryos or larvae in laboratory cultures
where large numbers of individuals can be evalu-
ated is not surprising. A low frequency of occur-
rence coupled with rapid rates of clone formation
(e.g., Vaughn, 2009) further reduces the likelihood
of detecting rare, fast-developmental events from
field samples. MacBride (1918) similarly com-
mented on the frequency of developmental abnor-
malities in coelomogenesis of echinoderm larvae:
“In this list of recorded instances of the occurrence
 A s e x ua l R e p r o d u c t i o n o f M ar i n e I n v ert e b rat e E m b ryo s a n d Lar vae    77
of two hydrocoels among echinoderm larvae, it will
be noticed that only one such specimen was found
among larvae fished from the open sea, although
hundreds of such larvae have been examined.” We
are hopeful that further field sampling will reveal
the presence of cloning larvae of echinoids and
holothuroids.
(3) How does the ability of larvae to clone evolve in
marine invertebrate lineages?
Currently, most descriptions of larval cloning
have been scattered throughout lineages, poten-
tially due to opportunistic observations or studies
in species that can be cultured in the laboratory. If
asexual reproduction is a component of a life his-
tory that is under selection, we would expect it to
be gained and lost in lineages in the same way as
any other character. Research in gains and/or losses
of asexual reproduction in sea anemones (Geller
and Walton, 2001) as well as regeneration in an-
nelids (Bely and Sikes, 2010) and other spiralians
(Bely et al., 2014) suggest that these developmental
processes can be quite dynamic within particular
lineages. The evolution of mechanisms of asexual
reproduction requires explicit comparison in a re-
solved phylogeny where the presence or absence of
“asexual reproduction” by embryos or larvae can
be mapped to determine if this is a primitive or a
derived life history character.
In asteroids, current evidence hints that larval
asexual reproduction may have evolved multiple
times. Cloning larvae have been collected from
opportunistic sampling from Barbados, Panama,
Beliz­e, Jamaica, Commonwealth of the Bahamas,
Gulf of Mexico, and throughout the Gulf Stream
system. Despite this regional geographic distribu-
tion and high seasonal abundances (Knott et  al.,
2003), the species identity of cloning asteroid larvae
remains poorly resolved. Using morphological fea-
tures of collected larvae, Bosch et al. (1989) assigned
cloning bipinnariae from the Sargasso Sea to the ge-
nus Luidia (order Paxillosida). Further samples from
the waters surrounding the Commonwealth of the
Bahamas and from the Florida Current of the Gulf
system revealed the presence of cloning larvae from
a non-paxillosid group (Jaeckle, 1994). Knott et  al.
(2003) compared tRNA gene sequences and recog-
nized three groups of cloning asteroid larvae (that
nested among species of the taxonomic families
Luidiidae, order Paxillosida, Ophidiasteridae, order
Valvatida, and Oreasteridae, order Valvatida), but
genus and species-level identities could not be re-
solved. More recently, Galac et al. (2016) sequenced
mitochondrial and nuclear genes from cloning and
aclonal bipinnariae and brachiolariae collected
from the Florida Current of the Gulf Stream sys-
tem expressing the same color phenotype. Through
their analysis they assigned these specimens to a
single, yet undescribed, species within the family
Oreasteridae. Their species was equivalent to “Lar-
val Group 1” reported by Knott et al. (2003). To our
knowledge the only species of asteroid echinoderm
known to produce secondary larvae by paratomy
is Luidia senegalensis (W. Jaeckle, personal observa-
tion). This specimen was identified based on the
number of arm rays of the juvenile.
(4) Will changing climate lead to increased asexual re-
production by larvae?
There is abundant evidence that climate change
will result in changes in ocean conditions that are
likely to have strong effects on microscopic larvae.
Given what is known about abiotic induction of pol-
yembryony and larval cloning (see earlier), there is
reason to believe that some potential outcomes of cli-
mate change will also lead to increased frequency of
asexual reproduction. First, phytoplankton blooms
may shift in their frequency, intensity, and location
under future climate change scenarios. It is likely
these changes will be uneven and likely difficult to
predict. For example, in the North Atlantic warming
sea surface temperatures have resulted in both in-
creased (in cooler regions) and decreased (in warmer
regions) phytoplankton abundance (Richardson and
Schoeman, 2004). If the overall levels of phytoplank-
ton increase, or if the speed and intensity of bloom
appearance increase, current research (Vickery and
McClintock, 2000; McDonald and Vaughn, 2010)
suggests that larval cloning may be facilitated.
Concurrent with ocean warming, there has also
been a freshening of the surface waters where the
development of marine invertebrates frequently
occurs. While freshening is not uniform globally,
major portions of the world’s oceans are freshening,
including the Gulf of Maine (notably on Georges
Bank), parts of the Indian and Pacific Oceans and
the Southern Ocean (Wong et  al., 1999; Drinkwa-
ter et al., 2009; Haumann et al., 2016). Since at least
78   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert e b rat e Lar vae
some cases of polyembryony are reported to oc-
cur in response to elevated temperature and con-
comitant reductions in salinity (Allen et al., 2015),
it may be of special interest for developmental
biologists and larval ecologists to examine field-
collected embryos and larvae for signs of polyem-
bryony and cloning in areas where the interactive
effects of temperature rise and salinity decline may
be strongest. Greater understanding of the physi-
ological tolerances and responses of embryos and
larvae, along with improved understanding of the
mechanisms of asexual reproduction are needed
before any substantive predictions can be made in
this regard.
5.9 Summary
1. Larvae in multiple phyla of marine invertebrates
undergo asexual reproduction, with most obser-
vations occurring under laboratory conditions.
2. Echinoderms, particularly asteroids and echi-
noids, are the most thoroughly described group
with respect to the diversity of modes of asexual
reproduction as well as the occurrence of clonal
reproduction in the natural environment.
3. Numerous abiotic and biotic factors have been
described that result in increased frequency
of asexual reproduction events in echinoderm
larvae, suggesting that larvae may respond to
diverse environmental signals by initiating a
cloning response.
4. The fitness costs of cloning to the “parent” larva
and how those costs may vary among different
forms of asexual reproduction are unknown and
warrant future study.
5. Future studies aimed at dissecting the evolution-
ary gains and losses of larval cloning in particu-
lar groups, as well as the frequency of cloning in
natural environments will provide essential data
to better understand how and why larvae of par-
ticular species have included asexual reproduc-
tion as part of their life history.
6. Future work on the developmental biology of
asexual reproduction is needed to determine if
the processes of asexual reproduction by free-
living larvae are different reiterations of de-
velopment from zygotes or if they represent
developmental novelties.
Acknowledgment
We acknowledge the assistance of Dr. Uwe Staerzfo­r
who provided a translation of Mortensen’s (1898)
description of “Auricularia paradoxa.”
AMR would like to thank the University of North
Carolina at Charlotte for support through a Junior
Faculty Development Award.
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 Ch a pt er 6
Section 1 Summary—Evolutionary
Origins and Transitions
in Developmental Mode
Abiotic variables and biotic interactions can act
on variation in life history traits, ultimately lead-
ing to divergence in reproductive mode. Marine
invertebrates have a remarkable diversity in such
strategies, sometimes even between closely related
species. It is this natural diversity that lends itself
to employing a powerful comparative approach,
both for particular morphological characteristics as
well as molecular signatures from developmental
genes. For example, complex life histories, where a
larval stage is interposed between the embryo and
juvenile, likely represent the product of numerous
selection pressures, historical and current, that have
shaped the diversity of larval stages in extant ma-
rine species. In fact, the very question about “what
is a larva?” has to be addressed, as it is so intimately
connected to bentho-planktonic life cycle and meta-
morphosis. Furthermore, novel larval types have
evolved in particular lineages and larvae have been
secondarily lost in others. This in itself creates an
interesting and exciting playground to test evolu-
tionary developmental hypotheses.
Chapter  1 by Claus Nielsen explores the evo-
lutionary origins of larvae and through that pro-
vides many examples for the diversity of larval
forms, as mentioned earlier. Similarly to Heather
Marlow (Chapter  2), Nielsen uses a compara-
tive framework to explore the question “What is
a larvae?” In contrast, Marlow discusses specific
developmental mechanisms involved in the for-
mation of the larval body. Marlow further dis-
cusses maternal investment strategies as a major
Carrier, T. J., Reitzel, A. M., and Heyland, A., Section 1 Summary—Evolutionary origins and transitions in developmental mode.
In. Evolutionary Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland:
Oxford University Press (2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0006
driving factor in the evolution of alternative life
histories. Dustin Marshall and colleagues discuss
the link between maternal investment and larval
diversity in Chapter 3. Chapter 4 by Rachel Collin
and Amy Moran exemplifies how specific predic-
tions can be derived from two main inferences ob-
served in larval diversity, i.e., the transition from
feeding to nonfeeding and the hypothesis that
complex characters that are not used are quickly
lost. Lastly, Chapter 5 by Jonathan Allen and
colleagues discusses polyembryony and larval
cloning, and presents many interesting new per-
spectives for future research.
In summary, this section addresses central ques-
tions on the origin of larvae, the diverse mechanisms
for how the larval stage is modified and how these
may relate the co-occurring changes in maternal in-
vestment and the environment, and how life histo-
ries have evolved from an indirect to direct mode.
In the process, these comparisons highlight the com-
mon balance and difficulties of discerning between
homology and homoplasy in evolutionary biology.
Decades of research using comparative research
and life history models have developed hypotheses
for correlates, and perhaps causes for how different
larval stages evolve. A particular focus in marine in-
vertebrates has been maternal investment strategies,
where smaller eggs develop into planktotrophic lar-
vae and larger eggs into lecithotrophic. Reappraisal
of the hallmark mathematical models coupled with
data from an increasing array of species suggest
additional factors that can be profitably included

to characterize these transitions in investment and
larval diversity. One of these factors is a shift in
ecology of the larva itself, which includes a transi-
tion from pelagic to demersal and from free-living
to parasitic. Lastly, larvae have been secondarily
S e c t i o n 1 S u m m a ry    83
lost in the life cycles for marine invertebrates many
times independently. The selection pressures and
functional limitations on the loss of the larval stage
represent fertile ground for how and why lineages
have evolved these major life history changes.
 C h a pt er 7
Larval Feeding: Mechanisms, Rates,
and Performance in Nature
Bruno Pernet
7.1 Introduction
Adults of many marine invertebrates make tiny
eggs, each provisioned with scant energy. These
typically develop into larvae that must acquire
energy and materials from the environment to
support growth and development through meta-
morphosis and to provision a successful juvenile.
To most biologists, larval feeding means the capture
of suspended particulate food by planktonic larvae
(planktotrophy), though planktonic larvae may also
acquire dissolved organic matter from seawate­ r and advection (Pineda and Reyns, this volume). It
(Jaeckle, this volume), and many larvae spend part
of their lives in capsules or masses, where they of-
ten acquire parentally supplied dissolved or partic-
ulate food. Here, I focus on suspension feeding on
particles by planktonic larvae, a process that occurs
in members of at least a dozen phyla.
How well feeding larvae perform the task of
capturing food has many potential consequences.
Food concentration is often so low that it limits
larval growth, extending the time larvae spend in
the plankton before metamorphosis (Fenaux et
al., 1994; Bos et al., 2006a; Pedersen et al., 2010).
Instantaneou­s mortality rates of planktonic larvae
are generally thought to be high (Morgan, 1995;
Pedersen et al., 2008). Thus larval feeding perfor-
mance may affect planktonic duration and risk of
mortality, as well as dispersal. The performance of
feeding larvae may also have long-lasting effects
on juvenile performance (Pechenik, this volume).
These ecological processes imply that there is selec-
tion on the form and physiology of feeding larvae
to maximize net rates of energy gain by feeding.
Pernet, B., Larval feeding: mechanisms, rates, and performance in nature. In. Evolutionary Ecology of Marine Invertebrate Larvae.
Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018). © Oxford University Press.
DOI: 10.1093/oso/9780198786962.003.0007
In most taxa feeding larvae are strikingly different
in form from nonfeeding larvae, consistent with the
idea that the requirement for food shapes the evolu-
tion of larval form (Emlet, 1991).
No matter the immediate interest of the biologist
studying larval feeding—functional morphology,
population ecology, or the evolution of life histories
and larval nutritional mode—an ultimate aim must
be to understand how well larvae feed in nature.
This is a challenge. Feeding larvae are generally tiny
(< 1 cm) and are subject to spread by “diffusion”
is thus difficult to study most aspects of larval biol-
ogy in the field. Most insights into larval feeding
are based on results from laboratory beakers, or
from rare studies in field-embedded mesocosms.
However, conditions in nature differ from those in
beakers in terms of hydrodynamics, the population
of edible and inedible particles, and temporal and
spatial heterogeneity in these and other features. As
noted by Strathmann (1987, p. 490), “The extension
of laboratory results to field conditions is fraught
with uncertainties.”
Such an extension, however, is needed. One way
that it might be made is via detailed mechanistic
models linking larval form and feeding mechanism
to feeding performance. With high spatial and
temporal resolution data on characteristics of the
feeding environment (in particular, temperature
and the abundance of particles of various types),
such models could be used to estimate larval in-
gestion rates over time, a parameter that can then
be integrated into broader bioenergetics models to
predict rates of growth in particular environments
88   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
(Bos et al., 2006a; Monaco et al., 2014). Such models
can be used not only to understand the biology of
larvae in the habitats in which they evolved, but
also to predict larval responses to novel conditions,
such as changes in seawater viscosity or in the
types of food particles available as a consequence
of ocean warming (Podolsky, 1994; Falkowski and
Oliver, 2007).
Here I review progress in our understanding
of larval feeding, focusing on three topics. First,
I describe the small set of physical processes that
larvae can use to concentrate particles from the
plankton, and the diverse morphologies and be-
haviors larvae use to implement these processes.
I focus especially on discoveries made in the past
30 years, since Strathmann’s (1987) comprehen-
sive review of larval feeding. Second, I discuss
examples of models that link larval form and
feeding mechanism to feeding performance, and
review some of the insights these models have
yielded. Finally, I identify several areas in which
advances are needed before we can achieve the
goal of integrating such models into a broader bi-
oenergetics context to predict how feeding larvae
perform in nature.
7.2  How Do Marine Invertebrate Larvae
Feed?
Planktonic larvae live in a dilute suspension of par-
ticles (Kiørboe, 2011), only some of which are edible
and nutritious. One way to acquire edible particles
is to rapidly drink the medium, but it would likely
be impossible for larvae to digest and assimilate
particles at the high flow rates required by this
strategy, and costs of pumping fluid through the di-
gestive system and rapidly losing secreted enzymes
would be high. Instead, feeding larvae concentrate
particles from suspension before processing them
in the gut. Our intuitions about how larvae do this
are not particularly useful, because larvae and their
feeding structures are very small and flow around
them is very slow. At that scale, viscous forces dom-
inate over inertial forces. Flow is laminar, and both
feeding structures and potential food particles are
surrounded by thick shells of sticky fluid (Hodin et
al., this volume).
7.2.1 The Limited Palette of Physical Processes
of Particle Encounter and Capture
Given these conditions, larvae can use only a few
processes to encounter and capture particles from
suspension. They can be divided into two classes:
those in which particles are trapped on a physical
filter, which does not require a larva to alter behav-
ior in response to individual particles, and those in
which particles are individually detected, stimulat-
ing an active response by the larva to direct the par-
ticle toward a capturing surface (LaBarbera, 1984;
Strathmann, 1987; Hart and Strathmann, 1995).
Filtering processes include hydrosol filtration
and sieving of particles from suspension. Hydrosol
filtration encompasses five distinct processes (di-
rect interception, inertial impaction, gravitational
deposition, diffusional deposition, and electrostatic
attraction) by which particles contact a capturing
surface (Figure 7.1A). The capturing surface can be
of any shape—e.g., a sphere (like a cell) or a cyl-
inder (a cilium or seta)—and can be made up of a
single component or an array of components (e.g.,
a band of cilia or an array of setae). Shimeta and
Jumars (1991) modeled encounter rates of particles
with idealized capturing surfaces for all five types
of hydrosol filtration. They distinguished between
encounter (physical contact of particle and captur-
ing surface) and retention (adhesion) of the parti-
cle on the capturing surface; together, they called
the two processes “capture” of a particle. For most
larvae, direct interception is the type of hydrosol
filtration that could yield high rates of particle cap-
ture, though any given capturing surface might en-
counter particles by multiple varieties of hydrosol
filtration. For a cylindrical capturing surface, par-
ticles are encountered by direct interception (and
captured, assuming 100% retention) at rates propor-
tional to their diameter (Shimeta and Jumars, 1991).
In sieving, the medium is forced through pores
in a mesh. Suspended particles larger than pores
are trapped on the mesh. Particles smaller than
pores may also be captured on mesh elements by
hydrosol filtration. Loudon and Alstad (1990) mod-
eled this process, including both sieving and direct
interception as mechanisms. Examples of larvae
that sieve food from suspension (e.g., cyphonautes:
Figure 7.1B) are rare, presumably because pore size
 L a r va l Fee d i n g    89

(A) (B) (C)

Figure 7.1  Examples of larvae that capture particles using hydrosol filtration, sieving, and active response. A. Veligers of gastropods capture particles
by direct interception on prototrochal cilia. Captured particles are transferred to a food groove between prototrochal and metatrochal bands, where
they are carried to the mouth. B. Cyphonautes of bryozoans use several particle capture mechanisms, one of which is sieving particles onto laterofrontal
cilia of the ciliated ridges in the mantle cavity. C. Echinoderm larvae use active responses in the locomotory and feeding ciliary band to capture particle.
Particles approaching the larva are detected by cilia in the band, and cilia near the particle reverse direction of beat, eventually driving the particle to
the mouth. Solid arrows indicate excurrent flow. Drawings modified from Fenchel and Ockelmann (2002), Strathmann and McEdward (1986), and
Strathmann (1971).

must be small to capture appropriately sized parti-
cles, but generating sufficient pressure to push fluid
through such a mesh is energetically costly.
In “active response” (also known as scan-and-
trap) processes, a larva detects an approaching par-
ticle and alters its behavior to move a parcel of water
containing the particle or to move the larval body
relative to the particle, so that the particle ends up
positioned near to a capturing surface (usually the
larval mouth) (Figure 7.1C). Detection may involve
chemical or hydromechanical cues. The intensity
of such cues vary with particle size, among other
characteristics, and particle detection depends on
the sensitivity of the larva’s sensors. Jumars (2003)
and Kiørboe (2008) provide models for predicting
encounter rates of particles by actively responding
predators. In larvae that feed in this way, the ma-
nipulation of flow to reposition the particle near the
larval mouth is often called particle capture (e.g.,
Hart, 1991), but the particle at this point has not
adhered to or even necessarily contacted a feeding
structure; it has really just come under the control of
the larva’s flow-management structures. It is then
directed toward a surface, where it is eventually
captured in the sense of hydrosol filtration.
Once captured, particles are rejected or trans-
ported to the mouth for ingestion. Captured par-
ticles may be rejected by larvae at multiple points
during the feeding process. If retained, particles are
transported to the mouth. Captured particles may
be lost during transport to the mouth, but little is
known about loss rates and whether or not these
rates differ between transport mechanisms. Particles
are either swallowed as they arrive at the mouth, or
held in a storage area (e.g., the esophagus of echino-
derm larvae) and swallowed in groups periodically.
7.2.2 The Diverse Array of Larval Forms
and Feeding Mechanisms
For the processes described previously, theoreti-
cians have modeled how rates of particle capture
by simple capturing structures depend on particle
size, concentration, and other variables. Larvae,
however, feed using complex and varied morphol-
ogies and behaviors. To understand how actual
larvae concentrate particles from suspension, we
need good descriptions of those morphologies and
behaviors. Biologists have been studying larvae for
more than two centuries (Young, 1990), but often
not at a resolution sufficient to make strong infer-
ences about mechanisms of larval feeding. In the
past 50 years, however, light and scanning electron
microscopy have allowed high spatial resolution
description of potential capturing surfaces, and film
and video microscopy have allowed researchers to
observe behavior and the kinematics of particle
capture in living larvae at relevant temporal scales
(Strathmann et al., 1972; Hart, 1996). Though these
tools have limitations (emphasized by Hart, 1996,
and Strathmann, 2005), they have greatly enhanced
our knowledge of suspension feeding.
Here I briefly summarize our knowledge of how
marine invertebrate larvae feed, focusing on recent
discoveries and on remaining gaps in our knowl-
edge. Strathmann (1987) provides more detailed
90   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e

descriptions of feeding in many of these larvae. have distinctive helmet-shaped pilidium larvae that
Several more general overviews of suspension feed- bear lateral lappets and an anterior and a posterior
ing also provide useful perspectives (Riisgård and lobe; a nearly continuous primary ciliated band
Larsen, 2010; Kiørboe, 2011). runs along the edges of the lappets and lobes. Pi-
lidia have long been known to feed on phytoplank-
Cnidaria. Planulae of medusozoans lack a mouth
ton, but their feeding mechanism has only recently
(Martin and Koss, 2002), and there is no evidence that
been elucidated in one species (von Dassow et al.,
they can feed. Anthozoan planulae, however, have a
2013). Pilidia of this species (Micrura alaskensis) use
mouth and gastrovascular cavity. Capture and in-
the motile cilia of the primary ciliated band to gen-
gestion of particles has been documented in several
erate a swimming and feeding current. Particles ap-
species, but almost always under unusual circum-
proaching the primary ciliated band appear to be
stances (Tranter et al., 1982; Schwarz et al., 2002). In
detected by periodically spaced non-motile cilia in
most cases, planulae have not been observed feed-
that band. When a particle is detected, the lappet
ing unless first offered a slurry of macerated crusta-
or lobe nearest the particle is rapidly moved (using
cean tissue. When a slurry is added, planulae often
muscles) so as to locally direct the current into the
stop swimming and begin feeding from the bottom
region between the lappets and lobes, the vestibule.
of the dish. Some continue swimming, trailing a
Once the particle is in the vestibule, cilia of the in-
mucus strand to which particles adhere. The mucus
ner ciliated band—which runs inside the lappets,
strand is then drawn into the mouth. Siebert (1974)
parallel to the primary ciliated band—are raised to
observed planulae of two sea anemones feeding
form a “fence” preventing its escape. The particle is
in the absence of macerated tissue. They captured
then carried to the mouth by cilia of the esophageal
plastic beads 1 µm in diameter and algal cells via
funnel. This is an active response type of particle
a trailing mucus strand, but apparently rejected the
capture, and thus chemical or hydromechanical sig-
algal cells at the mouth. There is need for additional
nals from approaching particles must be detected to
studies of feeding by anthozoan planulae in more
stimulate the response; this suggests that very small
realistic conditions, with known and appropriate
particles (e.g., bacteria) are unlikely to be captured
concentrations of potential food items.
by this mechanism. It is not yet known if other
Platyhelminthes. Larvae of many polyclads hatch pilidi­a feed in the same way.
bearing four or more lobes and a mouth leading
Bryozoa. Only one type of bryozoan larva, the
to a blind gut. These larvae, known as Götte’s or
cyphonaute­s, is known to feed. These larvae (Figur­e
Müller’­s larvae depending on the number of lobes
7.1B) pump water through the mantle cavity us-
they bear, are completely ciliated, with longer cilia
ing cilia on a pair of ciliated ridges that divide it
arising from the edges of the lobes (Rawlinson,
into inhalant and exhalant chambers. Each ciliated
2014). Some are able to capture and ingest phyto-
ridge bears three parallel ciliary bands (frontal, lat-
plankton (Ballarin and Galleni, 1987; Scarpa et al.,
erofrontal, and lateral, from inhalant toward exhal-
1996), and larvae of one species survive for longer
ant) that run perpendicular to the direction of flow.
when fed than when starved (Scarpa et al., 1996).
The lateral cilia beat to generate a current through
Feeding mechanisms of these larvae are unknown,
the mantle cavity. As water flows between the cili-
as is the size range of particles that they can cap-
ated ridges, some particles are retained by sieving
ture and ingest.
on the laterofrontal cilia, and others seem to trig-
Nemertea. Some members of all three major groups ger localized reversal of the direction of beat of the
of nemerteans are known or inferred to have feed- lateral cilia, pushing the particle back into the in-
ing larval stages. The uniformly ciliated planuliform halant chamber (Strathmann, 2006). Thus, cypho-
larvae of some Paleonemertea and Hoplonemer- nautes seem capable of using both filtration and
tea seem to capture and ingest large prey, by un- active response to capture particles. Captured par-
known mechanisms (Jagersten, 1972; Maslakova ticles may be transported toward the mouth on the
and Hiebert­, 2014). Most members of Pilidiophora frontal cilia. Others are apparently directed toward

the mouth by currents generated by reversal of the
lateral cilia, or, for particles sieved on laterofrontal
cilia, by flicks of the laterofrontal cilia toward the
inhalant chamber.
Brachiopoda. The larvae of lingulacean and disci-
niscid brachiopods feed on phytoplankton using
tentacles of the lophophore. Feeding is best known
in larvae of the lingulacean Glottidia pyramidata
(Strathmann, 2005). Its tentacles bear three parallel
ciliary bands: frontal, laterofrontal, and lateral. Lat-
eral cilia beat from anterior to posterior, generating
a swimming and feeding current. Particles passing a
tentacle can be retained on its frontal side by mech-
anisms similar to those used by cyphonautes larvae:
some are retained by sieving on the laterofrontal
cilia, and others seem to trigger localized reversal
of the direction of beat of the lateral cilia. Captured
particles are moved down the tentacle toward the
mouth, presumably by the action of frontal cilia.
Strathmann (2005) noted that a few particles caught
up in the feeding current moved directly into the
oral cavity without any direct interaction with the
ciliary bands of the tentacles.
Phoronida. Actinotrochs of phoronids have a mus-
cular hood over the mouth, and an arc of ten or
more larval tentacles extending ventrally from just
posterior to the mouth. Ciliation on the tentacles is
similar to that on the tentacles of brachiopod lar-
vae described previously. Lateral cilia beat from
anterior to posterior, generating a swimming and
feeding current (a ring of telotrochal cilia at the pos-
terior end of the larva assists in swimming). Film
and video of feeding actinotrochs show that some
particles passing the larval tentacles are retained on
their upstream sides briefly. This initial retention
of particles must be due to localized alteration of
beat of the lateral cilia (Riisgård, 2002) or to siev-
ing on the array of laterofrontal cilia, or both. Re-
tained particles are transported to the mouth via
the frontal cilia (Strathmann, 1973) or by localized
muscular expansion of the oral hood, presumably
generating suction that draws the particle into the
hood (Strathmann and Bone, 1997). Riisgård (2002)
sometimes observed tentacles flicking anteriorly;
such flicks might deliver retained particles to the
edge of the oral hood. Once in the oral hood, parti-
cles appear to be carried to the mouth by cilia.
L a r va l Fee d i n g    91
Mollusca. Veligers of gastropods and bivalves feed
using paired extensions of tissue, the velar lobes,
whose edges bear three parallel bands of cilia (Fig-
ure 7.1A). The long, compound prototrochal cilia
beat from anterior to posterior; metatrochal cilia beat
in the opposite direction; and food groove cilia beat
toward the mouth. Because prototrochal and meta-
trochal cilia beat in opposite directions, this is some-
times called “opposed band” feeding (alternatively,
“catch-up principle” feeding; Riisgård et al., 2000).
Cilia and particle movements in tethered veligers
suggest that as prototrochal cilia beat from anterior
to posterior they overtake suspended particles and
directly intercept them (Strathmann and Leise, 1979;
Gallager, 1988); evidence of adhesion between the
prototrochal cilia of a gastropod and captured parti-
cles is consistent with that interpretation (Romero et
al., 2010; but see Mayer, 2001). Particles smaller than
the metachronal length of the prototrochal ciliary
band are captured at high rates (Strathmann and
Leise, 1979; Gallager, 1988), suggesting that sieving
by prototrochal cilia is probably not important for
smaller particles; it may, however, be involved in
the capture of larger particles. The maximum size of
ingested particles may be determined by the length
of the prototrochal cilia or width of the food groove
(Hansen, 1991). The role of the metatrochal cilia in
particle capture in opposed band systems is un-
clear (Strathmann and Grünbau­m, 2006). Once cap-
tured, particles are transferred to the food groove
by unknown mechanisms. Food groove cilia may
bear stickier adhesive than do prototrochal cilia, fa-
cilitating transfer of particles, or the active strokes
of metatrochal cilia may scrape captured particles
off prototrochal cilia during the latter’s recovery
strokes. Once in the food groove, particles are car-
ried around the velar edge to the mouth, formed
into a bolus, and swallowed (Gallager, 1988).
Several authors have observed swimming ve-
ligers trailing mucus strands behind them. Such
strands might serve as an additional capture sur-
face that then must be ingested. Alternatively, they
might have an indirect role in feeding: the increased
drag associated with such structures would in-
crease shear around prototrochal cilia, enhancing
rates of direct interception (Emlet and Strathmann,
1985; Emlet, 1990; Fenchel and Ockelmann, 2002;
Strathmann and Grünbaum, 2006).
92   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
Annelida. Some trochophores in at least nine fami-
lies of annelids feed using an opposed band system
similar to that of veligers (Pernet et al., 2015). There
is substantial functional diversity in larval feeding
even within this set of taxa. For example, some feed-
ing larvae of Oweniidae and Capitellidae seem to use
a prototroch of simple, not compound, cilia (Emlet
and Strathmann, 1994; Pernet et al., 2015). Members
of two annelid families extend their opposed bands
on sinuous (Oweniidae: Emlet and Strathmann,
1994) or elongate (Amphinomidae: Jagerste­n, 1972)
lobes, but all other opposed band feeding annelids
bear relatively short opposed bands. Members of at
least two families, Opheliida­e and Echiuridae, use
opposed bands for feeding but also capture large
particles directly at the mouth (Miner et al., 1999).
Though the geometry and beat patterns of cilia in an-
nelids with opposed bands are similar to those of ve-
ligers, direct interception of particles by prototrochal
cilia has not been visualized in annelids. Assuming
that opposed band feeding annelids do capture par-
ticles by direct interception on prototrochal cilia, as
in molluscs, it is not known how captured particles
are transferred to the food groove.
Other feeding larvae of annelids lack food groove
and metatrochal cilia and so must feed by alterna-
tive mechanisms. One example is that of feeding lar-
vae in the large clade Phyllodocida (which includes
glycerids, hesionids, nephtyids, phyllodocids,
and polynoids, among others). Feeding has been
described in only one phyllodocidan, a polynoid
(Phillips and Pernet, 1996). Polynoid larvae bear a
tuft of long, compound cilia—the oral brush—on
the left side of the mouth. The prototroch generates
a swimming current. When large particles approach
or contact the larva’s anterior end (the episphere),
they move toward the mouth, perhaps under the
influence of episphere ciliary bands. The larva then
recoils, disengaging the particle from the episphere.
It then resumes rotating, with the particle now
held in front of the mouth, perhaps by the erected
oral brush. Eventually the particle is moved to the
mouth and swallowed. A second example is that of
the larvae of pectinariids. Larvae of at least one spe-
cies secrete and inhabit large, transparent, spheroid
“houses” whose walls are porous (Pernet, 2004).
The larva uses prototrochal cilia to pump water
through the house; particles retained in the house
are carried in a current to the elaborate oral hood
before ingestion.
Feeding mechanisms of some other annelid larvae
are largely or completely unknown (Strathman­ n,
1987; Pernet et al., 2002). Two taxa whose larval
feeding mechanisms are of particular interest are the
spionids and the sipunculids. Larvae of spionids are
often extremely common in coastal waters, and at
natural densities can sometimes control populations
of phytoplankton (Martin et al., 1996). Larvae of
one spionid use opposed bands for feeding (Pernet
and McArthur, 2006), but larvae of many other spe-
cies lack opposed bands and must feed in another
fashion. The pelagosphaera larva of sipunculids are
very different in form from other annelid larvae. Pel-
agosphaera larvae of some sipunculids are known
to feed, but the mechanisms by which they capture
particles are unknown (Strathmann, 1987).
Entoprocta. Feeding larvae of entoprocts have pro-
totrochal, food groove, and metatrochal ciliary
bands like those of feeding veligers and some an-
nelid trochophores (Mariscal, 1965; Jagersten, 1972).
Particle capture has not been described, but trans-
port in the food groove is as expected for an op-
posed band system.
Arthropoda. The larvae of crustaceans are strikingly
different from all other larvae described here, as
they lack locomotory cilia and instead swim and
feed using muscles to move pairs of jointed, setose
appendages. Three main types of crustacean larvae
are usually recognized: nauplii, which swim using
three pairs of cephalic appendages; zoeae, which
swim using thoracic appendages; and megalopae,
which swim using abdominal appendages (Harvey
et al., 2002). Free-swimming nauplii are widespread
in Crustacea, but zoeae and megalopae are found
only in Malacostraca.
Nauplii are often extremely common in the
plankton, but little is known about their feeding
mechanisms. Though they have only three pairs
of appendages, of which the posterior two (second
antennae and mandibles) probably play a primary
role in feeding, those appendages are morphologi-
cally complex and move at high frequency, making
it difficult to visualize their movements and those of
particles in relation to them. There is little consensus
on naupliar feeding mechanisms in either copepods

or cirripedes, the two best-studied groups. Strath-
mann (1987) suggests two areas of agreement: (a)
the proximal setae of the second antennae are used
as filters that retain captured particles, and (b) the
gnathobases of the second antennae and mandibles
aid in transport of captured particles to the mouth.
There has been little recent progress in under-
standing feeding by the nauplii of cirripedes, save
for the work of Stone (1988; 1989) suggesting that
the spacing of the setae and setules of the second
antennae is related to the size of cells that can be
captured and ingested efficiently. There has been
significantly more work on feeding by copepod
nauplii, with two studies of particular importance.
Paffenhöfer and Lewis (1989) studied both tethered
and free-swimming nauplii of three calanoids feed-
ing on diatoms. Nauplii of two of the species cre-
ated a feeding current using the second antennae
and mandibles, then responded actively to each
approaching particle by altering beat patterns of
the appendages nearest the particle. Eventually, a
combined inward motion of both mandibles moved
the particle to the mouth. Paffenhöfer and Lewis
inferred that detection of particles was by chemore-
ception when particles were immediately adjacent
to or in contact with setae of the feeding append-
ages, though mechanoreception seems a viable
interpretation as well. Nauplii of the third species
apparently did not create a feeding current, instead
swimming actively to encounter prey.
Bruno et al. (2012) examined prey detection and
capture in free-swimming nauplii of three cope-
pods. Nauplii of two species clearly detected ap-
proaching motile prey from a distance, presumably
by hydromechanical signals. Detection of a prey
item induced the nauplius to carry out an attack
jump (driven by movements of the two pairs of an-
tennae in a calanoid, and by movements of all three
pairs of cephalic appendages in a cyclopoid), swim-
ming past the prey to end with it slightly posterior
to the nauplius. The nauplius then used the second
antennae and mandibles to create a current toward
the mouth, moving the prey item in that direction.
The actual events of capture were not visualized,
but once the prey was captured, handling and in-
gestion involved additional movements of the sec-
ond antennae and mandibles. Nauplii of the third
species, another calanoid, captured particles from
L a r va l Fee d i n g    93
a feeding current. Approaching particles were de-
tected by contact with setae of an appendage, usu-
ally the first antennae. In some cases there was an
obvious response of the nauplius to the detected
particle, with a repositioning of the body before
capture, but in several other cases there was no ob-
vious response to particles before capture. These
alternative prey capture behaviors are associated
with distinct naupliar motility patterns, as demon-
strated by Titelman and Kiørboe (2003).
These studies show that there is diversity among
copepod nauplii in the behaviors associated with
prey encounter and capture, and that nauplii of at
least some copepods actively respond to detected
prey items. Some nauplii appear to be able to cap-
ture very small particles (e.g., bacteria: Turner and
Tester, 1989; Vogt et al., 2013) that seem unlikely to
provide chemical or hydromechanical cues detect-
able by nauplii, suggesting that filtering processes
might also be involved in feeding.
Later larval stages of malacostracans are ex-
tremely diverse in form (Harvey et al., 2002; Martin
et al., 2014). As noted by Strathmann (1987), though
the morphology of feeding appendages has been
described in detail, feeding mechanisms of zoeae
are not well known. Zoeae of some species are ca-
pable of some type of suspension feeding, captur-
ing relatively small phytoplankton cells, and even
tiny cyanobacteria (Factor and Dexter, 1993; Hinz et
al., 2001; Fileman et al., 2014), and most zoeae (es-
pecially later stages) also capture larger prey when
they encounter them. Sight does not seem to be
important in detection of large prey (Harvey and
Epifani­o, 1997). Once encountered, prey are grasped
using the endopods of the thoracic appendages, then
moved forward to be held by the maxillae and torn
and chewed by the mandibles. Megalopae are typi-
cally considered encounter feeders, though there is
some evidence of active response feeding on smaller
prey in feeding currents (McConaugha, 2002).
Echinodermata. Feeding larvae of echinoderms bear
a long sinuous band of simple cilia around the oral
field (Figure 7.1C). Cilia on this band beat away
from the oral field to generate a swimming and
feeding current. Particles appear to be captured
by two distinct mechanisms. Most commonly, par-
ticles entrained in the current and approaching
94   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
the ciliary band trigger localized reversal of cili-
ary beat, pushing the particle toward the larva’s
mouth (Strathmann, 1971). The stimulus trigger-
ing ciliary reversal is likely mechanical, which
suggests that very small particles should not be
detectable by this mechanism (Strathmann, 2007).
Redirected particles may be recaptured several
times, appearing to “bounce” toward the mouth,
before they are eventually carried into the buccal
cavity by cilia-generated currents (Hart, 1991). In
these active responses of cilia to detected particles,
particles are generally not contacted by cilia, but
instead local water currents are altered so that a
parcel of water containing the particle is redi-
rected toward the buccal cavity. The complex flows
around echinoderm larvae, as well as direct obser-
vation of captures, suggest that different parts of
the ciliated band may vary in their effectiveness at
capturing particles (Hart, 1991; Gilpin et al., 2016).
Hart (1991) estimated that ~95% of the captures he
observed occurred by ciliary reversal. The remain-
ing ~5% occurred without ciliary reversals, when
particles entrained in the feeding current simply
followed a path that took them directly into the
buccal cavity. Peristaltic contractions of the mus-
cular esophagus push collected particles into the
stomach (Strathmann, 1971).
Hemichordata. Some enteropneusts have feeding
tornaria larvae, which, like the larvae of echino-
derms, have a sinuous band of simple cilia around
the oral field. Young tornariae have a relatively
simply organized ciliary band, but as they increase
in size it becomes much more convoluted, some-
times developing small protrusions called tentacles
(see Hadfield, 2002). Tornariae also bear a posterior
ring of compound cilia, the telotroch, which is used
to generate a swimming current. Strathmann and
Bonar (1976) described feeding in larvae of two
species. Most captures involve localized ciliary re-
versal, as described for echinoderms. Detected par-
ticles are redirected to the food grooves, and then
carried by their cilia to the buccal cavity. As in echi-
noderms, a few particle captures occur when par-
ticles reach the oral field without being redirected
by ciliary reversal. Once in the oral field, captured
particles are carried in food grooves to the buccal
cavity. Particles are passed from the mouth to the
esophagus by ciliary activity; ciliary action appar-
ently drives them into the stomach, as well, in con-
trast to echinoderms, where particles move from
esophagus to stomach by peristaltic contractions of
esophageal muscles.
Hart et al. (1994) made observations of particle
movements around a living planctosphaera, which
is likely a giant (up to ~28 mm in diameter) entero-
pneust larva. The individual examined was 18 mm
in diameter, but its convoluted ciliary band was an
estimated 1.5 m in length. Particle movements sug-
gested that captures occurred by localized ciliary
reversal, followed by transport to the mouth in the
ciliated food grooves.
7.3  Predicting Larval Feeding
Performance from Form and Feeding
Mechanism
It would be ideal if we could combine knowledge
of the physical processes of particle encounter and
capture with data on the size, arrangement, and
kinematics of feeding structures to yield predictions
of larval feeding performance in a given environ-
ment. Though theoreticians have modeled physi-
cal processes of particle encounter using idealized
capturing structures, such models have not been
scaled up to the level of an individual larva, whose
many capturing structures are organized in three-
dimensional arrays and move in complex patterns
(e.g., rows of cilia beating in metachronal waves,
with distinct active and recovery strokes). Progress
toward this goal is being made, however (e.g., Ding
and Kanso, 2015). Though not quite as satisfying, it
is currently possible to make predictions of feeding
performance from coarser models that include only
information on the size and arrangement of feeding
structures, and estimates of water flow past them.
Before reviewing these, however, it is important
to define “feeding performance.” Two metrics of
performance are commonly considered in studies of
larval feeding. First is clearance rate—the volume of
water a larva clears of particles per unit time. With
data on particle concentrations in the environment,
clearance rates can be converted to ingestion rates.
Clearance rates can be estimated in a diversity of
ways. Maximum clearance rates can be estimated

by directly observing captures by free-swimming
larvae over brief periods of time (seconds–minutes)
or by quantifying ingestion rates during brief incu-
bations of larvae in suspensions of known concen-
tration. Steady-state clearance rates are estimated in
longer-term (hours) observations, usually by quan-
tifying rates of decline in particle concentration in
incubation chambers. Estimates are usually made
in suspensions of a single type of particle. These
are usually natural (e.g., laboratory-cultured uni-
cellular algae), but artificial particles (e.g., plastic
beads) are often used to estimate maximum clear-
ance rates. All methods of estimating clearance rate
except for direct observation should yield under-
estimates of maximum clearance rate (Appelmans,
1994). Maximum clearance rate is an appropriate
metric of the performance of larvae in food-limited
conditions, where ingestion rates are likely limited
by particle capture rates and not by downstream
digestive processes. Many studies report on steady-
state clearance rates of various invertebrate larvae;
fewer report on maximum clearance rates. A compi-
lation of maximum clearance rates of diverse larvae
is provided by Strathmann (1987; his Table 4.1).
A second metric is the size spectrum of catchable
particles, or, more specifically, the shape of the re-
lationship between clearance rate and particle size.
Potential food particles in the plankton vary in size
over at least four orders of magnitude, from bac-
teria and picoeukaryotes on the order of 0.1 µm in
maximum dimension to phytoplankton aggrega-
tions or the embryos or larvae of other species 1000
µm or more in maximum dimension. Further, parti-
cle size distributions vary over both space and time.
One might imagine that larvae capable of feeding at
high rates on a wide range of particle sizes would
perform better than those restricted to feeding only
on particles in a narrow size range. Though larval
feeding mechanisms undoubtedly do constrain the
size spectra of catchable particles, we know rela-
tively little about how clearance rate varies with
particle size for most larvae.
7.3.1 Predicting Maximum Clearance Rate
The maximum clearance rate of any suspension-
feeding larva may be modeled by estimating the rate
that water passes by filtering structures or through
L a r va l Fee d i n g    95
the perceptive volume of an actively responding
larva, and making assumptions about the propor-
tion of particles moving through this volume that
are captured and retained (in the absence of other in-
formation, investigators usually assume 100%). For
larvae that feed using bands of cilia, Strathmann et
al. (1972) pointed out that this could be estimated as:
C = lb (lc−lr) v
where C is maximum clearance rate, lb is the length
of the relevant ciliary band, lc is the length of the
cilia in that band, lr is a correction factor accounting
for the ciliary recovery stroke, and v is the velocity
of particles moving in the region of the ciliary band
between lr and lc.
Numerous authors have used models of this sort.
Strathmann and Leise (1979) created a slightly more
complex model incorporating empirically estimated
retention rates to predict clearance rates of veliger
larvae. Gallager’s (1988) estimate of “flow through
the capture zone” based on similar morphological
and kinematic criteria for a bivalve veliger can be
used as an estimate of clearance rate. Several authors
have used even simpler models, omitting the correc-
tion for the ciliary recovery stroke (e.g., McEdwar­d
and Strathmann, 1987; Hansen et al., 2010). As noted
by Strathmann et al. (1972), the most difficult param-
eters to estimate for such models are velocity pro-
files of particles through the ciliary band. There is
a close correlation between cilium length (whether
simple or compound) and maximum velocity of
particles through a feeding ciliary band (Emlet and
Strathmann, 1994, their Fig. 12.4), so it might be
possible to predict maximum velocities from cili-
ary length using this relationship. McEdwar­d and
Strathmann (1987) used half of ciliary tip velocity as
to approximate average velocity across the length of
cilia in a band; Pernet and Strathmann (2011) used
half of maximum particle velocity.
Though not maximally mechanistic, such models
yield useful generalizations about how larval form
affects feeding performance of larvae that feed with
ciliary bands. In echinoderm larvae, cilia in the
feeding ciliary band do not increase in length with
growth, but ciliary band length increases dramati-
cally with growth (McEdward, 1986); thus maxi-
mum clearance rate increases largely as a function of
ciliary band length (Strathmann, 1971; Hart, 1996).
96   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
In most larvae that feed with opposed bands, the
prototrochal band increases in length with growth
(Henderson and Strathmann, 2000), but cilia in that
band also increase substantially in length and angu-
lar velocity (Emlet and Strathman­n, 1994); in these
species, maximum clearance rate thus depends not
only on prototrochal band length but also on the
length of cilia in the band.
Because larvae with longer cilia in the ciliary
band should have higher clearance rates per length
of ciliary band (because of increased area of captur-
ing surface and higher angular velocities of cilia),
and because larvae that feed using ciliary reversal
tend to have short simple cilia compared to the often
long compound cilia of most opposed band feed-
ers, larvae that feed with opposed bands may often
have higher per band-length clearance rates than
larvae that feed by ciliary reversal. Such a pattern
is not evident in data on clearance rates compiled
from the literature by Strathmann (1987). Planned
comparisons are needed to test this hypothesis. If
high band-length specific clearance rates are char-
acteristic of opposed band feeders, this might help
explain the smaller egg sizes of many species with
larvae that feed with opposed bands, compared to
egg sizes of echinoderms, whose larvae feed using
ciliary reversal (Strathmann and Vedder, 1977).
McEdward and Strathmann (1987) found that the
maximum band-length specific clearance rates of a
cyphonautes and a pluteus were roughly similar,
but that the cyphonautes had a much shorter cili-
ated band relative to body size than did the pluteus,
suggesting that feeding capacity relative to body
size was lower for the cyphonautes than for the plu-
teus. The consequences of this are not clear. Cypho-
nautes develop from very small eggs, but co-occur
with plutei in similar habitats and seasons, develop
to a similar size at metamorphosis, and do not seem
to have extraordinarily long planktonic periods.
Similar models aimed at estimating the volume
of water sampled for food per unit time have been
used to predict clearance rate in crustacean nauplii,
with parameters of naupliar size, perception dis-
tance, and difference in velocity between nauplius
and prey (Titelman and Kiørboe, 2003; Bruno et
al., 2012). Predictions of such models are similar to
empirical estimates of clearance rate in at least one
species.
7.3.2 Predicting the Size Spectrum of Catchable
Particles
Knowledge of the physical processes of particle
capture used by a larva should permit us to predict
the rates at which it can capture of particles of dif-
ferent sizes. There may be systematic differences
in the size spectrum of catchable particles among
larvae with different feeding mechanisms. For ex-
ample, larvae that feed with opposed bands of cilia
appear to capture particles by direct interception on
prototrochal cilia. For an isolated cylindrical col-
lector, rates of encounter and clearance by direct
interception should be proportional to particle di-
ameter (Shimeta and Jumars, 1991). Thus one might
predict that clearance rates in larvae that feed using
opposed bands should be proportional to particle
diameter, at least up to a limit where increased drag
on large particles reduces retention on capturing
surfaces, or where particles are too large (e.g., larger
than the width of the food groove) to be transported
efficiently to the mouth after capture (Hansen, 1991;
1993; but see Romero et al., 2010). Recent efforts to
model particle capture by a prototroch-like band of
cilia suggest that the particle size-specific efficiency
of capture depends on the length of prototrochal
cilia as well as their beat patterns (Ding and Kanso,
2015), but these predictions have not yet been tested.
Likewise, in larvae that capture particles by siev-
ing (e.g., cyphonautes), all particles larger than the
spacing between sieve elements (laterofrontal cilia,
spaced ~3–4 µm apart; Nielsen, 2005; Strathmann,
2006) should be captured. This suggests that cy-
phonautes should capture particles ≤3 µm in diam-
eter at low rates, and particles above this size at
high rates. The only data relevant to this predic-
tion are those of McEdward and Strathmann (1987)
showing that cyphonautes capture 10 µm spheres
at clearance rates an average of 60 times higher
than those of 2 µm spheres (mean of three experi-
ments in their Table IV).
Many larvae (e.g., echinoderms) use cilia to de-
tect approaching particles in a current by chemical
or hydromechanical signals, then reverse the beat of
cilia near the particle to separate it from the feeding
current and move it toward the mouth. Smaller cells
will generate smaller signals (Visser, 2001; Strath-
mann, 2007; Kiørboe, 2008), so one might predict
 L a r va l Fee d i n g    97

cells below a size that will stimulate larval sensory
cells should not be captured. The sensitivity of the
relevant sensory cells is not known, so predictions
about the minimum size of particles that can be de-
tected are not yet possible. Nauplii of copepods also
may detect approaching particles by hydromechan-
ical or chemical signals, and thus should not react
to or capture particles too small to stimulate naup-
lius sensors (mechano- or chemosensory setae). The
sensitivity of the sensors of copepod nauplii has not
been assessed directly, though they may be similar
to those of adults (Yen et al., 1992). As noted earlier,
knowledge of the feeding mechanisms of cirripede
nauplii remains scant, but Stone (1988; 1989) sug-
gested that the spacing of the setae and setules of
the second antennae is related to the sizes of cells
that can be captured and ingested efficiently.
As indicated previously, we know rather lit-
tle about the size spectrum of catchable particles
for most types of larvae. There are even fewer
planned comparisons of this performance met-
ric among larvae with different feeding mecha-
nisms. One such comparison suggests that larvae
that feed using opposed bands of cilia (annelids
and molluscs) have their highest clearance rates
on relatively small particles, consistent with the
idea that the width of the food groove places an
upper limit on the size of particles that can be ef-
ficiently transported to the mouth, while larvae
that feed using ciliary reversal (echinoderms)
have their highest clearance rates on larger par-
ticles, consistent with the idea that large particles
are more likely to induce ciliary reversal events
than smaller particles (Figure 7.2).
Building explicit models of how larval form and
feeding mechanism affect feeding performance and
testing their predictions is an obvious way to make
progress in our understanding of the form and
function of feeding larvae. Hart and Strathmann
(1995) point out the value of this approach, and the
hazards of ignoring it. Such studies remain rare.
Feeding rates are often quantified without refer-
ence to larval form or function, which makes it dif-
ficult to interpret unexpected results. For example,
several workers have inferred high clearance rates
on bacteria by annelid, echinoderm, and crustacean
larvae (Rivkin et al., 1986; Turner and Tester, 1989;
Pile and Young, 2006). High clearance rates on such

1.0 echinoid

0.8 asteroid
relative clearance rate

0.6

0.4 gastropod

0.2
annelid

0.0
0 5 10 15 20 25 30
particle diameter (μm)

Figure 7.2 Relative clearance rate as a function of particle diameter for two larvae that feed with opposed bands (solid lines: the annelid Hydroides
gracilis and the gastropod Crepidula onyx), and two larvae that feed using ciliary reversal (dotted lines: the asteroid Patiria miniata and the echinoid
Dendraster excentricus) (B. Pernet and A. McKee, unpubl. data). Larvae of each species were offered uniformly flavored polystyrene divinylbenzene
spheres of a variety of sizes (0.45, 1, 3, 6, 10, 15, 20, and 30 µm diameter, one size at a time) at low concentrations and allowed to feed for 10 min at
16 °C. Ingestion rates were then estimated from gut contents. Clearance rates for each species are scaled to the maximum for that species.
98   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e L a r va e
small particles are not predicted by hypotheses on
the feeding mechanisms of any of these species,
suggesting either that measured rates are overesti-
mates or that our understanding of how those lar-
vae feed is incomplete.
7.4  Inferences on Larval Feeding
Performance in Nature
An ultimate goal of studies of larval feeding is to
understand how larval form and feeding mecha-
nism affect development rate, planktonic mortality,
and eventually recruitment to benthic populations
in nature. What steps are necessary to approach
that goal? I suggest two general areas where pro-
gress is needed.
First, as described earlier, we need more detailed
mechanistic models linking larval form and feeding
mechanism to feeding performance, especially on
the relationship between clearance rate and particle
size, as well as laboratory validation of the predic-
tions of these models. As noted, our knowledge of
these linkages is not yet strong for any type of larva.
If we are unable to successfully predict larval feed-
ing performance in simplified laboratory situations,
our chances of doing so in complex natural habitats
are slim. Progress in the development of models that
make specific predictions about the particle sizes that
can be captured by ciliary arrays of various sorts has
been made recently (Tripathi et al., 2013; Ding and
Kanso, 2015). Designing such models for the com-
plex geometries of actual larvae should yield testable
predictions on larval feeding performance.
One aspect of such models that especially needs
improvement is in the incorporation of thermal ef-
fects on larval feeding. Such data are necessary to
understand seasonal and geographic patterns in
feeding performance (e.g., feeding performance
in temperate vs. polar echinoderm larvae), and to
make predictions about the effects of changes in
ocean temperature on feeding performance (Byrne
et al., this volume). This is critical in part because
of the complex effects of temperature on feeding
larvae: not only does it affect rates of biochemical
processes, but it also affects the viscosity of seawa-
ter. These changes may have dramatic effects on
feeding performance. Podolsky (1994) and Bolton
and Havenhand (1998) found that reductions of
10 °C led to ~60% reductions in ingestion rate in
larvae of an echinoid and an opposed band feed-
ing annelid, respectively. In the echinoid, changes
in viscosity independent of temperature affected
rates of consumption of particles of different sizes,
with large particles making up a larger propor-
tion of the ingested particles in high-viscosity
conditions. Quantification of how temperature
affects ciliary beat frequency, angular velocity,
and water movement through the ciliary band in
various types of larvae would permit these vari-
ables to be included in models predicting feeding
performance.
Second, prediction of larval ingestion rates in
natural settings will only be possible with detailed
understanding of the diversity and distribution of
particles at spatial scales relevant to larvae. Our
knowledge of the particles larvae interact with is
most frequently derived from proxy indices like
chlorophyll concentration averaged over sample
volumes many orders of magnitude greater than
that of an individual larva. The low spatial resolu-
tion of such measurements, and the fact that they
ignore potential food particles that do not contain
chlorophyll but include chlorophyll-containing
particles that may not be suitable as food (Olson
et al., 1987; Reitzel et al., 2004), means that we can-
not use such data to make good predictions about
encounter rates of larvae with potential food items.
Another approach is to sample smaller volumes
(tens of ml) of seawater and identify and measure
all the particles they contain (Bos et al., 2006b). This
yields data of much higher resolution, but is labor
intensive. A diversity of automated devices able to
collect higher resolution data on the particle popu-
lation surrounding larvae are now available. These
include in situ flow cytometers (Almeda et al., 2009),
some of which are capable of imaging particles as
they pass through the flow cell (Olson and Sosik,
2007). Such high-resolution sampling has led to the
identification of spatial patterns in the concentra-
tion of particles at a variety of scales (e.g., in thin
phytoplankton layers; Durham and Stocker, 2012).
Although when characterizing particles in the
plankton the focus is typically on the abundance of
potential food items, it is also important to consider
that particles vary in their catchability, and that

some particles may actually interfere with feeding.
Many protists have escape responses, for example
(Jakobsen, 2001); we do not know if larval feeding
behaviors trigger these responses, or whether they
are effective in preventing capture. Larvae have up-
per limits on the sizes of particles they can capture
or swallow, but particles larger than these size limits
may frequently be common. Hansen et al. (1991) as-
sessed whether or not such large inedible particles
might reduce clearance rates by a veliger on edible
particles. They found that the presence of large in-
edible particles (plastic beads 45 µm in diameter) at
concentrations of 2,500•ml-1 or more reduced clear-
ance rates on edible particles. Additional studies
aimed at understanding what types of planktonic
particles are easily catchable by larvae, and what
types may actually hamper feeding, will be very
useful in interpreting rates of feeding in nature.
One can imagine combining models of larval feed-
ing performance as a function of temperature with
detailed descriptions of the larval feeding environ-
ment and physiological data on digestion, assimila-
tion, growth, and the attainment of competence in
bioenergetics models. Such models could be tested
by rearing cohorts of larvae in in-situ chambers or
mesocosms, with frequent and automated sampling
of temperature and chamber particle environment.
With such validated models in hand, we could
begin to make predictions about how larval form
and feeding mechanism affect growth and plank-
tonic larval duration in natural habitats, as well as
about how larvae might respond to challenges like
harmful algal blooms, ocean warming, and ocean
acidification.
7.5 Summary
1. Marine invertebrate larvae from a dozen phyla
must feed on suspended particles to fuel growth
and development to metamorphosis.
2. Larvae capture particles using just a few physi-
cal processes, but implement these processes in
diverse morphologies and behaviors.
3. Detailed knowledge of mechanisms of larval
feeding permit investigators to make predictions
about how rapidly larvae can capture particles of
different types.
L a r va l Fee d i n g    99
4. Mechanistic models of larval feeding can be
combined with detailed data on food avail-
ability in nature and integrated into broader
bioenergetics models to yield increased under-
standing of the biology of larvae in complex
natural habitats.
Acknowledgments
I thank D. Pace for helpful discussion, and B.
Hansen and the editors of this volume for sug-
gestions that helped improve an earlier version of
this manuscript. This work was supported by Na-
tional Science Foundation grants OCE-1060801 and
DEB-1257355.
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 C h a pt er 8
Phenotypic Plasticity of Feeding
Structures in Marine Invertebrate
Larvae
Justin S. McAlister and Benjamin G. Miner
Researchers have worked over the last thirty years
to examine and understand phenotypic plasticity
in larvae of marine invertebrates. These studies
range from documenting the association between
algal food availability and larval morphology in
different species to examining the ecological and
evolutionary implications of the plastic response.
However, investigators have used disparate sys-
tems (genera and species of echinoid echinoderms
mostly, but also other taxa), employed various
rearing and measuring techniques and levels of
genetic replication, and relied on different statisti-
cal methodologies for data analysis. They have also
obtained results ranging from non-plastic to plastic,
and reached different conclusions about the func-
tional, ecological, and evolutionary roles of pheno-
typic plasticity in marine larvae. Our goals with this
chapter are twofold: to review the literature in order
to help researchers more quickly understand the
extent and breadth of this field of research, and to
identify gaps in our understanding in order to pro-
vide guidance for future research. We begin with a
more general introduction of phenotypic plasticity,
and then focus on feeding-structure plasticity in lar-
vae of marine invertebrates.
8.1  Phenotypic Plasticity
Research on the expression and evolution of phe-
notypic plasticity is a rich field, and readers will
find valuable entries into the literature through
the books of West-Eberhard (2003) and DeWitt and
McAlister, J. S. and Miner, B. G., Phenotypic plasticity of feeding structures in marine invertebrate larvae. In. Evolutionary Ecology
of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018).
© Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0008
Scheiner (2004). Here we summarize some of the
main concepts in order to facilitate discussion of
plasticity in marine invertebrate larvae later in the
chapter. Phenotypic plasticity is defined as the abil-
ity of a genotype to produce different phenotypes
(developmental, physiological, morphological, be-
havioral, or life history traits) in response to differ-
ent biotic or abiotic environmental factors. Plasticity
can be adaptive, maladaptive, or effectively neutral.
When an organism possesses plasticity that confers
a fitness advantage, then plasticity is considered
adaptive. Researchers have focused on examples in
which there is evidence that plasticity is adaptive,
likely because of the evolutionary and ecological
consequences of phenotypes that improve an organ-
ism’s fitness. However, phenotypic plasticity can be
non-adaptive and still have important ecological
consequences (Miner et al., 2005). It is often difficult
to experimentally test whether plasticity is adaptive
because phenotype and environment are typically
confounded, and evidence often comes from both
experiments and functional arguments. In addition,
adaptation that leads to a fitness advantage (en-
hanced reproductive success of phenotypes across
generations) is difficult to demonstrate for larval
traits of organisms because of the time required to
rear organisms to the adult stage. Plastic pheno-
types that are inducible are also classified broadly
as either defensive or offensive (Miner et al., 2005).
Plastic phenotypes that protect an organism when
at risk of death or injury are inducible defenses,
whereas those that help an organism gain resources
104   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e La r vae
when resources are limiting are inducible offenses.
In some cases, inducible defenses and offenses both
occur and are integrated in an organism.
There are several general categories of pheno-
typic plasticity—developmental, morphological,
physiological, behavioral, or life historical. Re-
sponses assigned to the same category often have
similarities in the timing and reversibility of the
response. Morphological changes typically require
relatively more time to produce than physiological
and behavioral changes. Physiological, behavioral,
and morphological changes are often, although not
always, reversible when the stimulating environ-
ment is allayed. Changes to an organism’s develop-
mental trajectory or timing of transitions between
life-history stages can be difficult or impossible to
reverse. Though specifically defined responses are
relatively easy to fit into one of these categories,
organisms generally respond to their environment
with an integrated response of many traits that
are defined by more than one category. For exam-
ple, some species of plant elongate their stem and
grow taller when shaded by other plants (West-
Eberhard, 2003). The response is morphological
but is the direct result of underlying physiological
and developmental plasticities. In addition to cause
and effect relationships among plastic responses,
energy can link different plasticity responses in an
organism. Responses that require more energy can
cause changes in development or life history tran-
sitions, such as metamorphosis from larva to juve-
nile (West-Eberhard, 2003; Gilbert and Epel, 2015).
Often, delays in life history transitions result from
an initial energetically expensive morphological or
physiological plastic response.
In order for plasticity to evolve, organisms
must detect reliable cues of environmental change
(chemical, tactile, etc.) and respond at an appro-
priate temporal scale. Thus, plasticity must occur
after these cues are received and processed. In or-
der for the organism to properly match phenotype
to environment, these cues must be reliable, pro-
viding information of environmental variability
(e.g., magnitude and frequency). In addition, an
induced phenotype must be well timed with en-
vironmental change. To ensure that the induced
plastic response is effective, the period of time
(i.e., lag-time) between sensation of the cue and
production of the phenotype must be appropriate
to the scale of environmental change (Padilla and
Adolf, 1996).
The degree of plasticity for a given genotype is
often measured as either the difference in the mag-
nitude of phenotypes expressed between two envi-
ronments or the slope of the reaction norm across
environments. The reaction norm for a given geno-
type depicts graphically the relationship between
phenotype and a specific environmental factor. The
phenotype is plastic when the slope of the relation-
ship deviates from zero, and non-plastic (or con-
stant) when equal to zero. The degree of phenotypic
plasticity can vary among genotypes within a pop-
ulation. Natural selection can act on this variation,
and thus the degree of phenotypic plasticity for a
given trait is considered a trait in and of itself, with
the potential to evolve. Numerous studies have
documented the expression of phenotypic plastic-
ity by organisms in general, but fewer have dem-
onstrated variation for plasticity among genotypes
both within and among populations, and fewer
still have demonstrated that phenotypic plasticity
has evolved in response to environmental changes
and results in higher fitness. Research on pheno-
typic plasticity has a long history among terrestrial
plants and insects, and a relatively recent history
among marine invertebrates, particularly in early
life stages (e.g., larvae).
Marine invertebrates and their larvae face se-
lective pressures in their environments that are
similar in some ways to those faced by terres-
trial plants and the freshwater juvenile stages of
some terrestrial animals (e.g., predation, compe-
tition, and dispersal), yet are uniquely different
in others (e.g., salinity and CO2-induced ocean
acidification). Also, most freshwater inverte-
brate species are taxonomically different, de-
veloping directly and not through larval stages
like marine invertebrates, although a few excep-
tions exist. Thus marine invertebrates and their
larvae constitute a set of research systems that
provide a counterpoint to terrestrial and fresh-
water organisms for testing our ideas about the
biology of phenotypic plasticity. We believe that
food-induced phenotypic plasticity exhibited by
the planktotrophic (i.e., feeding) larvae of many
 P h e n ot y p i c P l as t i c i t y o f Fee d i n g S t r u c t u r es    105
marine invertebrates provides one of the most
comprehensive examples of phenotypic plasticity
among marine organisms.
8.2  Feeding Larvae of Marine
Invertebrates
The life histories of most species of marine inverte-
brates involve transitions through a series of suc-
cessive developmental and ontogenetic stages (see
Nielsen, this volume). Larvae are typically classi-
fied by whether they can acquire exogenous food
and complete larval development using endog-
enous reserves in an egg. Feeding planktotrophic
larvae must feed to gain nutrition from the plank-
tonic environment, as they do not possess enough
energy in an egg to complete development and
metamorphose into a juvenile, whereas lecitho-
trophic larvae utilize primarily egg energy and
might have the ability to feed, though feeding is not
required. The combination of these two classifica-
tion schemes results in four possible types of larvae,
although only three are evolutionarily viable: feed-
ing planktotrophic larvae, feeding lecithotrophic
larvae, and nonfeeding lecithotrophic larvae. The
fourth possibility, nonfeeding planktotrophic lar-
vae, are not viable because there is no way for lar-
vae to gain the energy needed to metamorphose.
The vast majority of species with larval forms are
either feeding planktotrophic, typically referred to
as just “planktotrophic” larvae, or nonfeeding lec-
ithotrophic larvae, typically referred to as just “lec-
ithotrophic” larvae (Thorson, 1950; McEdward and
Miner, 2001; Marshall and Keough, 2008). Less com-
mon are species with feeding lecithotrophic larvae,
typically referred to as “facultative planktotrophic”
larvae (Allen and Pernet, 2007). In this chapter, we
focus on plasticity of the feeding structures of feed-
ing larvae, however, plasticity of other life history
characters are likely to exist and play important
roles for larvae of all developmental modes. For ex-
ample, adults might preferentially, and plastically,
provision eggs with different types and amounts
of biochemical constituents, and thus energy, by
consuming different food resources or altering pat-
terns (e.g., rate and timing) of oogenesis. While this
type of plasticity will impact larval development
and growth, the consequences of plasticity of egg
organic and energy composition also affect adult
fecundity and lie outside the scope of this review.
The duration of larval growth for feeding larvae
is influenced by temperature and the availability
of energetic resources (see Pernet, this volume;
Jaeckl­e, this volume). In general, higher tempera-
tures increase metabolic and growth rates, shorten-
ing the amount of time individuals spend as larvae
(O’Connor et al., 2007). Most feeding larvae actively
acquire and assimilate energy and needed resources
by capturing and consuming unicellular algae us-
ing elaborate ciliated feeding structures, which vary
greatly among taxonomic groups (Strathmann,
1987). Exogenous energetic materials can also be
acquired passively through the uptake of dissolved
organic materials (Manahan et  al., 1983). The de-
gree to which feeding larvae must acquire exoge-
nous food and resources is correlated with egg size
(Herrer­a et  al., 1996; Miner et  al., 2005; McAlister
and Moran, 2013). Some species require no or very
little additional energy or resources beyond what is
present in the egg (Allen and Pernet, 2007). Other
species cannot survive beyond the stage when they
gain the ability to feed. Small eggs contain less en-
ergy than larger eggs (Jaeckle, 1995; McEdward and
Morgan, 2001), and presumably larvae that develop
from relatively smaller eggs likely require more
food for metabolism and morphogenesis than lar-
vae that develop from larger eggs. Feeding larvae
of some echinoid species also require the hormone
thyroxine to metamorphose, which they gain from
their diet (Heyland and Hodin, 2004; Heyland et al.,
2004). The possibility exists that small eggs might
also not contain enough of these non-energetic com-
pounds needed for metamorphosis, although this
remains to be more critically tested.
In preparation for the transition between larval
and juvenile life, and to ensure successful meta-
morphosis, feeding larvae from various taxa begin
constructing juvenile structures by metamorphosis
within the larval body. Taxonomic groups of marine
species vary in the timing and duration of this tran-
sition, as well as in the mechanisms underlying this
process. In many species larvae can metamorphose
rapidly and transition quickly between pelagic and
benthic environments (e.g., echinoderms), in other
species the transition is gradual (e.g., molluscs).
106   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e La r vae

However, in species in which larvae and juveniles

live in different environments (e.g., the plankton ~100 μm
and benthos), the ecological transition is almost
always quick, regardless duration of the morpho-
logical transition; thus metamorphosis represents
a critical developmental milestone for larvae. With
metamorphosis serving as a definitive end-point for
larvae, natural selection should favor mechanisms
that decrease the duration of the larval period—for
example, plasticity of structures associated with
food capture or processing or via increased mater-
nal provisioning, both of which ameliorate the ef- Low fed High fed
fects of oligotrophic conditions (see following and
Marshall et al., this volume). Figure  8.1 Low-fed (left) and high-fed (right) full-sib larvae of
Lytechinus variegatus at four days post-fertilization. Note longer larval
arms, both absolutely and relative to body length, of low-fed larva.
8.3  Plasticity of Feeding Structures
in Planktotrophic Larvae and holothuroid echinoderms, as well as in mol-
luscan veligers (Table 8.1), although there are many
8.3.1  Food Limitation, Resource Acquisition, phyla and larval types for which feeding-structure
and Energetic Trade-offs plasticity is undemonstrated or undocumented
(Table 8.2). By increasing the length of the ciliated
Larvae in the sea are presumably food limited be-
band and supporting body form, the larval surface-
cause the unicellular algae they eat are dilute and
to-volume ratio increases. This could also increase
patchily distributed in time and space (Conover,
the intake of dissolved organic matter (Manahan
1968; Paulay et al., 1985; Fenaux et al., 1988; 1994),
et al., 1983), albeit also increasing the surface area
and larvae have limited ability to track patchy
over which dissolved organics can leak, and thus
food. Thus, natural selection has likely favored
the net benefit is not clear.
strategies that improve the acquisition of scarce
Investing energy to lengthen larval feeding struc-
resources and reduce the amount of energy and
tures in low food conditions might result, how-
materials that need to be obtained from the envi-
ever, in decreased energetic investment in larval
ronment. Researchers in the late 1980s discovered
food-processing structures (e.g., the larval stomach;
an example of the first strategy; morphological
Miner, 2005) or juvenile structures required for met-
phenotypic plasticity in response to the amount
amorphosis (e.g., the juvenile rudiment; Boidron-
of food was demonstrated in feeding larvae from
Metairon, 1988; Strathmann et  al., 1992; Bertram
echinoid echinoderm species (Boidron-Metairon,
et al., 2009; see also Adams et al., 2011). In addition
1988; Fenaux et  al., 1988; Hart and Scheibling,
to increasing ciliary band length, larvae might also
1988). Larvae fed a dilute concentration of food
adjust capacity for capturing food relative to de-
respond by growing longer skeletal arms, which
mand for energy and materials for growth of post-
increases the length of the ciliated band used for
larval structures. For example, another response to
food collection (Strathman­ n, 1971; McEdward,
starvation might be to retain a given ciliary band
1986b; see Figure  8.1), thereby allowing larvae to
length but to reduce parts of the larval body not es-
capture more phytoplankton food particles from a
sential for larval life (e.g., the rudiment or epaulets).
greater volume of water (Hart, 1991). Furthermore,
This reduction of unessential larval tissues presum-
Hart and Strathmann (1994) demonstrated that
ably reduces demand for energy and materials
plasticity of ciliated band length was correlated
for maintenance, and uses the resorbed tissues as
with plasticity of arm length. Subsequent work
a source of both energy and materials, as demon-
has documented similar feeding-structur­e plastic-
strated in annelid trochophores (Pawlik and Mense,
ity in larvae of other echinoid species, ophiuroid,
 P h e n ot y p i c P l as t i c i t y o f Fee d i n g S t r u c t u r es    107

Table 8.1  Marine Invertebrate Species Tested for Larval Feeding-Structure Plasticity.

Phylum Plasticity? Statistical Methodology Genetic Replication Reference

Class Species Measure Type

Echinodermata
Asteroidea
Acanthaster planci Yes: 1, 2 ANOVA, PCA 2F, 1M Wolfe et al. 2015
Luidia foliolata Yes: 0 ANOVA 1F, 1M for 1 full-sib family George 1994
Pisaster ochraceus Yes: 1, 2 ANOVA, Canonical Unspecified; multiple F and M George 1999
Discriminant Analysis
Sclerasterias mollis Yes: 1 PCA, Wilcox 3-way 4 parents (presume 2F, 2M) per Poorbagher et al. 2010a
population
Echinoidea
Centrostephanus rodgersii No: 1, 2 ANOVA, ANCOVA, PCA 1F, 1M each for 3 full-sib families Soars et al. 2009
Clypeaster subdepressus Yes: 2 ANOVA 1F, 1M for 1 full-sib cross Reitzel and Heyland 2007
Dendraster excentricus Yes: 0 ANOVA Unspecified # of parents Boidron-Metairon 1988
Yes: 2, 3 ANCOVA 1F, 1M each for 2 full-sib families Hart and Strathmann 1994
Yes: 1 ANCOVA 1F, 1M each for 2 full-sib families Miner 2007
Diadema antillarum No: 1 Repeated measures ANCOVA 1F, 1M for 1 full-sib family McAlister 2008
Diadema mexicanum No: 1 Repeated measures ANCOVA 1F, 1M each for 4 full-sib families McAlister 2008
Echinometra lucunter No: 1 Repeated measures ANCOVA 1F, 1M each for 3 full-sib families McAlister 2008
Echinometra vanbrunti No: 1 Repeated measures ANCOVA 1F, 1M each for 3 full-sib families McAlister 2008
Echinometra viridis No: 1 Repeated measures ANCOVA 1F, 1M each for 3 full-sib families McAlister 2008
Encope michelini No: 0 ANOVA, MANOVA Unspecified # of parents Eckert 1995
Eucidaris tribuloides No: 1 Repeated measures ANCOVA 1F, 1M each for 3 full-sib families McAlister 2008
Eucidaris thouarsii No: 1 Repeated measures ANCOVA 1F, 1M for 1 full-sib family McAlister 2008
Evechinus chloroticus Yes: 1 ANOVA, MANOVA, PCA 1F, 1M for 1 full-sib family Sewell et al. 2004
Heliocidaris tuberculata Yes: 1 ANOVA, ANCOVA, PCA 1F, 1M for 1 full-sib family Soars et al. 2009
Leodia sexiesperforata No: 2 ANOVA 1F, 1M for 1 full-sib family Reitzel and Heyland 2007
Lytechinus variegatus Yes: 0 ANOVA Unspecified # of parents Boidron-Metairon 1988
Section 1.03
Yes: 1 ANOVA Unspecified # of parents McEdward and Herrera 1999
Yes: 1 Profile Analysis (Repeated 1F, 1M for 1 full-sib family Miner and Vonesh 2004
measures ANOVA and MANOVA)
Yes: 1 ANCOVA 18F, 6M breeding design for 29 McAlister 2007a
full-sib, half-sib families
Melitta tenuis Yes: 2 ANOVA 1F, 1M for 1 full-sib family Reitzel and Heyland 2007
Paracentrotus lividus Yes: 0 Not reported Unspecified # of parents Fenaux et al. 1988
Yes: 1, 2, 3 ANOVA 1F, 1M each for 3 full-sib families Strathmann et al. 1992
Yes: 3 ANOVA Full-sibling cultures, Unspecified # Fenaux et al. 1994
of parents
Pseudochinus huttoni Yes: 1, 2, 3 ANOVA, PCA 9–10F per parental diet Poorbagher et al. 2010b
treatment, 1M

(continued)
108   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e La r vae

Table 8.1  (Continued)

Phylum Plasticity? Statistical Methodology Genetic Replication Reference

Class Species Measure Type

Strongylocentrotus Yes: 1 PCA 4F, 4M pooled gametes Hart and Scheibling 1988

droechachiensis Yes: 1, 2, 3 ANCOVA 8F (4F, 2 locations), 1M for Bertram and
8 half-sib families Strathmann 1998
No: 1 ANOVA, PCA 3F, 3M pooled gametes Meidel et al. 1999
N/A (examined N/A (examined gene Unspecified # of parents Carrier et al. 2015
gene expression) expression)
Strongylocentrotus Yes: 1, 2 ANCOVA 3F, 1M pooled gametes Miner 2005
franciscanus Yes: 1 ANCOVA 2F, 5M pooled gametes McAlister 2007b
Strongylocentrotus Yes: 1, 2 ANCOVA 3F, 1M pooled gametes Miner 2005
purpuratus Yes: 1 ANCOVA 1F, 1M each for 2 full-sib families Miner 2007
Yes: 1 ANCOVA 2F, 7M pooled gametes McAlister 2007b
Yes: 0 ANOVA Unspecified # of parents Adams et al. 2011
Tripneustes gratilla Yes: 1, 2 ANOVA, PCA 1F, 1M for 1 full-sib family Byrne et al. 2008
Holothuroidea
Australostichopus mollis Yes: 2, 3 ANOVA, PCA Unspecified # of parents Morgan 2008
Apostichopus japonicus Yes: 2 PCA 1 F, 1M for 1 full-sib family Sun and Li 2013
Ophiuroidea
Macrophiothrix koehleri Yes: 1, 2 ANCOVA, Linear Mixed Model, 6 separate experiments, Podolsky and McAlister 2005
Cubic spline Unspecified # of parents
Macrophiothrix longipeda Yes: 1, 2 ANCOVA, Linear Mixed Model, 2 separate experiments, Podolsky and McAlister 2005
Cubic spline Unspecified # of parents
Macrophiothrix lorioli No: 1, 2 ANCOVA, Linear Mixed Model, 5 separate experiments, Podolsky and McAlister 2005
Cubic spline Unspecified # of parents
Macrophiothrix rhabdoti No: 1, 2 ANCOVA, Linear Mixed Model, 1 experiment, Unspecified # Podolsky and McAlister 2005
Cubic spline of parents
Mollusca
Bivalvia
Crassostrea gigas Yes: 3 ANCOVA 9F, 2M pooled gametes Strathmann et al. 1993
Gastropoda
Crepidula fornicata Yes: 3 ANCOVA 1F, unknown M Estrella Klinzing and
Pechenik 2000
Annelida*
Polychaeta
Phragmatopoma lapidosa Yes: N/A Not reported Unspecified # of parents Pawlik and Mense 1994
californica
Hydroides dianthus Yes: N/A ANOVA 15–25F, 1M pooled gametes Toonen and Pawlik 2001

(continued)
 P h e n ot y p i c P l as t i c i t y o f Fee d i n g S t r u c t u r es    109

Table 8.1  (Continued)

Phylum Plasticity? Statistical Methodology Genetic Replication Reference

Class Species Measure Type

Bryozoa*
Gymnolaemata
Membranipora Yes: 3 ANOVA Unknown. Wild caught larvae Strathmann et al. 2008
membranacea

Note. “Measure type” refers to measure of relative feeding-structure length to (1) midline body length or other structure functional only in larva, (2) stomach (functional
in both larva and juvenile), or (3) rudiments of juvenile structures (not functional in larva). A measure type of (0) indicates that data from absolute feeding-structure
length was used to assess plasticity, although data from body length, stomach length, or juvenile rudiment length may have also been reported. ANOVA = analysis of
variance; ANCOVA = analysis of covariance, MANOVA = multivariate analysis of variance, PCA = principle components analysis, F = female, M = male, N/A = not appli-
cable. Phyla in which plasticity of other larval structures, settlement and/or metamorphic competency (in association with feeding, starvation and subsequent re-feeding)
has been demonstrated, albeit not plasticity of feeding structures per se, are indicated by *.

Table  8.2  Phyla with Feeding Larvae for Which We Do Not Know rigorously tested in a field setting. Thus there exist
(or Negative Data Has Not Been Reported) Whether Larvae Possess several different, yet intricately connected exam-
Feeding-Structure Plasticity
ples of phenotypic plasticity in response to food
Phylum Feeding larva availability in this system: morphological plastici-
ties of feeding-structure size (larval arms and cili-
Cnidaria planula ated band lengths), food-processing structure size
Platyhelminthes müller’s larva (stomach length or volume), and developmental
Annelida trochophore* plasticities of the time to initial formation of the ju-
Mollusca trochophore venile rudiment and duration of the larval period
Sipuncula trochophore (time to metamorphosis). For the latter, the time pe-
riod is presumably shorter in low-fed larvae that ex-
Bryozoa cyphonaute*
hibit morphological plasticity of feeding structures
Phoronida actinotroph
as compared to low-fed larvae that do not exhibit
Arthropoda nauplius and zoea this form of plasticity. Difficulties in experimentally
Hemichordata tornaria preventing the expression of morphological plastic-
ity by larvae in low food conditions have prohibited
*Indicates larval types in which plasticity of other larval structures, settlement a direct test of this hypothesis. Studies that employ
and/or metamorphic competency (in association with feeding, starvation, and
subsequent re-feeding) has been demonstrated, albeit not plasticity of feeding targeted phenotypic engineering (e.g., through de-
structures per se. velopmental manipulation or genetic modification,
to “trick” larvae into expressing a short-arm pheno-
1994; Toonen and Pawlik, 2001) and bryozoan cy- type in a low food environment) could be used to
phonautes (Strathmann et  al., 2008) (see Table  8.1; test this assumption, and thus the adaptive signifi-
Table 8.2). cance of plasticity in this system. A study by Adams
As a consequence of these types of plasticity, et al. (2011) suggests that this type of manipulation
food-limited larvae exhibit delayed time to meta- or modification is feasible (see later).
morphosis, a potentially dangerous prospect for With respect to the second strategy, larger egg
planktonic-feeding organisms (Thorson, 1950; size typically reflects an increase in maternally
Rumrill, 1990; Morgan, 1995). Strathmann et  al. provisioned energetic materials, which are used
(1992) suggested that plasticity in larval arm length to build larger larvae and to fuel more rapid larval
provides information about larval feeding his- development (McAlister and Moran, 2012; 2013;
tory in the field, although this idea has not been see Marshall et al., 2008, and Moran and McAliste­r,
110   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e La r vae
2009, for reviews). Egg size has been linked to sev-
eral life history traits or events, including larval
form (McEdward, 1986a), developmental mode
(Strathmann, 1985), and the length of larval devel-
opment (Thorson, 1950; Vance, 1973; Strathmann,
1985). Herrera et al. (1996) demonstrated variation
in feeding period with egg size; development time
to metamorphosis is inversely related to egg size
among various echinoid species. Poor larval feed-
ing environments might have selected for the evo-
lution of large eggs to minimize high planktonic
mortality (Rumrill, 1990). Alternatively, condi-
tions of sperm limitation might have selected for
large eggs to increase fertilization success (Levitan,
1993; Podolsky and Strathmann, 1996), indirectly
affecting larval traits. Several studies indicate
that arm-length plasticity is primarily expressed
(or at least is detectable) during early larval de-
velopment (Boidron-Metairon, 1988; Hart and
Scheiblin­g, 1988; Eckert, 1995; Miner, 2007; Adams
et al., 2011; though see Hart and Strathmann, 1994;
George, 1999) and that the capacity for plasticity
of arm length early in development is associated
with the amount of maternally provisioned ener-
getic reserves, and thus with egg size (Bertram and
Strathmann, 1998; McAlister, 2007b; Reitzel and
Heyland, 2007; Bertram et  al., 2009; Poorbagher
et  al., 2010a; 2010b). These results suggest that
larvae utilize endogenous resources for the initial
production of food-collecting structures, and then
exploit exogenous resources for the development
of other, later-appearing structures.
Given that feeding-structure plasticity is common
in planktotrophic species of echinoids, coupled with
the general interest in egg size and evolution of ma-
rine invertebrates, it is not surprising that research-
ers have tested whether egg size is associated with
feeding-structure plasticity in larvae. Indeed, egg
size varies greatly (>80-fold difference in volume as
calculated from values below) among species of echi-
noids with planktotrophic larvae. In some species
mothers provision their offspring with a small por-
tion of the energy needed to complete larval develop-
ment and metamorphose into a juvenile (e.g., Arbacia
stellata, egg volume 0.13 nl, egg energy 1.0 mJ; Moran
et al., 2011); whereas in other species, mothers pro-
vide all of the energy needed to complete larval de-
velopment and metamorphosis (e.g., the facultative
planktotroph Clypeaster rosaceus, egg volume 10.77
nl, egg energy 110 mJ; Miner et al., 2002).
There are two general hypotheses for the relation-
ship between egg size and degree of morphological
plasticity. Embryos that develop from larger eggs
have access to greater endogenous materials and
energy and thus have the propensity to display a
greater degree of morphological plasticity than lar-
vae that develop from smaller eggs (Herrera et  al.,
1996; McAlister, 2007b). Alternatively, selection
might have favored a greater capacity for plasticity
in embryos that develop from smaller eggs to more
effectively utilize exogenous resources (McAliste­ r,
2007b). The results from other studies support both
of these hypotheses. Comparative measurements
between closely related species and physical ma-
nipulations of egg size in a single species demon-
strate that large egg size is associated with a greater
degree of plasticity (McAlister, 2007b). Alternatively,
comparative studies among multiple species of ophi-
uroids (Podolsky and McAlister, 2005) and echinoids
(Reitze­l and Heyland, 2007) have supported the alter-
nate hypothesis. Egg size is likely a coarse measure of
egg energetic content, however, it provides no infor-
mation about egg biochemical composition (Moran
and McAlister, 2009; Moran et al., 2011). Increases in
egg size can be obtained through increased maternal
provisioning or through simple hydration (Podolsky
and Strathman­n, 1996; McAlister and Moran, 2012).
Thus, outside of a controlled phylogenetic context,
using egg size alone to make assumptions about egg
composition and energetic content and their associa-
tion with other life history characters might be prob-
lematic (McAlister and Moran, 2012). We suspect
that the relationship between plasticity and egg size
is a combination of both hypotheses, and reflects a
nonlinear negative relationship between these vari-
ables for feeding larvae (Figure 8.2). For larvae that
develop from relatively small eggs (i.e., egg energy
runs out shortly after larvae gain the ability to feed),
low levels of endogenous energy likely limit the pro-
duction of long arms when food is scarce. At the other
extreme, larvae that develop from relatively large
eggs (i.e., egg energy fuels nearly all of larval devel-
opment) are not energy limited and thus plasticity
of arm length does not improve fitness if the main
role of arms is feeding. Studies that manipulate the
timing of exposure to exogenous food can elucidate
 P h e n ot y p i c P l as t i c i t y o f Fee d i n g S t r u c t u r es    111
Degree of Plasticity
Egg Size
Figure  8.2 Degree of phenotypic plasticity of feeding-structure
length relative to egg size. Feeding larvae that develop from small or
large eggs exhibit lower degrees of plasticity (less difference between
phenotype values of high- and low-fed larvae) than larvae that develop
from intermediate size eggs.
whether the degree of the plastic response changes
during the course of larval development and, further,
if the response is confined to specific developmental
windows or time periods.
8.3.2 Patterns of Expression and Environmental
Cues
Although several studies have focused on under-
standing the energetic trade-offs discussed pre-
viously, more research has involved functional
Plastic
High Fed Low Fed
Arm Length
Time
demonstrations and documentation of the expression
of feeding-structure plasticity. These studies have
included assessments of the presence, magnitude,
variability, and timing of the response within specific
taxa (Table 8.1). Other studies have investigated the
environmental cues that induce the ­feeding-structure
response, indicating that in addition to low algal
food availability, algal exudates (Miner, 2007) and
thyroxin compounds (Heyland and Hodin, 2004;
Heyland et al., 2004) can also play a role.
Many researchers measure plasticity early in lar-
val development and have demonstrated that for
the majority of species, larvae fed scarce amounts of
food after several days during the first week or two
of development exhibit longer larval arms than lar-
vae fed abundant food (see Table 8.1). Across stud-
ies, starved larvae exhibit on the order of an 8–30%
increase in arm length over their well-fed counter-
parts. Although the percentage difference in arm
length between well- and under-fed larvae can be
relatively small, because the relationship between
absolute arm length and food availability over time
is concave (both decreasing and decelerating; Miner
and Vonesh, 2004), morphological changes made
early in development will have a greater influence
on an organism’s overarching ontogenetic trajec-
tory than will changes made later in development
(Figure  8.3). Additionally, small differences made
early in development, coupled with adjustments of
metabolic rate, might also be very important if they
increase the threshold at which larvae starve. It is
important to note, however, that not all species may
Non-Plastic
Low Fed
Figure 8.3  Absolute arm length relative to
time for high-fed larvae (solid line), low-fed
larvae exhibiting plasticity (long-dash line),
and low-fed larvae not exhibiting plasticity
(short-dash line). Phenotypically plastic
low-fed larvae exhibit longer arms early
in development and theoretically enjoy a
fitness advantage of reaching metamorphosis
sooner than non-plastic low-fed larvae.
112   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e La r vae
be phenotypically plastic (Eckert, 1995; McAliste­r,
2008; Soars et al., 2009; see caveat later), which sug-
gests that the functional arguments for plasticity,
its magnitude, and the cost-benefit relationship
for maintaining the ability to express plasticity, are
needed for each case.
Another important metric for feeding-­structure
plasticity is the relative relationship between
feeding-structure length and a measure of body
size, such as midline body length. In a system in
which food is the inducing environmental vari-
able, as well as the resource necessary for growth,
measurements of feeding-structure length (e.g.,
arm length) relative to body length can indicate
whether low-fed larvae are expressing phenotypic
plasticity throughout the duration of larval ontog-
eny. In this scenario, graphical plots of arm length
vs. body length will exhibit a more steeply sloped
relationship for poorly fed than for well-fed larvae
(Figure  8.4). Slopes calculated for individuals fed
different food levels can then be used to assess the
degree of plasticity expressed by a given genotype,
by plotting and calculating the slope of the reaction
norm between these values.
Different genotypes are expected to vary in their
degree of plasticity, though differences in the degree
of plasticity among genotypes in marine invertebrates
remain largely unexplored (but see McAlister, 2007a).
Plastic
Low Fed
Arm Length
Body Length
This might be due in part to the logistical complica-
tions of simultaneously rearing multiple, replicated-
treatment cultures of larvae and to the large numbers
of genotypes that must be examined for valid sta-
tistical comparisons. Because phenotypic variation
among genotypes provides an opportunity for evo-
lution by natural selection, studies that examine the
genotypic-level variation in phenotypic plasticity
are needed. Data describing the across-environment
pattern of phenotypic expression (e.g., mean and var-
iance) among genotypes in a population could pro-
vide important clues about the evolutionary potential
of the population. Examinations of this type could
be extended to an inter-population level of analy-
sis. Although two studies have investigated differ-
ences in phenotypic plasticity between populations
(Bertram and Strathmann, 1998; Poorbagher et  al.,
2010b), these studies were concerned more with the
effects of nutrition on the expression of plasticity than
investigating differences in the degree of inter- and
intra-­population variation for plasticity. Researchers
have proposed that phenotypic plasticity varies by
latitude: species, populations, and genotypes from
higher latitudes will express a greater degree of phe-
notypic plasticity than those from lower latitudes
(McAliste­r, 2008; Soars et  al., 2009). This idea is un-
tested, but could be examined using multi-genotype,
common-garden analyses.
High Fed
Non-Plastic
Low Fed
Figure  8.4 Arm length relative to body
length. Phenotypically plastic low-fed larvae
(long-dash line) exhibit a more steeply sloped
relationship between these structures than
high-fed (solid line) or non-plastic low-fed
(short-dash line) larvae. High-fed larvae reach
metamorphosis sooner than plastic and non-
plastic low-fed larvae.
 P h e n ot y p i c P l as t i c i t y o f Fee d i n g S t r u c t u r es    113
Hart and Strathmann (1994) showed that poorly
fed larvae produced longer arms and ciliated bands
and exhibited approximately a 20% increase in
maximum clearance rates over well-fed larvae,
providing evidence that arm length plasticity is
functionally significant. Strathmann et  al. (1992)
demonstrated that full-sibling larvae exhibit het-
erochronic shifts in the relative timing of the de-
velopment of food-collecting arms and of the
juvenile rudiment. However, production of larger
food-collecting structures occurs at the expense of
food processing (stomach) or postlarval (rudiment)
structures (Strathmann et  al., 1992; Miner, 2005).
Whether the delay in the development of postlar-
val structures is due to larvae preferentially allocat-
ing energy, materials, or both to feeding structures
or a consequence of larvae having fewer resources
because they are fed less is currently unclear (but
see Adams et al., 2011, for some evidence for pref-
erential allocation of energetic reserves). These
results suggest that feeding-structure plasticity is
evolutionarily adaptive: larvae that alter feeding-
structure length to match environmental conditions
via plasticity will enjoy a fitness advantage rela-
tive to those that do not. Strathmann et  al. (1992)
hypothesize that plasticity in allocation toward the
growth and developmental timing of body parts is
associated with the evolution of lecithotrophy from
planktotrophy. Herrera et  al. (1996) demonstrated
that species whose larvae develop from larger eggs,
and thus possess greater endogenous energy stores,
attain later developmental stages (characterized by
the growth of successive pairs of larval arms to a
typical maximum of four pairs and lastly the ap-
pearance of a juvenile rudiment adjacent to the lar-
val stomach) than species from smaller eggs. Thus
the effects of maternal nutrition on larval develop-
ment and morphogenesis might mimic the effects of
greater endogenous food supplies.
Strathmann et  al. (1993) suggest that larval
feeding-structure plasticity has evolved at least
­ larval juvenile, and (3) to rudiments of juvenile struc-
twice in marine invertebrates. Larvae of both echi-
noderms and molluscs display feeding-structure
plasticity, but are distantly related and have very
different types of larvae with different feeding
mechanisms. Echinoderms are deuterostomes and
the juvenile develops as a separate structure within
the larval body. Feeding larvae of echinoderms
capture particles of food by reversing a short section
of their ciliary band, which is composed of simple
cilia. Molluscs are protostomes and many struc-
tures are unchanged during metamorphosis from
larva to juvenile, but the larval feeding structure is
resorbed or shed at metamorphosis. Feeding larvae
of molluscs capture particles with an opposed band
feeding mechanism and compound cilia. For these
reasons, larvae of echinoderms and molluscs are
thought to have evolved independently. Because
feeding-structure plasticity has evolved in both
groups, the implication is that plasticity is either
ancestral to both groups, or is a coevolved, homo-
plastic trait (Podolsky and McAlister, 2005).
Among echinoderms, phenotypic plasticity of
larval feeding structures has been examined in ap-
proximately 22 echinoid, four asteroid, two holo-
thuroid, and four ophiuroid species (Table 8.1). The
majority of species that exhibit no measured plastic-
ity in response to food-limiting conditions are tropi-
cal (Eckert, 1995; McAlister, 2008; Soars et al., 2009),
which suggests there are general differences in the
expression and evolution of plasticity between spe-
cies in temperate vs. tropical ecosystems. There also
might be differences in plasticity on smaller geo-
graphic scales, such as between more closely located
populations of a given species. For example, two
populations of the subtropical subspecies Lytechinus
variegatus carolinus, located approximately 185 km
apart (collection sites near Beaufort and Wilmington,
North Carolina) differ (McAlister, unpublished data),
with larvae from one population demonstrating arm-
length plasticity and the other seemingly not.
To date, studies examining feeding-structure
plasticity have utilized measures of absolute
­feeding-structure length as well as three different
types of relative measure: (1) to body length or some
other measure restricted to ephemeral larval struc-
tures, (2) to stomach, wherein the stomach is a
structur­e that is functional in both the larva and post-
tures that are not functional until after the larval stage
(see Section 8.3.4 for statistical considerations). While
feeding-structure plasticity may potentially be dem-
onstrated as present through any of these metrics,
the conclusion that plasticity is absent in some spe-
cies (Eckert, 1995; McAlister, 2008; Soars et al., 2009)
or populations may be premature unless a study
114   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e La r vae
specifically includes measures of feeding-structure
length relative to rudiments of juvenile structures
(see Table 8.1). For example, if there was no change
in arm or ciliary band length in response to low food,
but development of the juvenile rudiment was de-
layed until the larva had attained a stage with longer
arms or ciliary band, then that developmental delay
would confer some of the hypothesized benefits of
producing absolutely longer arms or ciliary band
earlier in the larval period. Demand for energy and
materials for growth would thus be postponed un-
til the capacity to capture food had been increased.
Although studies collecting absolute or relative
measures similar to (1) or (2) are certainly valuable,
and may be logistically more feasible, future stud-
ies should strive to include measures of both larval
structures and the rudiments of juvenile structures to
definitively confirm presence or absence of plasticity.
Furthermore, from the population and geo-
graphical patterns presented earlier, many new
questions arise: What environmental factors or or-
ganismal functions drive or constrain the expres-
sion of ­feeding-structure plasticity between taxa?
Are there measurable environmental differences be-
tween sites or do differing results reflect some type
of seasonal or individual sampling bias? Are dif-
ferences in expression proximally tied to ecological
differences between ecosystems or locales, or have
ultimate evolutionary responses occurred in line-
ages? Has there been a loss of chemosensory struc-
tures in non-plastic species? Is it difficult to regain
these structures once lost? Does absence of plastic-
ity merely reflect absence of measures that would
detect it? Answers to these questions will require
researchers to continue empirical analysis within
a comparative framework while collecting all rel-
evant metrics. At the same time, our understanding
of the expression and evolution of plasticity will be
deepened by delving into the developmental and
physiological mechanisms that are involved during
feeding-structure production.
8.3.3  Developmental Mechanisms of Feeding-
Structure Plasticity
Although there is currently limited research on the
developmental or physiological mechanisms that
cause feeding-structure plasticity in marine larvae,
researchers are well poised to rapidly advance this
area of research (reviewed by Miner, 2011; see also
Adams et al., 2011; Carrier et al., 2015). First, sea ur-
chin larvae have long been a model system for de-
velopment, and the developmental mechanisms for
some of the structures involved in feeding-structure
plasticity in echinoid larvae are well understood
(e.g., formation of the larval skeleton). Second, re-
cent advances in molecular techniques allow re-
searchers to measure changes in the timing and
quantities of gene expression in larvae exposed to
different concentrations of food. Last, the interest in
ecological development is rapidly growing and as
new techniques emerge, feeding-structure plasticity
in echinoids is very well suited to advance this field
(Hofmann et  al., 2005; Adams et  al., 2011; Carrier
et al., 2015; Gilbert and Epel, 2015).
Here we focus on four general questions that re-
quire answers to understand how larvae adjust their
morphology in response to algal concentration: (1)
How do larvae detect algal concentration? (2) How
do larvae adjust the size of the ciliary band, skel-
eton (in echinoids and ophiuroids), and stomach?
(3) How do larvae regulate the timing of production
of rudimentary juvenile structures? (4) How do lar-
vae coordinate these different responses? We focu­s
on several studies in which researchers have direct
answers about the developmental mechanisms of
feeding-structure plasticity in larvae, and draw
a few examples from a large body of research on
the development of echinoderm larvae to highlight
some genes and pathways that are likely involved
in feeding-structure plasticity.
There is indirect evidence about the type and
location of receptors that larvae use to detect algal
concentrations. Echinoid larvae can respond to al-
gal concentrations before they can even ingest al-
gae (Miner, 2005; Adams et al., 2011). These results
suggest that larvae use receptors expressed in epi-
thelial cells to detect algal concentrations (Shilling,
1995; Miner, 2007). At least in some echinoid spe-
cies, these receptors are unlikely mechanoreceptors
because larvae do not alter their morphology in re-
sponse to plastic beads that are similar in size and
concentration to algae (Miner, 2007). More likely
the receptors are chemoreceptors because larvae re-
duce the size of their feeding structures in response
to algal exudates (Miner, 2007) and amino acids
 P h e n ot y p i c P l as t i c i t y o f Fee d i n g S t r u c t u r es    115
(Shilling, 1995) in these species. There is some evi-
dence that thyroxine is a signal molecule between
algae and larvae (Heyland and Hodi­n 2004), though
researchers have not identified a chemoreceptor or
specific chemical signal. In other echinoid species,
larvae might use both mechanoreceptors and chem-
oreceptors in tandem to detect algal concentrations.
Larvae of a sand dollar only produce smaller feed-
ing structures in response to intact algae and not
algal exudates or plastic beads (Miner, 2007). It is
possible that larvae use the same receptors to de-
tect and capture algae and to alter the size of their
feeding structures. There is also evidence that sug-
gests larvae are more sensitive to environmental
cues when starved. Starved larvae up-regulate
cAMP response element-binding protein (CREB),
ETS domain-­containing protein (Elk), and an ex-
citatory amino acid transporter (EAAT), which in
other organisms are known to increase the sensitiv-
ity to environmental stimuli (Carrier et  al., 2015).
Interestingly, thyroxine receptor and dopamine re-
ceptor (see later) activity can cross-regulate CREB
(­Méndez-Pertuz, 2003; Neve et al., 2004).
After larvae detect an external signal from algae,
the nervous system and dopaminergic neurons are
likely involved in the signaling pathway. Adams
et al. (2011) demonstrated that dopamine is involved
in the signaling pathway that induces smaller feed-
ing structures, which occur when food is abundant.
The ability to manipulate the phenotype of larvae
through dopamine now allows researchers to de-
couple the effects of energy and materials from algae
from the effects of the induced response (e.g., Adams
et al., 2011). We look forward to future research on
the role of dopamine, especially in other plastic spe-
cies and related non-plastic species (see Table 8.1).
Compared to research on the developmental
pathways involved in creating the larval skeleton in
echinoids, there are fewer studies on the develop-
mental pathways that larvae use to specify the cili-
ary band. However, our understanding is rapidly
improving as researchers use new tools to improve
the developmental map of echinoids (e.g., Li et al.,
2014). In echinoids, the ciliary band is specified by
interactions between the oral and aboral ectoderm
(Davidson et al., 1998; Su et al., 2009), and the gene
products of forkhead g (FoxG), hepatocyte nuclear
factor 6 (Hnf6), and orthodenticle β ½ (Otxβ) are a
few of the genes likely involved specifying to the
ciliary band (Optim et  al., 2004; Tu et  al., 2006;
Yankura et al., 2013). Of these genes, Hnf6 and Otxβ
are not regulated by genes that pattern the embryo,
which might allow their expression patterns to be
altered without disrupting general morphological
development (Duboc et  al., 2004; Su et  al., 2009).
The length of the ciliary band might be regulated
by genes, like forkhead J1 (FoxJ1), which are asso-
ciated with ciliagenesis. Starved plutei up-regulate
FoxJ1 (Carrier et  al., 2015), which is a transcrip-
tional regulator associated with many aspects of
cilia development in mouse (Choksi et  al., 2014).
Despite our improving understanding of the path-
ways that specify the ciliary band, understanding
the mechanisms responsible for adjusting the size
of the ciliary band is critical, and an area that we
hope researchers investigate more in the future.
The pathways that control the length of the
larval skeleton in echinoids are well studied in
echinoids—though much less is known about ophi-
uroids, which also produce a larval skeleton (Dylus
et  al., 2016). Primary mesenchyme cells form the
larval skeleton by producing an extracellular ma-
trix and precipitating calcium carbonate onto this
matrix (Killian and Wilt, 2008), and involve many
hundreds of genes (Ettensohn 2009; Rafiq et  al.,
2014). The location and length of the skeleton are
regulated by interactions between the primary mes-
enchyme cells and ectoderm (Hardin et  al., 1992;
Armstrong et al., 1993; Guss and Ettensohn, 1997).
Vascular endothelial growth factor (VEGF), fibro-
blast growth factors (FGF), orthopedia (Otp), and
tetraspanin are involved in the interaction between
primary mesenchyme cells and ectoderm, and ap-
pear to affect size of the skeleton (DiBernardo et al.,
1999, Cavalieri et  al., 2007; Duloquin et  al., 2007;
Love et  al., 2007; Röttinger et  al., 2008)—the gen-
eral shape of the skeleton appears fixed early in
embryonic development (Armstrong and McClay,
1994). It is also possible that the transmembrane
protein P16, which is necessary for skeletal spicules
to elongate, is involved in the pathway (Cheers and
Ettensohn, 2005). In addition, the proteins spicular
matrix protein (SM 50), mesenchyme-specific pro-
tein (MSP  130), advillin, and carbonic anhydrase
are associated with primary mesenchyme cells,
and likely control, at least in part, the length of the
116   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e La r vae
skeleton (Carson et al., 1985; Leaf et al., 1987; Peled-
Kramar et al., 2002; Love et al., 2007). For example,
advillin and carbonic anhydrase appear to pro-
mote lengthening the skeleton at the tips, whereas
MSP 130 adds more calcium carbonate to the exist-
ing skeleton (Carson et  al., 1985; Leaf et  al., 1987;
Peled-Kramar et al., 2002; Love et al., 2007). Similar
to the pathways that specify the ciliary band, we
hope researchers also focus on mechanisms that can
regulate the length of the skeleton. It will be espe-
cially interesting to compare the pathways involved
in skeletal plasticity between echinoids and ophi-
uroids because their skeletons are likely convergent.
Carrier et  al. (2015) provide information about
the pathways that might regulate the development
of the rudiment. Plutei adjust a suite of genes asso-
ciated with metabolism when exposed to different
concentrations of unicellular algae. Starved plutei
down-regulate genes associated with growth and
mitochondrial activity and up-regulate genes as-
sociated with energy homeostasis. In particular,
starved larvae up-regulate forkhead O (FoxO) and
down-regulate target of rapamycin, which is associ-
ated with the formation of the rudiment. Given that
both of these genes respond similarly in distantly re-
lated model species, it is possible that this pathway
is conserved and involved in feeding-structure plas-
ticity in all echinoderms (Carrier et al., 2015). Unlike
adjusting of the length of the ciliary band and larval
skeleton in certain regions in response to food, the
response of the rudiment only requires a shift in the
overall timing of when to produce the rudiment,
which might indicate a simpler mechanism.
Electrical and chemical signals likely coordinate
the morphological responses among structures
(Adam­s et  al., 2011; Miner, 2011). The nervous sys-
tem is well developed in echinoderm larvae before
larvae can feed (reviewed in Burke et al., 2006; also
see Adams et al., 2011). Sensory neurons are located
on the epithelium and nerves are closely associated
with the ciliary band and stomach, especially the
oral field and mouth. The close proximity between
the skeletal rods, primary mesenchyme cells, and ec-
toderm makes it more likely that chemical signals co-
ordinate responses in the ciliary band and skeleton.
For example, dopamine is found in close associa-
tion with ectoderm and primary mesenchyme cells,
which produce the skeleton (Adams et  al., 2011).
Partitioning-defective protein localizes in both pri-
mary mesenchyme and stomach cells, and might co-
ordinate the response between these two structures
(Shiomi and Yamaguchi, 2008).
We encourage research specifically on the de-
velopmental and physiological mechanisms of
­feeding-structure plasticity in larvae. We antici-
pate that the current knowledge will allow rapid
progress for echinoids, and our understanding of
echinoids will facilitate research on other groups.
We especially encourage comparative studies
that will permit tests of hypotheses about the
evolution of larval feeding-structure plasticity
among metazoans.
8.3.4 Experimental Designs and Analyses
Investigators studying feeding-structure plastic-
ity of larvae are faced with a challenge because
manipulating food concentrations alters rates of
growth and development, as well as size of feed-
ing structures (Hart and Scheibling, 1988). As a re-
sult, it is not possible to simply compare a metric of
­feeding-structure size between treatments at a given
time. Fed larvae are typically much larger in gen-
eral, including the size of feeding structures, and
more developed than starved larvae at any given
time. In what follows we discuss how investigators
have tried to solve this challenge with the design
of their experiments and the analysis of their data.
Investigators often use a nested experimental
design, where larvae are measured from a few rep-
licate containers per treatment. The reasons for a
nested design are primarily practical because rear-
ing larvae in many replicate containers requires
considerable time for maintenance, as well as spe-
cialized equipment to control temperature (e.g.,
incubators or shallow tanks of water to partially
submerge containers)—containers typically only
hold one or two liters and can fluctuate in tem-
perature beyond the physiological limits of larvae
if temperature is not rigorously controlled. Most
studies have less than five replicates per treatment.
The small number of replicates, however, means
that statistical power of experiments is often low,
which is of concern because differences between
treatments range from 10% to 30%. To increase
the statistical power and thus the ability to detect
 P h e n ot y p i c P l as t i c i t y o f Fee d i n g S t r u c t u r es    117
plasticity, some investigators have analyzed data
treating each individual larva as a replicate, after
determining that containers were not significantly
different within treatments (e.g., Strathmann et al.,
1993; Hart and Strathmann, 1994), rather than us-
ing the mean for each container. This is a reason-
able approach, but one with inherent problems.
For example, if there are really differences among
containers within treatments but only a few larvae
are measured per container, then an investigator is
much more likely to incorrectly determine that con-
tainers are not different (a Type II error). Differences
among containers within treatments are likely com-
mon (Sewell et  al., 2004), and therefore investiga-
tors should not use larvae within a container as
independent replicates without a clear justification
and power analysis of the test to determine whether
containers within a treatment differ from one an-
other. We recommend that investigators work to in-
clude more containers, which are true independent
replicates, in their experiments.
Most investigators have tested two or three con-
centrations of algal food as experimental treatments
and code algal concentration as a categorical vari-
able. This approach allows researchers to determine
whether there is an effect of food concentration, but
provides little information about the relationship be-
tween feeding structures and algal ­concentrations—
the shape of this relationship has important ecological
and evolutionary consequences. An alternative de-
sign is to assign a different algal concentration to
each container within some range of concentrations
and code the algal concentration as a continuous
variable (Miner and Vonesh, 2004). Both approaches
allow for inferences about whether plasticity occurs,
but the latter approach allows for stronger inferences
about the shape of the reaction norm. Both linear and
generalized linear models can be used with either
approach, though generalized linear models provide
more flexibility because the researcher can indepen-
dently specify the relationship between the response
and predictor variable and the error structure (Quinn
and Keough, 2002)—however, they are more difficult
to interpret with complex designs where multiple
factors are manipulated.
Feeding interval can also affect the concentration
of algae and the interpretation of results. Two pro-
cesses can change the concentration of algae after
algae are added to a container: they can multiply, die
or sink to the bottom and become unavailable. Algae
are probably not very likely to divide much between
feedings every one to three days because researchers
rarely add nutrients to replicates or provide suffi-
cient lighting for algal growth. However, if nutrients
are added with algae and light is provided, then
alga­l growth might be of concern. Larval grazing
is likely a larger concern because larvae can greatly
reduce algal concentrations. For example, feeding
larvae of echinoids can clear 1 ml of water per day
of food (Hart and Strathmann, 1994). If there are 500
larvae in a container with 2 L of seawater (a concen-
tration of 0.25 larvae per ml), then the larvae will
reduce the concentration of algae by 25% per day.
There are two consequences that researchers should
consider when algal densities change between feed-
ings. First, the concentrations that are added to a
replicate container do not reflect the actual concen-
trations that larvae experience, and researchers will
incorrectly estimate the relationship between larval
morphology and algal concentration. Second, inves-
tigators manipulate a variety of aspects about the
food environment (e.g., mean, variance, maximum,
and minimum of algal concentration). Currently it is
unclear which parameters larvae use to adjust their
morphology (Miner and Vonesh, 2004). Changes in
algal concentrations among containers in a treat-
ment might explain in part why in some studies
larvae in different containers of a treatment differ in
phenotype (Sewell et al., 2004).
Investigators have used either larval arm length or
ciliary band length as a metric of feeding-structur­e
size and have employed a variety of different statis-
tical approaches to detect feeding-structure plastic-
ity in larvae. Because food concentrations also affect
growth, investigators often measure a covariate to
account for larval size (e.g., midline body length,
a skeletal component of the body, or shell length).
Investigators often adjust for larval size, to com-
pare larvae of an equivalent size, by dividing the
response variable by the covariate and testing for
differences with an analysis of variance (ANOVA;
e.g., Sewell et  al., 2004; Byrne et  al., 2008), or an
analysis of covariance (ANCOVA; e.g., Miner, 2005;
McAlister, 2007b). An alternative approach used by
some investigators is to measure a variety of met-
rics and use principle components analysis (PCA)
118   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt eb r at e La r vae
to reduce the number of variables before analyzing
the principle components with ANOVA (Hart and
Scheibling, 1988; Sewell et  al., 2004; Miner et  al.,
2005; Soars et al., 2009). All approaches have poten-
tial problems that investigators should be aware of.
Analyzing the ratio of the response variable to the
covariate with ANOVA confounds which variable,
the response or covariate, is causing differences. By
contrast, ANCOVA explicitly tests whether larvae
of a given size differ in the response variable and
the interaction between the response and covariate.
However, ANCOVA assumes there is no error in the
covariate (Quinn and Keough, 2002), which is never
true when measuring some aspect of larval size.
It is likely that the covariate will have similar er-
ror as the response variable. PCA is likely the most
problematic because transforming the original vari-
ables into composite variables mixes both among-
and within-group variation, which often reduces
the ability to detect differences among treatments
(McCoy et  al., 2006). McCoy et  al. (2006) provide
a more appropriate, albeit more complex, method
(common principal components analysis combined
with Burnaby’s back projection method—CPCA/
BBPM), and we recommend that investigators use
this approach in the future.
To complicate the analysis even further, investi-
gators often measure larvae from containers more
than once during the experiment. Repeatedly sam-
pling from containers presents the same challenges
as measuring multiple larvae from a container—data
collected at different times from a container are not
independent. The common solution is to use repeated
measures ANOVA or ANCOVA to test for differences
among treatments. Recently, mixed effects models
that utilize maximum likelihood are common in sta-
tistical packages and allow for random effects that
deal with the non-independence of repeatedly sam-
pling from containers and measuring more than one
individual from a container at a given time.
Lastly, and with respect to genotypic variation
and genetic diversity broadly, our knowledge of
feeding-structure plasticity across taxa derives
from data in which only a single female (and of-
ten only a single male for one full-sib family) was
used to produce the larvae examined in a given
study. This is the case for approximately 1/3 of the
species (14 of 37) listed in Table 8.1. The collective
lack of genetic diversity across studies is striking
and makes generalizing broad biological patterns,
such as the temperate vs. tropical argument pre-
sented earlier, from single-family data difficult and
potentially tenuous. Future studies should strive
to maximize genetic diversity by incorporating
as many full-sib families as is feasible, or alterna-
tively by maximizing the number of gametes from
a given sex, as appropriate for the hypotheses driv-
ing a particular study. For example, increasing the
number of males in a given study will increase the
amount of additive genetic variation among fami-
lies, whereas increasing the number of females
will diversify the effects toward growth of mater-
nal provisions available from the egg. Research-
ers should incorporate and statistically account
for these variables (numbers of males and females
used and full- and half-sib families reared) to the
degree possible given the space and time con-
straints associated with rearing multiple, replicate
larval cultures.
8.4 Summary
1. There is compelling evidence that feeding-­
structure plasticity in marine invertebrates is
adaptive, but we need a better understanding
of the genetic variation of plasticity within and
among populations. Researchers should strive to
maximize genetic diversity in future studies.
2. There are important trade-offs among morpho-
logical structures and the timing of life history
events that influence the evolution of feeding-
structure plasticity.
3. Maternal provisions influence the evolution of
feeding-structure plasticity; minimal provision-
ing limits the energy available to produce larger
feeding structures, whereas maximal provision-
ing removes the need to produce larger feeding
structures.
4. Feeding-structure plasticity has likely evolved at
least twice among marine invertebrates.
5. Researchers are starting to understand the mo-
lecular mechanisms of feeding-structure plas-
ticity in marine invertebrates. In light of our
understanding of development in echinoids,
this system is especially well suited for future
research.
 P h e n ot y p i c P l as t i c i t y o f Fee d i n g S t r u c t u r es    119
6. Researchers have used a variety of experimen-
tal designs and statistical analyses in studies of
larval feeding-structure plasticity. Some of these
designs are problematic and should be avoided
in the future. In all cases, all metadata, including
larval density, food concentration and feeding
frequency, should be reported.
Acknowledgments
The authors would like to acknowledge Adam
Reitzel, Andreas Heyland, and Tyler Carrier for
editing this book. We would also like to thank
Richard Strathmann, Dianna Padilla, David Leaf,
Bob Podolsky, Joel Kingsolver, and Steve Stancyk
for many insightful conversations. We particularly
thank Richard Strathmann, Dianna Padilla, and one
anonymous reviewer for providing insightful com-
ments that helped to improve this chapter.
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 Ch a pt er 9
Physiology of Larval Feeding
William Jaeckle
9.1 Introduction
The enormous diversity of form of marine inverte-
brate larvae has been long recognized (see Young
et al., 2002, for a compilation). As larvae represent
a transient stage in a species’ life histor­y, their
performance in situ has effects on survivorship
in the near term (larval life) and carryover effects
that are realized in juveniles (e.g., ­Marshall et al.,
2003; Pechenik, this volume); recent work also
indicates that such effects may exist across gen-
erations (Dupont et al., 2008; Ross et al., 2016). Ir-
respective of how larval performance is measured
(development rate, metabolic rate, growth rate,
etc.), it represents the summation of the physi-
ological activities of the individual, the interde-
pendence among cells that make up the whole,
and the correct positioning of these cells within
the larval body. Thus, the physiological properties
of larvae are constrained, in part, by their form,
and the phrase, “physiology is the handmaid of
anatomy,” although typically applied in other
contexts (e.g., human anatomy and physiology;
Robinson, 1847), aptly applies to marine inverte-
brate larvae.
The focus of this chapter is to examine the physi-
ology of marine invertebrate larvae from an organis-
mic perspective. The functional attributes of larvae
are, in part, a result of the integration of function(s)
of their different parts (cells, tissues, and organs).
Highlighting the importance of understanding the
relationship between structure and function in the
context of larval physiology is an intentional focus
of this document.
Jaeckle, W., Physiology of larval feeding. In. Evolutionary Ecology of Marine Invertebrate Larvae.
Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018).
© Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0009
9.2  Physiology of Invertebrate Larvae:
The Challenge
Owing to their small size and associated low rates
of physiological processes, the physiological and
biochemical properties of a larva is challenging to
measure, and groups of individuals are often com-
bined to ensure a detectable signal. Such aggregated
measurements result in an understanding of the
average response of larvae and limits appreciation
of variability that exists among individuals. Given
this unknown variability, statistical assessments of
experimental results benefit from inclusion of esti-
mates of effect size (Nakagawa and Cuthill, 2007) in
addition to or in lieu of traditional null hypothesis
testing.
When measures of variability among individu-
als are available (e.g., Marsh et al., 2001; Szela and
Marsh, 2005), considerable differences are evident.
Although the ultimate cause of inter-individual
differences is genetic, the underlying influence of
genotype on physiological differences was, until
recently, largely inaccessible. Work on genetically
defined strains of the Japanese oyster, Crassostrea
gigas, for example, has provided access into the
physiological and biochemical (Hedgecock et al.,
1996; Bayne et al., 1999; Hedgecock et al., 2007;
Meyer and Manahan, 2010) bases for differences in
larval and adult performance. Using adults from in-
bred, pedigreed “families,” Pace et al. (2006) dem-
onstrated that differences in growth among larvae
from specific crosses were correlated with variation
in size-specific feeding capacity; ≈50% of the ob-
served variation in growth was explained by rates

of ingestion. They hypothesized that the remain-
ing differences in larval growth were explained by
genotype-specific variation in “other growth ef-
ficiencies” such as efficiency of protein synthesis.
The value of understanding the effect of genotype
on physiological capacity of larvae increases as the
effects of current and future environmental condi-
tions on marine invertebrates and their larvae are
explored. Recent work on the effects of climate
change on marine invertebrates and their larvae
(see Byrne and Przeslawski, 2013; Przeslawski et
al., 2015; Byrne et al., this volume) has also revealed
considerable variation within and among species.
An ability to differentiate among the effect of treat-
ment or treatment level, genotype, and a geno-
type  × treatment interaction is necessary to more
accurately assess sensitivities and predict eventu-
alities (see Applebaum et al., 2014 for a discussion).
Generalizations about the physiology of marine
invertebrate larvae are inherently challenging ow-
ing to the diversity in form of larvae among groups.
Taxonomic sampling for studies of the physiologi-
cal properties of larvae is inevitably skewed to those
groups with high fecundity, large size, easily scala-
ble culture requirements, and economic value. Such
focused attention on certain species results in few,
well studied, experimentally tractable “models”
whose properties may or may not serve as accurate
exemplars of the physiology of planktonic larvae.
The evident morphological diversity among lar-
vae also challenges the development of general
patterns among groups. The simplest separation
among larval forms is the requirement for exog-
enous food. Feeding (= planktotrophic) larvae must
acquire food from the environment while nonfeed-
ing (= lecithotrophic) larvae can complete their de-
velopmental program in the absence of food. The
distinction between these two larval forms is cor-
related with egg size (Vance, 1973; McEdward and
Morgan, 2001; ­Moran et al., 2013), but the size cut-
off between the small eggs of feeding larvae and the
large eggs of nonfeeding larvae varies among groups
(see McAliste­r and Miner, this volume). Along this
continuum fall larvae of several taxa that are mor-
phologically competent to feed, but can complete a
significant percentage of their development program
without feeding (Herrera et al., 1996; reviewed in
Anger, 2001). Despite the resilience of these larvae to
P h ys i o l o g y o f L a r va l F e e d i n g    125
the absence of food, metamorphosis does not occur
without input of “new” energy. Still others can feed,
but can complete development in the absence of ex-
ogenous food (reviewed in Allen and Pernet, 2007).
The small size of larvae and their component
cells has, to a large degree, hindered precise dis-
section and description of their physiological prop-
erties and how cells and tissues interact. Further,
the presence of anatomic structures named for one
function may also contribute to other physiological
processes. For example, the presence of nephridia
in larvae does not necessarily predict a unique ex-
cretory function. Flows of fluid created during the
formation of “urine” may also contribute to distri-
bution of materials within the larval body. Whether
and how different regions of larvae are physiologi-
cally linked is also not well known. However, appli-
cation of recently developed molecular techniques
has provided opportunities for greater temporal
and spatial resolution about the physiological prop-
erties of larvae (Hui et al., 2014; Carrier et al., 2015;
Chen et al., 2015).
9.3  Acquisition of Materials
The organic materials that support the development
and growth of feeding larvae are acquired from the
environment (see Pernet, this volume). Food sources
consumed by feeding larvae are divided between
particulate organic materials such as bacteria, phyto-
plankton, and zooplankton (organic particles >~0.45
µm in size) and dissolved organic materials (DOM,
organic particles < ~0.45 µm in size and individual
or aggregated organic molecules). For feeding lar-
vae these materials can be acquired through (1) the
concentration and capture of suspended particles,
(2) the ingestion of other organic particles too small
to be removed from suspension by described mech-
anisms, and (3) by the direct assimilation of DOM
by the cells of the outer epithelium or the epithelium
that lines the digestive system (­DeBurgh and Burke,
1983; Manaha­n and Crisp, 1983). The cells that as-
similate these different sources of organic matter
may differ in their position, form, or contribution to
the nutrition of the larva as a whole. For nonfeed-
ing larvae—althoug­ h their development can be
completed in the absence of exogenous m ­ aterials—
the maternally provisioned organic content may
126   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e

be augmented by the uptake of DOM (Jaeckl­e and Table 9.1  Reported Values of the Gut Passage Time (GPT) for Marine
Manahan, 1989a; 1989b; Shillin­g et al., 1996). Invertebrate Larvae.
How the development, growth, and survivorship
Phylum, Species or Diet GPT Source
of larvae is affected by quantitative and qualita- Species (min)
tive changes in diet has been the subject of many
Mollusca
detailed studies (e.g., Sprung, 1984a-d; reviewed in
Anger, 2001). Similarly, how events that occur dur- Mytilus (Nannochloropsis +
galloprovincialis oculata polystyrene
ing a planktonic larval existence influence benthic
beads)
juveniles (“carryover” effect) has also received con-
2-day-old 120+ Gray et al., 2015
siderable attention (Emlet and Hoegh-Guldberg,
1997; Wendt, 1998). Yet how the anatomy and phys- 7-day-old 56
iology of organ systems within larvae serve to link Crassostrea Isochrysis galbana ~60 Reinfelder and
inputs of organic materials to functional responses virginica Fisher, 1994
is, at best, incompletely understood. Although rates Mercenaria Isochrysis galbana ~60 Reinfelder and
mercenaria Fisher, 1994
of particle capture and uptake of DOM are readily
quantified, the physiological processes involved Ostrea chilensis colored plastic ~360 Chaparro et al.,
particles 2001
with food digestion, assimilation, and distribu-
tion within larval bodies are not. Consequently, the Arthropoda, Crustacea
physiological events that link feeding to develop- Artemia Isochrysis galbana 291 Evjemo et al.,
mental or phenotypic changes in larvae are largely franciscana 2000
unresolved. Although reports of the function and Penaeus monodon
structure of the digestive system of larvae are phy- protozoea herbivore/ 12 from Le Vay
logenetically scattered, comparative assessments omnivore et al., 2001
can yield insights about the design and physiology mysis herbivore/ 20
of the digestive system (Karasov et al., 2011; Kara- omnivore
sov and Douglas, 2013). Penaeus setiferus Tetraselmis chuii 5–15 Lovett and
Felder, 1990
Homarus Carnivore 660 from Le Vay
9.4  Material Movement sp. stage 1 et al., 2001
The rates of material transport through the digestive Scylla serrata
system of feeding larvae are known for relatively Zoea “rotifers” 80
few taxa. Processing of ingested foods within the megalopa “rotifers” 120–135 Serrano, 2012
digestive systems of invertebrate animals has been Echinodermata
modeled as a chemical reactor (Penry and Jumars,
Lytechinus pictus (Rhodomonas lens + 60–90 Huvard and
1987; Jumars, 2000a; 2000b; see also Logan et al., colloidal carbon) Holland, 1986
2002). Yet to my knowledge, there are few tests of Patiria miniata 60 Huvard and
how larval digestive systems conform to these func- Holland, 1986
tional hypotheses (plugged-flow reactors, batch re-
actors, or continuously stirred tank reactors). Gray Note. GPT represents the transit time of ingested materials through the larval
et al. (2015) compared the gut passage time (GPT) digestive system, but the manner in which it is measured varies considerably.
Reported values of GPT rarely include a parallel measure of the length of the
of material through the digestive system of veliger digestive system and, as a result, the velocity of material movement through the
larvae of the mussel Mytilus galloprovincialis to the digestive system is unknown.
three “models” and reported that larvae match a 1Mean values reported by the authors.
continuously stirred tank reactor model in isolation
or in combination with a plugged-flow reactor. through the digestive system is accomplished by
Although not specifically evaluated in the context peristalsis, and currents created by cilia and may be
of a “reactor model,” intra- and interspecific differ- regulated by muscular sphincters. Regional compart-
ences exist in the GPT of larvae (Table 9.1). Transit mentalization and the ability to alter the residence

time of material within the digestive system allow
for digestion and assimilation to vary with rates of
food ingestion. More work is needed to assess the de-
gree to which the digestive systems of larvae change
in response to previous and current nutritional con-
ditions (e.g., difference in prey composition or abun-
dance, presence of dissolved solutes).
The types of food ingested, the ease by which
these materials are digested, and the type and
abundance of digestive enzymes are factors that
influence GPT (Fretter and Pilkington, 1971; Strath-
mann, 1971; Gallager, 1988; Karasov et al., 2011;
Karasov and Douglas, 2013). Variations in GPT ex-
ist among larvae (Table 9.1) and are predicted to in-
fluence the extraction efficiency (the proportion of
ingested material that is digested and assimilated
by the digestive epithelia) by altering the residence
time of ingested materials within digestive and ab-
sorptive regions of the digestive system. Larvae of
echinoderms (Strathmann, 1971) and veliger lar-
vae of gastropod and bivalve molluscs (Fretter and
Pilkington, 1971; Gallager, 1988) show an ability to
differentially process organic and inorganic parti-
cles. Inert particles are apparently sorted within the
digestive system to decrease their GPT. In a taxo-
nomically broad review of digestive physiology of
animals, Karasov and Douglass (2013) described
different digestive strategies for species that ex-
ploit types of food with different susceptibilities
to chemical digestion. Those species that ingest
materials largely refractory to chemical digestion
(e.g., cellulose-rich foods, phytoplankton) typically
possess a relatively rapid transit time through the
digestive system. In contrast, species that ingest
more easily digested materials (e.g., animal tissue,
zooplankton) generally have a relatively slow GPT,
which enhances chemical digestion.
The relationship between GPT and resistance to
chemical digestion for crustacean larvae follows
this predicted pattern (reviewed by Le Vay et al.,
2001). They found that those larvae exhibiting a pri-
marily herbivorous or omnivorous diet had shorter
transit times through the digestive system when
compared to larvae with a predominantly carniv-
orous diet. There is also evidence that changes in
the GPT occur during larval life. Among different
larval stages of some caridean shrimps there is an
inverse relationship between digestive enzyme
P h ys i o l o g y o f L a r va l F e e d i n g    127
activity and GPT (reviewed in Anger, 2001). Be-
cause of the inverse relationship, these larvae were
predicted to maintain a more-or-less stable extrac-
tion efficiency. Evidence also exists for plasticity in
GPT within species that is associated with the diet
composition. Serrano (2012) suggested that GPT of
artificial food was greater than for live food (the ro-
tifer Brachionus plicatilis) in larvae of the mud crab
Scylla serrata.
How GPT, length, diameter, and volume of the
digestive system, and enzyme activity covary dur-
ing the complete development of larvae is uncer-
tain. Evjemo et al. (2000) reported that ingestion
rate of the unicellular alga Isochrysis galbana by
Artemia franciscana increased among three different
stages (and sizes) of larvae and postlarvae; how-
ever, there were no significant differences in GPT
among these groups. As the larval groups differ
significantly in average length (1.09, 2.74, and 5.15
mm, respectively), the velocity of material move-
ment through the digestive system must be dif-
ferent. The differences in length of the digestive
system may explain how, despite a similar GPT,
ingestion rate and extraction efficiency were in-
versely related. The decrease in the proportion of
the ingested materials that is assimilated is appar-
ently compensated by the increased rate of deliv-
ery of food into the digestive system. For feeding
larvae, the rates and composition of ingested foods
and the GPT influence a larva’s capacity to assimi-
late organic materials.
9.5  Ventilation of the Digestive System
A flow of seawater passes through the digestive sys-
tem of crustacean larvae, and this ventilation occurs
whether or not the larvae are actively capturing par-
ticulate foods. The digestive system of crustacean
larvae is irrigated by an oral and, in some cases, an
anal flow (see Fox, 1952, for an early review). Seawa-
ter entered the mouth of larvae (protozoea and my-
sis stage) and postlarvae of Penaeus setiferus in the
presence or absence of food, and anal irrigation was
interrupted only when feces were voided (Lovett
and Felder, 1990). Lovett and Felder (1990) sug-
gested that these flows contribute to digestive pro-
cesses by hydrostatically expanding the tubules of
the hepatopancreas and by facilitating bidirectional
128   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e

movement of materials within the digestive system.
Navarro et al. (1993) measured the “drinking rate”
of nauplius larvae of Artemia franciscana by moni-
toring the appearance of 14C in larvae incubated in
water containing 14C-labeled polyethylene glycol.
The radioactive label appeared in larvae at every
tested temperature (15–35 °C, 5 °C intervals), and
the measured drinking rates (nL × hour-1—larva-1)
equaled (15 °C) or exceeded (≥ 20 °C) the measured
volume of the digestive system (0.55 nL).
More recently, Morais et al. (2004) exposed newly
hatched nauplii of Artemia sp. to a mixture of 14C-­
labeled amino acids and reported a rapid appear-
ance of the 14C label in larvae. Although the cuticular
exoskeleton of crustacean larvae and adults is gen-
erally thought to inhibit assimilation of DOM from
seawater (reviewed in Manahan, 1990), the major-
ity (≈85%) of the 14C label in Artemia nauplii was re-
covered as trichloroacetic acid–insoluble materials
(“proteins”). The anatomical location of the 14C-label
in these larvae was not identified, but assimilation
of these amino acids by cells of the digestive system
could explain this experimental result. More work
is needed to understand how seawater ventilation
influences digestive processes in crustacean larvae.
Ingestion of seawater by larvae is not unique to
crustaceans. For feeding larvae that concentrate
and clear particles from seawater, the movement of
material into the digestive system is accomplished
within a discrete bolus or stream of seawater. Gil-
mour (1985; 1986) described the movement of
streamlines of seawater into the digestive system
of feeding larvae of the sea urchin Lytechinus pic-
tus. The proportion of this flow that passed through
the esophagus into the stomach was not reported.
However, the ingestion of seawater by a variety of
feeding larvae is inferred by the delivery of minute
particles or organic molecules into the digestive
system (Figure 9.1). However, little is known about
how consumption of seawater impacts chemical di-
gestion and subsequent assimilation of particulate
(≥1 μm) foods (see Stumpp et al., 2013, for a study
of how ocean acidification may influence chemical
digestion by echinopluteus larvae).
Recent interest in the ecological and toxico-
logical effects of nanoparticles on aquatic ecosys-
tems has resulted in the discovery that feeding
larvae of several taxa (Echinodermata: Magesky
et al., 2016; Crustacea: Bergami et al., 2016; and
Mollusca: Chan and Chiu, 2015) can ingest these

E
T
S M

M
Mg

Figure 9.1  Light micrographs of larvae of Phoronis architecta exposed to the iron-containing protein ferritin (0.5 mg/mL) in seawater for four
hours. The darker shade denotes the location of Fe3 + (originally present in ferritin) within the digestive system and the bounding epithelia. Larvae
not exposed to ferritin (controls) contained significantly fewer and less intense spots (not shown). Left: Whole mount. Scale bar = 150 μm. Right:
1-μm transverse section at the level of the stomach. The darker-shade reaction product is contained within the epithelial cells of the larval stomach;
some of the apparent vesicles containing the reaction product are indicated by white triangles. Scale bar = 50 μm. Abbreviations: E = esophagus, M
= metasomal sack, Mg = midgut, P = proctodeum, S = stomach, T = larval tentacle (see Plate 7).

minute particles that are <200 nm in diameter. For
feeding larvae of echinoderms, particles of this
size do not induce behaviors normally associated
with the active capture of particles (Strathmann,
2007). Although the mechanism for ingesting
such tiny particles is not well explored (but see
Shimeta, 1993; Pernet, this volume), ingestion of
particle-containing seawater represents a possi-
ble mode of delivery into the digestive system.
After silver nanoparticles entered the digestive
system of pluteus larvae of the sea urchin Strong-
lyocentrotus droebachiensis, they were detected in
cells of the digestive epithelium (Ma and Lin,
2013; Beddoes et al., 2015) and were subsequently
translocated to other regions of the larval body
(Magesky et al., 2016).
9.6  Digestion and Absorption
Digestion of particles ingested by larvae occurs
intracellularly, extracellularly, or through a com-
bination of both processes. Intracellular digestion
begins with the internalization of particles by cells
through phagocytosis and is completed within
membrane-bounded phagosomes. Phagocyto-
sis of intact or fragments of algal cells has been
reported for larvae of molluscs (e.g., Kempf and
Hadfield, 1985; Muniai­n et al., 2001). In contrast,
extracellular digestion requires the delivery of en-
zymes into the lumen of the digestive system. Ex-
amination of the fine structure of the epithelium
lining specific areas of the digestive system of
feeding larvae has revealed cells containing elec-
tron-dense, membrane-bounded vesicles. These
“secretory” cells are hypothesized to be the source
of digestive enzymes released into the lumen of
the larval digestive system (echinoderms: Burke,
1981; Huvard and Hollan­ d, 1986; enteropneust
hemichordates: Dautov et al., 1994; crustacean­s:
Lovett and Felder, 1989).
Digestive enzymes from a number of different
larval forms have been reported and include a di-
verse array of endo- and exoproteases, carbohy-
drases, and lipases (reviewed in Jones et al., 1997;
Anger, 2001, for crustacean larvae). Earlier stud-
ies, where the activities of relatively few enzymes
within homogenates of whole animals were char-
acterized, have been replaced by more specific
and diverse chemical assays (e.g., Colli­ n and
P h ys i o l o g y o f L a r va l F e e d i n g    129
Starr, 2013). More recently, changes in the diges-
tive enzymes produced by larvae have been as-
sessed using mRNA-seq (Wei et al., 2014a; 2014b;
Li et al., 2015; Yang et al., 2015; Perillo et al., 2016).
Type and abundance of digestive enzymes re-
flect the dietary preferences of the larvae. Among
crustacean larvae, those that primarily consume
phytoplankton or detritus have lower abundances
of proteolytic enzymes than larvae that consume
zooplankton (see Anger, 2001, and Le Vay et al.,
2001, for reviews). For those species where a die-
tary shift occurs during larval life, there is evidence
for a compensatory shift in both the number and
type of released proteases (Johnston et al., 2004;
Andres et al., 2010).
The end products of extracellular digestion are
assimilated by cells that line the digestive tube
(stomach, intestine, or equivalent). Specific mol-
ecules, like amino acids, are assimilated by solute-
specific transport proteins (Applebaum et al., 2013;
Meyer and Manahan, 2009); nonspecific assimila-
tion can occur by diffusion and pinocytosis. Pino-
cytosis is a broadly distributed form of absorption
(Huvard and Holland, 1986) that is inferred ana-
tomically by the presence of invaginations of the
apical epithelium and membrane-bounded, elec-
tron-luscent vesicles within the apical cytoplasm
of cells and by the presence of a discretely visible
label within these vesicles (Huvard and Holland,
1986; Rivest, 1992).
Although quantitative data are few, there are
apparent region-specific differences in the abil-
ity of cells of the digestive system to absorb or-
ganic materials. Absorptive cells of the stomach
and intestine of feeding larvae of the sea urchin
Lytechinus pictus and the sea star Patiria miniata
(Echinodermata) can assimilate dissolved pro-
teins (Huvard and Holland, 1986); there was no
evidence of assimilation by esophageal cells.
DeBurg­h and Burke (1983) reported amino acid
uptake by the epithelia of the stomach and intes-
tine of pluteus larvae of Dendraster excentricus, but
again no esophageal assimilation was detected.
Similarly, stomach cells of actinotroch larvae of
Phoronis architecta (Phoronida) revealed an ap-
parently greater capacity to assimilate the iron-­
containing protein ferritin than epithelial cells
from anterior and posterior regions of the diges-
tive system (Jaeckle, unpubl., Figure 9.1).
130   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e

9.7  Extraction Efficiency

The proportion of ingested organic materials that
is assimilated by cells of the digestive system is
the extraction efficiency (assimilation efficiency,
Table 9.2). How this measure of digestive physi-
ology is calculated varies among researchers,
and the proportion of assimilated carbon that is
lost as CO2 is commonly not reported. Irrespec-
tive of how extraction efficiency is calculated, it
is sensitive to both the anatomic and physiologi-
cal characteristics of the digestive system and the
chemical composition of the ingested food. The
extraction efficiency is approximated by the fol-
lowing relationships (Karaso­v et al., 2011; ­Karasov
and Douglas, 2013) among properties of the diges-
tive system:
[ reaction rates of  chemical digestion] × 
[ food retention within the digestive system]
[ concentration of  materials within digestive system] × 
[ volume of  the digestive system]
The retention time of ingested food within the
digestiv­e system is predicted as
[ volume of  digestive system] 
[ fluid  flow rate through the digestive system]
Estimates of extraction efficiency (“retention effi-
ciency”; Pace et al., 2006) for invertebrate larvae are
typically made without knowledge of some of these
components. In the absence of such data, differences
in published values of extraction efficiencies among
larval forms (Table 9.2) are difficult to explain with
confidence.

Table 9.2  Variation in the Extraction (Assimilation) Efficiency of Elements within Particulate Foods Ingested by Marine Invertebrate Larvae.

Phylum, Species Element -species Extraction Efficiency (± SE) Source

Mollusca
Ilyanassa obsoletus C- Isochrysis galbana 40.5 (± 4.8) Pechenik, 1980
Ilyanassa obsoletus C- Isochrysis galbana 16.71 Pechenik and Fisher, 1979
C- Dunaliella tertiolecta 8.71 Pechenik and Fisher, 1979
C- Thalassiosira pseudonana 10.71 Pechenik and Fisher, 1979
Bittium alternatum C- Isochrysis galbana 40.8 (± 1.6) Pechenik, 1980
Crepidula fornicata C- Isochrysis galbana 69.82 Pechenik, 1980
Crassostrea gigas C- Isochrysis galbana 461 (150 µm); Pace et al., 2006
251 (300 µm)
Mercenaria C- Isochrysis galbana ~551 Gallager et al., 1994
mercenaria
Mercenaria C- Synechococcus sp ~551 Gallager et al., 1994
mercenaria
Crassostrea virginica C- Isochrysis galbana3 53.61 Reinfelder and Fisher, 1994
Mercenaria C- Isochrysis galbana3 62.61 Reinfelder and Fisher, 1994
mercenaria
Arthropoda, Crustacea
Artemia franciscana C- Isochrysis galbana (mg C/L) Evjemo et al., 2000
1
0.5 88.2
2.5 75.01
7.0 62.71
15 36.81
30 39.21

Note. All values included in this table were determined by experimentation.

1
Reported values are mean values reported by the authors.
2
Reported values represent a single measurement.
3
The extraction efficiency of other elements was reported in this work.

Examination of variation in the extraction ef-
ficiencies among stages or ages of larvae within
species, however, suggests some general patterns.
For example, the extraction efficiency of nauplii of
Artemia franciscana differed among stages and sizes
of larvae, but decreased with increasing prey ra-
tion for all larval stages (Evjemo et al., 2000). Pace
et al. (2006) measured the extraction efficiencies for
different-sized larvae of the Japanese oyster Crassos-
trea gigas from known pedigrees and specific crosses,
and found no statistically significant difference be-
tween inbred and hybrid larvae. However, they also
reported a significant difference in ingestion rates
between these two larval groups (inbred < hybrid)
and detected no difference in food retention (= GPT)
in the digestive system. In contrast, when data for
all crosses were combined, there was a significant
decrease in extraction efficiency as larvae grew from
150 µm (~46% extraction efficiency) to 300 µm (~25%
extraction efficiency) in shell length. Why the extrac-
tion efficiencies of hybrid and inbred larvae were
similar is unknown and may relate to an unmeas-
ured difference between the two groups of larvae.
Although larval feeding is generally modeled
as a two-step process of particle concentration and
capture (see Pernet, this volume), as mentioned ear-
lier, seawater enters the digestive system of inver-
tebrate larvae. Any DOM within this flow would
enter the digestive system and become available for
digestion and assimilation. A number of laboratory-
based experiments using autoradiographic tech-
niques have demonstrated the assimilation of DOM
by the epithelia of the digestive system of larvae.
When veliger larvae of the bivalves Crassostrea gi-
gas, Ostrea edulis, and Mytilus edulis larvae were in-
cubated with 3H-labeled glycine (1 µM), the majority
of the label was present in ectodermal cells directly
exposed to seawater (Manahan and Crisp, 1983).
However, when incubation periods were ≥100 min-
utes, the label was also detected within the cells of
the digestive system. Similarly, DeBurgh and Burke
(1983) also reported that the cells of the stomach and
intestine of pluteus larvae of the sea urchin Den-
draster excentricus became labeled after a 60-minute
exposure to a mixture of 3H-labeled amino acids
(6.74 µM). Here, too, the intensity of the label in
the cells of the digestive system was lower than
that found in the cells of the outer epithelium. In
P h ys i o l o g y o f L a r va l F e e d i n g    131
contrast, Meyer and Manahan (2009) did not detect
uptake of a 3H-label by the digestive system of plu-
teus larvae of Strongylocentrotus purpuratus after a
100-minute exposure to 1 µM 3H-alanine. Although
the cause(s) of these differences among larvae is un-
known, this variation is potentially explained by dif-
ferences in the rate of seawater ingestion relative to
the duration of the experiment, the surface area of
the absorptive epithelium, the concentration of the
label, and the volume of the digestive system.
Absorption of dissolved macromolecules in sea-
water by the digestive system of larvae has also been
described. Huvard and Holland (1986) detected
ferritin in pinocytotic pits and within membrane-
bounded vesicles of epithelial cells of the digestive
system of Lytechinus pictus and Patiria miniata, but no
ferritin was detected in the outer epithelium. More
recently, Skikne et al. (2009) reported that ephyra
larvae of the scyphomedusa Aurelia labiata when
incubated with a fluorescently labeled lysine poly-
mer (3 µM) rapidly accumulated the label within the
branched gastrovascular cavity. The distribution of
the fluorescence increased “over time” as the larval
body became uniformly labeled. It is unknown if
the movement of the label within these larvae accu-
rately reflects the translocation of the intact polymer.
9.8  Internal Transport Systems
The movement of materials (water and solutes) from
the lumen of the digestive system to other areas of
the larval body may occur through diffusive move-
ment between (paracellular transport) or through
(transcellular transport) cells. Among adult inver-
tebrates and their developmental stages, epithelial
resistance to paracellular transport is regulated
by subapical septate junctions (Itza and Mozingo,
2005; Jonusaite et al., 2016). The “tightness” of epi-
thelia in larvae was demonstrated by Dan-Sohkawa
et al. (1995), who reported that epithelia of embryos
and larvae of the sea stars Asterias amurensis and As-
terina pectinifera were normally impermeable to flu-
orescein isothiocyanate-labeled proteins. However,
exposure to hypertonic seawater decreased the
number of physical contacts (septa) between adja-
cent cells, and this structural change was correlated
with the appearance of fluorescence within the blas-
tocoel. The effect of the hypertonic treatment (and
132   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e
other treatments that destabilized septate junctions;
Kaneko et al., 1995) was reversed after the embryos
and larvae were returned to unaltered seawater,
and the physical integrity of the septate junctions
was re-established. The appearance of nanoparti-
cles within the epithelial cells of the digestive sys-
tem, blastocoelic mesenchyme cells, and coelomic
epithelia of sea urchin larvae (Magesky et al., 2016)
is explained best by transcellular processes.
The linkages between feeding and growth and
development of larvae are intuitive. However, the
pathways and regulatory mechanisms that distrib-
ute absorbed nutrients throughout the larval body
remain unknown and understudied. Distribution
of assimilated materials could be a consequence of
diffusion or the combination of diffusive and ad-
vective flows. Advection of fluid within and among
compartments within the larval body is created by
a pressure gradient formed by the beating of ne-
phridial cilia (Ax and Bartolomaeus, 1992; Ruppert
and Smith, 1988), contraction of body wall muscula-
ture, and the movement of appendages.
General morphological complexity of larvae and
the size of body compartments (blastocoelic or coel-
omic) vary among taxonomic groups. Yet, despite
structural variability, the presence of a distribution
system within feeding larvae must be common to
all. In the absence of such a system, ectodermal
and mesodermal cells would receive their nutrition
from source(s) independent of the digestive system.
Similarly, how cells that are part of the digestive sys-
tem—but lie anterior to the sites of nutrient assimi-
lation—receive nutrition is also unresolved. Under
these circumstances, spatially separated epithelia
would be nutritionally isolated from one another.
The elements of a nutrient distribution (circula-
tory) system are well known from studies on larger
animals (Schmidt-Nielsen, 1990). For feeding lar-
vae that lack a spacious internal body cavity (e.g.,
anthozoan planulae), assimilated materials could
simply be released from the basolateral surfaces
of the absorptive cells and diffuse across the me-
soglea to superficial cells. For feeding larvae with
internal body cavities, the release of assimilated
materials across the basolateral surfaces of the ab-
sorptive cells would be a prerequisite for a nutrient
distribution system. In those larvae with an excre-
tory system, the movement of blastocoelic fluid
during the formation of the primary (unmodified)
urine could serve as the motive force to distribute
organic molecules or deliver material to a coelomic
circulatory system.
For those feeding larvae with an expansive
blastocoel (some echinoderms and enteropneust
hemichordates), the “larval kidney” has been hy-
pothesized to contribute to the distribution of ma-
terials within larvae (Ruppert and Balser, 1986).
Analysis of morphological and experimental data
from feeding larvae of the sea star Asterias forbesi
and the tornaria larvae of Schizocardium brasiliense
supports the existence of a flow of blastocoelic
fluid across a permeable region of the mesothe-
lium into the left anterior coelom(s). According
to their hypothesis, the composition of this “pri-
mary urine” would be modified through reab-
sorption as it is propelled along the pore canal
that leads to a single pore on the dorsal surface of
the larvae. When present, such a flow provides a
means to propel and distribute materials through
the gel-like extracellular matrix of the blastocoel
(Strathmann, 1989). Although the presence of
a nephridial system is common in most feeding
larvae of triploblastic invertebrates (Ruppert and
Smith, 1988), if and how it contributes to the dis-
tribution of organic materials throughout the lar-
val body is not known.
Feeding larvae of asteroid echinoderms develop
a variably connected system of coelomic spaces; the
cilia of the apical surface of the mesothelial cells
propel the coelomic fluid within these spaces. As
the coeloms connect during development (Gemmil­l,
1914) and the apical surface of coelomic epithelium
is ciliated, a circulation of coelomic fluid could dis-
tribute materials throughout this common space;
Gemmill (1915) depicted such a coelomic circula-
tory system in these larvae. There is experimental
evidence for such a coelomic circulatory system in
feeding larvae of the sea urchin Paracentrotus depres-
sus. Unuma et al. (2009) reported the synthesis of a
180 kDa glycoprotein (“major yolk protein”) by epi-
thelial cells of the digestive system of late-stage plu-
teus larvae. The detection of this protein within the
blastocoel and the epithelium of the posterior-most
coeloms (somatocoels, metacoels) is consistent with
a connection between blastocoelic and coelomic cir-
culatory systems.

9.9  Alternative Sources of Organic
Materials—Dissolved Organic Materials
The organic material consumed by feeding larvae
exists as particles (bacteria, phytoplankton, and
zooplankton) that are actively collected from sea-
water. Yet other sources of organic materials exist
in seawater that are too small to be captured by
commonly acknowledged mechanisms (see Per-
net, this volume). DOM in seawater represent a
large pool of organic carbon that is heterogeneous
in size, composition, and biological availability to
larvae. Recognized components of DOM include
specific dissolved organic molecules (e.g., amino
acids, monosaccharides, nucleic acids), polymeric
macromolecules, gels, colloids, and bacteriophages
(­Passow, 2002; Ogawa and Taneou, 2003; Verdugo et
al., 2004; Azam and Malfatti, 2007; Verdugo, 2012).
For feeding larvae, absorption of DOM may occur
across the outer epithelium or by the activity of
the cells of the digestive epithelium. The degree to
which “dissolved” materials in seawater contribute
to the survivorship, growth, and development of
marine invertebrate larvae is a consequence of their
form and abundance and the differences that exist
among the anatomical and physiological properties
of larvae (Manahan, 1990).
Studies of DOM uptake by larvae have histori-
cally been focused on rates of transport of specific
molecules. Invertebrate larvae of almost all exam-
ined groups have revealed a capacity to transport
organic molecules from seawater (Manahan, 1990).
Larvae of the Crustacea represent an apparent ex-
ception, as the presence of a continuous exoskeleton
impedes direct exposure of the outer epithelium to
seawater. Earlier reports of DOM uptake by crusta-
ceans were attributed to the presence of epicuticular
bacteria (Anderson and Stephens, 1969). An inabil-
ity to detect uptake of dissolved solutes by feeding
larvae of crustaceans is somewhat surprising in
light of reports of their ingestion of seawater (see
Section 9.5). If seawater containing DOM enters the
digestive system of crustacean larvae, then varia-
tion in a larva’s ability to assimilate DOM contained
within this flow is explained by factors including
the composition and concentration of the dissolved
solutes, the rate of delivery of these materials into
the digestive system, the rate of movement of water
P h ys i o l o g y o f L a r va l F e e d i n g    133
through the digestive system, and physiological
properties of the absorptive cells. However, Souza
et al. (2010) demonstrated the presence of cuticular
pores in phyllosoma larvae of the Japanese spiny
lobster Panulirus japonicus. These perforations pro-
vide epithelial cells access to seawater, facilitating
their exposure to and uptake of DOM. They also
demonstrated 3H-glycine uptake by cells of the
anterior portion (oral) of the digestive system in
these larvae. The hypothesis that feeding larvae of
crustaceans can assimilate DOM from seawater is
supported by other studies (e.g., Morais et al., 2004)
reporting the appearance of a radioactive label in
crustacean larvae.
The cells responsible for the assimilation of spe-
cific organic solutes have been identified using
autoradiography for feeding larvae of molluscs
(Manahan and Crisp, 1983), echinoderms (Karp
and Weems, 1975; DeBurgh and Burke, 1983; Meyer
and Manahan, 2009), and crustaceans (Souza et al.,
2010). In every instance, the label accumulated in
cells directly exposed to seawater. After short expo-
sures, the cells of the outer epithelium contained the
radioactive label, but with extended incubations the
label also appeared in epithelial cells of the diges-
tive system. The temporal lag in the appearance of
the label between the two epithelia can be attributed
to differences in exposure to the solutes, the surface
area of the absorptive cells, and the biochemical
properties of transport proteins (i.e., substrate affin-
ity and transport capacity).
The uptake of small organic molecules by lar-
vae appears to be the result of cellular assimilation
rather than passive diffusion of materials between
ectodermal epithelial cells. The connections that ex-
ist between cells of the outer epithelium of larvae
from different taxa consist of an apical zonula ad-
herens and subapical septate junction (see Bereiter-
Hahn et al., 1984; Harrison et al., 1990–1999). The
apparent impermeability of the outer epithelium of
embryos and early larvae of the sea stars Asterias
amurensis and Asterina pectinifera is mediated by
the integrity of septate junctions between epithe-
lial cells (Dan-Sohkawa et al., 1995; Kaneko et al.,
1995). These older, microscopy-based descriptions
of membrane impermeability are in contrast to later
work by Stumpp et al. (2011), who reported a “rapid
equilibrium” between seawater and the blastocoelic
134   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e
space in pluteus larvae of the sea urchin Strongylo-
centrotus droebachiensis. Using microelectrodes and a
pH-sensitive indicating dye, they reported that the
pH of the blastocoel mirrored that of the surround-
ing seawater, suggesting the existence of a “leaky”
outer epithelium. When these larvae were exposed
to fluorescein-labeled dextrans (4–40 kDa), fluores-
cence appeared within the blastocoelic space after
incubations of 15 and 60 minutes. These results
were interpreted as evidence of paracellular move-
ment of materials through the outer epithelium.
Such epithelial permeability argues against the
presence of a nutrient distribution system in these
larvae. If small molecular weight materials are re-
leased by absorptive cells, then they would poten-
tially diffuse across the blastocoelic space and exit
the larval body in these spaces between cells. An
understanding of the permeability of all epithelia
exposed to seawater is necessary before this appar-
ent paradox can be resolved.
Assimilation of these materials by nonfeeding
larvae results from active or passive transport of
materials uniquely across the outer epithelium.
Although nonfeeding larvae take up specific mol-
ecules from seawater (Jaeckle and Manahan, 1989a;
Jaeckle, 1994; 1995; Shilling et al., 1996; Ben-David-
Zaslow and Benayahu, 2000), there are few data
from which to generalize about the sites of DOM
uptake and potential translocation to deeper (in-
terior) cells by nonfeeding larvae. Jaeckle (1995)
reported that parenchymulla larvae of the demos-
ponge Tedania ignis assimilated the fatty acid pal-
mitic acid (3H-labeled) from seawater. With longer
incubation periods, the radioactive label appeared
in deeper areas of the larval body, suggesting the
presence of a distribution system of organic mate-
rial within this larva.
Most studies of DOM uptake by larvae reveal
their capacity to assimilate a specific molecule(s).
For both feeding and nonfeeding larvae, rates of
amino acid transport from seawater can be suffi-
ciently high to represent a substantive proportion
of aerobic metabolism (Manahan, 1990). In con-
trast, transport of simple sugars by larvae does
not (Welborn and Manahan, 1990). Johnson and
Wendt (2007) used a different, holistic approach
to assess the effect of DOM on larvae. They reared
nonfeeding larvae of the bryozoan Bugula neritina
in seawater containing a fatty acid or a mixture
of amino acids, and monitored the effect of each
treatment to reduce the energetic cost created by
a prolonged larval life (detected as differences in
juvenile performance). Juveniles from larvae held
in augmented seawater experienced lower mortal-
ity, greater metamorphic success, and produced
larger feeding structures than juveniles from larvae
that were reared in a reduced DOM treatment. The
rates of oxygen consumption by larvae experienc-
ing these treatments did not differ, and Johnson
and Wendt (2007) suggested that the observed dif-
ferences were explained by the transport of DOM
from seawater.
Given the diversity in composition and size of
DOM in seawater, such solute-specific studies do
not reveal how larvae exploit other undefined com-
ponents of DOM in seawater. For example, the abun-
dance of bacteriophages in seawater (~108–1011/L;
Suttle, 2007; Danovaro et al., 2011) and the amount
of organic carbon they represent (~400 nM; Jover et
al., 2014) suggest that this form of DOM could be an
unrecognized source of organic material for feed-
ing larvae. Despite their abundance, if and to what
degree bacteriophages and other minute (<0.45 µm)
organic particles in seawater directly contribute to
the nutrition of feeding and nonfeeding larvae is
unrecognized.
However, for some feeding larvae, there are exam-
ples where the ability to transport DOM from seawa-
ter could explain resiliency to an apparent absence
of particulate foods. Moran and Manahan (2004)
reared feeding larvae of the Japanese oyster Cras-
sostrea gigas in filtered seawater (0.2 μm pore size)
and reported ~75% survivorship after two weeks
and ~15% survivorship after three weeks. Rates of
aerobic metabolism for these larvae remained un-
changed throughout the experiments. When the
metabolic rates of larvae were converted to a rate of
self-consumption (as predicted from the absence of
particulate foods), the larvae were persisting approx-
imately two weeks beyond the time when the initial
content in the egg would be consumed. This result
is not unique, as experiments on feeding larvae of
polar (Sterechinus neumayeri: Marsh et al., 1999; Od-
ontaster meridionalis: Ginsburg and Manahan, 2009)
and temperate (Patiria [Asterina] miniata: Pace and
Manahan, 2007; Strongylocentrotus purpuratus: Meyer

et al., 2007) echinoderms have also reported a similar
and unexpected resiliency to deprivation from par-
ticulate food (phytoplankton). In each of these cases,
during those periods when phytoplankton were
absent, the maintenance metabolism of these larvae
is hypothesized to be supported by an external, un-
characterized source(s) of organic material.
Such resiliency to periods of food deprivation
is not seen in all feeding larvae. The stasis of size
and metabolism of larvae reared in filtered seawa-
ter is not reported for feeding larvae of crustaceans.
When crustacean larvae are reared in seawater
without added food, there is a decrease in biomass,
biochemical composition, and aerobic metabolism
(reviewed in Anger, 2001). If starvation conditions
extend beyond a certain time (variable among and
within species), the reintroduction of food will not
affect the ability of larvae to complete development.
Anger and Dawirs (1981) described this as a “point
of no return” (PNR50), the time period of nutrient
deprivation after which >50% of the larvae cannot
progress to the juvenile stage. After larvae of crus-
taceans have exceeded their PNR, if access to and
ingestion of food occurs, they remain incapable of
completing the next molt cycle.
For larvae that grow continuously (i.e., not incre-
mentally), determining the PNR requires culturing
larvae through metamorphosis. When “unfed” (≤17
days) larvae of C. gigas were provided phytoplank-
ton, the resulting rates of growth (shell length),
metabolism, and accumulation of organic material
approximated those of newly formed veliger lar-
vae (Moran and Manahan, 2004). This capacity to
respond to food availability after 17 days of an un-
fed treatment suggests that, although development
was retarded, these larvae have the ability to con-
tinue and potentially complete their developmental
program. Fu et al. (2014) monitored the viability of
ephyra larvae of Aurelia aurita that were deprived of
food, and the PNR50 for these larvae ranged from 34
days (15 °C) to 59 days (9 °C). In contrast to the sta-
sis in size seen in larvae of C. gigas, during extended
periods of nutrient deprivation, larvae of A. aurita
experienced significant self-consumption (42%–51%
of initial carbon content). Although the physiologi-
cal basis for the PNR is not fully understood, the du-
ration of the PNR50 is related to a number of factors,
including the initial size of the larvae at the onset
P h ys i o l o g y o f L a r va l F e e d i n g    135
of starvation (Xiwu et al., 2009), the metabolic rate
of the larvae (Fu et al., 2014), the availability of al-
ternative food resources, and the ability of larvae to
exploit these materials.
9.10  An Example of Physiological
Responses to Environmental
Change—Ocean Acidification
Ocean acidification caused by increasing atmos-
pheric CO2 can affect invertebrate larvae in a
number of different ways, including changes in
transcription, larval shape, and rates of develop-
ment, growth, and metabolism (Hofmann et al.,
2010; Gazeau et al., 2013; Ross et al., 2016; Byrne et
al., this volume). The effects of ocean acidification
on embryos and feeding larvae of sea urchins have
been detected as changes in morphology (Chan et
al., 2011), gene expression (Pespeni et al., 2013), and
a number of measures of larval metabolism and di-
gestive physiology (Stumpp et al., 2011; 2013; Yu et
al., 2011; 2013; Pan et al., 2015).
Any negative effect on development rate and
growth rate of larvae after exposure to acidified
seawater may be associated with a decrease in the
digestive efficiency. Although the feeding rate of lar-
vae does not seem strongly influenced by environ-
mental acidification (Stumpp et al., 2011; 2013), this
treatment decreased the pH of the stomach contents
in larvae of the sea urchins Strongylocentrotus droe-
bachiensis by 0.3–0.5 pH units (Stumpp et al., 2013).
This change in the internal environment of the diges-
tive system is predicted (Stumpp et al., 2013; 2015) to
impair the function of digestive enzymes. Assuming
that the GPT is insensitive to acidification, then this
decrease in digestive function alone could reduce
the extraction efficiency of these larvae.
Alternatively, an increase in metabolic demand
in response to exposure to acidified seawater may
reduce the amount of energy or materials avail-
able for larval development and growth (Pan et al.,
2015). The nutritional history of larvae, however,
may also influence their metabolic response to acidi-
fied seawater, but the direction of the responses was
different between larvae of the sea urchins Strongy-
locentrotus purpuratus and S. droebachiensis (Stumpp
et al., 2011; Pan et al., 2015; Byrne et al., this volume).
136   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e
The aggregated work on the effects of seawater
acidification on the biology of Strongylocentrotus
spp. larvae highlights the importance of study-
ing both the structural and functional attributes
of larvae. A decrease in development or growth
rate may be explained, in part, by change(s) in a
number of different processes (e.g., feeding rate,
efficiency of digestion, GPT, extraction efficiency)
that link the structure of larvae to their physiologi-
cal properties. The importance of integrating the
functions among spatially distant, differentiated
cells in “large” animals is readily acknowledged
(e.g., Schmidt-Nielsen, 1990). Despite their small
size, larvae share many of these properties, and
understanding how a larva “makes its living” re-
quires an appreciation of its structural and func-
tional attributes.
9.11 Summary
1. Any measure of performance (e.g., growth, me-
tabolism, development) of larvae represents the
sum of the activities of the independent and in-
terdependent cells that make up the whole.
2. Conversion of ingested particulate and, in part, dis-
solved organic materials into growth and develop-
ment of feeding invertebrate larvae is dependent
upon the anatomical and physiological properties
of cells that make up the digestive syste­m.
3. Delivery of assimilated foods to physically dis-
tant cells requires the presence of a distribution
(circulatory) system.
4. Ingestion of (drinking) water may represent
a mechanism common to feeding larvae that
deliver­s dissolved organic materials into the di-
gestive system.
5. Within and among feeding larvae of inverte-
brates, variation and inducible plasticity species
of secreted digestive enzymes, transit time of food
through the digestive system, and mechanisms of
assimilation are understudied or unknown.
Acknowledgments
I appreciate the efforts of Crystal Boyce (Ames
Library) in tracking down early use of “physiol-
ogy is the handmaid of anatomy.” The quality of
this document was significantly improved by the
comments of Richard Strathmann and two anony-
mous reviewers; any errors in this document are of
my own making. This work would not exist with-
out the support of Nisa Blackmon and Jolie Hoff-
mann; their patience is greatly appreciated.
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 Ch a pt er 10
Section 2 Summary—Functional
Morphology and Ecology of Larval
Forms
Larvae are intermediate life history stages between
embryos and juvenile and/or reproductive stages,
but this characteristic is about the only feature that
unites the incredible diversity of larval forms. The
majority of larval forms evolved in the sea and ex-
hibit tremendous morphological, physiological,
and molecular variation, many of which are po-
tential adaptations to match form and function in
the context of the aquatic environment. The three
chapters in this section review how larvae from dif-
ferent taxonomic groups sort through and ingest
exogenous nutrients and how environmental vari-
ation elicits morphological variation.
Planktotrophic larvae from various phyla of ma-
rine species have been studied to characterize the
diverse mechanisms for food capture and how these
relate to other features of larval shape. In Chapter 7,
Bruno Pernet reviews the methods which larvae
with diverse morphologies use to feed. Specifi-
cally, this chapter synthesizes feeding mechanisms
across  phyla and larval types of marine inverte-
brates. ­Using empirical data and model-based pre-
dictions, he then compares rates of feeding and the
size spectrum of exogenous food to understand the
performance of particular larval types in nature.
The movement of studies from simple experimental
designs (e.g., single algae species, one concentration
of food) to conditions more representative of natural
Carrier, T. J., Reitzel, A. M., and Heyland, A., Section 2 Summary—Functional morphology and ecology of larval forms. In. Evolutionary
Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press
(2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0010
pelagic environments is uncovering additional
complexity of larval morphology and physiology.
For feeding larvae, the planktonic environment
is heterogeneous with respect to food availability
over space and time. Over the past few decades,
considerable progress has been made in showing
how larvae, like adults, can have tremendous plas-
ticity in their morphology depending on the food
environment. In Chapter 8, Justin McAlister and
Benjamin Miner review the phenotypic plasticity
of feeding structures in marine invertebrate larvae.
They discuss how careful consideration of experi-
mental design and associated statistical analyses
are necessary for interpreting data from studies
of environmentally induced structures. They con-
clude with a discussion on the evolution and mo-
lecular underpinnings of morphological plasticity
in larvae.
Finally, in Chapter 9, William Jaeckle synthe-
sizes the extensive literature on the physiology of
larval feeding, beginning with the acquisition of
exogenous particles or dissolved organic material
and ending with the delivery and assimilation of
biomolecules. He provides a summary of the move-
ment of materials in the digestive system of larvae
and patterns of extraction efficiency in different
taxa. These physiological processes are placed in
the context of ongoing climate change.
 C h a pt er 11

Larval Transport in the Coastal Zone:

Biological and Physical Processes
Jesús Pineda and Nathalie Reyns

But the most frequent mode of transportation, probably, consists in their buoyancy of their eggs…
These…may be swept along by a wave that breaks upon a coral-reef, and may then be borne by a
current to a distance. Charles Lyell (1832, p.112)

11.1 Introduction Larval transport is a concept that can also be de-

The life cycle of most benthic marine metazoans is
characterized by large fecundities and a two-phase
life cycle, where individuals produce hundreds to
millions of eggs. Typically, fertilized zygotes de-
velop into minute, swimming larvae, and larvae
travel away from adult habitats as they develop. Af-
ter developing for minutes to several months, larvae
must return to suitable adult habitats to settle, grow,
and reproduce; this migration is mediated by larval
transport (see review of concepts by Young, 1990).
Larval transport can play a key role in population
dynamics by determining settlement rates (Rough-
garden et al., 1988). Further, the local circulation
regime (Zimmer et al., 2012), in conjunction with
behavioral and morphological characteristics of the
larvae, may cause local population fecundities to
decouple from local settlement rates (reviewed in
Pineda et al., 2010), such as when larvae produced
at one location are advected away from natal habi-
tats and must settle on distant habitats. Thus, larval
transport is a key biological-physical process in the
ecology of benthic populations and communities.
Further, larval transport is funda­mental to the dy-
namics of metapopulations (Botsford et al., 2001),
with implications for fisheries management and
conservation.
scribed as a measurement and, more specifically,
quantified as a flux, a quantity flowing across a sur-
face per area and time (e.g., an “Eulerian” approach).
For example, larval flux across a lagoon’s mouth dur-
ing the incoming tide (e.g., DiBacco and Chadwic­k,
2001) can be quantified using fixed nets at the la-
goon’s mouth, together with measurements of flow
across the mouth (with units # larvae/L2 · time).
To be most relevant to understanding population
processes, larval transport should be measured dur-
ing the entire settlement season, including periods
when the bulk of larvae are transported. Measuring
larval transport as larval flux in these cases is not
typically practical, because larval transport tends to
be sporadic, larvae are patchily distributed (Haury
et al., 1978; Natunewicz and Epifanio, 2001), and
the settlement season may be long (several months).
For example, plankton pump and trap data (Reyns
and Sponaugle, 1999; Queiroga et al., 2006), as well
as settlement, suggest that larval transport of crabs,
barnacles, bivalves, gastropods, and polychaetes
tends to be unpredictable, including long periods
of negligible transport (weeks to months) punctu-
ated by daily or even hourly events (Garland et al.,
2002; Porri et al., 2006; Reyns et al., unpub.) when
the bulk of the larvae are transported. For meas-
ures of larval flux to be effective in demarcating

Pineda, J. and Reyns, N., Larval transport in the coastal zone: biological and physical processes. In. Evolutionary Ecology
of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018).
© Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0011
146   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v ert e b r at e L a r va e
the relevant components of larval transport, con-
tinuous measurements resolving short timescales
(minutes to hours) and sustained for months would
be necessary. For most open coastline applications,
however, such time-series biological measure-
ments would be impractical, requiring fixed sam-
pling structures to measure larval fluxes through a
depth layer or the entire water column. Moreover,
with current methodologies, the effort required to
process larval samples would be immense. Despite
the potential advantages of measuring larval trans-
port as larval flux, so far these measurements are
prohibitively taxing for the majority of benthic sys-
tems, and therefore marine ecologists have resorted
to other descriptions of larval transport.
Larval transport has been defined simply as “the
horizontal translocation of a larva between points
x1,y1 and x2,y2, where x and y are horizontal axes, say,
perpendicular and parallel to the coastline” (Pineda
et al., 2007). In this definition, larval transport is meas-
ured in length units (e.g., meters). To more closely re-
late the meaning of larval transport to “Lagrangia­n”
descriptions of flow, which describe the trajectory of
a parcel of water with respect to a temporal dimen-
sion, a more refined definition is needed.
We define larval transport as the mean horizon-
tal translocation of larvae between points along a speci-
fied one-dimensional axis per unit time (with units L/
time). This definition does not consider timing and
magnitude of spawning, larval abundance, mortal-
ity, or settlement (discussed in Pineda et al., 2007). It
acknowledges the separation of pelagic and benthic
habitats, and accommodates the mechanistic resolu-
tion of the fundamental biological-physical processes
accounting for larval translocation along a spatial di-
mension of interest (e.g., alongshore, cross-shore).
11.2  Scales of Larval Transport
While understanding the processes that account for
larval translocation appears straightforward, sev-
eral processes are involved which range over vast
spatial and temporal scales. At one extreme, biolog-
ical-physical accumulation processes with scales of
centimeters to meters and seconds to minutes play
an important role. At the other extreme, processes
with thousands of kilometers and interannual
scales, such as the El Niño–Southern Oscillation,
modulate local biological and physical processes
that can determine larval transport, including sur-
face gravity waves (Seymour et al., 1989) and ther-
mocline depth (Filonov and Tereshschenko, 2000).
The spatial scales of larval transport—that is, the
horizontal distances covered by the larvae—must
vary in different systems (e.g., tropical estuaries,
seamounts, temperate intertidal). However, the
dominant scales of larval transport are imperfectly
known. There is some consensus that dispersal
distances in many coastal invertebrates are on the
order of 5–25 km (Shanks, 2009), but transport dis-
tance over hundreds to thousands of kilometers
have been inferred in species with a long larval
duration (Scheltema, 1968). Temporally, lunar and
seasonal variability in larval transport, inferred
from settlement data, are important in many sys-
tems, but in most cases, the relative contribution of
biological (reproductive cycles) and physical (e.g.,
water stratification, seasonal variability) processes
is not well resolved, since rarely has larval trans-
port and timing of reproduction been measured
concurrently (but see Hurlbut, 1992; Barbosa et al.,
2016). One exception is for many hermatypic corals
and other reef fauna that spawn according to the
lunar cycle and season (Harrison et al., 1984), and
thus predictably initiate larval transport. A further
complication to understanding larval transport is
that the physical processes that might be involved
are themselves imperfectly known (Lentz and
Fewings, 2012). For these reasons, a mechanistic
understanding of the processes involved in larval
transport is challenging, and the variability among
systems is likely high.
This chapter reviews larval transport, empha-
sizing the advances in the last 25 years. A full
discussion of the different processes and oceanic
regions is beyond the scope of this chapter. Thus,
we emphasize first-order phenomena, where
mechanistic understanding has been achieved
and on the topics that may need more attention.
Whereas the processes we discuss apply to most
habitats, we focus on open coastal environments,
which the bulk of studies on invertebrates are
concerned with and for which process-oriented
understanding is most advanced. We use litera-
ture for fish larvae and other environments when
it helps clarify the issues at hand.

11.3  Components of Larval Transport
and Other Relevant Phenomena
What are the components of “larval transport”—the
first-order processes responsible for the translocation
of a larva from point a to point b? And what other
general processes are relevant to larval transport it-
self, and to benthic population dynamics, including
processes determining how many larvae transport,
and the spatial and temporal variability in the num-
ber of larvae that transport? We suggest that the com-
ponents of larval transport include the following: (1)
larval behavior, particularly behaviors influencing
vertical distribution, and (2) the physical transport
mechanisms accounting for advection, diffusion, and
their variability. In instances where larval transport
is associated with propagating fronts, an additional
component is (3) the mechanism of larval accumula-
tion in those features. Knowledge of all the compo-
nents is necessary. For example, if one knows that the
cross-shore physical transport mechanism a features
an accumulation mechanism b which requires a be-
havior c for accumulation and transport, then one
could make a larval transport prediction depending
on whether an untested species possesses behavior c.
11.3.1  Larval Behavior
Horizontal flow velocities vary with depth, from
slowing down toward the bottom due to friction,
to current reversals in different layers in some
flows, and larvae may exploit this variability by
swimming vertically. The hypothesis that minute
planktonic larvae can control their horizontal trans-
port by exploiting behaviorally the layered vertical
structure of marine flows was suggested over a cen-
tury ago by J. Nelson (1912):
The swimming powers of the fry are too feeble to
change their distribution in the current directly.
But they have the power to change this distribu-
tion indirectly. For they can rise and sink, they
have the power to seek the bottom and to rise to
the top. . . . Then the larvae could control their
own distribution. (p. 307)
Thus, the essence of the “behavioral control of
larval transport” hypothesis is (a) given that hori-
zontal ocean current magnitudes are much larger
La r va l Tr a n s p o rt i n t h e C o a s ta l Z o ne    147
than non-oscillatory vertical flows (e.g., cm/s vs.
mm/s), and that (b) small invertebrate larvae are
weak swimmers and cannot generally determine
their horizontal distribution by swimming against
horizontal currents, then (c) by swimming verti-
cally, larvae can exploit the layered structure of the
horizontal currents, and in this way control their
horizontal distribution.
In its simplest and best-known version, a larva
encounters a layer where the flow is predominantly
in one direction, resulting in transport in that direc-
tion. This concept has been applied to two-layer es-
tuarine and coastal lagoon (Nelson, 1912; Bousfiel­d,
1955), nearshore and coastal (Rothlisberg and
Miller, 1983; Cowen et al., 2000; Paris and Cowen,
2004), and open-ocean (Scheltema, 1968) flows.
What triggers larvae to swim vertically to exploit
depth-dependent flows? Studies invoking horizon-
tal control of larval transport by vertical swimming
often explain vertical positioning as a fixed-schedul­e
behavior promoting return to natal habitats. For ex-
ample, crab larvae can migrate vertically by enter-
ing the water column during one phase of the tidal
cycle and remaining near bottom during the other
tidal phases, resulting in horizontal movement
termed selective tidal stream transport (Forward
and Tankersley, 2001). Another frequently invoked
view in coastal lagoons and other systems is ontoge-
netic vertical positioning, where early stage larvae
occupy a depth layer promoting their transport
away from shallow adult habitats, followed by mi-
gration of developing larvae to another depth layer
with flows promoting their return to adult habitats
(Bousfield, 1955). In each case, behavioral responses
to environmental cues, in the form of taxis or kine-
sis, mediate vertical larval migrations. Cues may
include changes in temperature, salinity, gravity,
light, turbulence, currents, chemicals, and sound,
among other factors, and have been extensively ad-
dressed elsewhere (Metaxas, 2001; Fuchs et al., 2004;
Weidberg et al., 2015; Epifanio and Cohen, 2016).
Given the times scales of larval development,
which vary from minutes to years, with a mode
of ~1–8 weeks for temperate species (Levin and
Bridges, 1995), the implication is that there must
exist mean Eulerian flows connecting juvenile and
adult benthic habitats to the “larval pool,” the pe-
lagic grounds where larvae develop. Moreover, the
148   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v ert e b r at e L a r va e

temporal scales of these flows should be of similar
order or longer than those of invertebrate larval du-
ration. However, whereas these assumptions may
hold in some estuarine systems and oceanic islands
and reefs, they are problematic for most continen-
tal open-coast nearshore systems, because mean
cross-shore currents are small or negligible. While
vertical migration at fixed tidal and ontogenetic
schedules may work in many systems, settlement
data (a proxy for larval transport) indicate that at
least some larval transport is associated with rapid
and unpredictable (in the short term) changes in hy-
drodynamic conditions, sometimes associated with
small-scale frontal features (Garland et al., 2002).
How can larvae exploit depth-dependent flows
in unpredictable features? Pineda (1999) found that
the vertical distribution of barnacle larvae switched
from a deeper distribution at sites away from an
internal bore warm front to a near-surface distribu-
tion at the front. It was speculated that cyprid larvae
responded to the convergent frontal downwelling
currents by swimming up against the downwelling
circulation, and in this way accumulated and translo-
cated with the front. A later study found that cyprid
larvae indeed respond to downwelling currents by
swimming up, indicating that this mechanism is
plausible (DiBacco et al., 2011). Here we use “adap-
tive behavior” in the context of larval transport to
denote near-instantaneous individual responses to
a change in a physical field (Figure 11.1). Adaptive
behavior could lead to shifts in vertical distribution
or to swimming that reduces vertical displacement.
The appearance of adaptive behaviors might coin-
cide with developmental changes in morphology
and could be used to exploit event-type processes
with short temporal scales, such as increased wave
height, sea-level change (pressure), intensification
of winds, upwelling relaxation, and internal tidal
bores. Larvae possessing adaptive behaviors would
have a higher success rate in reaching the juvenile
habitat, settling, and growing to reproduce.
11.3.2 Physical Transport Mechanisms
and Hydrodynamic Variability
Ecologists aspire to identify the advective physi-
cal mechanisms associated with larval transport to
understand the spatial and temporal modulation of
larval supply, settlement, recruitment, and popu-
lation dynamics by physical processes, as well as
the larval traits that mediate transport. Advection
and diffusion determine planktonic larvae separa-
tion with time (Scheltema, 1986), and a plethora of
small- to large-scale physical processes cause the
water to move, from eddy diffusion, to the slow
Stokes drift associated with surface gravity waves
(but see Lentz and Fewings, 2012), to fast western
boundary currents such as the Gulf Stream.

(A) (B)

Figure 11.1  Hypothesis of adaptive behavior. A rapid change in an environmental property, for example, currents (arrows on top of the panels)
leads to a sudden change in larval behavior in barnacle cyprids, such as swimming up in response to downwelling currents (DiBacco et al., 2011),
influencing vertical distribution and leading to larval transport in some taxonomic groups. Panels A and B are Time 1 and Time 2. Short-term changes
in vertical distribution in barnacle cyprids have been observed in response to internal tidal bore warm fronts and alongshore current reversals (Reyns et
al., unpub.). In this example, other type of larvae, such as bryozoan cyphonautes, gastropod veligers, and anemone planulae, do not change behavior;
shade gradients represent stratification (see Plate 8).

The fundamental attribute of advection in
larval transport is variability. If currents would
flow in only one direction, with similar magni-
tude at all depths and no temporal variability,
weak larval swimmers would have no chance to
return to their natal habitats. Temporal current
variability is frequently associated with tides, but
wind-forced, buoyancy-driven flows and other
processes (e.g., waves, internal waves) also cause
current fluctuations. Thus, spatial and temporal
hydrodynamic variability are fundamental in lar-
val transport.
Is larval transport intrinsically linked to certain
processes? Are mass transport, diffusion, and be-
havior necessary for larval transport to occur? In
principle, it appears that larval transport has no
strict dependencies, as it can occur with or without
mass transport and diffusion in the case of strong
larval swimmers, and in the absence of any particu-
lar behavior, when transport would be entirely de-
pendent on advection and diffusion.
It is impossible for maximum larval distance
to exceed the limits set by two factors: the upper
limits of larval duration and the maximum sepa-
ration between initial and ending positions set by
advection and diffusion (i.e., maximum horizon-
tal particle dispersion), plus distance added by
horizontal larval swimming. Further, larvae must
be present to be transported; yet this obvious con-
sideration becomes non-trivial when testing links
between physical transport and larval supply or
settlement using time-series analyses (discussed
in Pineda, 2000). Diffusion is often used as an im-
portant source of larval loss (Hill, 1991), and it
also plays a role in the distribution and transport
of invertebrate larvae (Okubo, 1994); however, its
role remains to be investigated in field conditions.
In one of the few observational studies addressing
horizontal larval distributions, Natunewicz and
Epifanio (2001) tracked larval crab patches and hy-
pothesized that patch coherence was maintained
behaviorally. Larvae may aggregate in conver-
gences, and these features limit larval dispersion.
Given that swarming behavior may be associated
with proficiency in swimming capabilities and that
larger larvae tend to be better swimmers, diffusion
may be more important in larval transport of the
smallest and weakest swimmers.
La r va l Tr a n s p o rt i n t h e C o a s ta l Z o ne    149
11.4  Advective Physical Mechanisms
in the Coastal Ocean Associated With
Larval Transport
Winds, tides, and buoyancy-driven flows (flows as-
sociated with horizontal differences in water den-
sity) are the three main forcing mechanisms in the
coastal ocean, and, unsurprisingly, most physical
mechanisms associated with larval transport in-
voke flows forced by these processes, a perception
that has changed little in the past 25 years (Epifanio
and Garvine, 2001). What has changed is a broader
use of physical measurements to study larval trans-
port, more attention to nearshore and small-scale
flows in observational studies, and increased so-
phistication in considering potential mechanisms
at work. Waves, the main forcing mechanism in
the surf zone (Lentz and Fewings 2012), have only
been recently considered by larval ecologists as
a component of larval transport (Fujimura et al.,
2014; Röhrs et al., 2014).
The importance of the three main forcing mech-
anisms varies depending on the domain (i.e.,
cross- vs. alongshore). Some of the coastal physical
processes that tend to be energetic in the alongshore
axis, such as tidal currents, tend to have smaller var-
iability in the cross-shore (Figure 11.2). Conversely,
the internal tide can have larger cross-shore vari-
ability (Winant and Bratkovich, 1981). This asym-
metry in cross- and alongshore processes, however,
does not always hold in other systems where larval
transport is studied, including coral reefs, where
tidal currents play an important role (Wolanski et
al., 1989), and in coastal lagoon mouths and chan-
nels, where tidal currents are also energetic (Epifanio,
1988; DiBacco and Chadwick, 2001). Much interest
in larval ecology in coastal areas is focused on larval
transport perpendicular to the shoreline. Nearshore
water cooling during winter (Checkley et al., 1988),
thermocline shallowing associated with the internal
tide (Pineda, 1994), and strong offshore winds can
lead to the appearance of a cool, dense body of wa-
ter in the nearshore with corresponding cross-shore
density gradients. If the density field relaxes on
short timescales, these density gradients can create
gravity currents, where the dense nearshore water
flows and sinks offshore, while a buoyant, lighter
parcel of water advances shoreward.
150   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v ert e b r at e L a r va e

(A)
70000 cross-shore currents
60000

# of 1-minute samples
near-surface
50000
mid-depth
40000
near-bottom
30000
20000
10000

0
25

25

20

15

10

05

00

05

10

15

20

25

25
.

.
0.

0.

0.

0.

0.

0.

0.
–0

–0

–0

–0

–0

–0

>
<

(B)
70000 alongshore currents
60000
# of 1-minute samples

50000
40000
30000
20000
10000

0
5

25
5

5
00

05

20

25
10

15
.2

.2

.2

.1

.1

.0

0.
0.
–0

0.

0.

0.
–0

–0

–0

–0

–0

0.

0.

>
<

m/s

Figure 11.2 Frequency distribution of 1-minute averaged cross-shore (A) and alongshore (B) currents (m/s), collected in an open-coast nearshore
environment using an Acoustic Doppler Current Profiler deployed at an 8m, ~ 900m from Bird Rock, La Jolla, California, USA, during April–July
2014. Positive are southward and onshore currents. Near-surface, mid-depth, and near-bottom currents were collected at 6.7, 4.7, and 1.5 m above
bottom, respectfully. Cross-shore currents are less energetic and have a narrower distribution in speed than alongshore currents. Additionally, near-
surface currents have the largest range, and speeds decrease with depth (mid-depth and near-bottom have progressively smaller ranges in currents),
particularly in the alongshore domain (Reyns et al., unpub.).

During the past 25 years, studies have high-
lighted evidence of larval transport or settlement
associated with upwelling relaxation/downwelling
(Farrell et al., 1991; Wing et al., 1995; Shanks et al.,
2000; Garland et al., 2002), internal tides (Pineda,
1994; 1999), storm systems (Eggleston et al., 1998;
2010), mesoscale eddies (Sponaugle et al., 2005),
and surface gravity waves (Pfaff et al., 2015). Many
of these studies underscore the switch in focus from
consideration of open-ocean processes to nearshore
processes (discussed in Pineda, 2000), such as sur-
face gravity waves (Pfaff et al., 2015) and shallow-
ing of the internal tide (MacTavish et al., 2016).
Moving forward, there need to be more studies ex-
amining systems where the previously mentioned
physical processes are not dominant (Hrycik et al.,
2013; Daigle et al., 2014).
Another recent development is the realization
that large-scale and remote physical processes may
modulate nearshore flows and larval transport (dis-
cussed in Pineda et al., 2007). For example, both
coastally trapped waves (Brink, 1991) and mesoscale
eddies (Sponaugle et al., 2005) influence nearshore
thermocline depth. The thermocline can shallow in
response to these processes, allowing the internal
tide to penetrate the nearshore and settlement to
increase (Pineda and López, 2002; Sponaugle et al.,
2005). Thus, modulation of small-scale nearshore
flows by large-scale and remote processes provides a
way for linking small to larger scales (Mullin, 1993).

11.5  Other First-order Phenomena
and Processes Relevant to Larval Transport
11.5.1  Swimming Proficiency and Size
Larval swimming proficiency, a potentially key
trait for the behavioral control of larval transport, is
partially dependent on whether larvae use cilia or
muscles for locomotion (Young, 1995), and how this
varies with respect to phylogeny, size (from microns
to a few centimeters), and ontogenetic stage. For ex-
ample, late-stage bivalve larvae tend to be small (less
than a few hundreds of microns), with some excep-
tions (e.g., teleplanic Planktomya henseni maximum
size 1.5 mm; Allen and Scheltema, 1972). Viscous
forces dominate the environment of the smallest
plankton, so while these larvae may attain relatively
large horizontal swimming speeds in body-lengths
per second, these speeds may be insignificant for
larval transport because they are negligible relative
to the ambient flow velocity (see Hodin et al., this
volum­e). Even for the larger larvae (e.g., ~>1 millime-
ters to few centimeters), swimming efficiency ranges
from excellent (e.g., crustacean larvae; Phillip­s and
McWilliam, 1986) to weak and at the mercy of the
flows (e.g., ascidian larvae; Olson, 1985).
11.5.2  Larval Duration
Is there a simple relationship between larval trans-
port distance and larval duration? Most marine in-
vertebrates develop through either a lecithotrophic
(nonfeeding) or planktotrophic (feeding) larval
phase, and it is generally accepted that feeding larvae
spend more time in the plankton than nonfeeding lar-
vae (e.g., Strathmann, 1985). This development time,
or larval duration (we use larval duration instead of
the commonly used pelagic larval duration because
many invertebrate larvae are demersal and neritic,
not pelagic), constrains the upper limit of larval trans-
port, with potentially greater spatial scales of larval
transport for longer-lived species (Scheltema, 1989),
and clearly, species with very short larval life cannot
translocate long distances (Gerrodette, 1981). There-
fore, a simple relationship between larval duration
and dispersal distance is expected (Scheltema 1989),
and plots describing such relationships have been
offered in the literature (Siegel et al., 2003; Shanks,
La r va l Tr a n s p o rt i n t h e C o a s ta l Z o ne    151
2009) and used in modeling studies (discussed in
Shanks, 2009; Leis et al., 2011). Scheltema (1989)
asked whether “these assumptions and intuitions are
really correct” and tested whether geographic range
correlated with larval duration in coastal larvae.
Although he found no correlations, long-lived tel-
eplanic larvae tended to have distributions on both
sides of the Atlantic Ocean, whereas species with 2–4
week larval duration did not. Several biological and
physical factors negate a simple relationship between
larval transport distance and larval duration. These
include larval behavior; taxon-specific swimming
capabilities; physiological condition; impacts of vari-
able environmental conditions, such as temperature;
capacity for delaying settlement; as well as sheared
flows, including reduced velocities at the bottom of
the benthic boundary layer; local physiography; the
coastal boundary (Shanks, 2009); and regional circu-
lation singularities.
Further, recent research indicates that larvae of
some invertebrate and fish taxa may disperse less
from adult habitats and have more constrained dis-
tributions (Jones et al., 1999; Tapia and Pineda, 2007;
Morgan et al., 2009) than anticipated by larval du-
ration (Efford, 1970); these findings could not have
been anticipated by simple relationships between
larval transport scale and larval duration. Thus, the
existence of an upper limit in larval transport dis-
tance determined by maximum larval duration is
arguably a useful construct; however, there is a need
for rigorous observational data on larval transport
and dispersal distance, and larval duration.
11.5.3 Accumulation
Several studies have suggested that larval transport
is associated with front propagation, including in-
ternal bore warm fronts and upwelling-relaxation
fronts (Pineda, 1999; Shanks et al., 2000). Accumu-
lation processes are likely particularly important in
buoyancy-driven flows. For example, accumulation
and transport associated with the propagation of
gravity currents have been considered in the case of
larval transport by internal tidal bore warm fronts. In
these gravity flows, surface currents behind the lead-
ing edge of the gravity current, the front, are faster
than the propagating speed of the front (Simpso­n
and Britter, 1979; Helfrich and Pineda, 2003).
152   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v ert e b r at e L a r va e

As fronts propagate, buoyant particles and plankton
behind the front reach the downwelling convergent
frontal area, and those that swim up in response to
the frontal downwelling convergent currents will ac-
cumulate near the surface and transport in the direc-
tion of front propagation. Those lacking the swim-up
behavioral response will also be transported, but to a
lesser extent. Larval transport with accumulation in
fronts can be more efficient than pure advection in
cases where the buoyant body advances all the way
to near the juvenile benthic habitat. After the front
reaches the juvenile habitat, mean distance between
juvenile habitat and larvae that accumulated in the
front would be shorter than for those larvae that are
advected but did not accumulate (Figure 11.3). Im-
portantly, a variety of phenomena can result in grav-
ity current propagation, including nearshore cooling
by cold fronts (Checkley et al., 1988), internal tidal
bores (Pineda, 1994), and relaxation from cross-shore
winds (Shanks et al., 2000; Tilburg, 2003). In all cases,
as well as in transport in trapped cores of depression
and elevation in nonlinear internal waves, where
current speed can be larger than the propagation
speed (White and Helfrich, 2008), relevant larval be-
havior to transport efficiently is to swim against the
dominant vertical currents (Figure 11.4).

(A) (B)

Figure 11.3  Representation of larval transport associated with advection, with accumulation (panel A) and without accumulation (B). The shallow
adult habitat is on the left side of each diagram. Accumulation can occur when the current speed u associated with a parcel of water behind
(seaward side) a front (i.e., light arrows on the top of the panels) is faster than the propagation speed C of the front (thick black arrows), and larvae
swim against the dominant vertical current (curved arrows). Larvae that do not swim against the dominant vertical currents at the front will also be
advected and swept toward the front, but they will not accumulate at the front (panel B). Larvae that accumulate will reach the vicinity of the adult
habitat, while larvae that do not accumulate will be, on average, more distant (Pineda, 1999) (see Plate 9).

(A) (B)
ρ1

ρ2

ρ1

ρ2

Figure 11.4  Hypothesis that swimming behavior against vertical currents can lead to more efficient larval transport in a variety of physical
processes. Panel A: Onshore larval transport associated with gravity currents, including relaxation after nearshore cooling associated with cold fronts
(Checkley et al., 1988), in internal bore warm fronts (Pineda, 1999), upwelling relaxation/downwelling (Shanks et al., 2000), and after cross-shore
winds (Tilburg, 2003). Panel B: Larval transport associated with bottom-propagating gravity currents (Simpson and Britter 1979), such as internal
tidal bore cold surges. Sigma is density, and arrows denote the propagation direction of the gravity currents. Hypothetical onshore larval transport
associated with recirculating trapped cores in nonlinear internal waves of depression and elevation (not depicted; White and Helfrich, 2008) would
also be aided by the behavior of swimming against the vertical currents (see Plate 10).

11.5.4 Patchiness and Episodic Transport
Patchiness is an aspect of spatial variability and is
pervasive in all animal distributions, and in under-
standing larval transport, the combination of patchy
larval distributions and episodic physical transport
have conspired for decades against fundamental
understanding. Put another way: while analyses of
time series of dependent biological variables such
as larval settlement and independent physical vari-
ables associated with advection can be used to hy-
pothesize functional relationships between larval
transport and physical forcing, these correlations
may be weak or fail due to larval patchiness (dis-
cussed in Pineda, 2000). One way to circumvent
this problem is to use statistical analyses that filter
the independent variable, keeping instances where
there is a response in the dependent variable such
as settlement (Prager and Hoenig, 1989). Another,
more powerful approach, is using a hypothetic-
deductive method and sampling in response to the
hypothesized larval transport events.
11.5.5  Spatial Variability in Larval Abundance
As noted previously, elucidating the vertical dis-
tribution of larvae is essential to resolve transport
processes. Larval transport hypotheses and re-
search that disregards the layered structure of the
flows and the dependence of larval transport with
depth may eventually prove wrong. Spatial scales
of variability are expected to be larger on horizontal
scales where larval patches are separated by greater
distances (kilometers) than on vertical scales. This
is due to the constraining effects of the sea surface
and sea floor in addition to physical (e.g., currents,
temperature) and chemical (e.g., oxygen) gradients
that are much sharper in the vertical than horizon-
tal dimensions. The vertical and horizontal scales
of variability might be more comparable in benthic
deep-sea environments because vertical gradients
near the bottom may be less pronounced than in
shallow coastal environments that are influenced
by sunlight and surface heating. However, some
benthic deep-sea gastropod larvae can be found in
surface waters (Killingley and Rex, 1985).
Horizontal variability in larval abundance may
relate to proximity of adult habitat, to habitat phys-
iography (e.g., “coastal retention zones”), and to
La r va l Tr a n s p o rt i n t h e C o a s ta l Z o ne    153
water column stratification. If increased stratifica-
tion leads to a more sheared environment, sites with
stronger stratification might promote the retention
of larvae that behaviorally exploit these flows, as
speculated by Pineda and López (2002). Spatial
variability in benthic population abundance, an-
other expression of patchiness, is a hallmark of
many marine populations, and ecologists have
asked for decades whether this variability is related
to variability in larval transport, settlement, and
recruitment (Effor­d, 1970; Eggleston et al., 1998).
Horizontal variability in larval transport has also
been related to coastal physiography (Morgan et
al., 2011; von der Meden et al., 2012) and potentially
larval swimming proficiency (Daigle et al., 2014).
In coastal environments, cross-shore crustacean
larval distributions are sometimes structured by
stage (Bousfield, 1955) and reflect distance from na-
tal habitat, with early stages adjacent to nearshore
habitats. These distributions might also suggest
changes in behavior with stage. For example, Tapia
and Pineda (2007) found early nauplii stage inter-
tidal barnacle larvae near the shore with mid-stages
further offshore; yet larvae of the terminal cyprid
stage were found nearshore, suggesting an ontoge-
netic change in behavior.
11.6  Challenges and Recent Approaches
to Understanding and Measuring Larval
Transport
11.6.1 Challenges
The importance of larval movement and dispersal
for the ecology of bottom organisms has been ac-
knowledged for over one and a half centuries (Lyel­l,
1832; Young, 1990). For example, discussing the
mechanisms accounting for fish size-class strength,
Hjort (1926) offered two explanations, one based on
food quality and the other on larval transport.
Despite decades of interest, larval transport
mechanisms for the majority of systems remain
unresolved. Spatial and temporal scales of vari-
ability in chemical and physical variables can be
addressed with time-series analyses using off-the-
shelf sensors (e.g., temperature loggers and acoustic
Doppler current meters) and statistical techniques,
such as spectral analyses (discussed in Haury et
al., 1978). So far, these approaches are limited for
154   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v ert e b r at e L a r va e
resolving larval transport and distribution because
high-frequency, species-specific sampling and pro-
cessing capability of up to thousands of samples is
practically impossible. Moreover, variability in the
larval pool may result in failure in identifying the
forcing physical processes, as discussed previously,
and zooplankton distributions are non-stationary
(Haury et al., 1978), violating the assumptions of
spectral analysis.
Ideally, development of high frequency and spa-
tially dense sampling, and development of tech-
niques for analyzing large numbers of samples
would resolve dynamic larval distributions, and in
this way advance our understanding of the larval
transport process. Pragmatically, costs associated
with the development of such sampling and anal-
ysis techniques at current cost and funding levels
may hinder their further development.
The main challenges for resolving larval trans-
port include the following: (a) patchy spatial dis-
tributions, (b) episodic larval transport events, (c)
enormous ranges of spatial and temporal scales of
the potential processes, (d) knowledge of larval be-
havior during transport, (e) imperfect knowledge of
physical transport processes, (f) the arduous nature
of observational approaches to measuring and in-
ferring larval transport, and (g) the variety of spe-
cies and settings to be understood. Next we discuss
these challenges, and contemporary and promising
approaches that might help resolve the larval trans-
port processes. We also caution about potential pit-
falls of some methodologies.
To address patchiness, episodic transport, and the
wide range of scales, (a)–(c), we suggest that, when
possible, a hypothesis-based approach and “adap-
tive sampling” be used to test specific processes.
An explicit hypothesis-based approach identifies
advective mechanisms underlying larval transport
events, with expectation on how physical fields
such as temperature and currents behave with or
change in response to the advective event. It tests
predictions of temporally and spatially constrained
larval abundance, and horizontal and vertical dis-
tributions. In adaptive sampling, changes in specific
environmental conditions initiate larval sampling.
Obviously, this approach requires contrasting
larval distributions during hypothesized trans-
port against larval distributions in non-transport
circumstances. Ideally, adaptive sampling should
be accompanied by other types of measurements,
including settlement time series. This sampling ap-
proach also requires real-time communication to
remote environmental sensors (e.g., temperature
or current velocity data). Technology for adap-
tive sampling, including satellite, radio, and cell
network communications, is mature and, because
some of this technology is used commercially, it is
relatively inexpensive. Adaptive sampling has been
used successfully in addressing various event-type
phenomena in biological oceanography, including
the study of larval transport by internal tidal bore
warm fronts (e.g., Pineda, 1999). Adaptive sampling
is challenging to research teams because it requires
the formulation of a specific hypothesis and a crew
to be in standby for extended periods.
Larval behavior during transport, (d), may be
more economical to address after other larval trans-
port processes are identified, because the larval
behavioral repertoire is very large (Metaxas, 2001;
Epifanio and Cohen, 2016). For example, hypoth-
eses of larval behavior during transporting events
can be posed after identifying the physical pro-
cesses and the vertical distributions associated with
the event. Improvement in understanding near-
shore physical advective processes associated with
larval transport, (e), will require progress in under-
standing fundamental nearshore hydrodynamics
(Lentz and Fewings, 2012), as well as knowledge
of the nearshore and coastal circulation associated
with unpredictable events, such as storms.
Observationally, larval transport can be ad-
dressed with field measurements of physical fields,
larval distribution, and settlement (f). In situ cir-
culation and larval distribution studies can unam-
biguously identify larval transport processes, but
sustained and targeted studies of larval distribu-
tions are very challenging. Thus, researchers often
resort to study settlement, where long-term sus-
tained measurements are possible. These observa-
tions can reveal characteristic spatial and temporal
patterns and suggest larval transport mechanisms
(Eggleston et al., 1998; Reyns and Sponaugle, 1999;
Pfaff et al., 2015). Settlement, however, is necessar-
ily inferential, and results require cautious interpre-
tation (Pineda, 2000). Simultaneous measurement
of larval distribution and settlement is arguably the

most powerful approach to address larval trans-
port processes. Finally, for addressing (g), the large
diversity of systems, the hope is that resolution
of a number of larval transport cases will lead to
some generalities, for example, swimming behavior
against the dominant vertical currents (Figure 11.4).
11.6.2 Recent Approaches
Recent technology and methodological approaches,
including computer-technology miniaturization,
improved computing capability, development of
imaging techniques, and remote sensor develop-
ment (e.g., satellite SAR and spectroradiometer
sensors), have revolutionized areas in ecology and
oceanography. Some of the recent technology and
approaches, including inexpensive sensors with im-
proved endurance, and numerical modeling, have
been used to address larval transport. These devel-
opments represent opportunities for larval ecolo-
gists, yet there are also nagging issues to consider.
11.6.2.1  Autonomous and Remote Sampling
Autonomous and remote observations of physi-
cal and biological properties are revolutionizing
fields in oceanography (Clark and Isern, 2003).
One promising area for larval transport is the de-
velopment of remote, autonomous sampling of
larval distributions by building and mounting
larval samplers to autonomous underwater vehi-
cles (AUVs). Researchers have demonstrated AUV
sampling of larvae in coastal (e.g., Ryan et al., 2014;
Govindaraja­n et al., 2015) and benthic deep-sea en-
vironments (Billing­s et al., 2016). The advantages
of these devices over plankton pumps and nets
include (a) the ability to sample very close to the
bottom; (b) increased spatial resolution (i.e., few
meters), particularly for deep sampling, where net
resolution is coarse and maneuvering a pump in-
take is very challenging; and (c) the possibility to
sample larvae adaptively, in response to changing
physical and biological fields (e.g., temperature,
fluorescence). These devices, however, take only a
few samples (two to less than a handful; Ryan et al.,
2014; Billing­s et al. 2016), or sample a small volume
of water (~2 to 55 L; Ryan et al., 2014; Govindarajan
et al., 2015). Further, they are expensive to build and
operate. Therefore, while the devices have already
La r va l Tr a n s p o rt i n t h e C o a s ta l Z o ne    155
helped resolve larval abundance and distribution
fields, improvements in sample number, sampling
volume, and cost reduction remain necessary. Other
recent advances include the development of drift-
ers that can be tracked and programmed for differ-
ent vertical positioning (T. Wolcott and S. Morgan,
pers. comm.; Jaffe et al., 2017) The next step in AUV
development, particularly to address issues of lar-
val patchiness, might include swarms of AUVs
that communicate and adaptively sample larval
distributions.
11.6.2.2  Imaging in Distributional Studies
and Species Identification
Imaging of plankton using towed vehicles can re-
veal fine- to large-scale (centimeters to thousands of
kilometers) field distributions, and a variety of ap-
proaches have been developed in the last 25 years
(Wiebe and Benfield, 2003). Use of this approach has
revealed small (few centimeters) to basin-scale vari-
ability in plankton distribution (Davis et al., 1992;
Davis and McGillicuddy, 2006), including distribu-
tions of diverse trophic groups (Greer et al., 2013),
and spacing among and orientation of individual
zooplankton (Cowen and Guigand, 2008). A caveat
of some of these devices, small sample size, has been
addressed with new systems that can sample larger
volumes (Cowen and Guigand, 2008), although at
the expense of sampling rate, towing speed, and
plankton size resolution. These approaches have
enormous potential for resolving plankton and lar-
val distribution, but automatized identification of
rare or small and “difficult” invertebrate larvae re-
main a problem (Wiebe and Benfield, 2003), and the
cost of the technology makes it prohibitive for large
segments of the research community.
Automated larval species recognition in field
samples, one of the nagging issues in larval studies,
was addressed with the development of an imag-
ing approach based on automated recognition of
species-specific coloration of larval bivalve shells
(Tiwari and Gallager, 2003; Thompson et al., 2012a).
This approach has been used successfully in field
studies of larval bivalve dynamics (Thompson et al.
2012b; Goodwin et al. 2014). This system is, so far,
limited to bivalve shells and requires a reference
­library with the exact set of known species to be
identified (Thompson et al., 2012a).
156   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v ert e b r at e L a r va e
11.6.2.3  In Situ Species Identification
and Robotic Techniques
The -omics approaches are revolutionizing the
biological sciences, including select areas in ma-
rine ecology, which are addressed in Williams
and Carrie­r (this volume). Here, we point to two
developments. First, use of barcoding to identify
minute larvae to species (e.g., Ryan et al., 2014;
Govindarajan et al., 2015; Billings et al., 2016). This
development is p ­ articularly important for identify-
ing problematic larvae (e.g., small bivalve larvae).
A related approach is in situ species identification
using robotic systems and sandwich hybridization
methods ­(Goffredi et al., 2006). With improvement
in sample processing time, necessary for resolving
high-frequency transporting events, these devices
will be able to remotely detect species of interest
and perhaps initiate adaptive sampling. The wide-
spread use of these devices might only be feasible
once costs decrease.
11.6.2.4  Numerical-Modeling and Observational
Approaches to Larval Transport
Taking observations to measure larval transport is
arduous and expensive, equipment can be lost and
vandalized, and researchers get seasick. Moreover,
biological observations in systems where processes
at multiple spatial and temporal scales operate tend
to be sparse. Despite these impediments, observa-
tional approaches have revealed almost all phe-
nomena relevant to larval transport and dispersal,
including the following: (a) the realization that larval
vertical migration is the main mechanism determin-
ing horizontal transport (Nelson, 1912); (b) aware-
ness of “wastage” during the larval phase, including
a geometric decrease of larval concentration with
distance from natal habitat related to mortality and
diffusion (e.g., Hjort, 1926; Scheltema, 1986), and the
role of large-scale features in larval wastage (Porri et
al., 2014); (c) the existence of long-lived larvae and
potentially large dispersal distances (Scheltema,
1968); (d) that upwelling (Garland et al., 2002), up-
welling relaxation/downwelling (Farrel­l et al., 1991;
Wing et al., 1995), internal tidal bore warm fronts
(Pineda, 1999), and eddies (Sponaugle et al., 2005)
are associated with onshore larval transport to adult
habitats; (e) unambiguous evidence that larvae can
return to natal habitats (Jones et al. 1999); and (f) the
finding of constrained larval distributions in near-
shore temperate settings (Barnett and Jahn, 1987;
Tapi­a and Pineda, 2007; Morgan et al., 2009).
Simulation numerical models of larval transport
and dispersal, on the other hand, are relatively inex-
pensive to run, can readily fill any spatiotemporal
domain and simulate larval transport and dispersal
processes at regional scales, link a variety of dis-
parate processes (Werner et al., 2007), and rapidly
generate results for unsuspecting researchers. Here,
we restrict our discussion to numerical simulation
models at geophysical scales in the sense of Oreskes
et al. (1994), where these models are used “to eval-
uate large-scale or complex physical processes”
(p. 641). We do not deal with models based on inter-
polated observations (Paris and Cowen, 2004), with
“heuristic” models (Cowen et al., 2000; Botsford et
al., 2001; Tilburg, 2003; Pringle et al., 2011), where
fundamental relationships are used to illustrate a
given dynamics, or with models of small-scale do-
mains (Scotti and Pineda, 2007), where the physical
processes are relatively well understood and the pa-
rameter space is constrained.
Compared to observational approaches, how-
ever, fundamental new insights by numerical mod-
els are very rare, even considering their relatively
recent emergence in oceanography (late 1960s;
Kantha and Clayson, 2000). For example, combined
numerical modeling and observational studies
have suggested that dispersal among coral reefs
could be both limited and feasible (Wolanski et
al., 1989; cited in Black et al., 1991). Other types of
modeling (e.g., “heuristic” models, not simulation
models), have also contributed. A heuristic model
highlighted that diffusive and mortality processes
caused larval abundance to decrease geometrically
with distance from larval release, contributing to
the realization that marine populations were more
closed that originally thought (Cowen et al., 2000).
Researchers are gravitating toward regional simu-
lation numerical models, such as the regional ocean
modeling system (ROMS) and the finite volume
community ocean model (FVCOM) to link hydro-
dynamic and larval behavior processes with lar-
val transport and dispersal patterns. Simulation of
physical fields (e.g., density, currents) by numerical
modeling is a key component in many disciplines,
and these models are well accepted in situations

where the systems to model are relatively simple
and the processes involved well understood and
parameterized (e.g., White and Helfrich, 2008). In
systems where models have been thoroughly tested
(Gilg and Hilbish, 2003), or the behavioral and dis-
tributional processes accounting for the transport of
larvae are understood, and the hydrodynamic pro-
cesses relatively well resolved (North et al., 2008),
numerical modeling can offer valuable insights to
larval transport and dispersal issues. Research com-
paring model predictions and field observations is
also valuable, as modeling results can be tested
against some observable property (Wolansk­i et al.,
1989; Sponaugle et al., 2012).
Unfortunately, many systems studied by marine
ecologists are complex (e.g., nearshore open-coast),
biological and physical processes are understudied
or unknown, and models go untested. Ecologists
should hold numerical-modeling studies to the
same standards as experimental and observational
studies, but this practice is very rare. Simulation
numerical models, like other hypotheses, should be
tested observationally, against “Lagrangian” obser-
vations (e.g., Hrycik et al., 2013), or at least against
other observed properties (Trindade et al., 2016).
Further, in some larval transport and connectivity
studies, numerical output is equated to an accurate
depiction of the physical fields, with little objec-
tive evaluation of the assumptions or validity of
this premise. In other cases, modeled and observed
“Eulerian” fields are compared, and if the compari-
son is favorable, the model is said to be “validated,”
justifying using the numerical output as observa-
tions. However, simulation numerical models can-
not be validated (Oreskes et al., 1994); they remain
hypotheses. As such, it is untenable to use them to
test other observational data. Another complication
is the many and poorly characterized biological
variables integrated in these models (Metaxas and
Saunders, 2009). Finally, because model parameters
can be tweaked to represent a range of results (e.g.,
limited or wide dispersal distance), research report-
ing modeling results should present a clear descrip-
tion of the iterative parameter choice or a range of
results (e.g., Ross et al., 2016).
No simulation numerical model features all the
potentially relevant mechanisms that move the
water and are associated with larval transport in
La r va l Tr a n s p o rt i n t h e C o a s ta l Z o ne    157
coastal and nearshore settings—another issue that
is not often addressed in the larval transport and
connectivity literature. For example, these models
often do not capture the small-scale processes that
may account for nearshore larval transport (e.g.,
transport by surface gravity waves and propagating
convergences such as upwelling fronts and internal
tidal bore warm  fronts). This is highlighted by re-
cent advances in our understanding of the dominant
processes driving the cross-shelf circulation over the
nearshore region (for a review, Lentz and Fewings,
2012). Historically, studies of wind-driven shelf cir-
culation have focused on along-shelf wind forcing
and coastal upwelling or downwelling. The domi-
nant role of surface waves and cross-shelf winds in
driving cross-shelf circulation over the nearshore has
been highlighted in a sequence of process-oriented
studies (Tilburg, 2003; Lentz et al., 2008), and, no-
tably, surface wave forcing has only recently been
incorporated into regional hydrostatic circulation
models (e.g., Newberger and Allen, 2007). Thus, the
commonly invoked premise that numerical models
can be used to test the role of larval behavior in lar-
val transport and dispersal by comparing numerical
model output with observations of dispersal ob-
tained by other means does not hold, because the nu-
merical model does not include all relevant processes
that account for passive dispersal, and therefore the
“real” dispersal of passive particles is unknown.
Regional hydrostatic models are not a substitute
for process-oriented understanding of circulation.
Particularly for research addressing complex set-
tings such as the nearshore and coastal ocean, with
a large range of scales and where multiple mecha-
nisms might play a role, it should be obvious that
some level of process-understanding of larval trans-
port should be achieved first, followed by simula-
tions by numerical modeling. Knowledge of the
process reduces the dimensional complexity of the
modeling problem, limiting the number of possible
scenarios, and focusing the modeling research. Blind
modeling is risky and inefficient, yet, to date, few
modeling studies addressing larval transport and
dispersal operate with knowledge of the specific
larval transport processes involved, and even fewer
compare model predictions with observations.
In summary, process-oriented understanding
of larval transport and model testing is key for
158   E v o l u t i o n a ry E c o l o g y o f M a r i ne In v ert e b r at e L a r va e
achieving an understanding of larval dispersal and
connectivity, and basic process understanding of bi-
ological and physical processes is necessary before
credible simulation numerical modeling.
11.7  Conclusion and Next Steps
Understanding larval transport processes in the
coastal ocean has increased considerably during
the past 25 years, due to improvements and cost
reduction in technology, approaches often based
in hypothesis testing, a more sophisticated under-
standing of the behavioral and physical processes,
a more critical appraisal of the evidence in obser-
vational studies, and a mechanistic resolution of a
handful of larval transport cases. Advances include
an understanding that larval supply and settlement
observations measure different processes, apprecia-
tion of the importance of accumulation processes
and of how larger-scale processes might modulate
local factors that influence larval transport, a reali-
zation that propagules and larvae have generally
a more constrained distribution than originally
thought, and insight about the distinction of the
nearshore and open-ocean domains. Although pro-
cess-oriented understanding of larval transport has
increased incrementally, more understanding of the
dominant mechanisms of larval transport is needed.
On the other hand, interest and necessity in un-
derstanding larval transport and connectivity at
regional scales, coupled with the difficulty in re-
solving observationally these processes, has led to
the growth of simulation numerical modeling. So
far, this growth has been disproportionate relative
to model testing and fundamental knowledge of the
processes. Thus, if these models are to be useful to
understanding natural systems, simulation numeri-
cal models should be accompanied by testing (e.g.,
Hrycik et al., 2013) and constrained by understand-
ing of specific larval transport processes at work.
Technological development, broader sensor us-
age, and reduction of costs foretell a future where the
coastal ocean will be seeded with a multitude of sen-
sors that aid in resolving larval transport. Nonethe-
less, intensive field and laboratory studies addressing
process-level understanding of larval transport will
still be sorely needed. Simulation numerical models
are growing in sophistication, yet our understanding
of physical processes in complex systems is still in-
complete, and our knowledge of the relevant larval
processes to include in these models is meager.
We conclude by suggesting that the next step
in larval transport understanding should include
wider use of emergent technologies and mecha-
nistic resolution of larval transport in multiple
systems. Numerical models studies will be useful
if they incorporate the relevant larval transport
processes, and are well tested; unfortunately, so far
these investigations are sorely lacking.
11.8 Summary
1. Larval transport is defined and discussed, both
as a concept and as a measurement.
2. The components of larval transport include
larval behavior, primarily those behaviors in-
fluencing larval vertical distribution, physical
advective and diffusive processes, and their vari-
ability.
3. Other first-order phenomena in larval trans-
port include larval accumulation in propagating
fronts, larval swimming proficiency and size,
larval duration, larval patchiness and episodic
larval transport, and spatial variability in larval
transport.
4. There are multiple challenges for understanding
larval transport: (a) patchy spatial distributions,
(b) episodic larval transport, (c) enormous range
of spatial and temporal scales of the potential
processes, (d) knowledge of larval behavior, (e)
imperfect knowledge of physical transport pro-
cesses, (f) the arduous nature of observational
approaches, and (g) the multitude of species and
settings to be understood.
5. Recent approaches addressing larval transport
include autonomous and remote sampling, im-
aging approaches, in situ species identification
and robotic techniques, and numerical models.
6. The advantages of simulation numerical models
are minimal cost and the ability to address re-
gional scales and link disparate processes. Some
drawbacks include lack of testing, use of nu-
merical output as observations, and uncertainty
surrounding whether relevant larval transport
processes are included.
 La r va l Tr a n s p o rt i n t h e C o a s ta l Z o ne    159

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 CH A PT ER 12

Genetic Analysis of Larval Dispersal,

Gene Flow, and Connectivity
Peter B. Marko* and Michael W. Hart*
*These authors contributed equally to this chapter

“We have to remember that what we observe is not nature in itself but nature exposed to our
method of questioning.” – Werner Heisenberg (1958, p. 78)

12.1 Introduction (fewer settling larvae), but we cannot easily describe

the processes that affect the destination of larvae that
One of the major hallmarks of marine species is
disperse from a particular source, or the source of lar-
that many produce large numbers of small pelagic
vae that settle or recruit into a particular destination.
larvae that drift in the ocean for varying periods of
As with the study of all unobservable processes,
time. For these species, establishing the degree to
the methods of inquiry will determine, to some
which different populations are connected by larval
extent, the apparent properties of the process. For
dispersal is a fundamental goal for larval ecologists
example, not long ago, observations of marine lar-
interested in understanding the influence of plank-
vae far offshore (Scheltema, 1971) combined with
tonic processes and larval supply on ecological and
widespread genetic homogeneity at allozyme loci
evolutionary processes within populations. Assess-
(Buroke­r, 1983; Saunders et al., 1986) led many ma-
ing and predicting local population and commu-
rine ecologists to the reasonable conclusion that
nity dynamics, spread of invasive species, patterns
marine larvae regularly traveled vast distances,
of local adaptation, spread of advantageous alleles,
such that many marine populations were likely well
maintenance of local biodiversity, sustainability of
mixed on spatial scales of thousands to tens of thou-
fisheries, and effective marine reserve design all re-
sands of kilometers (Palumbi, 1992). When Palumbi
quire some knowledge of rates and patterns of lar-
(1995) reviewed the evidence for associations be-
val exchange among populations.
tween variation in larval dispersal potential (such
However, the tiny size of most marine larvae and
as among species with long or short pelagic larval
the variable length of time they spend in the plank-
duration) and the geographic distribution of genetic
ton present obvious and significant obstacles for
variation, nearly all comparative studies analyzed
identifying the geographic origins and destinations
small numbers of populations and loci (typically al-
of dispersing larvae. The fate of marine larvae in the
lozymes and mtDNA) with a limited number of ana-
plankton may be likened to a black box (Buston and
lytical approaches. In the intervening years, the size
D’Aloia, 2013): for any local population, we can esti-
of datasets and the diversity of methods of analysis
mate its contribution to the pool of individuals in the
have grown dramatically, and significant progress
planktonic darkness (many dispersing larvae) and
has been made toward understanding the scope and
its harvest of individuals that emerge into the light

Marko, P. B. and Hart, M. W., Genetic analysis of larval dispersal, gene flow, and connectivity. In. Evolutionary Ecology of Marine
Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018).
© Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0012

scale of larval dispersal. Many tools have been used,
but much of this progress has come from using ge-
netic methods. In this chapter, we describe some
of the most commonly used analytical genetic ap-
proaches and then discuss how these methods have
improved our understanding of larval dispersal.
12.2  Genetic Approaches to the Study
of Larval Dispersal
Rates and patterns of larval dispersal have been stud-
ied using a wide variety of methods, including direct
observations of larval movement (e.g., Gerrodett­e,
1981; Olson, 1985; Knowlton and Kelle­r, 1986), mark-
and-recapture (Jones et al., 1999; Thorrol­ d et al.,
2001), natural environmental markers (DiBacco and
levin, 2000), oceanographic modeling (Largier, 2003;
Siegel et al., 2003), hybrid zones (Gilg and Hilbis­h,
2003), and the expansion of geographic ranges
caused by anthropogenic introductions (Kinlan et
al., 2005) or by climate change (Sunday et al., 2015).
Techniques that employ artificial tags (i.e., to otoliths
or calcified structures) or that estimate dispersal
from inferred parent-offspring relationships may be
considered direct measurements of net dispersal, but
these methods do not necessarily measure larval dis-
persal alone, but instead some combination of larval
and adult movement. We consider only direct obser-
vations of advection or diffusion of individual larvae
in the plankton as yielding direct measurements of
larval dispersal. Although direct observations give
immediate insight into dispersal, they are limited
to species with large, short-lived larvae that can be
followed on small spatial scales. Indirect methods
based on experimental or natural marking of larvae
are similarly limited to species with larval structures
that can be marked and that are retained in adults,
and they share with direct observations some other
important limitations (especially the inability to in-
fer average effective rates of dispersal integrated
over longer periods of time into the ecological or ge-
ological past). Consequently, indirect methods that
use genetic data have become the most widely used
approach for inferring patterns of larval dispersal.
Although many methods have been developed to
infer patterns of larval dispersal from genetic data,
we think a more useful and important categoriza-
tion of genetic approaches is based on the theoretical
G e n e t i c A n a lys i s    165
framework used to infer patterns of larval dispersal.
The oldest and most familiar framework is based on
data in the form of allele frequencies and gene gene-
alogies in explicit population genetic models. These
population models include one or more parameters
that represent migration (m), the proportion of indi-
viduals or gene copies (i.e., alleles or haplotypes)
in a population that are new immigrants each gen-
eration (and that successfully reproduce), and use
a diverse range of assumptions and calculations,
and either optimization (e.g., maximum-likelihood)
methods or simulations to find the best estimate of
migration and other model parameter values fitted
to the genetic data. When combined with an esti-
mate of the effective population size (Ne or simply
N), the product of the two parameters together can
be used to characterize the population migration
rate (Nm), usually interpreted as the number of im-
migrant individuals (i.e., organisms) per generation
or the number of immigrant gene copies (2Nm, for
diploid loci) per generation (Wright, 1969). Because
these population-based estimates represent the
number of gene copies moving between popula-
tions, Nm is often called gene flow (Slatkin 1987;
1993; but see Mallet, 2001).
The second, newer framework for these ap-
proaches is based on data in the form of multilocus
genotypes for individual organisms without speci-
fying a demographic model with explicit param-
eters that represent migration or gene flow. Instead,
these individual-based approaches use multivariate
clustering, similarity measures, or inferred parent-
age to assign sampled individuals to groups (i.e.,
nominal populations consisting of similar geno-
types, or families consisting of parents, offspring,
and siblings). Simple Mendelian inheritance rules
or population parameters are used to identify the
most likely number of groups or clusters that mini-
mizes within-group differences (Pritchard et al.,
2000; Wilson and Rannala, 2003), or to assign indi-
viduals to likely families including parent-offspring
pairs or groups of siblings (Marshall et al., 1998;
Wang, 2004). These individual-based approaches
infer larval dispersal from counts of migrant individu-
als, including individuals that are strongly clustered
with individuals from a different sample, and indi-
viduals that have a high likelihood of being closely
related to parents or siblings in a different sample.
166   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r va e
One of the most important distinctions between
population-based methods and individual-based
methods is the timescale over which they provide
information about migration. All population-based
approaches assume that the spatial distribution of
alleles, the frequency of alleles in populations, and
the structure of gene genealogies evolve slowly, and
that this long timescale has an important effect on
the observed patterns of genetic similarity or dif-
ference among populations. Therefore, population-
based approaches estimate migration and gene flow
as the time-averaged cumulative effects of varia-
tion in larval dispersal, larval retention, population
growth, and other demographic or ecological pro-
cesses integrated over relatively long evolutionary
timescales (on the order of hundreds to thousands
of generations). Individual-based approaches also
make an important but different assumption about
the temporal scale of genetic variation that can be
used to detect migration. Strong allele- or genotype-
sharing between members of the same genetic clus-
ter or members of the same family is expected to
be rapidly broken down by random mating with
local mates (from other clusters or families) after
migration. Consequently, recombination among
immigrant and local alleles will degrade the sig-
nal of group membership or family identity among
the descendants of an immigrant after one or a few
generations. Counts of immigrants based on such
approaches can only identify new migrants or their
recent descendants on short ecological timescales,
on the order of one or two generations (Wilson and
Rannala, 2003). A corollary of this limitation is that
individual-based methods may not easily distin-
guish between recent immigrants that reach sexual
maturity and reproduce (and contribute to gene
flow) and those that do not.
With the recent incorporation of individual-based
approaches into the genetic toolkit of larval biolo-
gists, researchers can potentially compare counts
of immigrant individuals to historical patterns of
gene flow inferred from more complex population-
based approaches. In cases where both approaches
give similar estimates of genetic connectivity, lar-
val dispersal may have been consistently high (or
low) over both long and short (recent) timescales,
and those concordant measures of dispersal on
both timescales may give reliable insight into the
strength (or weakness) of population connectivity
(Pinsky et al., 2016). Numerous factors can poten-
tially explain contradictory results from these two
approaches (Palstra et al., 2007), such as natural
year-to-year variability, recent changes in larval
dispersal caused by human impacts (and an oppor-
tunity for conservation ecologists to mitigate that
change), genotyping errors, or violation of assump-
tions of either method. Distinguishing among these
hypotheses may be difficult and will likely require
repeated studies.
As with other approaches to the study of lar-
val dispersal, those that use genetic techniques
have strengths and weaknesses, and the choice of
methods will depend on what a researcher wants
to know about larval dispersal: an understanding
of migration rates over relatively long evolutionary
timescales (population-based methods) or docu-
mentation of extremely recent migration events
(individual-based methods). In the next section, we
briefly discuss the theory and practice of studying
larval dispersal of marine species using genetic data
in either population- or individual-based methods.
We then highlight some specific areas of progress
in applying both individual- and population-based
methods, and consider how those approaches give
concordant or discordant insight into genetic and
demographic connectivity based on larval dispersal
among marine populations.
12.3  How to Estimate Larval Dispersal
from Genetics
12.3.1  Population-Based Methods
The use of genetic methods to estimate larval dis-
persal requires a realistic model of the processes
that cause allele frequency changes and the evolu-
tion of allelic differences between populations. The
primary processes are mutation, genetic drift, and
natural selection. By contrast, gene flow is a ho-
mogenizing evolutionary force that slows, erodes,
or prevents the buildup of genetic difference be-
tween populations. Most of the population genetic
theory that has been developed to understand the
movement of genes and individuals does so by fo-
cusing on the interaction between gene flow (which
introduces alleles to populations) and genetic drift

(which eliminates alleles from populations). Be-
cause genetic drift is a stochastic evolutionary force
caused by random mating in a finite-sized popula-
tion, it will work randomly and independently in
different populations such that, in the absence of
gene flow between populations, genetic drift is ex-
pected to cause allele frequencies to diverge. Given
sufficient time (generations), drift will cause the
fixation of different alleles in different populations,
meaning that different alleles will reach a frequency
of 1.0 in each of the individual populations.
12.3.1.1  Neutral genetic markers
To concentrate exclusively on gene flow and genetic
drift, population geneticists focus on neutral poly-
morphisms: allelic differences that are expected to
have no (or few) direct effects on fitness. To focus on
neutral loci, population geneticists can study genes
or nucleotide sites that appear to have few functional
constraints, such as microsatellite loci, anonymous
DNA, and synonymous third codon polymorphisms
in protein-coding DNA (Karl and Avise, 1992). One
important consideration in identifying candidate
neutral polymorphisms is their possible linkage to
other polymorphisms under selection. More impor-
tantly, however, population geneticists can identify
neutral polymorphisms for analysis using data from
multiple, unlinked genetic loci. Unlike migration
and genetic drift, natural selection is expected to
cause idiosyncratic patterns of differentiation at indi-
vidual loci, such that loci affected by selection can be
identified and potentially excluded as outliers with
respect to a larger sample of loci from across the ge-
nome (Schopf, 1974; Koehn et al., 1976; Johnso­n and
Black, 1984). The important corollary of this idea is
that similar spatial patterns of differentiation across
multiple loci are best explained by the action of gene
flow and genetic drift, forces that are expected to
affect all loci across the genome in the same way.
The statistical power for identifying outlier loci (in-
fluenced by selection) has increased significantly as
data from very large numbers of loci can now be
gathered and analyzed, potentially within the frame-
work of a linkage map (Bradbury et al., 2013).
12.3.1.2  Classical population genetics
With respect to estimates of dispersal and gene
flow, an important point of departure for classical
G e n e t i c A n a lys i s    167
population genetic methods is the Hardy-Weinberg
equilibrium (HWE) principle: in the absence of any
evolutionary forces acting on a genetic locus, allele
frequencies at that locus will remain constant over
time. This deterministic theory also predicts that,
for a locus with two alleles A and a with frequen-
cies p and q, respectively, the proportion of AA ho-
mozygotes, aa homozygotes, and Aa heterozygotes
is expected to be p2, q2, and 2pq, respectively. De-
viations from the expected genotype proportions in
natural populations provide evidence that at least
one force of evolution is influencing allele frequen-
cies. In the absence of selection, Hardy-Weinberg
deviations can be caused by several processes that
lead to non-random mating among individuals,
and can provide insight into the genetic structure
of populations and, potentially, patterns gene flow.
For example, consider a neutral genetic locus in
two isolated populations (Figure 12.1). Because there
is no larval dispersal (and no gene flow) between
eastern and western populations, genetic drift has
caused allele frequencies to diverge, such that they
have become fixed for different alleles. In this situ-
ation, the HWE principle provides a null model of
high gene flow by predicting that, if eastern and
western individuals were freely exchanging migrants
and completely interbreeding with one another, half
of the individuals (i.e., 2pq) should be heterozygotes.
Although an extreme example, any divergence in al-
lele frequencies between populations will result in a
deficit of heterozygotes compared to expected HWE
genotypic proportions under the null model of high
gene flow between populations.
Detection of heterozygote deficiencies forms the
basis for the most common measure of population
genetic divergence, Wright’s (1978) FST = (HT−HS)/
HT, the difference between the expected HWE het-
erozygosity for the “total” population (HT) and the
average expected heterozygosity among “subpopu-
lations” or individual populations (HS) scaled by
HT. Other measures of genetic differentiation have
been developed, but are like FST in that they describe
how genetic variation is partitioned among popu-
lations or samples from different geographic loca-
tions (Excoffier et al., 1992). Genetic differentiation
measured as FST and its analogs can also be used to
infer the rate of gene flow among populations; in
the simplest case, FST = 1/(4Nm + 1) under a set of
168   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r va e

West East

All AA All aa

Figure 12.1  A hypothetical example in which a large difference in allele frequency between two populations results in a deficiency of heterozygotes
if all individuals are assumed to be members of a single population with p = 0.5 and q = 0.5. The eastern and western populations are completely
isolated by a strong northward flowing current that prevents larvae from being exchanged between east and west. Expected heterozygosity for the total
population is HT = 2pq = 0.5, but the average expected heterozygosity for each of the two individual populations is HS = 0. Therefore, FST = (HT—HS)/HT
= 1.0. Adapted from Hartl and Clark (1997) (see Plate 11).

assumptions known collectively as Wright’s (1951)
“island model.” The island model assumes many
equally sized populations (each of size N) in which
a fixed proportion (m) of every population are im-
migrants each generation, and that m is relatively
small. The model also assumes each population
has been separated for long enough that gene flow
among populations and genetic drift within popu-
lations have reached an evolutionary equilibrium.
An important consequence of these assumptions is
that FST can only be used to estimate the compound
parameter Nm, a useful parameter for understand-
ing the impact of gene flow on allele frequencies,
but of less value in understanding the demographic
impact of migration (see later).
Because many real populations likely do not
conform to island-model assumptions (Beerli and
Felsenstein, 1999; Whitlock and McCauley, 1999;
Neigel, 2002), estimates of Nm from FST are now rare
in the literature (Marko and Hart, 2011). However,
understanding the theoretical relationship between
Nm and FST highlights several important facts. First,
Wright’s equation FST = 1/(4Nm + 1) demonstrates
that a single migrant per generation has the same
impact on allele frequencies in a large population
as a single migrant per generation has in a small
population. This counterintuitive result is explained
by the fact that, in a large population, the relatively
small impact of a single migrant on allele frequen-
cies is opposed by relatively weak genetic drift; by
contrast, in a small population, a single migrant has
a much larger effect on allele frequencies, but the im-
pact of gene flow in a small population is opposed
by much stronger genetic drift. Second, the rela-
tionship between FST and Nm emphasizes how lit-
tle gene flow is necessary to keep allele frequencies
similar among populations (Figure 12.2) and how
difficult it can be to precisely measure migration
when Nm >10 migrants per generation, especially
considering that the error associated with estimates
of FST is often as large as the estimate itself when FST
is small (Waples, 1998). This “gene flow problem”
(Waples, 1998) makes real population differentiation
very difficult to distinguish from random noise in
marine species with high gene flow, and makes gene
flow that is sufficient to homogenize allele frequen-
cies very difficult to distinguish from panmixia (i.e.,
all individuals are potential mating partners).

0.8
0.6
FST
0.4
0.2
0 Figure 12.2 The relationship between FST
0 2 4 6 8 10 12 14
Nm
12.1.3.3  Coalescent population genetics
Rather than modeling how allele frequencies are
expected to change moving forward in time, coa-
lescent population genetics focuses on the genea-
logical history (i.e., a gene tree) for a sample of gene
copies moving backward in time: if two individual
gene copies have the same common ancestor in a
previous generation, those two copies are said to
have coalesced. The earliest applications of coa-
lescent theory were used to make inferences about
demographic parameters for a single population,
but the theoretical framework of the coalescent has
been expanded to incorporate other demographic
parameters. For example, gene flow between popu-
lations can be estimated with gene trees by infer-
ring the rate at which gene copies in one population
coalesce in an ancestor in another population (Nath
and Griffiths, 1993; Beerli and Felsenstein, 1999;
Nielsen and Wakeley, 2001).
The primary advantage of coalescent gene flow
estimators is that they often employ a much more
realistic model of gene flow and population history
than Wright’s island model (Beerli and Felsenstein,
1999). Most coalescent gene flow estimators use a
Bayesian statistical framework, in which a posterior
distribution is estimated for each demographic pa-
rameter by simultaneously “sampling” (searching
G e n e t i c A n a lys i s    169
16 18 20 and Nm based on Wright’s (1978) equation
FST = 1/(4Nm + 1) that assumes Wright’s
island model.
among) tree topologies and parameter values.
Coalescent methods typically use computationally
intensive Markov chain Monte Carlo (MCMC) sam-
plers, in which small random changes are repeat-
edly applied to gene trees. The likelihood of each
gene tree and parameter estimate is calculated at
each step in the search until the search converges
on a sample of highly likely gene trees and associ-
ated parameter estimates. This capability to calcu-
late likelihoods for multiple individual parameter
values (including those associated with gene flow
and other demographic processes) is an important
source of the increased realism of coalescent gene
flow estimators, in contrast to the insights gained
from single summary statistics (such as FST). Either
Nm or m can be estimated with coalescent samplers,
but estimates of m are typically scaled by mutation
and can only be converted into demographically
meaningful values with an estimate (or assump-
tion) about the mutation rate of the markers.
Coalescent methods have important limitations
(Marko and Hart, 2011). First, despite lacking sev-
eral of the unrealistic assumptions of Wright’s
island model, each coalescent estimator has an
underlying demographic model that still makes
some assumptions about the history and struc-
ture of the sampled populations that may or may
170   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r va e
not match reality. Second, given the high among-
locus variance in the coalescent, robust and con-
sistent answers from coalescent estimators require
data from multiple loci (Karl et al., 2012). Third,
even with high-performance computing clusters,
it is impractical to apply coalescent methods that
use MCMC samplers to genomic datasets consist-
ing of thousands of individual single nucleotide
polymorphisms (SNPs) in which each SNP has its
own gene tree (although many population genetic
questions often do not require thousands of loci).
Lastly, coalescent theory is based on mathematical
approximations that assume that effective popula-
tion size is large and that migration rates are low.
So, like FST, coalescent methods are not expected to
perform well when Nm is very large (>10) or when
N is very small (<100).
12.1.3.4  Approximation methods
A less computationally intensive alternative, ap-
proximate Bayesian computation (ABC) meth-
ods (Beaumont, 2002; Lopes and Beaumont, 2009;
Csilléry et al., 2010), estimate posterior distribu-
tions of demographic parameters from simulated
datasets and summary statistics (e.g., number of
alleles, genetic diversity, genetic distances) rather
than from samples of likely gene trees. Instead of
assuming a single demographic model (as with
Nm from FST and from coalescent estimators), ABC
typically starts by simulating data under several
alternative demographic models (e.g., with and
without migration) and then either accepting or
rejecting models by comparing summary statistics
for each simulated dataset to the observed values.
The posterior distributions for demographic pa-
rameters of interest are then approximated from the
distribution of parameters values from the accepted
models (like model selection and model averaging
approaches used in some coalescent methods based
on likelihoods). Like other population-based meth-
ods, even though the posterior for m is estimated
in demographic quantities in most ABC methods,
the value of m depends on the priors for mutation
rates used in the simulations. Because ABC meth-
ods do not make full use of sequence data (i.e., the
coalescent), they typically do not provide estimates
of demographic parameters as precise as those from
MCMC methods (Beaumont et al. 2002). However,
the practical advantages of ABC lie in the capabil-
ity to consider very large genome-wide datasets
and to make direct comparisons among complex
demographic models defined by the investigator
(Rougemont et al., 2016).
Alternatively, when estimating migration rates
between two or more populations, the joint site fre-
quency spectrum (SFS) can be used instead of sum-
mary statistics. The SFS is more informative than
any single summary statistic (all summary statistics
can be calculated from allele frequencies, but allele
frequencies cannot be calculated from summary
statistics) and is advantageous in that increasing
the number of SNPs or individuals does not pro-
portionally increase the computational time, but
greatly increases the power of the analysis (Excoffier
et al., 2013). Another relatively new computation-
ally efficient approach for inferring demographic
parameters (including migration) combines dif-
fusion approximation (Fisher, 1922; Kolmogorov,
1931; Kimura, 1964) of the expected SFS under al-
ternative demographic models with maximization
of the similarity between the observed SFS and the
simulated SFS across simulated parameter values in
the demographic model (Gutenkunst et al., 2009).
12.3.2  Individual-Based Methods
In contrast to population-based methods, the rate or
direction of ongoing dispersal can be estimated by the
classification of individual genotypes as immigrants
(or the recent descendants of immigrants). This ap-
proach has great appeal because these insights into
the dispersal history of individual organisms pro-
vide estimates of net dispersal and therefore come
closer to the kind of direct insight into connectivity
that would be gained from direct observations of
larval movements in the plankton. Here we focus on
three methods and their implementation that have
been used by empiricists studying marine larvae.
All three of these methods use transient, short-lived
effects of immigration on inter-individual genetic
variation to detect recent or ongoing migration
events. This feature also sets the individual-based
methods apart from population-based methods that
draw inferences from allele frequencies, coalescent

times, or other population genetic variables that are
associated with changes in genetic variation on long
timescales. Thus, individual-based methods give
insights into connectivity among populations that
are complementary to the results from population-
based methods, but like population-based methods,
they have their own distinctive limitations on those
insights (Jones et al. 2005; Planes et al., 2009; Chris-
tie et al., 2010; Harrison et al., 2012; Saenz-Agudelo
et al., 2012; Pusack et al., 2014).
12.3.2.1  Clustering individuals from different
samples
The most intuitively appealing individual-based
methods use clustering of individual genotypes from
multiple samples (e.g., from several geographic loca-
tions) into one or more genetically similar groups;
immigrants can then be identified as genotypes from
one sample that are more confidently clustered with
genotypes from some other sample or location.
The most widely cited clustering method is
the suite of algorithms in the program called
STRUCTUR­E described by Pritchard et al. (2000).
Such clustering methods are sometimes described
as making fewer assumptions about demographic
structure and history in comparison to population-
based methods (Pearse and Crandall, 2004). In the
case of STRUCTURE, each cluster of genetically sim-
ilar individuals is assumed to have its own demo-
graphic history in which genotypes at a single locus
are in HWE (due to random mating in a large popu-
lation), and alleles at different loci are expected to
be in linkage equilibrium (due to recombination and
independent assortment). Optimization by MCMC
is used to find the individual assignments to clus-
ters that minimize linkage disequilibrium (LD) and
maximize HWE within each cluster. Thus, although
clustering and other individual-based methods do
not directly estimate migration rates or population
size in an explicit population model, some of those
model parameters enter the individual-based meth-
ods under the guise of the quantities to be optimized
in the search for immigrant genotypes.
Under these assumptions, recent immigration
events are expected to cause both transient LD
among loci (by adding unusual combinations of
­alleles at different loci) and deviations from HWE
G e n e t i c A n a lys i s    171
for single loci (by adding unusual genotypes).
These effects are expected to be short-lived because
random mating and recombination will break up
linkage groups and restore HWE within one or
a few generations after each immigration event.
Consequently, STRUCTURE results are sensitive
mainly to the genetic signal from recent or ongoing
immigration. In the original STRUCTURE model,
first-generation immigrant individuals could be
identified as those multilocus genotypes from one
geographic region (or destination) that had a high
probability of assignment to a cluster that was com-
mon in a different geographic region (or source). In
subsequent versions of the model, recent descend-
ants of immigrants could also be identified as those
individuals with an admixture of alleles character-
istic of both the source and destination populations
(Falush et al., 2003).
Although STRUCTURE can be thought of as a
method to “let the data define the populations”
(Pearse and Crandall, 2004), an important assump-
tion of STRUCTURE is that the total sample consists
of one or more genetically discrete clusters, each of
which is internally homogeneous, and that the true
number of clusters (K) is known and specified by the
researcher. In other words, K is not a model param-
eter value estimated from the data by optimization,
but rather a variable in the optimization algorithm.
Incorrectly specifying K can lead to errors in assign-
ment, and thus errors in inferring dispersal from the
distribution of clusters among different geographic
locations. The limitations on inferring both number
of clusters and assignment of genotypes to clusters
in the same optimization are well known (Pritchard
et al., 2000), and several heuristic solutions to esti-
mating K have been proposed (Evanno et al., 2005;
Kalinowski, 2011; Puechmaille, 2016).
A second important limitation of the STRUCTUR­E
method is the assumption that gene flow is low
(Pritchard et al., 2000) and immigrants are rare.
This assumption is inherent in all individual-based
methods (and like the assumptions underlying
population-based methods), which depend on the
occurrence of recognizable clusters or differentiated
populations that could be the source of distinctive
immigrant genotypes. This leads to the surprising
expectation that as the true migration rate (and the
172   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r va e
expected occurrence of immigrants in population
samples) increases, the sensitivity of individual-
based methods to count immigrants and quantify
migration rates may greatly decline. Given this con-
straint, can clustering methods be used to discover
ecologically meaningful gene flow, or can these
methods discover only gene flow that is trivially
low? Simulations (Waples and Gaggiotti, 2006) sug-
gest that there may be “situations where Nm is high
enough that a realistic population sample would
contain enough immigrants to shed light on immi-
gration patterns, yet where there remained enough
differentiation between populations to endow ge-
netic assignment methods with adequate power for
F0 [first-generation] immigrant detection” (Paetkau
et al., 2004, p. 56). It seems uncertain whether such
situations are common among systems of marine
animal populations. However, this sensitivity to
the homogenizing effects of long-term gene flow
suggests caution in the interpretation of individual-
based estimates of ongoing gene flow when those
estimates are high (Saenz-Agudelo et al., 2009;
Lowe and Allendorf, 2010).
12.3.2.2  Assigning immigrant individuals
to source populations
This second approach includes some of the same
model parameters from population-based meth-
ods (including the migration rate, m), but estimates
those model parameters by identifying recent im-
migrants and their population of origin (rather than
by characterizing long-term rates of gene flow).
The most widely used of these methods is called
BayesAss (Wilson and Rannala, 2003). Each sam-
pling location is assumed to constitute a population
that may include some first-generation immigrants
from one or more source populations, as well as
recent (second- or third-generation) descendants
of immigrants; allele and genotype frequencies at
each locus can vary among populations (and to
vary away from HWE conditions); and different
pairs of populations may exchange migrants at dif-
ferent rates. Unlike clustering methods, BayesAss
starts with the assignment (Paetkau et al., 1995)
of each individual genotype to the sample where
that genotype’s expected frequency is the great-
est (based on the observed distributions of alleles).
Then, like clustering methods, the fit of the model
to multilocus genotype assignments is evaluated by
maximum likelihood, and optimization is used to
find the most likely values of m that can account for
the number and source of immigrant genotypes (or
descendants of recent immigrants one or two gen-
erations into the past). As with clustering methods,
the assignment tests are sensitive only to recent im-
migration because additional generations of mat-
ing with non-immigrant genotypes will erode the
signal of immigrant ancestry beyond the second-
generation descendants of immigrants.
A significant limitation of this assignment ap-
proach may be its ability to resolve complex pat-
terns of migration among a biologically realistic
(large) number of populations (Faubet et al., 2007;
Mardulyn et al., 2008). Meirmans (2014) showed
that estimates of migration rates from BayesAss
analyses may be biased by computational limita-
tions on the ability to optimize model parameter
values (migration rates into many populations)
from limited data (small numbers of individuals
and loci). In general, the quality of BayesAss results
improves with larger samples of organisms and
deeper sampling of genomes, but declines with lar-
ger numbers of populations.
12.3.2.3  Assigning individuals to families
A third—and conceptually distinct—approach to
counting migration events includes the fewest popu-
lation model parameters. This approach uses genea-
logical methods to infer parent-offspring relationships
among sampled genotypes, and infers migration
from the discovery of close family members in differ-
ent population samples. One highly cited method is
called CERVUS; the original version was designed to
assign paternity to offspring given genotype data for
those offspring and their known mothers (Marshall
et al., 1998); extensions of the method allowed for the
effects of genotyping errors, and for more accurate
assignment of parentage given only genotypes of off-
spring and candidate parents (and without a known
maternal or paternal genotype; Kalinowski et al.,
2007). Some candidate parents can be excluded by al-
lelic mismatches with offspring genotypes; like clus-
tering and assignment methods, CERVUS then uses
likelihood scores to assess nonexcluded candidate
parents and identify the most likely parent for each
parent-offspring pair (based on the frequencies of the

shared alleles, and heterozygosity of the parental gen-
otypes). Unlike other methods, which fit population
model parameters (and characterize confidence in the
parameter value estimates) by optimization, parent-
age methods are based on simple Mendelian inher-
itance rules; instead of optimization, the confidence
in the identification of a specific parent-offspring pair
in CERVUS is assessed by comparison to simulations
that use empirical allele frequencies from the sam-
pled populations.
Although methods like CERVUS are designed
to assign parentage and identify parent-offspring
pairs, some studies that identify the same parent(s)
for more than one offspring can thus also identify
full- or half-sibling pairs, including siblings that
were collected in different population samples.
This is a significant but largely untapped strength
of parentage methods: they provide the only indi-
vidual- or population-based genetic approach that
can quantify migration specifically caused by ad-
vection of offspring away from their parents (e.g.,
a planktonic cohort of sibling larvae that disperses
away from the parental population in an ocean cur-
rent), and distinguish this from migration caused
by diffusion of siblings away from each other (e.g.,
spread of siblings of the same cohort or different
cohorts due to spatial or temporal variation in cur-
rent speed and direction). Both advection and dif-
fusion in the plankton contribute to observed levels
of migration and gene flow, but the two modes of
dispersal have different ecological and evolution-
ary consequences (Palmer and Strathmann, 1981)
on both small (Grosberg, 1991) and large (Largier,
2003) spatial scales.
Parentage methods (especially those that use
exclusion to screen out most candidate parent-
offspring pairs) also have a significant weakness:
they may be sensitive to the effects of genotyping
errors that cause non-parental alleles to be ob-
served in true offspring of a parent (e.g., so-called
stuttering of microsatellite allele sizes; Bonin et al.,
2004); conversely, as the size of studies grow, a
large number of pairwise comparisons can cause
unrelated individuals to share alleles by chance.
A counterintuitive effect of the sensitivity to geno-
typing error is that the number of mistakes in par-
entage assignment may increase with the number
of sampled loci in assignment methods based on
G e n e t i c A n a lys i s    173
exclusion (because non-parental alleles observed
in offspring may be sufficient to mistakenly ex-
clude a true parent-offspring pair), in contrast to
other individual- or population-based methods
where confidence in clustering or population as-
signment should increase with the number of loci
sampled (Sobel et al., 2002). Genotyping errors can
be included in probabilistic models for identifying
parent-offspring pairs (Kalinowski et al., 2007), but
decreasing the sensitivity of the models to genotyp-
ing error also decreases the accuracy with which
true parent-offspring pairs may be distinguished
from other genetic similarities between individuals
(Christie, 2009). This sensitivity to genotyping er-
rors may impose a significant limitation on the use
of some parentage methods in genome-scale stud-
ies of larval dispersal and gene flow.
12.4  Improved Understanding of Larval
Dispersal and Gene Flow
Here we highlight several areas of advancement
since Palumbi’s (1995) review, especially those ar-
eas that have benefited from the application of new
coalescent population models or individual-based
methods.
12.4.1  Biological Correlates of Larval Dispersal:
Planktonic Larval Duration
Because most marine larvae cannot be followed di-
rectly in the plankton, ecologists have long searched
for useful proxies for dispersal potential. One com-
mon and accessible proxy—the duration of the
planktonic larval stage (PLD)—can be estimated by
rearing larvae in the laboratory or by observing cal-
ibrated growth marks (such as daily increments in
growth of fish otoliths) in larvae collected from the
plankton. Estimates of dispersal potential based on
PLD vary among species from several minutes to
several years (Strathmann and Strathmann, 2007).
Palumbi (1995) reviewed the early evidence for
variation in PLD. He asked whether PLD covaries
with (and statistically accounts for) realized disper-
sal measured as differentiation itself (e.g., FST) or as
a pattern of increased differentiation among popu-
lations separated by larger geographic distances
174   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r va e
known as isolation-by-distance (IBD), in which the
strength of IBD is characterized by the slope of a
regression of FST against geographic distance be-
tween pairs of populations (Slatkin, 1993; Rousset,
1997). This definition of IBD among populations in
a steppingstone model of multiple habitat patches
is slightly different from Wright’s (1943) original
definition of IBD among individuals in a single
habitat patch in which typical dispersal distances
are less than the dimensions of the habitat (what
Wright called “local inbreeding in a continuous
area”). However, the two definitions share a similar
concept of limited or localized dispersal leading to
greater genetic differences on larger spatial scales.
Several early and important comparative stud-
ies of congeneric marine gastropod species with
or without planktonic larvae established support
for the specific and intuitive idea of an inverse
relationship between PLD and population gen-
etic differentiation (e.g., Berger, 1973; Snyder and
Gooch, 1973; Gooch, 1975), and the for the general
idea that PLD can be used as a proxy for realized
or typical larval dispersal distances (Crisp, 1978).
Other comparative genetic studies of related and
co-distributed molluscs (Hoagland, 1986; Kyle and
Boulding, 2000; Collin, 2001), fishes (Waples, 1987;
Doherty et al., 1995), echinoderms (McMillan et al.,
1992; Arndt et al., 1998), corals (Hellberg, 1996), and
crustaceans (Duffy, 1993) provide additional evi-
dence of a strong negative correlation between PLD
and population genetic differentiation.
However, a steady accumulation of counterex-
amples cast some doubt on the generality of this
pattern (Burton, 1983; Palumbi, 1995). Some of the
exceptions are more difficult to evaluate given that
they are generally not comparative studies of either
closely related or co-distributed taxa, but are instead
studies of population structure in single species that
revealed unexpectedly high or low genetic differen-
tiation relative to the authors’ expectations based
on PLD estimates (Saunders et al., 1986; Watts et al.,
1990; France et al., 1992; Todd 2003; Planes, 1993;
Wares et al., 2001; Taylor and Hellberg, 2003; Rocha
et al., 2005; Baums et al., 2006; Bowen et al., 2006;
Marko et al., 2007). Several literature reviews and
meta-analyses have attempted to resolve this issue,
but have come to substantially different conclusions,
with some studies reporting a strong relationship
between PLD and metrics of population differentia-
tion (Bohonak, 1999; Shanks et al., 2003; Siegel, 2003)
and others reporting a much weaker relationship
(Bradbury et al., 2008; Ross et al., 2009; Weersing
and Toonen, 2009; Kelly and Palumbi, 2010; Selkoe
and Toonen, 2011; Riginos et al., 2011).
Several factors may account for this uncertain or
contentious relationship between PLD and popula-
tion genetic differentiation. First, although meta-
analyses of large numbers of studies have great
power, they also confound variation in the biology
of the study organisms with variation in the meth-
odological approaches and shortcomings of the in-
dividual studies (Selkoe and Toonen, 2011; Dawson,
2014). Sample sizes, spatial scales of sampling, bio-
geographic region, genetic marker choice, and the
metric of realized larval dispersal (especially the
choice of FST vs. the IBD slope) can affect the ap-
parent relationship between PLD and population
genetic differentiation (Weersing and Toonen, 2009;
Selkoe and Toonen, 2011).
Second, the strength of early comparative studies
lay in phylogenetically controlled comparisons of
co-distributed taxa. However, these studies focused
on comparisons between species with relatively
large, qualitative differences in dispersal potential
(i.e., planktonic larvae vs. non-planktonic larvae).
Although species that lack planktonic larvae are rel-
evant to predictions about the effect of PLD on dis-
persal, a strongly bimodal distribution of PLD (with
one mode at zero for species without a planktonic
larva) biases the perceived strength of the overall
relationship between time spent in the plankton
and genetic differentiation (Bay et al., 2006; Ross
et al., 2009; Weersing and Toonen, 2009; Kelly and
Palumbi, 2010; Riginos et al., 2011).
Third, the use of PLD as a proxy for dispersal po-
tential is itself fraught with difficulty. Laboratory
measures of PLD do not easily account for seasonal
and annual variation (especially in temperature and
food availability) in nature. The capabilities of some
larvae to greatly extend their time in the plankton
by the uptake of dissolved organic matter (Moran
and Manahan, 2004) or by developmental arrest
(Pradillon et al., 2001), and the abilities of other
larvae to enhance or limit their advection by active
swimming (Kough et al., 2014) or by orientation to
physical and chemical cues in the ocean (Mouritsen

et al., 2013) may also contribute to a mismatch be-
tween laboratory measurements of PLD, actual
time spent in the plankton, and the realized effects
of larval duration on dispersal and gene flow.
Lastly, FST-based metrics of realized dispersal in-
voke Wright’s island-model assumptions. Are these
assumptions justified, or would other measures of
migration and gene flow (McGovern et al., 2010;
Crandall et al., 2012) more closely reflect the dispersal
capabilities of larvae? Most of these assumptions are
difficult to test. The assumption of a drift-migration
equilibrium is supported by (but not a requirement
for) observations of significant IBD in a system of
populations (Wares, 2002). Gene flow and genetic
drift are also more likely to be near equilibrium at
small spatial scales between neighboring popula-
tions (Slatkin, 1993; Hellberg, 1995). Consistent with
that view, Selkoe and Toonen (2011) found a stronger
relationship between realized dispersal distances (the
IBD slope) and PLD when analyzing FST measured
on relatively small spatial scales (<650 km). It was
surprising, then, that the relationship between PLD
and FST was not improved by analyzing only datasets
with significant IBD (Selkoe and Toonen, 2011).
Given these conflicting results, it seems fair to
ask: does planktonic larval duration predict the
scale and magnitude of marine larval dispersal as
estimated from measures of population genetic dif-
ferentiation? Clearly, species that lack planktonic
larvae typically show much stronger patterns of
population genetic differentiation than species
with planktonic larvae. However, the overall quan-
titative relationship between PLD and genetic dif-
ferentiation is much less strong, probably because
dispersal potential of planktonic larvae is difficult
to estimate from laboratory PLD, FST is sometimes a
poor proxy for gene flow, FST itself is sensitive to the
overall level of polymorphism of the genetic mark-
ers (Meirmans and Hedrick, 2010), and because
many other factors probably affect the realized dis-
persal of marine larvae.
One proposed solution to that unsatisfying con-
clusion is to ask the same question in a more spe-
cific and restricted form. Evidence of a fundamental
mechanistic relationship between PLD and real-
ized dispersal is most often clear in comparisons
between co-distributed species that have occupied
the same seascape for the same amount of time (as
G e n e t i c A n a lys i s    175
measured by similarly aged coalescents), so that ge-
netic drift has had the same opportunity to cause
the evolution of population differentiation propor-
tional to PLD differences among species (Dawson,
2014; Dawson et al., 2014). Focusing only on such
synchronously diverging and co-distributed taxa
may give the clearest view of the mechanistic rela-
tionship between PLD and FST. Unfortunately, this
approach is less useful for interpreting population
differentiation in studies of single or idiosyncratic
species (lacking a co-distributed species of a similar
coalescent age for comparison), for which popu-
lation genetic structure may be important and in-
teresting but for which a comparative context is
unavailable for interpreting the geographic extent
or magnitude of differentiation relative to PLD.
A second proposed solution is to ask the ques-
tion in a broadly expanded form that includes PLD
plus the many other factors that influence realized
dispersal, such as the regional oceanography or
“seascape” (Barshis et al., 2011; Sunday et al., 2014;
Liggins et al., 2016), habitat availability (Crandall
et al., 2012), population size and fecundity (Saenz-
Agudelo et al., 2012; Dawson et al., 2014), tem-
perature (Kelly and Eernisse, 2007; Bradbury et al.,
2008), adult behavior during spawning (Carson
et al., 2010), and larval behavior in the plankton
(Gerlac­h et al., 2007).
This multifactorial approach seems promising.
For example, numerous studies have reported
highly variable genetic differentiation on small spa-
tial scales, much less than the expected geographic
scale of larval dispersal. This pattern is in some ways
the opposite of IBD, and has been dubbed “chaotic
genetic patchiness” (Johnson and Black, 1982; Da-
vid et al., 1997; Selkoe et al., 2010; Cornwell et al.,
2016). One proposed cause of such patterns is strong
temporal variation in both the source of plank-
tonic larvae and in availability of recruitment sites
among destination populations (Johnso­n and Black,
1984). A second proposed cause of such patterns
is high variation in reproductive success among
broadcast-spawning adults, leading to low genetic
diversity within cohorts of offspring (relative to
diversity among all adults) and strong differentia-
tion among cohorts (Hedgecoc­k, 1994; Hedgecoc­k
and Pudovki­n, 2011). This type of sweepstakes re-
production (with few winners and many losers)
176   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r va e

can reduce local genetic effective population size
within each reproductive event and may promote
population differentiation. However, migration of
larvae—and especially the diffusion of larvae from
a single cohort away from each other—should lead
to the homogeneous sharing of offspring of the
sweepstakes winners among many populations,
and should prevent the evolution of genetic dif-
ferentiation among populations with overlapping
generations derived from multiple cohorts of larvae
(Hedgecock, 1994). Broquet et al. (2013) used simu-
lations to show that a simple additional factor—a
small reduction in diffusion, or a small tendency for
larvae to aggregate and to disperse together—could
produce strong local population differentiation that
was quantitatively like empirical measures of cha-
otic patchiness in nature (also see Eldon et al., 2016).
Broquet et al. (2013) conceded that aggregation and
collective dispersal of cohorts seems unlikely, but
is consistent with greater than expected kinship
among some larvae and new recruits (Selkoe et al.,
2006; Hedgecock et  al., 2007; Iacchei et al., 2013;
Ottmann et al., 2016; Selwyn et al., 2016), and could
be readily tested using data from the quantification
of dispersal kernels (Figure 12.3) using individual-
based methods.
12.4.2  Estimates of Dispersal Distances
Given that few generalities can be made about larval
dispersal from large meta-analyses, the attention of
some marine ecologists has shifted toward under-
standing how far individual larvae travel between
fertilization and metamorphosis, and to character-
izing the frequency distribution of those dispersal
distances. That frequency distribution is also known
as the dispersal kernel (Largier, 2003; Botsford et al.,
2009), and it is expected to be related to many of the
most important ecological and evolutionary quanti-
ties associated with pelagic larval development.
12.4.2.1 Isolation-by-distance
One approach to measuring characteristic larval
dispersal distances has used parameter estimates

30 Dispersal kernel:
P(distance) = 0.36e–0.36(distance)

25

20
Number of offspring

15

10

0
0 5 10 15 20
Dispersal distance (km)

Figure 12.3 Frequency distribution of dispersal distances by larvae away from parents for 120 parent-offspring pairs of the neon goby Elacatinus
lori from the western Caribbean. The dispersal kernel was estimated by fitting a negative exponential function to the frequency distribution, with a
best estimate of the decay parameter λ = 0.36; the inverse of the decay parameter is the average dispersal distance (~2.8 km). Data and analysis
from D’Aloia et al. (2015); larval goby image from Smithsonian Belize Larval-Fish Group 2002, image ID C2-19 (see Plate 12).

from population-based methods such as FST. How-
ever, this fixation index is an unreliable measure
of genetic differentiation in species with high gene
flow (Figure 12.2; Waples, 1998). The IBD slope is
likely a better proxy for realized dispersal in high-
dispersal marine species (Palumbi, 2003) because
there should be no relationship between FST and
geographic distance if error associated with FST is as
large as the estimates of FST.
In an influential study, Kinlan and Gaines (2003)
used a population genetic simulation of larval dis-
persal (from Palumbi, 2003) to characterize the rela-
tionship between variation in simulated dispersal
distance and variation in the resulting IBD slope
among simulated populations. They then applied
that function to estimate dispersal distances from
empirical studies of IBD patterns among popula-
tion genetic samples of marine organisms. Kinlan
and Gaines (2003) used this simulation approach to
circumvent the difficulties in obtaining direct esti-
mates of larval dispersal distances for species with
prolonged planktonic development and large pop-
ulation sizes (Cowen and Sponaugle, 2009). As with
Palumbi (2003), they inferred surprisingly short
dispersal distances of only approximately 0.01–100
km per generation, and proposed that a large part
of that extraordinary variation was related to PLD
and other intrinsic biological differences in disper-
sal potential among marine animal species.
Many other marine animals have geographic
ranges that greatly exceed the dispersal distances es-
timated from IBD patterns. A plausible mechanism
leading to that difference is steppingstone migration
(Kimura and Weiss, 1964) over many generations of
short-­distance dispersal events that link far-flung
populations (Planes and Fauvelot, 2002). Crandall et
al. (2012) reviewed examples of IBD, and noted that
genetic evidence for steppingstone migration (and
its implications for larval dispersal) is rare relative to
many examples of spatial genetic variation that are
more consistent with the effects of recent range ex-
pansion, rare long-distance dispersal events, or other
nonequilibrium processes. Crandall et al. (2012) used a
clever comparative approach to distinguish the effects
of steppingstone gene flow from the contributions of
other processes to population genetic variation. They
sampled mtDNA population variation across Indo-
Pacific island archipelagos for neritid snails in which
adults of some species live in marine habitats, but
G e n e t i c A n a lys i s    177
adults of some other species have evolved to live only
in freshwater streams on high islands. Both types of
species have swimming planktonic marine larvae,
and adults of both types can live on high islands (in
freshwater or marine habitats), but the two types of
species differ in their opportunities for stepping stone
gene flow: species with marine adults can disperse
between high islands over several generations by
using the marine habitats of low atolls as stepping-
stones; in contrast, amphidromous species can only
disperse between high islands directly in a single
generation (because adults are unable to live on atolls
that lack freshwater habitats). By using a coalescent
population-based method to model migration rates
independent of other population model parameters,
Crandall et al. showed that steppingstone dispersal
led to greater gene flow between high islands for one
marine species but not for amphidromous species.
A physical circulation model of ocean currents pro-
vided important context for that comparison: in some
cases, high islands were connected by strong currents
that allowed direct dispersal between populations
of both marine and amphidromous species; only in
cases without a strong direct oceanographic connec-
tion did the two types of species differ in migration
rates (due to their different abilities to make use of
steppingstone dispersal via atolls).
This example has several interesting implications.
First, it shows that careful dissection of multiple
factors (including ocean currents and adult habitat
requirements) may often be needed to reveal the
specific circumstances that lead to a correlation
between genetic differentiation and geographic
distance, and to estimate the contribution of step-
pingstone larval dispersal to the direction and mag-
nitude of gene flow. Second, the neritid example
suggests that coalescent population models (rather
than summary statistics such as FST from Wright’s
island model) may be needed to estimate migration
rates from genetic data in a complex biogeographi-
cal setting where many other processes also affect
larval movements in the plankton. Third, the exam-
ple illustrates a potentially important limitation on
the use of individual-based methods and dispersal
kernels to understand larval migration, gene flow,
and population structure. Because individual-based
methods detect only the effects of ongoing or recent
immigration over one or a few generations, they
may be inherently incapable of capturing a complete
178   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r va e
view of larval migration if steppingstone dispersal
over many generations is an important mechanism
linking far-flung parts of a metapopulation. Dis-
tant populations in a stepping stone model may
show little genetic connectivity on short timescales
(measured by individual-based methods), but may
show substantial connectivity on longer timescales
(measured by population-based methods), espe-
cially if stepping stone gene flow is episodic and
influenced by temporally varying patterns of ocean
circulation. In this sense, the characterization of dis-
persal kernels from data using clustering methods
or assignment tests (see later) may be a necessary
but insufficient basis for linking larval biology and
dispersal to ecological or evolutionary processes,
and may require the addition of population-based
methods that integrate the effects of gene flow over
many generations via stepping stone migration.
12.4.2.2  Dispersal kernels
In contrast to the IBD approach that uses popula-
tion-based methods, a second approach to measur-
ing characteristic larval dispersal distances is the
dispersal kernel or the frequency distribution of
distances between parents and their offspring. This
distribution can be characterized using individual-
based methods to identify and count immigrants,
assign them to source populations, and describe the
frequency distribution of larval dispersal distances.
An elegant example from a coral reef fish illustrates
both the promise and the limits of this approach.
D’Aloia et al. (2014) used microsatellite polymor-
phisms and mitochondrial DNA sequences to char-
acterize population genetic variation in the neon
goby Elacatinus lori from several populations sam-
pled on the continuous barrier reef system and on
isolated atolls along ~150 km of the coast of Belize
in the western Caribbean. Adults of this species live
in large sponges where males brood developing
embryos; hatched larvae leave the sponge (and the
parents) to spend three weeks or more in the plank-
ton before recruiting to a new sponge host. Despite
the potential for long-distance larval dispersal dur-
ing that long planktonic period, population differ-
entiation (e.g., mtDNA FST up to 0.46) was strong
on both small (<  20 km) and large spatial scales;
differentiation was especially strong between atolls
and the barrier reef; and there appeared to be little
evidence of IBD.
To better understand those patterns, D’Aloia et
al. (2015) then intensively sampled >105 single-locus
microsatellite genotypes from thousands of E. lori
adults and newly recruited offspring along a 41-km
transect of the barrier reef. The parentage method
in CERVUS identified 120 parent-offspring pairs,
which allowed D’Aloia et al. (2015) to characterize
the dispersal kernel from that high-resolution fre-
quency distribution: the most common dispersal
distance was ≤1 km, the mean distance (estimated
by model-fitting; Figure  12.3) was <3 km, and the
largest distance traveled by any offspring was only
~16 km. Many (24) offspring were members of 11 dif-
ferent full- or half-sib families, including eight fami-
lies in which siblings diffused away from each other
and were advected different distances away from
their parents; two families in which siblings were ad-
vected the same distance from the parent(s) (without
diffusion away from each other); and one family in
which all siblings recruited to the same location with
the parent (and without advection or diffusion). A
single family included both the longest observed dis-
persal distance from the parent (16.4 km) for one of
the half-sibs, as well as the modal dispersal distance
from the parent (0.8 km) for the other half-sib. That
variation within and among families suggests that
advection (away from parents), diffusion (away from
siblings), and local retention of larvae all contribute
to patterns of larval dispersal and gene flow in E. lori.
Additional modeling (D’Aloia et al., 2015) sug-
gested that population genetic differentiation along
the barrier reef was caused by a form of isolation-
by-distance among mating groups (associated with
different host sponges) that were separated by dis-
tances greater than a few kilometers, like Wright’s
(1943) original definition of IBD. Although D’Aloia
et al. (2014) emphasized a lack of isolation-by-
distanc­e among populations, a relatively strong
pattern of IBD was evident among the barrier reef
populations only (Figure 12.4), and that pattern was
consistent with the expected effects of short-distance
larval dispersal along the continuously distributed
barrier reef habitat. In contrast, D’Aloia et al. (2015)
argued that populations on atolls are demographi-
cally isolated from the barrier reef (and from each
other) by deeper water and the absence of stepping-
stones (the host sponge species) in the intervening
habitat. That interpretation was supported by the
absence of IBD on any geographic scale for pairwise
 G e n e t i c A n a lys i s    179

differentiation among atoll populations or between
atolls and the barrier reef (Figure 12.4).
Comparable patterns of spatial genetic variation
have been found in other Elacatinus species (Taylo­r
and Hellberg, 2003; 2006), and similar dispersal
kernels have been found in a handful of other stud-
ies that used individual-based genetic methods to
characterize dispersal distances (e.g., clownfish;
Buston et al., 2012; Almany et al., 2013 Pinsky et al.,
2016). These results seem important because they
suggest that realized dispersal distances of many
coral reef fish may be considerably shorter than ex-
pected (Kinlan and Gaines, 2003; Cowen and Spo-
naugle, 2009; Shanks, 2009). They implicate larval
behavior in the plankton (e.g., swimming activity,
orientation, vertical migration) and steppingstone
gene flow between habitat patches that are sepa-
rated by distances less than the typical larval dis-
persal distance (Crandall et al., 2012) as critically
important factors that mediate the relationship
between potential and realized dispersal over both
short timescales (reflected in the dispersal kernel)
and over long timescales (reflected in population
model parameters such as pairwise population FST).
This detailed treatment of a specific example from
gobies illustrates both the potential importance of
individual-based genetic methods and the limits on
their application for understanding marine larval
dispersal. In a pilot study focused on a single E. lori
population from a 0.5 km section of barrier reef,
D’Aloia et al. (2013) were able to assign almost 5% of
offspring to parents; when the geographic scale of the
study was extended to capture the full dispersal ker-
nel, the assignment rate was reduced by about half
(D’Aloia et al., 2015), in spite of the large sample size
of adults and offspring (>7,000), because the broader
study area included many more individuals of which
a smaller proportion were sampled, genotyped, and
assigned to families. That difference suggests that it
may be realistic to use parentage (or other individual-
based) methods to characterize dispersal kernels only
on the smallest spatial scales in small populations
of organisms with long-lived adults. For example,
Stockwell et al. (2016) sampled thousands of SNP loci

0.016

0.012
Microsatellite pairwise F st

R2 = 0.00265

0.008

0.004

R2 = 0.25814

0.000
0 50 100 150 200
Euclidean distance (km)

Figure 12.4  Patterns of isolation-by-distance (IBD) in the neon goby Elacatinus lori from the western Caribbean (D’Aloia et al., 2014) based on 13
microsatellite loci sampled for 20–30 adults from five populations along a continuous barrier reef and five populations from isolated atolls. Relatively
strong IBD is detectable among barrier reef populations (open symbols; high coefficient of determination R2 ~0.26) that are connected by stepping
stone gene flow. D’Aloia et al. (2014) found no IBD among populations from atolls, or between atoll and barrier reef populations (closed symbols;
R2 ~0.0026). Adapted (with permission) from data in D’Aloia et al. (2014).
180   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r va e

from Indo-Pacific parrotfish (Scarus niger) collected or spatial variation in the frequency distribution of
at several population locations in the Philippines dispersal distances would require spatial replication
separated by >100 km. They discovered convincing and temporal repetition. Those sampling constraints
evidence of larval dispersal and gene flow on that seem to put the dispersal kernel for S. niger and many
larger spatial scale (two pairs of siblings in different other species (with broad geographic ranges, exten-
populations, identified using an assignment method sive larval dispersal, and large populations subject to
for identifying family members called ML-RELATE; spatial and temporal variation in dispersal) out of the
Kalinowski et al., 2006). That discovery suggests that reach of individual-based methods, even for studies
the method can in principle be applied to widely like that of Stockwell et al. (2016) that use automated,
separated populations connected by planktonic lar- high-throughput genomic data collection.
val dispersal on scales much greater than observed
in species like E. lori. However, the low assignment 12.5  Consequences of Larval Dispersal:
rates for S. niger individuals grouped into families Genetic Connectivity vs. Demographic
(~2%) implies that a vast collecting and genotyping
Connectivity
effort might be needed to find large numbers of sib-
ling pairs (and dispersal distances) and to fully char- The distinction between population- and individual-
acterize the dispersal kernel to the same precision based methods, including the diversity of specific
achieved by D’Aloia et al. (2015; Figure 12.3). Moreo- parameters that each may estimate (e.g., Nm vs. m),
ver, such studies can provide only individual snap- has become increasingly important for marine ecolo-
shots of the dispersal kernel; accounting for temporal gists, who have emphasized understanding patterns

marine & “gene flow” marine & “gene flow”

& larva & dispersal
200 200
Web of Science entries matching four search terms

NOT connectivity

150 150

NOT connectivity

100 100

50 50 AND connectivity
AND connectivity

0 0
1995 2000 2005 2010 2015 1995 2000 2005 2010 2015
Publication date (five-year intervals)

Figure 12.5  Trends in the use of the keyword “connectivity” in studies of marine larval dispersal and gene flow; data are counts of citations that
use different keyword combinations in searchable fields of records in the Web of Science database for the years 1996 through 2015 in five-year
increments following the review by Palumbi (1995). Results are shown for two alternative keyword searches using the terms “larva*” or “dispersal”;
in each alternative search, the results for items without “connectivity” (closed symbols) are contrasted with results for items including “connectivity”
(open symbols). In both cases the use of “connectivity” has dramatically increased since 2005.

of marine “connectivity” (Figure 12.5) via larval dis-
persal to resolve fisheries and conservation problems
such as the design of effective networks of marine
protected areas (MPAs; Gaines et al., 2010, Figure
12.1). In this context, it is useful to distinguish among
several types of connectivity that reflect different
population processes and can be estimated from ge-
netic data under different model assumptions.
First, marine ecologists and resource managers are
typically interested in characterizing demographic
connectivity, the degree to which population per-
sistence or growth rates are affected over ecological
timescales by larval dispersal among populations
(Waples, 1998; Waples and Gaggiotti, 2006; Lowe and
Allendorf, 2010). How much dispersal is necessary
to provide significant demographic linkage among
individual populations is not well understood and
highly context-dependent. One proposed standard
is that a migration rate of ~10% (e.g., m =  0.1) is
needed to maintain demographic synchrony (simi-
lar population dynamics and growth rates) among
separate populations linked by dispersal (Hastings,
1993; Waples and Gaggiotti, 2006).
By contrast, biogeographers and population ge-
neticists have emphasized the degree to which lar-
val dispersal affects evolutionary processes within
populations and evolutionary divergence among
them (Lowe and Allendorf, 2010). These effects
can include both positive (e.g., preventing inbreed-
ing depression) and negative consequences (e.g.,
preventing the fixation of locally adapted alleles
via gene swamping) of gene flow. Wright (1951)
concluded that Nm >1 was sufficient to prevent
the harmful effects of inbreeding and genetic drift
within populations. As a consequence, many popu-
lation geneticists have used this one-migrant-per-
generation (OMPG) “rule” as a threshold value
defining genetic connectivity, the amount of gene
flow necessary to homogenize allele frequencies
among populations over evolutionary timescales.
However, close inspection of the relationship be-
tween FST and Nm (Figure 12.2) reveals that much
greater gene flow (Nm >10) is necessary to prevent
divergence in allele frequencies. The confusion in
the literature over the OMPG rule and the expected
effects of gene flow led Lowe and Allendorf (2010)
to recognize three distinct levels of genetic connec-
tivity: drift connectivity (gene flow that prevents
G e n e t i c A n a lys i s    181
divergence in allele frequencies, Nm >10); inbreeding
connectivity (gene flow that prevents the harmful
loss of genetic diversity, Nm >1); and adaptive con-
nectivity (gene flow that is sufficiently high to pro-
mote the spread of adaptive alleles, Nm >0.1).
The differences among population- and individ-
ual-based genetic methods for analyzing larval dis-
persal have obvious parallels with these distinctions
between demographic connectivity (focused on per
capita migration rates, m) and genetic connectivity
(focused on gene flow, Nm), as well as implications
for the choice of methods to meet different study
objectives or goals. Clustering and assignment
methods characterize migration distances between
populations, and parentage methods characterize
migration distances away from parents; if sampling
is sufficiently intensive, some methods can estimate
the migration rate as the proportion of migrants
entering a destination population (m) through the
identification of migrant individuals, but not nec-
essarily the evolutionary impact of immigration in
the destination population (Nm) unless the effective
population size is known. Therefore, almost by defi-
nition, individual-based methods have the potential
to characterize demographic connectivity on short
(ecological) timescales (Christie et al., 2010); few give
insight into effective population size, such that indi-
vidual-based methods offer little insight into genetic
connectivity on longer (evolutionary) timescales.
By contrast, most population-based methods
(FST and coalescent methods) provide estimates of
the compound parameter Nm (the number of mi-
grants per generation), which provides insight into
the evolutionary impact of larval dispersal. This is
an important limitation because estimates of Nm
provide little insight into demographic connectivity
unless N or the population growth rate is known
(Lowe and ­Allendorf, 2010). This shortcoming of
population-based methods may often be moot for
some marine species with large population sizes be-
cause Nm can be most reliably estimated when it is
small (Nm <10), and for many marine species with
large population sizes, Nm <10 implies m << 0.1 and
is likely to be demographically insignificant. The
same cannot be said for cases in which Nm >10, be-
cause population-based methods have little power
to distinguish drift connectivity from strong demo-
graphic connectivity (Figure 12.2). However, even
182   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r va e
in cases where Nm can be reasonably interpreted
in the context of demographic connectivity (or if m
can be calculated with a known mutation rate), the
parameter estimates from most population-based
methods are time-averaged over long (evolution-
ary) timescales, and may not be similar to present-
day migration rates, particularly for species whose
populations may be declining or growing, or whose
distributions may be fragmented and rapidly
changing in response to anthropogenic effects.
12.6 Conclusion
For the last 20 years, population-based genetic ap-
proaches have been the go-to analytical methods for
studying marine connectivity, whereas the develop-
ment and use of individual-based methods is in its
infancy. Do the differences among the two classes of
methods and their variants matter for genetic analy-
ses of larval migration? Population-based methods
have proven to be most useful for identifying very
strong genetic breaks that often reflect a long-term
lack of both genetic and demographically significant
connectivity among populations, and genetic data
can provide crucial complementary data for ecolog-
ical studies that find evidence of demographic asyn-
chrony (Peterson et al., 1996; Marko and Barr, 2007).
However, if gene flow is high, population-based
methods are not expected to return clear and reli-
able answers about either genetic or demographic
connectivity. Individual-based methods may have
greater potential and show considerable promise
to uncover evidence of demographic connectiv-
ity among populations, but, like population-based
methods, are not expected to perform well when
migration rates are high (Waples and Gaggiotti,
2006; Paetkau et al., 2004; Lowe and Allendor­f, 2010)
and rely fundamentally on intensive sampling of all
potential source and sink populations. In short, ge-
netic methods can provide strong inferences about
either an absence of (either genetic or demographic)
connectivity or high genetic connectivity, but they
are, at present, less useful for demonstrating strong
demographic connectivity.
Clearly, no single approach will always tell a
complete story of realized dispersal and connec-
tivity. Comparing results from different analyses
may generate insights or reveal problems inherent
in any single approach, particularly for predicting
marine connectivity and its implications for fisher-
ies management or the design of networks of MPAs
in a rapidly changing environment. However, even
though individual- and population-based estimates
of connectivity can be inferred from the same data,
direct comparisons of migration rates over evo-
lutionary timescales to migration on more recent
ecological timescales have been rare (e.g., Pusack
et al., 2014; D’Aloia et al., 2015; Pinsky et al., 2016),
possibly because gene flow is often too high to ex-
pect informative population-based results on the
spatial scale at which individual-based methods
are most often applied (but see Pinsky et al., 2016),
and because the types of genetic markers that are
best for individual-based methods (microsatellites
and SNPs, but see Rougemont et al., 2016) are also
not ideal for some of the most powerful population-
based methods (DNA sequences). In addition to in-
ferences about larval dispersal from genetic data,
direct measurements of the impacts of dispersal on
demographic processes are likely crucial for dem-
onstrating meaningful demographic connectivity
for the purposes of resource management (Waples,
1998; Lowe and Allendorf, 2010). The complemen-
tary combination of these methods seems likely to
illuminate both the scope and scale of larval disper-
sal in marine systems.
12.7 Summary
1. Genetic data can be used to characterize the scale
or magnitude of connectivity via larval dispersal
in the plankton as the per capita migration rate
(m), the rate of gene flow (Nm), or counts of im-
migrant individuals.
2. Population-based methods infer average ef-
fective rates of connectivity on long timescales
(hundreds to thousands of generations), and
those estimates will be influenced by many pro-
cesses (including larval dispersal).
3. Individual-based methods based on clustering
or assignment of individual genotypes to popu-
lations or families are suitable for estimating
connectivity on short timescales.
4. The typical or characteristic larval dispersal dis-
tance for any one system of populations may
best be characterized by isolation-by-distance

patterns (using population model methods) or
by the dispersal kernel (using parentage-based
methods).
5. Migration rates estimated from individual-based
methods may be more relevant to ecologic-
al studies of demographic connectivity (e.g.,
among demes in a network of marine protected
areas) compared to rates of gene flow estimated
from population-based methods.
Acknowledgments
The ideas expressed in this chapter have been de-
veloped and honed through conversations with
many individuals. We are especially grateful to
Mark Christie, Joe Neigel, and an anonymous re-
viewer for numerous suggestions and constructive
criticism. Thanks to Cassidy D’Aloia for sharing
data and unpublished analyses of gobies.
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 CH A PT ER 13
I Feel That! Fluid Dynamics
and Sensory Aspects of Larval
Settlement Across Scales
Jason Hodin, Matthew C. Ferner, Andreas Heyland, and Brian Gaylord
13.1 Introduction
Standing on a wave-swept shore, it’s tempting to
imagine that the myriad microscopic larvae beneath
the surface are simply at the mercy of oceanic forces,
so-called “passive particles” being hurled to and
fro by the tremendous energy of tides, waves, and
currents. In this conception, a larva that eventually
would settle in the nearshore has three key tasks:
(1) to survive long enough and be lucky enough to
be passively carried to a suitable adult habitat;
(2) to recognize such a habitat when it arrives
there; and
(3) to attach or burrow into that habitat so as not to
be swept away by impinging flows.
In this sense, even a larva that might appear “pas-
sive” with respect to typical flow regimes could be
in some ways master of its own fate. For example,
engaging larval defenses could increase its odds
of survival in the presence of predators, adjust-
ing its feeding mechanisms could allow it to grow
faster and more efficiently, detecting conspecifics
or a favored adult food source could increase its
likelihood of settling in an appropriate location,
and quickly deploying well-developed adhesive
structures could allow it to withstand agents of
dislodgment when it arrives there. Furthermore,
our larva’s mother (and in some cases its father)
could have stacked the deck in its favor. For ex-
ample, she might have protected the embryo and
larva for a time, endowed it with extra energy in
Hodin, J., Ferner, M. C., Heyland, A., and Gaylord, B., I feel that! Fluid dynamics and sensory aspects of larval settlement across scales.
In. Evolutionary Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland:
Oxford University Press (2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0013
the form of yolk, or provided it with chemical de-
fenses to deter planktonic predators. She may also
have released her offspring during a specific sea-
son, lunar phase, or time of day that could offer
it the maximum available planktonic food, fewest
potential predators, and most favorable oceanic
flow conditions to retain it near to shore (see On-
line Supplementary Material).
But a growing body of evidence indicates that lar-
vae are best considered as other than passive par-
ticles (see Morgan, 2014). Although the maximum
swimming speed of the larvae of most invertebrates
(<1 cm·sec-1 or far less; Chia et al., 1984; Fuchs and
Gerbi, 2016) are too slow to make headway in
strong oceanic currents (10s of cm·sec-1) or within
wave-driven flows (meters·sec-1), these larvae nev-
ertheless have a behavioral repertoire that they
can exploit to increase their odds of finding food,
avoiding predators, and being carried back to shore
when they are ready to settle into benthic habitat.
Stronger swimming larvae of crustaceans and fish
can swim against and at least partially resist such
currents, and are thus even less passive.
Based on the notion that larvae of benthic spe-
cies have an underappreciated capacity to influence
their locations in space and thus their arrival into
specific habitats, in this chapter we will draw upon
examples from disparate marine invertebrates to
describe the following:
• the fluid environment that larvae experience in
the pelagic and benthic realms;
 F l u i d Dy n am i c s a n d S e n s o ry A s p e c t s o f La r va l S e t t l e m e n t    191
• what larva can sense in their fluid environment
and how they do so; and
• what cues larvae utilize, and how their responses
to such cues vary depending on the scale relative
to suitable settlement locations.
The main focus in this chapter is to review these
­topics from the perspective of larvae maxi­­mizing
their chances of surviving to settle at an appropri-
ate time and place. While addressing these issues,
we will often connect to subjects of other chapters
in this volume, which we will cross-reference for
their more extensive consideration of such mate-
rial. We also will highlight the tremendous progress
made in larval ecology in the last 50 years, and in
particular in the two decades since the publication
of Ecology of Marine Invertebrate Larvae (McEdward,
1995), the multi-authored work that inspired the
current edited volume. And, finally, we will look to
the future of the field, where new techniques and
interdisciplinary integration offer the promise of
deeper understanding of the surprisingly common
yet remarkably diverse complex life cycles of ma-
rine organisms.
13.2  What Does It Feel Like to be a Larva?
Although we do not know the complete answer to
this question, fluid dynamics offers some clues. A
bottlenose dolphin swimming through the wate­r
experiences its fluid environment much differ-
ently than does a coral planula larva. At the scale
of the dolphin, inertial forces predominate (think of
a boat continuing to glide long after the engine is
cut); at the scale of the planula, the dominant forces
are viscous (akin to a human swimming in a vat of
honey). The relative importance of inertial and vis-
cous forces can be described in terms of a parameter
called the Reynolds number (Re):
Re =
( ρU l )
µ cidians, and some crustaceans; McHenry et al., 2003)
The factors in the numerator contribute to larger
inertial forces (ρ—the density of the fluid; U—the
fluid velocity; and l—a characteristic length of the
organism in flow), whereas the surrounding fluid’s
dynamic viscosity μ is in the denominator. The units
cancel one another out, so the Reynolds number is
a dimensionless metric, useful across scales from
planktonic (millimeters or less) to oceanic (1000s of
kilometers), whereby two organisms with the same
Re can be thought of as experiencing a similar fluid
dynamic environment.
Because the Reynolds number depends on a
length scale, two organisms that differ vastly in size
but occupy the same habitat (like a bottlenose dol-
phin and a coral planula in a tropical lagoon) will ex-
perience quite distinct fluid dynamic environments:
while the density and viscosity of the ambient sea-
water are more or less the same for the two organ-
isms, the length of the dolphin (~3 m) is four orders
of magnitude greater than the length of the planula
(~0.3 mm), and the swimming speeds of these two
animals also differ by about three orders of magni-
tude (~3 m·sec-1 vs. ~3 mm·sec-1, respectively). At
the scale of the dolphin, the value in the numerator
of the Re equation is thus very large, and the inertial
forces override the viscous ones. At the scale of the
larva the opposite is true (see Vogel, 1994; and for a
more recent review, Weissburg et al., 2014).
In terms of the relative flow experienced by each of
these animals, this difference could not be more pro-
found. Flow at the scale of a dolphin (Re ~106) is cha-
otically turbulent: as it swims through the water, the
dolphin leaves a wake with swirling eddies behind
it (Vogel, 1994). In fact, the streamlined body of the
dolphin is well adapted to limit the size of the eddy-
filled wake since it increases drag, and thus impedes
forward progress. By contrast, the planula (Re ~1 or
less), due to its small size, does not create a turbu-
lent wake as it moves through the lagoon by ciliary
propulsion. Instead, the larva’s movement induces
strong local gradients in velocity that are character-
ized by adjacent layers of fluid slipping smoothly
past one another, with little mixing-type motions
(Figure 13.1). Less well studied are flow fields sur-
rounding organisms operating at intermediate
Reynolds numbers (Re in the 1–100 range), which is
relevant for many larger larvae (such as in fish, as-
as well as during certain burst swimming modes in
smaller larvae, such as in diving bivalves (e.g., Fuchs
et al., 2015). At such intermediate Re values, the flow
characteristics transition from viscous-dominated
to a domain where inertial forces are more promi-
nent, and the particular shapes of the larvae can
have an increasing effect on the flow characteristics
192   E v o l u t i o n a ry Ec o l o g y o f M a r i n e I n v e rt e b r at e La r va e

(A)

1m

flow field
(B) turbulent
eddies

0.3 mm

Figure 13.1 Turbulent and laminar flow at different Reynolds numbers (Re). As discussed in the text, a bottlenose dolphin (A) and a coral planula
larva (B) in the same habitat experience very different flow regimes, due to their vastly different sizes and corresponding Re. (A) At high Re, flow
(dashed black lines) even around a streamlined organism like a dolphin is broken up by turbulent eddies (dashed gray lines) in its wake, which
impedes forward progress. (B) At low Re, by contrast, typical flow around the larva is smooth, with no turbulent eddies. As such, any turbulent
intrusion (e.g., due to wave action) into the larval flow field would stand out against the background flow regime. Figure modeled after Weissburg
et al. (2014). Pocillopora damicornis planula photo by Bob Richmond. (see Plate 13)

compared to what is seen in lower Re conditions (see,
e.g., Koehl, 1995; McHenry et al., 2003).
The former example of eddies produced by flow
around larger and faster-moving objects is repre-
sentative of turbulent flow: parcels of water moving
in random directions on average relative to that of
the mean flow. The smooth flow around smaller ob-
jects is an example of laminar flow. Or, put another
way: at larger organism sizes and higher Re (as in
dolphins), flows are typically turbulent, whereas at
smaller organism sizes and smaller Re (as in planu-
lae), flows tend to be more laminar. Characterization
of laminar vs. turbulent flow regimes (and the tran-
sitions between them) based solely on Re should
be undertaken cautiously, as local geometries and
boundary conditions modulate such regime shifts
(see Denny, 1988; Vogel, 1994). In general, however,
flows at Re >105 tend to be turbulent, while flows at
Re <10 tend to be laminar.
For the purposes of this chapter, the character-
istics of the flow regime at the larval scale have
several implications. But to appreciate these impli-
cations, we first need to consider one more concept:
that of the boundary layer (reviewed in Nowell and
Jumars, 1984; Butman, 1987). Flow over smooth sur-
faces creates a boundary effect, where flow speed
decreases on average the nearer that flow is to the
surface. This principle holds across scales: it is why
taller wind turbines are more efficient (wind speed
is slower near the ground), and it is why so many
benthic filter feeders—from tube worms to barna-
cles to brittle stars—extend their feeding append-
ages above the substrate into the flow to increase
the rate of encounter with particles. Fast flow, es-
pecially over rough surfaces, creates turbulent vor-
tices that enhance transport of materials across the
boundary layer, and can expose organisms within
the boundary layer to instantaneous bursts in veloc-
ity (Nowell and Jumars, 1984).
Given these trends, a problem would emerge for
a larval-sized organism that relies on material ex-
change from the surrounding fluid, that operates at
low Re, and where surrounding flow is slow and
turbulent mixing is absent: such an organism itself
has a boundary layer. This boundary layer would
tend to interfere with its ability to interact with
more distant portions of its surrounding fluid envi-
ronment. For our larva, then, such limitations could
 F l u i d Dy n am i c s a n d S e n s o ry A s p e c t s o f La r va l S e t t l e m e n t    193
cause significant challenges: the unicellular algae
that our larva needs to eat, the oxygen that it needs
to absorb, and the cue molecules that it would use
to locate a suitable settlement location can be rap-
idly depleted adjacent to its body, and could take
considerable time to replenish via diffusion alone.
However, larvae have evolved mechanisms to
counter such limitations; in particular, behaviors
to ensure mixing across their boundary layers (see
Strathmann, 1995; Karp-Boss et al., 1996). In many
feeding larvae (see Pernet, this volume)—such as in
echinoderms, molluscs, and annelids—ciliary ac-
tion creates currents and locomotory movements
that replenish the water alongside the larval body
in a manner much more efficient than diffusion
alone (Gilpin et al., 2016). Likewise, larvae with
movable appendages—such as in arthropods, as-
cidian tadpoles, and possibly brachiolaria-stage sea
stars (Bashevkin et al., 2016)—can also break up the
boundary layers around their bodies, aiding in fluid
and material exchange.
A second implication of larval-scale flow, this one
more beneficial for our larva, relates to its entry into
the benthic boundary layer that forms over the sea-
floor. This boundary layer becomes relevant when
our larva attempts to settle at the end of its pelagic
life. In this context, solid surfaces within the bound-
ary layer—near which average flow speeds are
slower and (in the case of turbulent benthic bound-
ary layers) lulls in velocity occur with more regu-
larity—could afford precious refuge to our larva so
that it can attach strongly and reduce its chances
of being dislodged (Mullineaux and Butman 1991,
Crimaldi et al., 2002).
A third implication for our larva of the pre-
dominantly laminar flow that moves past its body
(Figure 13.1) is that any turbulent eddies that im-
pinge upon it, could, in a sense, “stand out” above
the typical smooth background flow regime. Such
turbulence could come from flow across rough sub-
strates, as mentioned earlier, from wind-generate­d
white-capping at the ocean surface, from the wate­r
movement created by potential predators, and
could also come from crashing waves in the surf
zone. The chaotic water movement produced from
each of these processes is translated down through
ever-smaller eddies to the smallest scales of fluid
motion where that turbulent energy is “dissipated”
(i.e., converted into heat) due to viscosity: more in-
tense turbulent flows result in higher levels of en-
ergy dissipation and a broader energy cascade that
sustains eddies of tinier size. Under the exception-
ally intense turbulence of the surf zones of rocky
shores (Gaylord et al., 2013), and to a somewhat
lesser extent in the other turbulence-generating
contexts mentioned earlier, the smallest turbulent
eddies operate at scales that are smaller than that
of a typical larva. As a consequence, such flow
structures could conceivably be sensed by larvae
as gradients in velocity across the dimensions of
their bodies (Jumars et al., 2009; Fuchs and Gerbi,
2016). Furthermore, because turbulence is so strong
in shoreline areas where waves break, the local
level of turbulence could be potentially utilized by
larvae as a reasonable—though not entirely diag-
nostic—proxy for their approach to benthic habitat
(Gaylor­d et al., 2013; Fuchs and Gerbi, 2016). This
ability would have profound implications for lar-
vae settling into nearshore locations, and we will
return to this point in some detail later.
A fourth implication of flow for our larva also
relates to boundary layers, but at much broader
scales. Unlike our previous examples of flow
around individual larvae, we here scale up to con-
sider flow that can affect the transport of entire
cohorts of larvae, thus possibly impacting connec-
tivity among populations. Adjacent to coastlines,
there is an area of slower alongshore flow known
as the coastal boundary layer (CBL). Several kil-
ometers offshore, depending on the bathymetry,
the alongshore (“free-stream”) flow is the fastest;
nearer to the coastline, the prevailing alongshore
flows decrease markedly due to the CBL. Larvae
released on the shoreline can also be retained near
to shore by reduced cross-shore mixing within the
CBL (Nickols et al., 2013), representing one possible
mechanism of the “larval retention” that data from
recent years (e.g., Morgan et al., 2009) has suggested
is much more common than previously thought.
In sum, understanding what it feels like to be a
larva involves understanding fluid dynamics at
multiple scales. As we will see, larvae are not al-
ways purely at the mercy of these flows. In some
situations, they can manipulate the local flow re-
gime to their advantage, and in others they can uti-
lize specific behaviors which increase the likelihood
194   E v o l u t i o n a ry Ec o l o g y o f M a r i n e I n v e rt e b r at e La r va e

that prevailing flows will carry them to suitable set-
tlement habitat: a critical need for every larva with
a benthic adult.
13.2.1  What Can a Larva Sense in Its Fluid
Environment and How Does It Do So?
The ocean is a rich sensory environment for the or-
ganisms within. Sound, gravity, pressure, organic
and inorganic chemicals, flow, light, salinity, pH,
and temperature are sensed by marine organisms
(Dusenbery, 1992; Young, 1995). In many cases, evi-
dence for the sensory response of marine organisms
to these cues, and the cellular mechanisms by which
they do so, come from studies on adults (and in some
cases their terrestrial relatives, such as insects and
nematodes). But whether larval forms in animals
predated the origin of their corresponding adult
body plans or the reverse (Strathmann, 1985), adults
and their larvae share the same genomes. As such, it
seems reasonable to hypothesize that selection could
efficiently lead to the acquisition of sensory modali-
ties in larvae that are known to occur in adults.
A full exploration of the sensory capabilities
and fluid dynamics of marine larvae—much less
so their adults—is beyond the scope of the current
review (see Crisp, 1974; Young, 1995; Yen, 2000;
Kingsford et al., 2002; Epifanio and Cohen, 2016;
Fuchs and Gerbi, 2016). Instead, we will focus on
well-studied examples where larvae utilize charac-
teristic features of the fluid environment to either
identify potential settlement locations or determine
their location relative to flow features that might
preferentially carry them to such locations. While
doing so, we will briefly describe some of the cel-
lular mechanisms that larvae use or might use to
monitor their external environment. We will con-
clude this section by speculating how these cellular
mechanisms might be integrated hierarchically not
only to maximize the probability of successful set-
tlement, but indeed to prevent the kinds of errors
that would often be fatal for larvae making what
is usually their irreversible decision to leave the
plankton (see Table 13.1 for definitions of terms).
Planktonic animals in general—and larvae
specificall­y—have been shown to have the ability

Table 13.1  Metamorphosis, Attachment, Settlement, Recruitment and “Continuous Settlement”.

Term Definition

Metamorphosis A more-or-less drastic morphological change between two multicellular phases (e.g., larva and juvenile),
often involving major changes in physiology and feeding. As such, the process can take from days to weeks to
complete, and can begin while the larva is still swimming (Chia, 1978).
Settlement The point at which the dispersive larval period ends in those marine organisms that undergo a shift between
the plankton and the benthos. As such, settlement is rapid (minutes to hours) and generally irreversible (though
there are a few exceptions to this; Richmond 1985). The notion that metamorphosis is distinct from settlement
is exemplified by crabs, whose metamorphosis occurs between the zooeal and megalopal stages, before
settlement occurs.
Attachment Typically the first step in the settlement process (though infaunal juveniles may burrow at settlement,
not attach). Care should be taken in using attachment as a proxy for settlement, since unlike settlement,
attachment can be and often is reversible. Larvae sampling the substrate might attach and release repeatedly
before finally settling.
Recruitment An ecological term describing the successful entry of a settled juvenile into a population of conspecifics. The
distinction between settlement and recruitment can be exemplified as follows: a larva that either settles in a
totally inappropriate location, far away from any conspecific adults—or a newly settled larva that is immediately
eaten—will never successfully recruit. A complication with the use of this term in the literature is that recruitment
is defined relative to a particular census time following settlement, which varies among studies.
Continuous A term proposed by Navarrete et al., (2015) to describe their observation of mussel “postlarvae” settling in
settlement-relocation one location and then tumbling along the substrate until they encounter their definitive adult (i.e., potential
recruitment) location.

Note. We here provide definitions of key terms involving the planktonic-benthic transition in marine invertebrates. We are compelled to do so due to the widely varied
(and often contradictory) definitions of these terms that have characterized the literature for over a hundred years right up to the present day.
 F l u i d Dy n am i c s a n d S e n s o ry A s p e c t s o f La r va l S e t t l e m e n t    195

to detect a wide range of environmental stimuli
(Figure 13.2). The majority of this evidence comes
from crustaceans (reviewed by Yen, 2000; Epifanio
and Cohen, 2016); however, several other phyla
have been studied as well (reviewed by Young,
1995; Kingsford et al., 2002). In many cases, these
cues have been hypothesized or demonstrated to
be employed by larvae to assist them in locating
settlement locations, and these are the ones we will
briefly describe here.
As mentioned earlier, most larvae swim too
slowly to be able to control their horizontal pos-
ition directly: their main strategy is to enter and exit
horizontal flows by adjusting their vertical position.
Larvae can detect their depth by sensing pressure
(Young, 1995), and can potentially tell if they are

(A) SENSORY INPUT-BEHAVIORAL OUTPUT GRAPHICAL MODEL

habitat (sensory inputs) possible behavioral outputs

larva’s developmental/ SETTLE

physiological state SWIM
ATTACH
FEED
(D) PICTORIAL KEY TO ICONS
RETRACT/
DEFEND DVM
salinity
SINK/DIVE
temperature
(B) IMMATURE LARVA OFFSHORE
relevant sensory inputs behavioral outputs
gravity
young larva with
immature nervous system SWIM
turbulence
FEED
olfaction
RETRACT/
DEFEND DVM
touch

light
(C) COMPETENT LARVA APPROACHING SHORE
relevant sensory inputs behavioral outputs
sound
competent larva with SETTLE
mature nervous system
ATTACH pressure

SINK/DIVE

Figure 13.2  Graphical model of multisensory inputs and behavioral outputs as they relate to settlement. Here we consider only those sensory
modalities and larval behaviors that have direct relevance to settlement itself, or that increase the likelihood that larvae get retained near or carried to
suitable settlement locales. (A) In the basic model, a generic larva (here a trochophore) can detect a wide range of sensory inputs—the combination
of those inputs can be thought of as a representation of the habitat that the larva is in. The physiological and developmental state of the larva can
be thought of as a lens (pictured in the center) through which the larva interprets these inputs. The larval nervous system (pictured at right) then
integrates those sensory inputs to elicit specific behavioral outputs. (B) An example of an immature larva (here, a sea star bipinnaria) detecting a
series of sensory cues that inform upon its depth, flow regime, and position relative to fronts and clines. Integration of those cues can provoke specific
swimming behaviors that could increase its likelihood of arriving at suitable settlement locations later in ontogeny. (C) An example of a mature,
competent larva (here a sea star brachiolaria, with a very well-developed juvenile rudiment) ready to settle in a favorable locale. Now, additional cues
can aid the larva in making the final phase of its journey to settlement on the seafloor, via specific behaviors such as sinking and attaching in flow.
(D) Pictorial key to the sensory icons shown in the left half of panels A–C.
196   E v o l u t i o n a ry Ec o l o g y o f M a r i n e I n v e rt e b r at e La r va e
sinking, stable, or rising by monitoring light in-
tensity, pressure, and their acceleration relative to
gravity vectors over time (Figure 13.2). Although
there is widespread behavioral evidence for these
sensory capabilities across phyla, direct physio-
logical and morphological evidence is more limited
(Kingsford et al., 2002; Epifanio and Cohen, 2016).
To adjust their vertical position in response to these
cues, larvae can either swim upward or downward,
sink passively if they are negatively buoyant, adjust
their buoyancy, or deploy or retract devices—like
threads or mucus—or appendages to either in-
crease or decrease resistance to sinking.
Using one or a combination of these mechanisms,
many larvae undergo daily migrations (so-called
diel vertical migrations; DVM) from depths up into
surface waters at night, at a time when visual pred-
ators are less of a problem, and prevailing winds
tend to blow toward the shore, and hence poten-
tially carry larvae there (reviewed in Queiroga and
Blanton, 2005). Other larvae undergo reverse DVM
into surface waters during the day, which would
tend to offer higher levels of their phytoplanktonic
food and a potential refuge from non-visual inver-
tebrate predators undergoing DVM, but could si-
multaneously expose larvae to visual predators and
potentially wind-driven offshore flows (Ohman
et al., 1983, Pennington and Emlet, 1996). It may
be that larvae undergoing reverse DVM are well
defended against visual predators, though we are
aware of no compelling evidence that tests this idea
in a comparative context. Larvae in estuaries are
known to undergo tidal migrations, which is best
studied in various crabs (reviewed in Queiroga
and Blanton, 2005). Depending on the species and
developmental stage, these migrations can either
retain or flush larvae from estuaries on ebb tides,
and carry them up-estuary on flood tides. Finally,
many larvae undergo so-called ontogenetic migra-
tions, in which earlier stages behave differently
than later ones (reviewed in Queiroga and Blanton,
2005). Such ontogenetic shifts may manifest as dis-
tinct tidal or DVM/reverse-DVM behaviors, or the
ontogenetic patterns might be consistent at a given
stage throughout the day or tidal cycle. The classic
ontogenetic migration is to sink at late stages, which
is thought to be an adaptation for approaching po-
tential settlement habitat (McCarthy et al., 2002).
It should be noted that late-stage larvae of many
taxa (e.g., echinoderms, gastropods, cladocerans,
brachiopods, bryozoans, crustaceans) acquire shells
or skeletal structures that are retained as these or-
ganisms enter the benthic juvenile stage. At some
point, such structures are likely (and in a few cases
have been shown) to make these larvae negatively
buoyant (Chia et al., 1984; but see Pennington and
Emlet, 1986). If so, we would argue that this is likely
an example of an exaptation (in the sense of Gould
and Vrba, 1982; often, but less precisely, called
“co-option”): the likely selective advantage of pre-
settlement skeletal development is protection from
predators, either in the benthos (e.g., newly settled
echinoderms) or in both the plankton and benthos
(e.g., gastropod larvae and corresponding juveniles).
The usefulness of such shells in helping larvae sink
would, in this conception, be a beneficial side effect.
After depth regulation, the next best-studied be-
havioral capacity of larvae is olfaction (Figure 13.2).
Detecting and responding to dissolved chemicals
could be useful to some larvae in feeding and avoid-
ing toxicants (see Corsi and Marques-Santos, this
volume; Yen, 2000; Zimmer and Butman, 2000), but
the most intensively researched olfactory behavior
of larvae is surely in identifying potential settle-
ment cues and deterrents (reviewed in Pawlik, 1992;
Young, 1995). In the cases where the existence and
activity of such dissolved cues have been demon-
strated, larval behavior in response to such cues can
be quite complex. For example, in the coral-grazin­g
sea slug Phestilla sibogae, entering and exiting
plumes of the dissolved coral-derived cue causes
larvae to sink and resume swimming, respectively
(Koehl et al., 2007). Interestingly, larval responses to
settlement-inducing olfactory cues are modulated
during ontogeny: they manifest more or less sud-
denly when a larva becomes “competent” to settle.
Indeed, response to settlement cues is the way com-
petence has traditionally been defined (see Table
13.1; Hodin et al., 2015).
The cellular and molecular mechanisms of ol-
faction are very well studied in fish and terres-
trial organisms, including flies, roundworms, and
mammals. The similarities in olfactory mecha-
nisms among these taxa (e.g., the involvement of
G-protei­n coupled receptors; Kaupp, 2010) make
it plausible that similar mechanisms are used by
 F l u i d Dy n am i c s a n d S e n s o ry A s p e c t s o f La r va l S e t t l e m e n t    197
aquatic organisms in general, and diverse larvae
at settlement in particular (Baxter and Morse, 1992;
Amador-Cano et al., 2006).
Whatever are the cellular mechanisms, the afore-
mentioned observation—across marine phyla—of
the sudden acquisition of competence and hence
responsiveness to olfactory settlement cues is most
consistent with the following scenario: olfactory
responsiveness is actively repressed in immature
and precompetent larvae. This is sensible, given
that de-repressing (or “unmasking”; Chia, 1978) an
intact olfactory signaling system is more efficient
than assembling the transcripts and proteins in-
volved de novo. Indeed, one potential global regu-
lator of settlement is nitric oxide/cyclic guanosine
monophosphate (NO/cGMP) signaling, which is
an active repressor of settlement in multiple phyla
(reviewed in Bishop and Biggers, 2014). Inhibition
of NO/cGMP signaling then represses the repres-
sor, thus uncovering the capacity of a larva to settle,
possibly in part via unmasking an already intact ol-
factory response. In sea urchins, histamine has been
identified as another such regulator of settlement
(Swanso­n, 2007; Sutherby et al., 2012), and, in the
context of fertilization, has been shown to activate
NO signaling (Leguia and Wessel, 2006).
A few notes of caution about larval olfaction are
warranted. First, the majority of studies of larval
responses to settlement inducers are undertaken
in dishes in the laboratory in still water. This is a
highly artificial situation (see, e.g., Metaxas, 2013),
and there is evidence that the olfactory system in-
tegrates with the larval response to water motion in
natural settings (Zimmer and Butman, 2000; Wood-
son et al., 2007). As such, more studies examining
olfactory and other settlement responses under re-
alistic flow conditions would surely be welcome.
Second, the levels of inducer that are needed to
stimulate settlement are often orders of magnitude
higher than concentrations measured in the field
(but see, e.g., Swanson et al., 2007). In such cases,
we should be circumspect in ascribing ecological
relevance to those cues and/or the mechanisms by
which larvae respond to them. Third, it has been
argued by several authors that dissolved chemical
cues are unlikely to be effective in most natural set-
tings (and especially in high-flow environments)
beyond a few centimeters from the source of the cue
(e.g., Denny and Shibata, 1989; Koehl et al., 2007).
Therefore, if larvae are responding to dissolved
cues, this is typically only going to be effective once
they have already managed to arrive extremely
close to a potential settlement habitat. This is one
of the reasons that we maintain the a priori expec-
tation that larvae also respond to other (non-olfac-
tory) types of cues that would be effective at scales
from centimeters to kilometers.
A third class of sensory modalities that larvae
could use to aid their successful settlement—and
one that could potentially act at much broader scales
than dissolved chemical cues—relates to water
movement. Larvae being carried in horizontal flows
of a given velocity would generally have no frame
of reference to detect that flow. By contrast, attached
larvae on the benthos are in some cases known to
react to different rates of flow going past them, and
this can influence their decision to either settle per-
manently in that location or to continue their search
(see Table 13.1; Figure 13.2). In addition, some larvae
are known to respond to turbulence, and because
turbulence and wave motions are often stronger in
shoreline regions, they could be useful indicators
to larvae attempting to return to nearshore settle-
ment habitats, as we discuss in more detail later.
The mechanisms by which larvae detect turbulence
and wave motions are unknown, and furthermore,
there are several aspects of water motion to which
larvae could be responding (Fuchs et al., 2015), in-
cluding translational acceleration, fluid rotation (via
statocysts), various gradients in velocity (via defor-
mation of cilia or activation of stretch receptors), or
some combination (Fuchs and Gerbi, 2016).
Additional sensory capabilities of larvae that
could aid their progression toward settlement are
the abilities to detect and monitor sound, touch,
temperature, and salinity (Figure 13.2). Response
to temperature (e.g., via transient receptor poten-
tial channels) and salinity (via sodium and potas-
sium channels) are widespread and likely generic
features of marine larvae. Behaviorally, responding
to temperature and salinity can be adaptive for lar-
vae entering or avoiding estuaries, or for entraining
into or exiting upwelling (colder, higher salinity) vs.
downwelling (warmer, lower salinity) flows, as we
will discuss briefly later. Mechanosensation is also
likely generic, even if it has not been widely studied.
198   E v o l u t i o n a ry Ec o l o g y o f M a r i n e I n v e rt e b r at e La r va e
Examples include some fouling organisms like co-
lonial ascidian larvae, as well as some non-animal
propagules (e.g., kelp spores), which will seemingly
settle when contacting virtually any solid surface
(Grosberg, 1981; Gaylord et al., 2006). Sound has
only recently been appreciated as an important cue
that larvae can use to recognize the overall features
of their adult habitat. In the last few years, compel-
ling evidence has been presented that some larval
reef fish and invertebrate larvae respond positively
to sound recordings of waves impacting coral reefs,
and late-stage oyster larvae likewise respond spe-
cifically to recordings made over an intact oyster
habitat, but not to control recordings from other
nearby locations (see Lillis et al., 2013).
We expect that larvae deciding where to irrevers-
ibly settle would draw on a rich and diverse array of
sensory information that could provide details about
the suitability of its potential adult habitat. In this
sense, our larva might be expected to use a process
salinity
temperature
gravity
turbulence
olfaction
touch
light
sound
pressure
0.1 mm 1 cm
Spatial Scale (relative to settlement location)
larval
akin to an analytic hierarchy process (Saaty, 2008), in
which larval experience could modulate the relative
importance and strength of certain cues. For exam-
ple, the presence of planktonic predators might lower
a larva’s threshold sensitivity to a dissolved chemical
inducer. More to the point, we might expect different
taxa in different types of habitats to prioritize certain
cues over others. For example, larvae settling in high-
energy habitats might prioritize turbulence cues over
chemical cues, those settling in mangrove estuaries
might prioritize salinity and temperature cues over
turbulence cues, and those settling on a specific spe-
cies of coral might prioritize a specific dissolved
chemical cue even in the temporary absence of char-
acteristic reef sounds. In particular, the hypothesized
hierarchically arranged signaling pathways might
be more-or-less organized in a fashion parallel to
the scale over which the cue acts (Figure 13.3). For
example, salinity, turbulence, and sound cues could
operate at a broader “habitat” scale of meters to
Figure 13.3 Relevant scales of sensory input
for settlement. As in Figure 13.2, we focus on
those sensory modalities that larvae might
use to identify suitable settlement locations
or that might trigger behaviors that would
increase the likelihood that they are retained
near or carried to such sites. For example, we
do not here consider the many sensory inputs
that larvae use (or likely use) for feeding. Note
the logarithmic distance scale on the x-axis,
denoting the larva’s distance from a suitable
settlement site. The four scale bins shown
below the x-axis (macro, meso, local, and larval)
mirror our treatment of these scales in the text.
Grayscale gradients indicate our approximation
of the relative importance of a given sensory
1m 100 m 10 km modality at a range of scales; dashed regions
indicate probable gaps in our knowledge of
the importance of specific sensory inputs for
local meso macro settlement at those scales.
 F l u i d Dy n am i c s a n d S e n s o ry A s p e c t s o f La r va l S e t t l e m e n t    199
kilometers, dissolved chemical cues and turbulent
flows over rough surfaces at millimeters to centim-
eters, and mechanosensory cues like surface topog-
raphy on sub-millimeter scales (Whalan et al., 2015).
Presently, the evidence for such hierarchical cue strat-
egies for settlement is limited (Kingsford et al., 2002),
and their arrangement by scale is pure speculation.
Still, it seems a fruitful area for future comparative
investigations.
In more general terms, the behavioral integration
of multiple sensory modalities (multisensory inte-
gration) is a concept that has received much atten-
tion in vertebrates and insects in recent years. It is,
in brief, the interaction or synergy among the dif-
ferent senses and the compilation of their informa-
tion content (Stein et al., 2014). In mammals, such
integration can typically be coordinated at multiple
levels of the nervous system (Stein et al., 2014). In
flies, the integration between olfactory, visual, and
mechanosensory input guides flight in three di-
mensions (Duistermars et al., 2009), but the under-
lying mechanisms of this integration remain to be
explored. Similarly, planktonic organisms maneu-
ver in a three-dimensional environment rich with
sensory cues, such as those outlined earlier (Wood-
son et al., 2007). Although larval nervous systems
are generally less centralized (and certainly less
well studied) than those of vertebrates and insects,
larvae from disparate phyla have concentrated neu-
ronal structures which in some cases are thought to
function in sensory integration during settlement
via the action of familiar neurotransmitters, which
act on single target cells, and neuromodulators,
which can have multiple targets (Hadfield, 2011;
Bishop and Biggers, 2014; Sutherby et al., 2012).
The small size of marine larvae makes functional
neurophysiological studies challenging. Neverthe-
less, modern methods examining the full comple-
ment of proteins and metabolic profiles (proteomics
and metabolomics, respectively) that are expressed
over time and under different conditions can and
are being employed in studies of larvae and their
metamorphoses (Song et al., 2016; Williams and
Carrier, this volume). Furthermore, targeted gene
manipulation methods have begun to be applied
to metamorphic stages of marine larvae as well
(Heylan­d et al., 2014). As such, the coming years
offer great promise for further elucidating the
detailed mechanisms of sensory perception in lar-
vae, and how—on a mechanistic level—settlement
decisions are made.
13.3  How Larvae Find Their Way Home:
Scales of Flow and Larval Behavior
We have considered the flow regimes that larvae ex-
perience as well as the sensory capacities that larvae
use to detect cues in their environment. Now we
move back out to larger scales to try to address the
following question: where do larvae go, and how
do they find their way back?
In recent decades, our understanding of where
larvae go and how they find their way back to shore
has grown enormously, with hundreds of papers
each year published on various aspects of this topic.
Nevertheless, there remain surprisingly fundamen-
tal disagreements in the field about the predomi-
nant oceanographic mechanisms that deliver larvae
to coastal habitats (reviewed in Pineda et al., 2010).
Are the numbers of larvae in the plankton (the so-
called larval supply) a good predictor of the number
of eventual settlers or not? Do larvae by and large
get advected far off shore by large-scale coastal pro-
cesses like upwelling, only to be returned to shore on
the occasional reversal events, or are most larvae ac-
tually retained very close to shore throughout their
entire larval life? (Morgan et al., 2009). If so, is this
pattern of nearshore retention due to active larval be-
haviors or passive responses to oceanographic forces
beyond their control? Do larvae concentrate in clinal
fronts, oceanographic eddies, or even flotsam as a
possible means of remaining close and/or transport-
ing to shore? Do larvae easily transit through the surf
zone, or do they remain in the waters just seaward of
the surf—like a sailing ship becalmed within sight of
port—with the surf zone as a semi-permeable barrier
(Rilov et al., 2008) to onshore delivery?
We will not attempt to offer definitive answers to
any of these questions. We instead defer to Pineda
and Reyns (this volume), who treat these questions
in much greater detail. For our purposes, we will
briefly describe some of these oceanographic mech-
anisms of transport as they relate to the likelihood
of larvae returning successfully to settlement lo-
cales, and we will provide some of the evidence for
and against them from specific case studies. Finally,
200   E v o l u t i o n a ry Ec o l o g y o f M a r i n e I n v e rt e b r at e La r va e
we will indicate where we think there are gaps in
the literature that could enrich our understanding
of how disparate larvae in discrete geographic or
oceanographic situations might enhance the prob-
ability of surviving to settle in the right place and
time. In so doing, we will follow our larva across
multiple scales from offshore waters back to the
nearshore, stressing the behavioral adjustments
that larvae could make to maximize the likelihood
of successful settlement in suitable habitats.
13.3.1  The Macro Scale: ~1–100 Kilometers
Much effort in recent decades has been directed at
developing increasingly realistic oceanographic cir-
culation models, and using them to predict the dis-
persal patterns of larvae, other propagules, and the
plankton in general. With respect to settlement and
the delivery of larvae to the shoreline, many of these
aforementioned models have made the optimistic
assumption that sampling of larvae at various dis-
tances from the shoreline will give direct insight
into their settlement on the shore. Or, to put it an-
other way: larval supply is the main driver dictating
settlement and ultimately recruitment. This view is
not without support (see Pineda et al., 2010; Pineda
and Reyns, this volume), but many additional stud-
ies have shown that the patterns are not quite so
simple, revealing situation-specific disconnects be-
tween larval supply and recruitment. For example,
pre-settlement processes like density dependence
of settlement itself and the association between
tidal height and settlement timing can profoundly
impact recruitment success and location (Grosberg
1981; 1982). Post-settlement processes such as sec-
ondary movement of settled larvae and heavy pre-
dation or environmental stress on settlers can also
break the simple connection between larval supply
and recruitment (reviewed in Pineda et al., 2010).
With respect to larval transport, the following
question seems deceptively simple, but is fraught
with complexity and controversy: where do larvae
go? One attractive scenario in upwelling-dominated
regions such as the eastern Pacific and west coast of
South Africa is that predictable wind-driven cur-
rents cause large-scale, coordinated movements
of larvae on a seasonal timescale. For example,
along the California coast, the prevailing California
Current flows from the north in the spring. Co-
riolis forces (driving Ekman transport) deflect the
prevailing currents offshore, with these displaced
waters subsequently replaced through upwelling
of deep, nutrient-rich waters up onto the continen-
tal shelf. These upwelled nutrients drive famous
seasonal plankton blooms, and Ekman transport is
predicted to send these plankton—and the larvae
therein—offshore. Occasional relaxations in the
prevailing winds cause temporary reversals in the
direction of the cross-shore currents, transporting
larvae shoreward.
Many observations have substantiated the pre-
dictions of this upwelling-relaxation hypothesis
(Roughgarden et al., 1991), finding enhanced set-
tlement and/or recruitment (see Table 13.1 for the
distinction) associated with relaxation conditions
and lower settlement/recruitment with upwelling
events. Nevertheless, there have also been numer-
ous studies in recent decades finding just the oppo-
site: no association between upwelling-relaxation
events and onshore recruitment (see Pineda et al.,
2010; Pineda and Reyns, this volume). Instead,
these and other studies have identified or proposed
additional oceanographic mechanisms for onshore
transport, including internal tides, fronts, wave-
driven flows, and even suspended materials like
flocs, flotsam, and surface slicks.
In some cases, the seemingly contradictory re-
sults just outlined could simply be due to differ-
ing sampling methodologies and intervals (Pineda
et al., 2010; Pineda and Reyns, this volume). Nev-
ertheless, there are a growing number of studies in
which the same species at different times or loca-
tions—or, more commonly, different species sam-
pled in the same location and time period—yield
conflicting support for relaxation-associated set-
tlement, even within the same publication. In one
recent example from an upwelling region of Brazil,
Mazzuco et al. (2015) found contrasting results for
mussel and barnacle settlement: barnacles settled
in association with times of predicted relaxation
events, whereas mussel settlement patterns showed
no such relationship.
One particular set of observations that runs
counter to a central prediction of the upwelling-
relaxation hypothesis is the surprising finding that
many larvae are not advected far offshore at all.
 F l u i d Dy n am i c s a n d S e n s o ry A s p e c t s o f La r va l S e t t l e m e n t    201
Instead, through various oceanographic and be-
havioral mechanisms (such as the coastal boundary
layer and vertical migration patterns, respectively,
as discussed earlier), larvae in multiple locations
and contexts appear to complete all or most of their
planktonic period very close to the shore at which
they were released. Thus a new paradigm has
emerged regarding so-called larval retention, which
would of course seem to make it far more likely that
larvae could make their way back to appropriate
settlement locations (presumably using the onshore
transport mechanisms previously referenced; see
Pineda et al., 2010; Pineda and Reyns, this volume).
One caveat with the majority of studies that have
addressed these issues to date is that they have gen-
erally taken a rather limited taxonomic focus, with
barnacles and other crustaceans, mussels, and fishes
as the subject of nearly all of the published work.
In certain respects, this focus is understandable.
First, most heavily studied taxa either have associ-
ated fisheries (crustaceans, mussels, fishes) or are
dominant in fouling communities (barnacles) as
adults. Second, studying larvae in the plankton can
be painstaking work, as larvae of many taxa appear
highly episodically in the plankton and in recruit-
ment events. By contrast, mussels, crabs, and bar-
nacles, in particular, are dominant shoreline and
estuarine invertebrates in many locales, and like-
wise dominate larval zooplankton assemblages in
corresponding offshore regions. Nevertheless, an ex-
panded taxonomic focus seems necessary to give a
more complete picture of where larvae go and when.
But research focused even within the well-stud-
ied taxonomic groups has yielded contradictory
findings, as mentioned earlier. It may be that the
taxonomically, spatially, and temporally diverse re-
sponses of larvae to a given set of oceanographic
conditions is consistent with the sweepstakes repro-
ductive success (SRS) hypothesis (see Hedgecoc­k
and Pudovkin, 2011). The SRS hypothesis posits
that larvae within populations demonstrate physi-
ological and behavioral diversity with respect to
their context-dependent growth and survival in the
plankton, and that successful recruitment can be
seen as a process akin to winning a sweepstakes,
where all of the right circumstances come together
for that improbable win. In a variable environment,
no one “strategy” would be consistently favored,
thus maintaining diversity in the way larvae re-
spond to oceanographic conditions. Such diversity
might be predicted to manifest in inconsistent geo-
graphic and temporal patterns of larval dispersal
mechanisms both among and within species.
Whether or not the SRS hypothesis is supported,
we conclude that biological and physical factors
that vary geographically, seasonally, tidally, daily,
and taxonomically can and do impact the relation-
ship between supply and recruitment, and there-
fore we are still seeking a holistic understanding of
these processes that might offer predictive power.
For example, we await a comprehensive meta-
analysis of the myriad published studies on larval
supply and recruitment to help illuminate the path
forward for the field.
13.3.2  The Meso Scale: <0.1–1 Kilometers
At this point, by whatever mechanism has brought
or retained our larva in the coastal zone, the diffi-
culties are far from over. A larva seeking shoreline
habitat still needs to traverse the surf zone to arrive
on shore, and once it does so, to recognize that it has
indeed arrived there. Is it possible that the surf zone
itself can provide such cues to larvae?
Recent and growing evidence suggests that the
answer is “yes” (Figure 13.3). First, larvae of mul-
tiple taxa have recently been shown to respond to
recordings of habitat sounds by increasing their
likelihood to settle (see Lillis et al., 2013). Likewise,
there may be chemical cues in some specific situ-
ations that are enhanced in broader-scale habitats,
like breakdown products of kelp in the nearshore,
and mangrove-derived chemicals in tropical es-
tuaries. Finally, the surf itself may be a cue. Stud-
ies by Gaylord et al. (2013) and Hodin et al. (2015)
show that sea urchin and sand dollar larvae with
nearshore adults exhibit enhanced settlement in re-
sponse to high levels of turbulence: specifically, lev-
els indicative of those seen under crashing waves.
Interestingly, turbulence is not a settlement “cue”
per se, since it does not directly induce larvae to
settle. Instead, exposure to turbulence primes these
larvae to settle: a greater proportion of turbulence-
exposed larvae will settle when subsequently
provided with a strong localized (i.e., chemical) set-
tlement inducer. Because such settlement inducers
202   E v o l u t i o n a ry Ec o l o g y o f M a r i n e I n v e rt e b r at e La r va e
are how competence is traditionally defined, these
results lead to the intriguing conclusion that tur-
bulence exposure actually causes larvae to become
competent to settle. Other studies demonstrate that
exposure to turbulence and waves can have another
seemingly advantageous impact on late-stage lar-
vae in some taxa: it causes them to either actively
or passively sink (see, e.g., Fuchs et al., 2015; but see
Wheeler et al., 2013). This behavior could provide
a selective advantage for nearshore-destined larvae
in the water column by increasing their chances of
contacting the seafloor (Denny and Shibata, 1989),
or at least arriving nearby.
13.3.3  The Local Scale: 10s of Centimeters–10s
of Meters
Our larva has now—through what was likely a
combination of luck and directed behaviors (such
as sinking in turbulence or association with surface
slicks)—arrived tantalizingly close to potential set-
tlement sites. What larval behaviors in association
with the properties of the fluid environment at these
local scales might make the difference between
reaching such benthic sites or being advected away?
Here our larva is approaching the benthos, and
much research effort has been directed at how flow
over complex substrates can impact the likelihood
of larvae—and non-animal propagules such as sea-
weed spores—entering the benthic boundary layer
and contacting the substrate. In one classic mod-
eling study, Denny and Shibata (1989) showed that
on wave-swept shores, turbulence alone (in the ab-
sence of directed larval behaviors) can carry larvae
efficiently and quickly to the substrate, and that
rapid (>1 mm•sec-1) sinking or downward swim-
ming of larvae can enhance this effect (Figur­e 13.3).
Likewise, multiple studies have shown that re-
alistic flow over complex surfaces (including
conspecific adults) can increase the likelihood of
larvae contacting the substrate, though in very
high flows, larvae may not be able to effectively at-
tach (Crimald­i et al., 2002). Manipulation of flow
dynamics in field settings show that increasing
flow over settlement plates can increase recruit-
ment rates (Palardy and Whitman, 2011), but it is
unclear if this result is due to settlement or post-
settlement processes.
In estuaries, salinity and temperature can
vary on the local scale, and competent larvae are
known to adjust their swimming behavior in re-
sponse, for example, to drops in salinity (Epifanio
and Cohe­n, 2016; Figure 13.3). Also, characteristic
habitat sounds (such as urchins scraping the sub-
strate; Radford et al., 2010) may indicate to larvae
that they have arrived close to preferred habitat.
Although most studies on larval sound perception
and settlement responses to date have considered
the meso scale to be the effective scale, stud-
ies contrasting more localized sound cues seem
warranted.
With respect to olfaction, as at the meso scale,
there are certain flow regimes in which one could
imagine larvae detecting and responding to local-
scale chemical signatures associated with specific
settlement sites, but definitive evidence is scant.
In some reef fish, larvae are known to settle in re-
sponse to the smell of conspecific adults, and in one
case (the humbug damselfish, Dascyllus aruanus),
previous eye contact of the adults with the juve-
niles changes the nature of the adult odor to make it
more inductive to larvae: these adults thus actively
recruit conspecific larvae (Roux et al., 2015).
There are other classes of potential local cues that
remain unexplored or largely so. For example, tide
pools are known to undergo dramatic diurnal fluc-
tuations in pH (Daniel and Boyden, 1975; Jellison
et al., 2016). The possibility that larvae settling in
the intertidal zone might respond positively to such
pH shifts has not to our knowledge been directly
addressed experimentally. Likewise, although tem-
perature can also fluctuate dramatically in inter-
tidal habitats, only a few studies have examined the
potential modulatory effect of temperature on other
classes of settlement cue (Pechenik, 1984).
13.3.4  The Larval Scale: <1 Millimeter–A Few
Centimeters
This is the spatial scale over which a larva experi-
ences its environment on short timescales (seconds
to minutes). At this point, our larva has successfully
been carried to benthic habitat that—based on cues
already received at broader scales—seems like a po-
tentially beneficial place to settle. But the final de-
cision about whether or not to irreversibly commit
 F l u i d Dy n am i c s a n d S e n s o ry A s p e c t s o f La r va l S e t t l e m e n t    203
to this settlement location could still be modulated
by the environment on very fine, even microscopic
scales (Figure 13.3).
Based on modeling of odor dispersion, this is
the scale at which dissolved chemical cues would
likely exist at sufficient concentrations to be de-
tected above background by—and thus elicit a be-
havioral response in—larvae (Koehl et al., 2007).
Furthermore, many chemical inducers of settle-
ment have been shown to be substrate-bound,
including what is perhaps the most generic chemi-
cal cue used by disparate marine larvae: chemi-
cals associated with surface biofilms (reviewed in
Hadfield, 2011); it is therefore only at the scale of
the larva that such cues are available to them. On
the other hand, chemical deterrents of settlement
(such as toxic compounds or the smell of a preda-
tor) could also be detected at the larval scale, and
may effectively inhibit the settlement process be-
fore it is too late (Woodin, 1991).
There are also several other classes of cues that
could inform our larva at the larval scale, includ-
ing light, fluid shear, and microtopography (Crisp,
1974). For example, certain coral and sponge lar-
vae preferentially settle on settlement tiles that
have holes drilled in them: tiles with 0.4 mm holes
(approximately the width of the larvae) show en-
hanced settlement relative to flat plates or those
with 0.7–1.0 mm holes (Whalan et al., 2015). Other
corals settle on the undersides of experimental set-
tlement surfaces, and clever manipulative experi-
ments demonstrated that this was due specifically
to the inhibitory nature of red (but not blue/green)
spectrum wavelengths striking the upper surfaces
(Strader et al., 2015). Finally, decisions on the larval
scale could be important for our larva to avoid be-
ing carried away by currents and strong, turbulent
flows during the initial stages of making its attach-
ment permanent (Reidenbach et al., 2009).
In sum, by the time our larva reaches the larval
scale in a potential settlement location, it likely has
already received broader-scale indicators of suitable
habitat. Depending on the resulting juvenile’s abil-
ity to move post-settlement—as well as its vulner-
ability to predation, grazing, or fouling soon after
settlement—cues at the larval scale might not only
provide valuable information to enhance growth
and survival (see Pechenik, this volume), but may
indeed be the last chance that our larva has to abort
the settlement process and seek purchase elsewhere.
13.4 Conclusion
These are exciting times for larval ecology and evo-
lution. Oceanographic models and survey meth-
ods are greatly improving our ability to determine
where larvae go and how they return to coastal
regions (see Pineda and Reyns, this volume). Bar-
coding methods will soon make it possible to obtain
rapid information on plankton assemblages that
previously required painstaking manual sorting (see
Marko and Hart, this volume). Laboratory methods
can challenge larvae with increasingly realistic flow
conditions and sensory experiences, coupled with
imaging techniques to visualize resulting behaviors.
And molecular methods are making it possible, in
almost any taxa, to interrogate and manipulate the
detailed cellular and neurophysiological mecha-
nisms underlying complex behaviors like larval set-
tlement (see Williams and Carrier, this volume).
Thus there is great promise for addressing issues
in larval ecology that have puzzled and inspired re-
searchers for many decades. For example, examin-
ing the sensory context of settlement behaviors in
realistic environmental conditions would give im-
portant insight into the evolution of contrasting set-
tlement strategies, how disparate larvae prioritize
diverse sensory inputs, and what this prioritization
means neurophysiologically. Elucidating the pre-
dominant migration pathways that larvae under-
take may inform on a second generation of marine
protected area design, one that more deliberately
couples critical nearshore locales to their offshore
“nursery” grounds (see Weissburg et al., 2014).
More generally, as we hope to have demonstrated,
settlement in marine invertebrates is an ideal subject
for integrative biology, combining oceanography,
fluid dynamics, sensory ecology, animal behavior,
and developmental, cellular, and molecular biol-
ogy. Furthermore, the likely independent origins of
larvae in diverse phyla, as well as the sometimes
extreme contrasts in settlement locales even among
closely related species, offer abundant compara-
tive material for detailed evolutionary studies into
this key life stage transition for animals and non-
animals alike.
204   E v o l u t i o n a ry Ec o l o g y o f M a r i n e I n v e rt e b r at e La r va e
13.5 Summary
1. Flow at the larval scale is fundamentally differ-
ent than flow at the scale of larger organisms
(such as ourselves): the former is dominated by
viscous forces, the latter by inertial forces.
2. Larvae are not merely “passive particles”: al-
though many larvae cannot make headway
against strong oceanic currents, they can adjust
their vertical position in the water column, and
this can have profound influences on where lar-
vae go and how they can find their way back to-
ward suitable settlement locations.
3. Larvae engage a wide range of sensory modal-
ities in order to ascertain and advantageously
adjust their vertical position in the water column,
as well as their proximity to potential settlement
locations; an overlapping but unique set of sen-
sory inputs characterizes their habitat at differ-
ent scales.
4. We hypothesize that larvae integrate these vari-
ous cues in a hierarchical fashion; disparate taxa
in different contexts will demonstrate alternative
arrangements and strengths of the cues that they
use to locate settlement sites.
5. Modern techniques offer the promise of answer-
ing several longstanding, vexing questions in lar-
val ecology that bear on topics as wide-ranging
as mechanisms of metamorphosis, life history
evolution, conservation biology, and the origins
(and losses) of larvae.
Acknowledgments
We are grateful to our institutions and Hopkins Ma-
rine Station of Stanford University for facilities and
library access, and Bob Podolsky, Richard Strath-
mann, and Bruno Pernet for helpful discussions.
The comments of Richard Strathmann, Steven
Morgan, the editors, and two anonymous review-
ers greatly improved the manuscript. Although we
were not able to include many relevant references,
we direct the interested reader to our Online Sup-
plementary Material, where we provide an anno-
tated list of scores of additional references. This
work was supported by National Science Foun-
dation grants (OCE-1356966, OCE-1357077 and
OCE-1357033 to B.G., M.C.F., J.H., and Christopher
J. Lowe), an award under the Federal Coastal Zone
Management Act, administered by the National
Oceanic and Atmospheric Administration’s Office
for Coastal Management (to San Francisco State
University), and by an NSERC Discovery Grant
(400230 to A.H.).
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 CH A PT ER 14
Latent Effects: Surprising
Consequences of Embryonic
and Larval Experience on Life
after Metamorphosis
Jan A. Pechenik
14.1  Introduction and Definitions
Metamorphosis is typically a time of rapid
­transition—in body plan, physiology, lifestyle, and
habitat. An old world and lifestyle are left behind,
and a new world is opened. That is certainly how
metamorphosis was presented to me as a student:
a new life, a new beginning. As Lahiri (2015, p. 35)
wrote, “Metamorphosis is a process that is both
violent and regenerative, a death and a birth.” But,
as it turns out, there is growing recognition of the
links between experiences in one life history stage
and phenotypes exhibited in later stages (Marshall
et  al., 2003; Allen et  al., 2006; Padilla and Miner,
2006; Marshall and Morgan, 2011; Flores et  al.,
2013; Burggren, 2014; Burton and Metcalfe, 2014;
Ross et al., 2016). In some cases, for example, juve-
nile growth, survival, or other performance criteria
are relatively poor following short-term, sublethal
stresses experienced by larvae, even when the lar-
vae seem to have completely recovered from the
stress before metamorphosing (e.g., Pechenik, 2002;
Chiu et al., 2008). Additional examples of such “la-
tent effects” (Pechenik, 2006) are given in the ma-
terial that follows. There is growing evidence that
such effects are not simply artifacts of laboratory
rearing: latent effects have now been documented
for juveniles transplanted into the field, for a num-
ber of species in a number of taxonomic groups
(e.g., bryozoans: Ng and Keough, 2003; ascidians:
Pechenik, J. A., Latent effects: surprising consequences of embryonic and larval experience on life after metamorphosis.
In. Evolutionary Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland:
Oxford University Press (2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0014
Marshall et  al., 2003; barnacles: Emlet and Sadro,
2006; Thiyagarajan et  al., 2007; gastropods: Chiu
et al., 2007; bivalves: Hettinger et al., 2013a). Such
responses are of growing importance as the aqua-
culture industry expands, and as the quality of life
in the marine environment changes due to global
warming, ocean acidification, changing rainfall pat-
terns, and increased pollutant input. They may also
help to explain at least some of the variation seen in
growth rates among individuals of the same species
recruiting to the same locations, and give us new
incentive to determine the degree to which parental
care (e.g., brooding) can protect against such im-
pacts. As discussed later, brooding can sometimes
expose developing embryos to some environmental
stresses even while protecting them from exposure
to others (Chaparro et al., 2014; Segura et al., 2014).
The delayed responses just referred to have been
defined as “latent effects” because they show up later
in development, after metamorphosis (reviewed by
Pechenik, 2006). To the extent that stress-induced
poor performance in the larval stage simply car-
ries over to poor performance of the same sort (e.g.,
growth rates) in juveniles, these can also be consid-
ered to be “carryover effects” (Marshall et al., 2003;
Wendt and Johnson, 2006; Hettinge­r et  al., 2012),
although such terminology has been used for a
very different purpose both in the medical litera-
ture (reviewed by Pechenik, 2006) and the ecologi-
cal literature (reviewed by O’Connor et  al., 2014).
 C o n s e q u e n c e s o f E m b ryo n i c a n d L a r val E x p e r i e n c e    209

But what do we call the effects of stress that cause
reduced larval growth throughout the larval stage
and also result in reduced juvenile growth under
ideal conditions after metamorphosis, when meta-
morphosis involves a pronounced switch in the
food-collecting machinery—for example, when
the food-collecting velum of a gastropod larva is
replaced after metamorphosis by a food-collecting
gill, or a food-collecting radula? If the reduced
juvenile growth rates are caused by flawed or
slowed development of the new feeding appara-
tus, I would refer to these as latent effects as well.
If they are caused by flaws (e.g., in digestive physi-
ology) initiated in the larval stage and maintained
in the juvenile, I would refer to those as carryover
effects. In all of these cases, the responses could
also be called “legacy effects” (Padilla and Miner,
2006), as they represent cases of juvenile or adult
performance being influenced by experiences in
previous stages of the life history: the juveniles or
adults bear the legacy of earlier experience. The
term “legacy effects” could also apply to the effects
of maternal or paternal experience—often relating
to nutritional stress or to the presence of predators
or predator cues—on offspring performance (Alle­n
et  al., 2006; Burggre­ n, 2014; Soubry, 2015; Long
et  al., 2016; Ross et  al., 2016); studies concerning
such transgenerational effect­s (e.g., Marshall, 2008)
will not be discussed in this review.
This review primarily concerns latent e­ffects—
effect­s that show up after metamorphosis but that
result from stresses experienced before metamor-
phosis—and focuses exclusively on the marine in-
vertebrate literature. Related findings have also been
reported for birds, lizards, amphibians, insects, and
even humans (Bresnahan and Susser, 2007; Stamper
et al., 2009; Anderson et al., 2013; Zambonino-Infante
et  al., 2013; Jonsson and Jonsson, 2014; Bouchard
et al., 2015; Whiteside et al., 2015; Wang et al., 2016a).
I reviewed the literature on this topic about ten
years ago (Pechenik, 2006). Of the 45 entries listed
in that paper’s Table 1, nearly one-third (14 studies)
concerned vertebrates, and two concerned insects.
For the remaining studies on marine invertebrates,
15  concerned latent effects resulting from delayed
metamorphosis, ten concerned the post-metamor-
phic effects of reduced food levels experienced dur-
ing larval development, three concerned the effects of
pollutants, and one paper each concerned the effects
of reduced salinity and differences in food quality.
The present review includes studies on the effects of
several newly tested stressors, including ocean acidi-
fication, hypoxi­a, and dietary quality (Table  14.1).
I also now include several recent studies on the latent

Table 14.1  Summary of Recent Research on Latent Effects in Marine Invertebrates.

Taxon Species Stress Parameter measured Latent effect Reference

Cnidaria Porites astreoides H2O2 oxidative stress, 24 h Survival in field No Ross et al., 2012
Thermal stress: 3°C, 24 h Yes
Montastraea faveolata Low salinity stress Juvenile survival Yes Vermeij et al., 2006
Acropora tenuis Delayed metamorphosis: 2, Colony development, No Graham et al., 2013
4, 6 wk survival
Gastropoda Crepidula fornicata Phytoplankton species Juvenile growth Sometimes Pechenik and Tyrell, 2015
Food level Juvenile growth Yes
Crepidula fornicata Low salinity: 12, 24, 48 h Juvenile growth No Diederich et al., 2011
Crepidula fornicata Low salinity and thermal Juvenile growth Sometimes Bashevkin and
stress Pechenik, 2015
Crepidula onyx Low salinity: 12, 24, 48 h Juvenile growth No Diederich et al., 2011
Food level Juvenile growth Yes Chiu et al., 2007
Juvenile survival Sometimes
Juvenile filtration rates Yes
(continued)
210   E v o l u t i o n a ry Ec o l o g y o f Ma r i n e I n v e rt e b r at e L a r va e

Table 14.1  (Continued)

Taxon Species Stress Parameter measured Latent effect Reference

Juvenile carbon assimilation No

Short-term starvation Juvenile survival No Chiu et al., 2008
Juvenile growth Yes
Hypoxia at 2 food levels, Juvenile growth, filtration Yes (low food) Li and Chiu, 2013
8–10 d rates
Crepipatella dilatata* Hypoxia: 24, 48, 72 h Juvenile survival Yes Segura et al., 2014
Juvenile growth Yes
Crepipatella peruviana Low salinity: 12, 24, 48 h No
Crepipatella peruviana Low salinity: 6 h Size at metamorphosis Yes Montory et al., 2016
Juvenile growth No
Feeding, respiration rates No

Bivalvia Haliotis diversicolor Delayed metamorphosis, Juvenile growth, survival Yes Onitsuka et al., 2010
3–11 d
Polychaeta Capitella teleta* Hypoxia: 24, 36, 48, 72, Juvenile survival No, negatively Pechenik et al., 2016
or 96 h
Juvenile growth Possible positive
Salinity: 24, 48, or 96 h Juvenile survival Sometimes
Hydroides dianthus Food level, 10 d Juvenile mortality Yes Allen and Marshall, 2010
Juvenile growth Yes
Crustacea Balanus amphitrite Delayed metamorphosis, Juvenile growth Yes Thiyagarajan et al., 2007
0–3 d
Low salinity, 0–3 d Yes

Balanus glandula Food level
Carcinus maenas Differences in presumed food Juvenile survival, growth
levels experienced in the field
Echinodermata Echinometra (4 spp.) Delayed metamorphosis, 5
months
Urochordata Ciona intestinalis Delayed metamorphosis, 3 d
Delayed metamorphosis, 7 d
Molgula socialis Delayed metamorphosis, 2 d
Ascidiella aspersa Delayed metamorphosis,
3–4 d
Juvenile growth Some Emlet and Sadro, 2006
Some
Yes Rey et al., 2016
Juvenile growth, survival Yes Rahman et al., 2014
Post-metamorphic growth No Jacobs et al., 2008
Yes
Post-metamorphic growth Yes (increased) Jacobs et al., 2008
Post-metamorphic growth No Jacobs et al., 2008

*indicates studies in which brooded embryos, rather than larvae, were stressed

impact of stresses experienced by brooded embryos 14.2  Latent Effects of Exposure

(Table 14.1). Interest in latent effects has been grow-
ing. Perhaps even more importantly, some marine
invertebrates may make especially good models for
understanding the mechanisms through which latent
effects are mediated.
to Toxicants
Only two papers concerning latent effects result-
ing from pollution stress were cited in my previ-
ous review, and surprisingly little additional work
 C o n s e q u e n c e s o f E m b ryo n i c a n d L a r val E x p e r i e n c e    211
has been published for marine invertebrates on
that topic in the intervening ten years. Expos-
ing young larvae of the oyster Crassostrea gigas to
the widespread aquatic pollutant nonylphenol at
the initial low concentrations of 1 µg l-1 and 100
µg l-1 impacted adult reproductive biology many
months later, and in unexpected ways (Nice et al.,
2003). Seven-day-old larvae were exposed to the
pollutant for 48 h and then returned to control
conditions for the rest of their larval development.
The spat were then reared to adulthood under con-
trol conditions and examined ten months later, af-
ter which eggs and sperm were mixed to obtain a
next generation of larvae. Although there was no
measureable impact of previous larval exposure at
either concentration on juvenile growth rates over
the 10-month rearing period, there was a marked
increase in the proportion of hermaphrodites in
the population, and a significant and dramatic re-
duction in larval survival in the next generation
48 h after successful fertilization, even when one
of the parents included in a cross was a control
individual. On the other hand, exposing larvae
of the Mediterranean sponges Crambe crambe and
Scopalina lophyropoda to cadmium (5 µg l-1) or cop-
per (30 µg l-1) for up to seven days had no effect
on juvenile survival over the following six months
(Cebrian and Uriz, 2007).
More studies seem warranted, using a greater
variety of toxicants and a greater range of species
and taking into consideration the impact of other
stressors experienced during and after larval de-
velopment (Byrne and Przeslawski, 2013). Future
studies should also investigate the potential for la-
tent effects caused by the ingestion of microplastics
during larval development (Galloway and Lewis,
2016). In addition, not all toxicants are produced or
released by humans; the frequency of harmful al-
gal blooms seems to be increasing in many parts of
the world (Gilbert et al., 2014). Future studies could
examine the potential impact of exposing larvae
to sublethal concentrations of algal toxins on post-
metamorphic fitness.
14.3  Latent Effects following Hypoxia
As hypoxic conditions become increasingly wide-
spread in coastal marine environments around the
world, due to climate change and the increasing
output of organic wastes and agricultural nutri-
ents (Wu, 2002; Zhang et  al., 2013), interest in the
potential consequences of spreading hypoxic stress
continues to grow. For the calyptraeid gastropod
Crepidula onyx, exposing well-fed (2 × 105 cells ml-
1
) larvae to oxygen levels as low as 2 mg O2 l-1 for
eight to ten days had no significant impact on the
growth rates of field-transplanted juveniles (Li and
Chiu, 2013). However, when larvae of this species
were exposed to the same level of low-oxygen stress
but at reduced food levels (1 × 105 cells ml-1), subse-
quent juvenile growth rates and filtration rates were
significantly reduced compared with those exhib-
ited by control individuals, even though there were
no significant differences in the size or lipid content
of juveniles from the different treatment groups at
metamorphosis (Li and Chiu, 2013). Thus latent ef-
fects were observed following the exposure of larvae
to the combined stresses of low food and reduced
oxygen, but were not observed if the larvae had
been well-fed when exposed to the same levels of
hypoxic stress.
Although most studies documenting effects
that cross the metamorphic boundary concern
stresses experienced during larval life, several re-
searchers have recently begun examining the post-­
metamorphic impact of environmental stresses
experienced by encapsulated or brooded embryos
(e.g., hypoxia and increased ammonia accumu-
lation in brood chambers). Although brooding
is generally viewed as protective (Gillespie and
McClintoc­ k, 2007; reviewed by Pechenik, 1979;
Noisett­e et al., 2014), brooding can, in some situa-
tions, expose developing embryos to stresses they
would otherwise avoid by being free-living, some-
thing that has only recently been considered for
marine invertebrates (Chaparro et  al., 2009; 2014;
Segura et al., 2014).
Calyptraeid gastropods, for example, brood their
encapsulated embryos within the female mantle cav-
ity for at least several weeks before releasing larvae or
metamorphosed juveniles, depending on the species,
into the surrounding seawater (Colli­n, 2004). Nor-
mally the females bring seawater into the mantle cav-
ity and flush out older seawater and wastes through
the actions of gill cilia (reviewed by Chaparr­o et al.,
2002; Shumway et al., 2014). However, in the Quem-
pillén River estuary in southern Chile, ambient salin-
ity frequently drops below a level (22 psu) at which
212   E v o l u t i o n a ry Ec o l o g y o f Ma r i n e I n v e rt e b r at e L a r va e
females completely seal themselves off from the am-
bient environment (Chaparro et al., 2009); such isola-
tion from the surroundings occurs frequently during
peak reproductive activity at this site, and can last for
at least three days at a time (Chaparro et al., 2009).
Although such isolation by the mother protects de-
veloping embryos from exposure to severe salinity
stress (Chaparr­o et al., 2009), it soon exposes them to
greatly reduced oxygen levels. Indeed, with only one
to two milliliters of fluid in the adult mantle cavity,
oxygen levels typically fall below 1 mg O2 l-1 within a
few hours of isolation from the external environment
(Chaparro et  al., 2014). When brooded embryos of
the direct-developing gastropod Crepipatella dilatata
were exposed to oxygen levels <1 mg O2 l-1 for 24 to
72 h, there was no effect on the average size at which
juveniles emerged from the female, but there was a
pronounced decline in subsequent juvenile growth
rates under control conditions over the subsequent
four-week monitoring period, regardless of how ad-
vanced the embryos had been when exposed to the
stress; indeed, final mean juvenile shell length was
approximately 20% less than that of control indi-
viduals for those exposed to severe hypoxia for even
just 24 h as brooded embryos, and more than 50%
less for those that had been exposed to hypoxia for
72 h (Segur­a et al., 2014). A single 24 h exposure of
juveniles to hypoxic stress was also enough to signifi-
cantly reduce juvenile survival over the first 30 days
after their release from the mothers.
In contrast, exposing brooding females of the
­deposit-feeding polychaete Capitella teleta to oxygen
concentrations <1 mg O2 l-1 for up to 96 h (the maxi-
mum period tested) had no effect on mean numbers
of larvae emerging from brood tubes, mean time to
emergence, mean juvenile survival or growth rates,
or on subsequent fecundity or time to reproduc-
tive activity in the next generation (Pechenik et al.,
2016). Whether these results reflect the benefits of
brood protection or simply a high embryonic toler-
ance to these stresses is an open question.
14.4  Latent Effects from Food
and Nutrient Limitation
A number of papers have considered how food
availability in the larval stage might impact post-
metamorphic performance. For the calyptraeid
gastropod Crepidula onyx, for example, starving
larvae of certain ages for two days or rearing the
larvae to metamorphic competence at a low phy-
toplankton concentration (1 × 104 cells ml-1 instead
of 20 × 104 cells ml-1) both resulted in significantly
reduced juvenile growth: final juvenile shell sizes
were up to 54% smaller than those of control indi-
viduals that had been reared as larvae at the higher
food concentration, for both laboratory-reared
and field-transplante­d juveniles (Chiu et  al., 2007;
2008). In neither study did larval nutritional expe-
rience influence subsequent juvenile survival in the
laboratory, but feeding the larvae at the lower food
concentration did result in higher mortality for field-
transported juveniles (Chiu et al., 2007).
Similarly, rearing nauplius larvae of the barna-
cle Balanus glandula on a low concentration of the
diatom Skeletonema costatum decreased mean juve-
nile growth rates and also decreased the survival of
juveniles that had been transplanted to field sites
for at least the first two to six days after metamor-
phosis, when most of the mortality occurred (Emlet
and Sadro, 2006). Field-transplanted juveniles of
the tube-dwelling polychaete Hydroides diramphus
also showed reduced survival if the larvae had
been reared at lower food concentrations (Allen and
Marshall, 2010).
When larvae of the gastropod Crepidula fornicata
were reared at a low concentration of the flagellate
Isochrysis galbana (clone T-ISO, 1 × 104 cells ml-1) for
two or four days early in development, no latent
effects on subsequent post-metamorphic growth
rates were detected (Pechenik et  al., 2002). How-
ever, when larvae of this species were reared at this
same low food concentration for their entire larval
development, juveniles fed ad libitum as their re-
ward for metamorphosing grew about 37% more
slowly than control juveniles for the first three days
after metamorphosis, and nearly 45% more slowly
for the first six days after metamorphosis (Pechenik
and Tyrell, 2015).
The post-metamorphic impact of differences in
nutritional quality during larval development has
recently been explored for marine invertebrates,
apparently for the first time. In contrast to either
starving larvae for a period of time or rearing them
at a greatly reduced food concentration, Pechenik
and Tyrell (2015) reared the larvae of Crepidula
 C o n s e q u e n c e s o f E m b ryo n i c a n d L a r val E x p e r i e n c e    213
fornicata at high concentrations of three different
algal diets that generally supported good survival
but at different rates of growth: Isochrysis galbana,
(clone T-ISO; the diet producing the fastest larval
growth), Dunaliella tertiolecta (clone DUN), and
Pavlova lutheri (clone MONO). Following metamor-
phosis, all juveniles were reared on the diet produc-
ing the fastest growth, T-ISO. In two experiments,
larvae reared on one of the nutritionally poorer
diets resulted in substantially reduced mean post-
metamorphic growth rates. However, in other ex-
periments larval diet had no effect on mean juvenile
growth rates despite substantial differences in the
mean growth rates of larvae that had been reared
on the different diets. Implications of these stud-
ies seem especially intriguing, as climate change
and ocean acidification appear to be shifting both
phytoplankton species ranges and the nutritional
composition of individual phytoplankton species
(Hinga, 2002; Tortell et al., 2002; Hayes et al., 2005;
Rossoll et al., 2012; Hettinger et al., 2013b; Leu et al.,
2013; Wynne-Edwards et al., 2014).
It should be noted that even though some re-
searchers have transplanted juveniles to field sites
for further monitoring once metamorphosis has
occurre­ d (Emlet and Sadro, 2006; Thiyagarajan
et  al., 2007; Allen and Marshall, 2010; Diederich
et  al., 2011; Graham et  al., 2013; Hettinger et  al.,
2013a), the larvae have always been stressed in
the laboratory. Typically, feeding larvae are reared
on one or several different phytoplankton species.
Even high food concentrations of phytoplankton
species producing the best growth in the laboratory
may be imposing stress on larvae. In the field, lar-
vae will probably have access to many dozens of
phytoplankton species at any one time. Thus, what
appears to be an ideal diet in the laboratory may
in fact be suboptimal, compared to what larvae are
ingesting in nature. Indeed, larvae of Crepidula for-
nicata collected from the field typically grew much
faster in the laboratory over the next several days
than larvae that had hatched in the laboratory and
were then reared on the same unigal diet (Isochrysi­s
galbana, clone T-ISO) (Pechenik and Levin­e, 2007),
suggesting that the field-collected larvae were
somehow healthier than those obtained and reared
in the laboratory. Hatchery operators might try
rearing juveniles obtained from field-collected
larvae. It would also be interesting to examine the
consequences of nutritional stress for larvae reared
under field conditions; this might be feasible for
­larvae that are large at hatching, such as those of
Crepidula fornicata, which typically hatch at shell
lengths between about 400 to 450 µm (Pechenik,
1984; ­Pechenik et  al., 1996a; b; Henry et  al., 2010;
Pecheni­k and Tyrell, 2015). Along these lines, larvae
of the green crab Carcinus maenas were collected in
the field during upwelling events of different mag-
nitudes, ­allowed to metamorphose in the labora-
tory, and were then reared in the laboratory through
the fifth juvenile instar (Rey et al., 2016). Larvae that
seem to have experienced better food conditions in
the field showed higher juvenile survival and better
juvenile growth in the lab than those that had been
collected during other larval supply events and
reared under the same conditions (Rey et al., 2016).
14.5  Latent Impact of Salinity Stress
In a study by Thiyagarajan et  al. (2007), cyprids
of the barnacle Balanus amphitrite were exposed to
the low salinity of ten psu for 24 h before the lar-
vae were allowed to metamorphose. Even though
there was no measureable impact of the treatment
on cyprid lipid reserves, subsequent mean juvenile
growth rates were up to 70% lower than those of
control animals for at least the next five days, in
both the laboratory and the field. Exposing larvae to
reduced salinities also impacted post-metamorphic
development for the South American suspension-
feeding gastropod Crepipatella peruviana (Montory
et al., 2016). Veligers of that species were exposed in
the laboratory to salinities as low as 15 psu for only
six hours, matching the salinity reductions recorded
from local tide pools during a heavy rain during a
single low tide. Clearance rates were reduced for
the first five days after metamorphosis for individu-
als exposed to 15 psu as larvae, and mean juvenile
shell sizes were still significantly smaller than those
of control individuals ten days later. Those individ-
uals that had been exposed to a salinity of 15 or 20
psu as larvae also showed an approximately 20%
decrease in subsequent juvenile survival.
In contrast, Diederich et al. (2011) exposed larvae
of three calyptraeid gastropods (Crepidula fornicata,
Crepidula onyx, and Crepipatella peruviana [formally,
214   E v o l u t i o n a ry Ec o l o g y o f Ma r i n e I n v e rt e b r at e L a r va e
Crepipatella fecunda]) to salinities of 30 (control), 20,
15, or 12 psu, for 24 or 48 hours, and found no ef-
fects on subsequent mean juvenile growth rates for
any of the three species, even when larval growth
rates failed to recover to control levels after the
stress period had ended. There was no juvenile
mortality for any of the three species tested in that
study. Longer exposures of larvae to reduced salin-
ity did produce significant latent effects on juvenile
growth in Crepidula fornicata, however, in four of
the six experiments conducted by Bashevkin and
Pechenik (2015), and the effects were generally
negative. In one of their experiments, however, the
effect was positive: juveniles grew faster than con-
trols at reduced salinity (20 psu) if the larvae had
been reared at that same reduced salinity. Again,
larval and juvenile mortality were both very low in
those studies.
When brooding females of the deposit-feeding
polychaete Capitella teleta were exposed to salini-
ties as low as ten psu for 24, 48, or 96 h, dramati-
cally fewer larvae subsequently emerged from the
brood tubes over the following weeks, suggesting
substantial pre-hatching mortality resulting from
exposures as short as 24 h at salinities as high as 20
psu (Pechenik et al., 2016). Surprisingly, however,
all but the longest duration of low-salinity expo-
sure (and, thus, the most intense level of salinity
stress) imposed on brooding females had no sub-
sequent detrimental effects on juvenile survival or
juvenile growth, for those larvae that did survive
to metamorphose. Indeed, exposing brooding
females to a salinity of 25 psu for 96 hours sig-
nificantly increased post-metamorphic survival of
the larvae that were released. In contrast, stress-
ing the larvae of C. teleta at low salinities for as
little as 24 h significantly reduced post-settlement
survival and juvenile growth rates in a previous
study (Pechenik et al., 2001).
Exposing brooding females of the direct-develop-
ing gastropod Crepipatella dilatata to a reduced am-
bient salinity of ten psu for 72 h resulted in reduced
juvenile oxygen consumption rates, clearance rates,
and growth rates that were still apparent four weeks
after metamorphosis (Chaparro et al., 2014). How-
ever, these latent effects were most likely due to the
impact of mothers sealing their mantle cavities off
from the surrounding seawater, with consequent
changes in mantle cavity oxygen levels, ammonia
concentrations, and pH, rather than to a direct effect
of reduced salinity.
14.6  Latent Effects of Delayed
Metamorphosis
Planktonic marine invertebrate larvae must typi-
cally develop for a time, usually ranging from hours
to weeks, before becoming competent to undergo
metamorphosis (reviewed by Pechenik, 1990). Once
competent, metamorphosis can be delayed for
hours, days, weeks, or months in different species
in the absence of appropriate environmental cues,
or in the presence of inhibitory factors (Pecheni­k,
1990; Marshall et al., 2003). Nearly half the studies
concerning latent effects that were reviewed previ-
ously (Pechenik, 2006) dealt with effects of delayed
metamorphosis. The following five more recent
papers add to the previous theme that delayed
metamorphosis produces substantial latent effects
in some species but not others, and that effects are
sometimes seen among the progeny of some par-
ents but not among the progeny of other parents
within the same species.
Delaying metamorphosis of the abalone Hali-
otis diversicolor (Onitsuka et  al., 2010) for up to 12
or 17 days and of four sea urchin species in the ge-
nus Echinometra (Rahman et  al., 2014) from one to
five months decreased both juvenile survival and
juvenile growth rates significantly. Larvae of Hali-
otis diversicolor are lecithotrophic; in contrast, those
of Echinometra are planktotrophs, and were fed the
diatom Chaetoceros gracilis during the delay period.
Similarly, delaying cyprid metamorphosis for the
barnacle Balanus amphitrite by just three to four
days resulted in reduced growth rates of juveniles
that were transplanted to two different field sites
(Thiyagarajan et al., 2007). In contrast, whether the
nonfeeding larvae of the coral Acropora tenuis were
allowed to metamorphose two, four, or six weeks
after fertilization had no significant detrimental im-
pact on post-settlement survival or time to initiate
colony formation in field-transplanted individuals
(Graham et al., 2013); indeed, budding began sooner
in the four-week-old cohorts than in those that had
been induced to metamorphose two weeks earlier.
Finally, delaying metamorphosis for seven days
 C o n s e q u e n c e s o f E m b ryo n i c a n d L a r val E x p e r i e n c e    215
reduced mean juvenile growth rates for the solitary
seasquirt Ciona intestinalis in studies by Jacobs et al.
(2008), but had no effect on the two other ascidian
species tested in the same study.
14.7  Latent Effects of Ocean Acidification
The data indicating declines of approximately 30%
in pH in the world’s oceans during the past several
hundred years have become increasingly convinc-
ing, and alarming (Doney et al., 2009; Byrne, 2011;
Barton et  al., 2012; Kroeker et  al., 2013), and the
cause seems unassailable: excessive release of CO2
into the atmosphere from a range of human activi-
ties including power production; manufacturing;
fuel-consuming air, sea, and land transportation;
cement production; and deforestation (Doney et al.,
2009; Friedlingstein et  al., 2014). Approximately
one-third of that excess CO2 has been absorbed by
the world’s oceans, resulting in a marked increase
in seawater acidity despite the presence of a bi-
carbonate buffering system (Pörtner, 2008; Doney
et al., 2009). By the year 2100, the average pH in the
world’s oceans is expected to drop from 8.1 to about
7.7– 7.8 (Doney et al., 2009; IPCC, 2013).
Although a good number of studies have examined
the effects of ocean acidification (OA) on aspects of
larval growth and development (reviewed by Byrn­e
and Przeslawski, 2013; Hettinger et  al., 2013b; see
also Byrne et al., this volume), and a number of stud-
ies have considered the effects of maternal exposure
to reduced pH on offspring responses (e.g., Parker
et  al., 2012; Pansch et  al., 2014; reviewed by Ross
et al., 2016), few studies have so far considered how
experiencing OA stress early in development might
influence post-metamorphic fitness. In one study of
such latent effects, larvae of the sea urchin Strongy-
locentrotus droebachiensis were reared to metamor-
phosis at pH 7.7 (control = 8.1) (Dupont et al., 2013).
The resulting juveniles grew more quickly over
the next three months when reared at the reduced
pH than when reared at the control pH (~ 0.29 mm
month-1 vs. ~0.16 mm month-1). Post-metamorphic
mortality was especially high (about 95%) for ani-
mals in that treatment, however, so it’s not clear
whether larval experience really promoted a greater
tolerance of reduced pH after metamorphosis or
whether the results reflect the preferential survival
of faster-growing juveniles. Similar results were
obtained for the bay scallop Argopecten irradians,
but again, the higher rates of subsequent juvenile
growth following larval exposure to the stress
may reflect differential survival of particular geno-
types rather than a true latent effect (Gobler and
Talmage, 2013).
In contrast, clear detrimental latent effects re-
sulting from larval exposure to acidification stress
were reported by Hettinger et  al. (2012; 2013a)
for the Olympic oyster Ostrea lurida. When larvae
were reared to metamorphosis at pH 7.8, juvenile
growth rates in the laboratory for the first week
after metamorphosis were about 26% lower than
those of control juveniles that had been reared
as larvae at pH  8.1, regardless of whether juve-
niles were reared under the control or reduced
pH conditions (Hettinger et  al., 2012). Similar ef-
fects of larval pH experience were subsequently
documented in juveniles that had been trans-
planted to field situations following metamorpho-
sis (Hettinge­r et  al., 2013a). The impact of being
reared at reduced pH during larval life persisted
for at least four months after metamorphosis in
that latter study, with no indication of compensa-
tory growth by individuals that had experienced
the stressful pH as larvae (Hettinger et al., 2013a).
Intriguingly, the effects on juvenile growth were
not mitigated by transferring individuals to more
benign field conditions following metamorphosis.
Additional studies investigating the potential for
latent effects following larval exposure to ocean
acidification seem warranted.
14.8  Latent Impact of Thermal Stress
Few studies have examined the effects of early ther-
mal stress on the post-metamorphic development
of marine invertebrates. Ross et al. (2013) exposed
larvae of the coral Porites astreoides to elevated tem-
perature (elevated by 3 °C, to 30 °C) for 24 hours,
and later transplanted the metamorphosed spat
to a field site. Temperature stress experienced
during the larval stage significantly increased
post-­metamorphic mortality (p = 0.044) over the
following 24 days, although mortality of control in-
dividuals was also quite high (90%).
216   E v o l u t i o n a ry Ec o l o g y o f Ma r i n e I n v e rt e b r at e L a r va e
14.9  Caution in Interpreting Latent
Effects
Interpreting data from latent effects studies can
be complicated by developmental mortality, as
alluded to earlier and as noted by Marshall and
Morga­n (2011). If larval or post-metamorphic mor-
tality is substantial, then what appear to be latent
effects on juvenile growth, adult fecundity, or other
measures could reflect differential survival of cer-
tain genotypes rather than a shift in individual
morphological development or physiology. Mor-
tality of laboratory-reared larvae and juveniles can
be quite high, even under what appear to be ideal
rearing conditions (Yund and McCartney, 2016),
and if that mortality is not random then results
may be difficult to interpret. Indeed, as mentioned
in a previous section, the favorable effects of rear-
ing larvae at reduced pH on the growth of juveniles
reared at the same low pH reported for the sea ur-
chin Strongylocentrotus droebachiensis (Dupont et al.,
2013) and the bivalve Argopecten irradians (Gobler
and Talmage, 2013) could have been mediated by
genotype-specifi­
­ c differential juvenile mortality
before growth measurements were made; juvenile
mortalities for individuals in this treatment were ex-
tremely high (up to 95%). Thus, the observed more
rapid growth of juveniles that had been stressed in
that treatment as larvae might reflect the differen-
tial survival of the most rapidly growing juveniles,
which is certainly an intriguing possibility.
However, latent effects have also been docu-
mented in cases where there was negligible larval
or juvenile mortality, in which case the effects must
be due to something more interesting. Such species
may be especially useful models for studying the
underlying basis for latent effects. The larvae and
juveniles of Crepidula fornicata, for example, can be
routinely reared in the laboratory with 0 to 7% mor-
tality (Pechenik and Lima, 1984; Pechenik and Tyrell,
2015; Bashevkin and Pechenik, 2015); the larvae are
large at hatching (typically 400–450 µm in shell
length) and grow quickly; can be easily induced to
metamorphose when competent (Pechenik and Gee,
1993); and grow rapidly following metamorphosis
(up to about 200 µm per day for the first week or so
after metamorphosis) with high survival (Pechenik,
1984; Pechenik et al., 1996a; Bashevkin and Pechenik,
2015; Pechenik and Tyrell, 2015). This is not the first
time that C. fornicata has been suggested as a promis-
ing research model (Henry et al., 2010).
Juvenile mortality for the Olympia oyster Ostrea
lurida was also low in the laboratory experiments
of Hettinger et  al. (2012), again implying that the
decreased mean post-metamorphic growth rates
observed for individuals that had experienced re-
duced pH stress as larvae have a basis other than
differential juvenile mortality, although it does not
rule out the possibility of selective mortality dur-
ing larval development. In a subsequent study
(Hettinger et al., 2013a), the researchers found that
the pH experienced as larvae significantly reduced
juvenile growth rates in field-transplanted individ-
uals even four months post-settlement, again with-
out the larval treatment having had a significant
differential impact on juvenile mortality.
14.10  Consequences of Larval Stress
are Not Always Negative
Most of the latent effects of larval experience that
have been reported to date have been negative,
with juvenile or adult survival or performance
being reduced in some way. The previously men-
tioned study with corals (Graham et  al., 2013), in
which budding began sooner in a cohort from lar-
vae that had had their metamorphosis delayed for
two weeks longer than those from another cohort,
is one exception. Similarly, in one experiment with
the polychaete Capitella teleta (Pechenik et al., 2016),
brooded embryos that had been exposed to hypoxic
stress (1 ml O2 l-1) for 36 hours grew more quickly
following metamorphosis than those that had expe-
rienced shorter periods of stress during brooding.
A similarly intriguing result has been reported for
male zebra finches, the first result of its kind for any
vertebrate: male birds that were stressed as nest-
lings were later found to have a higher reproduc-
tive fitness than those that were not stressed during
early development (Crino et al., 2014).
In some other experiments, the outcome of being
stressed in the larval stage depended on whether
juveniles were also reared under stressful condi-
tions. For example, in one experiment with Crep-
idula fornicata, larvae that had been reared at the
 C o n s e q u e n c e s o f E m b ryo n i c a n d L a r val E x p e r i e n c e    217
reduced salinity of 20 psu showed significantly
faster growth as juveniles when maintained at the
same low salinity after metamorphosis, compared
with the growth of juveniles that had been reared
as larvae at 30 psu and then transferred to the lower
salinity after metamorphosing (Bashevkin and Pe-
chenik, 2015); it appears that rearing larvae at that
low salinity pre-acclimated individuals for success-
ful development at that salinity. As mentioned ear-
lier (see Sectio­n 14.7), similar results were obtained
for the sea urchin Strongylocentrotus droebachiensis
when larvae were reared at the low pH of 7.7 and
juveniles were reared under the same conditions
(Dupont et al., 2013), and for the scallop Argopecten
irradians (Gobler and Talmage, 2013), although
the extent to which the results of those two stud-
ies reflect selective post-metamorphic mortality
rather than true latent effects is not clear. A num-
ber of studies with insects and vertebrates have
also found that stresses experienced during early
development can induce adult phenotypes to toler-
ate similar environmental stresses better (reviewed
by Wang et al., 2016b). Future studies with marine
invertebrates might focus specifically on the extent
to which stressful larval experiences pre-adapt ju-
veniles or adults for stressful conditions after meta-
morphosis. The likelihood of such findings would
seem to be greater the more that larval environmen-
tal conditions anticipate those likely to be experi-
enced by juveniles or adults.
14.11  Mechanisms Accounting for Latent
Effects
The mechanisms underlying the latent effects that
have now been documented for various marine
invertebrates have not been well explored and are
poorly understood (Williams and Degnan, 2009).
There are two major suggestions in the marine in-
vertebrate literature: (1) an effect on energy avail-
ability or allocation following metamorphosis, and
(2) epigenetic shifts in gene expression patterns.
The two may not be mutually exclusive.
A number of researchers have suggested or im-
plied a prominent role for depleted larval nutri-
ent reserves in producing latent effects (Emlet and
Sadro, 2006; Vermeij et  al., 2006; Chiu et  al., 2007;
2008; Johnson and Wendt, 2007; Thiyagarajan et al.,
2007; Hettinger et  al., 2012; Li and Chiu, 2013;
Rahma­n et  al., 2014). For example, in a study re-
ferred to earlier (Emlet and Sadro, 2006) in which
rearing nauplius larvae of the barnacle Balanus
glandula at a low food concentration led to reduced
post-metamorphic survival and growth, those re-
ductions were correlated with reduced lipid and
protein content of the pre-metamorphosed cyprids
(Emlet and Sadro, 2006), supporting a simple ener-
getics-limitation hypothesis for the observed reduc-
tion in juvenile growth rates. Similar results were
reported for the gastropod Crepidula onyx when
larvae were starved for portions of larval develop-
ment (Chiu et al., 2007). Vermeij et al. (2006) noted
increased swimming activity of nonfeeding coral
larvae (Montastrae­a faveolata) under conditions of
decreased salinity, and hypothesized that increased
rates of energy expenditure might have accounted
for the observed decrease in post-metamorphic
survival.
Along these same lines, the study by Li and Chiu
(2013) that was referred to earlier showed that ex-
posing the larvae of Crepidula onyx to hypoxic stress
reduced post-metamorphic growth and filtration
rates, but only when the stressed larvae had also
been reared at a reduced food concentration; larvae
exposed to hypoxic stress under high food condi-
tions showed normal post-metamorphic growth,
suggesting that having abundant nutrients prior
to metamorphosis ameliorated the latent impact of
hypoxic stress. Perhaps abundant embryonic nutri-
ent stores also account for the remarkable lack of
latent effects seen when brooded embryos of the
polychaete Capitella teleta were exposed to severe
hypoxia for up to 96 hours (Pechenik et al., 2016).
Nutrient limitation during larval development may
alter energy allocation and thus impede the nor-
mal development of feeding structures (Chiu et al.,
2007) or digestive machinery. Even so, what might
cause such specific shifts in morphological devel-
opment, and are they indeed specific to feeding-
related morphology or physiology?
Some of the best direct evidence for a role of
nutritional limitation in generating reduced post-
metamorphic growth rates comes from laboratory
studies with the nonfeeding larvae of the bryozoan
Bugula neritina (Wendt and Johnson, 2006; Johnson
and Wendt, 2007). Larvae whose settlement was
218   E v o l u t i o n a ry Ec o l o g y o f Ma r i n e I n v e rt e b r at e L a r va e
delayed by 24 h in a medium with depleted dis-
solved organic matter (DOM) showed a reduced
ability to metamorphose successfully, and those
that did metamorphose had smaller than normal
feeding lophophores; those effects of delayed meta-
morphosis were substantially (but not completely)
offset for a parallel group of larvae in seawater en-
riched with DOM (Johnson and Wendt, 2007).
On the other hand, post-metamorphic growth
rates at normal salinity (34 psu) were reduced for
the barnacle Balanus amphitrite after cyprids were
exposed for 24 hours to a salinity of only 10 psu,
despite the fact that there was no measureable ef-
fect on cyprid lipid content prior to metamorphosis
(Thiyagarajan et al., 2007). Does this argue against a
direct nutritional mechanism for the observed latent
effects, or were those latent effects caused by the
reduced availability of carbohydrates, proteins, or
some other non-lipid nutrient or micronutrient? De-
terring metamorphosis of this species for three days
also resulted in reduced post-metamorphi­c growth
for field-transplanted individuals, and allowing the
newly metamorphosed, field-transplante­d individ-
uals to grow in an area with higher food availability
“did not fully compensate for the negative effects
of delayed metamorphosis” (Thiyagarajan et  al.,
2007, p.  183); indeed, mean juvenile feeding rates
were significantly higher for individuals whose
metamorphosis had been delayed, again suggesting
that something more than a deficiency in juvenile
feeding ability was at play in reducing the juvenile
growth rates of these individuals (Thiyagaraja­ n Chiu, 2013); to date nobody has determined exactly
et al., 2007).
Although post-metamorphic feeding limitations
may play a role in causing observed latent effects
in at least some species, there is reason to believe
that other, largely unexplored factors are probably
also involved in at least some cases (reviewed by
Pecheni­ k, 2006; Chiu et  al., 2007). Consider, for
example, the impact of delayed metamorphosis
on colony development of the seasquirt Diplosoma
listerianum: branchial baskets were significantly
smaller not just in the newly metamorphosed an-
cestrulae, but also in individuals subsequently pro-
duced asexually weeks later in field-transplanted
colonies (Marshall et  al., 2003). Similarly, expos-
ing larvae of the bryozoan Watersipora subtorquata
to sublethal levels of copper for only a few hours
reduced colony survival months after colonies were
transplanted to field sites (Ng and Keough, 2003),
while forcing larvae of the bryozoan Bugula neritina
to swim for an extra 23 to 24 hours significantly re-
duced colony growth rates and fecundity of field-
transplanted colonies over the next two weeks,
and delayed the onset of reproduction (Wendt,
1998). It is also worth remembering that effects on
juvenile growth rates of the Olympia oyster Ostrea
lurid­a also persisted for months after individuals
were transplanted to field sites (Hettinger et  al.,
2013a). Such effects are unlikely to be caused sim-
ply by reductions in initial feeding ability follow-
ing metamorphosis. In addition, some studies have
failed to detect latent effects following stresses that
have diminished larval energy reserves (e.g., delay-
ing metamorphosis of the ascidian Styela plicata;
Thiyagaraja­n and Qian, 2003) or which are likely to
have diminished such reserves (e.g., salinity stress
in three species of calyptraeid gastropod; Diederich
et al., 2011).
What molecular mechanisms might account for
the persistent latent effects that have been observed
in so many studies? It is thought that some pollut-
ants could be acting as potent endocrine disrupt-
ers (Nice et  al., 2003), while the reduced juvenile
growth rates documented for several suspension
feeders following a variety of larval stresses are
possibly due to reduced rates of functional gill de-
velopment for at least the first few days or weeks
after metamorphosis (Pechenik et al., 2002; Li and
what about the gill is not working properly—is it
simply a reduced size, or does the reduced func-
tioning have a more interesting and subtle me-
chanical basis? But behind even these mechanisms
there may often be a more fundamental cause of
changes in juvenile and adult performance caused
by events experienced much earlier in develop-
ment. As suggested earlier (Pechenik, 2006), there
is growing evidence that patterns of gene expres-
sion can be altered by the environment to modify
phenotypes without making any changes in the ac-
tual gene sequences themselves. Indeed, Williams
and Degnan (2009) have demonstrated distinct
differences in patterns of gene expression, persist-
ing for at least 40 hours after metamorphosis, for
juveniles of the abalone Haliotis asinine that had
 C o n s e q u e n c e s o f E m b ryo n i c a n d L a r val E x p e r i e n c e    219
been triggered to metamorphose by contact with
different coralline algal species. Such epigenetic ef-
fects have a number of different potential mecha-
nisms (reviewed by Jablonka, 2013; Burggren,
2014; Skinner, 2015); to date the most attention has
been focused on the selective methylation of DNA
nucleotides (especially 5′ cytosine rings), namely,
DNA methylation which then blocks the expres-
sion of the targeted genes (Suzuki and Bird, 2008;
Williams and Degnan, 2009; Burggren and Crews,
2014), and histone modification (Gibson et al., 2012;
Robichaud et al., 2012) which can either silence or
activate gene expression. De-methylation can also
be involved in regulating gene expression patterns
(Kesäniemi et al., 2016). Few studies (e.g., Williams
and Degnan, 2009) have so far sought to identify
the molecular mechanisms that are responsible for
the latent effects that have been documented in
marine invertebrates, but the technology to do is
becoming increasingly available (Williams and De-
gnan, 2009; Lyko et al., 2010; Robichaud et al., 2012;
Flores et al., 2013; Gavery and Roberts, 2014), and
interest in doing so is growing (Gibson et al., 2012;
Robichaud et al., 2012).
Intriguingly, food levels and other nutritional
deficiencies can be especially important causes of
epigenetic shifts in patterns of gene expression in
vertebrates (Mazzio and Soliman, 2014), as can tem-
perature stress and exposure to toxicants (reviewed
by Skinner, 2015). Understanding the mechanisms
behind latent effects should enable us to explain
some important questions about the organismal
data collected to date. For example, why do we
see clear latent effects in some experiments but not
others using the same species, even when the lar-
vae have experienced the same stress treatments
at the same levels for the same amounts of time?
If susceptibility is genetically determined in such
cases, what makes the offspring of some parents
more susceptible than those from other parents?
Another especially intriguing question is why do
we sometimes not see any latent effects following
metamorphosis even when larval growth rates do
not recover to control levels after the stress period
has ended (Diederich et al., 2011)? Why do certain
stresses produce clear latent effects in some species
but not in other species? Also, to what extent is the
variability we currently see in juvenile growth rates
in the field due to direct genetic effects and to what
extent to the effects of previous experience during
embryonic or larval development?
Once the mechanisms lying behind the latent ef-
fects documented for marine invertebrates are un-
derstood, it should be possible to understand why
the larvae of some species show these responses to
certain stresses but not others, why the larvae of
some species don’t show latent effects in response
to the same stresses, and why the larvae of some
species may be more susceptible to stresses at cer-
tain stages of development.
Remarkably, there is increasing evidence, par-
ticularly from work with vertebrates, that environ-
mentally induced epigenetic alterations in patterns
of gene expression—particularly those involving
consequences of nutritional stress—can at least
sometimes be passed on to future generations; that
is, environmentally induced alterations in patterns
of gene expression involving nutritional stress
can be transgenerational and play key roles in fa-
cilitating evolutionary change (Jablonka and Raz,
2009; Burggre­n, 2014; Burton and Metcalfe, 2014;
Mendizabal et  al., 2014; Skinner, 2015). Whether
any of the latent effects discussed in this chapter are
ever transmitted to future generations has not yet
been assessed for any marine invertebrate (Marshal­l
and Morgan, 2011; Gavery and Roberts, 2014), but is
something that should be explored in future stud-
ies. Burggren (2014) has noted that what have been
referred to as maternal or paternal “transgenera-
tional effects” could in many cases be mediated by
direct effects on gametes or early embryos in spe-
cies with internal development, rather than being
transmitted to the next generation from effects on
adults. But such interpretive difficulties would not
apply to the sorts of latent effects discussed in the
present review.
14.12  Impact and Implications
Some of the immediate consequences of latent ef-
fects in the field are easy to imagine. Reduced
growth rates, in particular, are likely to increase
vulnerability to predation: to the extent that in-
dividuals become less vulnerable to predators as
they grow (Gosselin and Qian, 1996; 1997; Hunt
and Scheibling, 1997), slower growth should cause
220   E v o l u t i o n a ry Ec o l o g y o f Ma r i n e I n v e rt e b r at e L a r va e
longer periods of vulnerability. The population-
level impact of latent effects may be especially great
for organisms that are sessile after metamorphosis,
as juveniles and adults of such species typically
have fewer options for avoiding exposure to envi-
ronmental stresses. Also, some stresses experienced
early in development appear to increase juvenile
mortality directly (e.g., in the gastropod Crepipatella
dilatata: Chaparro et  al., 2014). Similarly, latent ef-
fects that involve delaying time to reproductive
maturity and reducing fecundity (e.g., Wendt, 1998)
may well negatively impact local population dy-
namics, or at least the likelihood of successful per-
petuation of the parental genotype.
On the other hand, to the extent that larval ex-
periences of ocean acidification, nutritional limita-
tion, hypoxia, or sublethal pollutant exposure can
improve juvenile or adult function under similarly
stressful conditions, latent effects could promote
species persistence in a changing ocean.
The frequency with which the larvae of marine
invertebrates experience delayed metamorphosis
and other potentially stressful experiences prior to
metamorphosis in the field, and the actual conse-
quences of those experiences on juveniles in natural
field situations, are unknown. Individual juveniles
and adults in natural populations certainly differ
greatly in behavior, growth rate, timing of sexual
maturity, and fecundity for all organisms studied
to date.
But the degree to which those differences reflect
the impact of pre-metamorphic experience, rather
than differences in underlying standard Mendelian
genetics or local differences in physical conditions
(e.g., Helmuth and Hofmann, 2001), is not clear.
What we need to find is an internal black-box re-
corder of some sort that would provide a record
of larval experience in field-collected juveniles or
adults that exhibit differences in key measures of
individual fitness. Something like that has been
found for fish (otoliths) (McCormick, 1999), but
we are still looking for something comparable for
studies of marine invertebrates (Levin et al., 2015).
Even if we find that a particular stressor causes
detectable and reproducible changes in isotope
ratios, trace element composition, or other com-
ponents of molluscan larval shell composition in
the laboratory (Levin et al., 2015), we also need to
make sure that such changes are not also induced
by any other factors.
To the extent that latent effects are indeed occur-
ring commonly for marine invertebrates in the natu-
ral world, there is tremendous potential for climate
change, pollution, hypoxia, and other environmen-
tal stresses to impact the geographic distributions
of species and local community structure through
those subtle effects. Thus, a widespread and in-
creasing occurrence of latent effects in the field will
make it even more difficult to predict the future
impact of environmental change on future popula-
tions and communities. The potential for multiple
environment-individual interactions adds further
complexity: studies to date are limited to testing the
effects of one to three multiple stressors at a time
(e.g., ocean acidification, pollutants, temperature
stress, hypoxia, salinity stress, or nutritional stress)
(Table 14.1), whereas in the real world, embryos and
larvae are probably subjected to many stresses si-
multaneously. The precise actual impact of human
activity on marine populations defies prediction:
there is probably only one way to conduct this par-
ticular experiment, and we are all participating.
14.13 Summary
1. A variety of stresses, including hypoxia, reduced
pH, and food limitation, experienced during de-
velopment can influence growth rates, survival,
and other fitness characteristics following meta-
morphosis.
2. The brooding of embryos may protect against
exposure to some environmental stresses during
development, but can sometimes expose devel-
oping embryos to stresses they would otherwise
have avoided by being free-living, resulting
again in reduced fitness in later life.
3. The mechanisms through which such “latent ef-
fects” are mediated are unclear: energy-balance
issues and epigenetic factors—in which gene ex-
pression patterns are altered without any chang-
es in DNA sequences—seem to be involved.
4. The extent to which documented variability
in factors such as growth, survival, and repro-
ductive output in benthic field populations is
explained by stresses experienced early in devel-
opment remains to be determined.
 C o n s e q u e n c e s o f E m b ryo n i c a n d L a r val E x p e r i e n c e    221
The coming years will apparently bring increas-
ing water temperatures; changing rainfall patterns
and consequent shifts in coastal salinities and in
the magnitudes of salinity fluctuation; increasing
ocean acidity; increasing levels of pollution (in-
cluding the spread of microplastics); increasing
incidences of hypoxic events; and shifting patterns
of phytoplankton abundance, species composition,
and nutritional quality; all of which have the po-
tential to increase physiological stresses on devel-
opmental stages and increase the energy costs of
development through metamorphosis. This pre-
sents us with a good number of important ques-
tions: (1) To what extent do these stresses—alone
and in combination—have latent impact on marine
invertebrates, and what are those impacts? (2) Do
they ever carry over to subsequent generations? (3)
To what extent are some species more vulnerable to
particular stresses than other species? (4) What ac-
counts for differences in vulnerability? (5) To what
extent does brooding protect offspring from expo-
sure to environmental stress and from exhibiting
latent effects later in development? (6) And what
are the underlying mechanisms causing latent ef-
fects? It seems likely that many of these questions
will be addressed, and possibly answered, over the
next ten years.
Acknowledgments
Thanks to Andreas Heyland, Tyler Carrier, and
Adam Reitzel for undertaking this project, for invit-
ing my participation, and for their helpful editorial
suggestions. Thanks also to my colleagues Oscar
Chaparro, Vengatesen Thiyagarajan (“Rajan”),
Richard Strathmann, Sam Bashevkin, and Sarah
Gilliand for thoughtful comments and suggestions
on a draft of this manuscript.
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280: 20123022.
Zhang, J., Cowie, G., and Naqvi, S.W.A. 2013. Hypoxia in
the changing marine environment. Environmental Re-
search Letters 8: 1–3.
 CH A PT ER 15
Section 3 Summary—Larval Transport,
Settlement, and Metamorphosis
Life cycles for broadcast-spawning marine inver-
tebrates are characterized by a pelagic (larval) and
benthic (juvenile and adult) stage, which are dif-
ferentiated by the process of settlement and meta-
morphosis. While settlement and recruitment are
concepts that primarily deal with the ecological
aspects of this transition, they are often accompa-
nied by a drastic morphological and developmental
transition. Larval transport, the means of reaching
suitable settlement sites, is difficult to study, but in-
sights from genetics, behavior, sensory ecology, and
oceanography have provided important insights.
In Chapter 11, Jesús Pineda and Nathalie Reyns
discuss larval transport as both a concept, includ-
ing its many components and as a measurement, as
well as outline the many challenges of taking these
measurements and recent approaches for doing so.
Next, in Chapter 12, Peter Marko and Michael Hart
discuss the utility of genetic approaches in studying
larval dispersal and how they may characterize the
scale and/or magnitude of the connectivity of pop-
ulations. In Chapter 13, Jason Hodin and colleagues
spatially examine the interaction between fluid dy-
namics and sensory processes, and how this enables
larvae to navigate the sea. Finally, in Chapter 14, Jan
Pechenik assesses and postulates how and why abi-
otic and biotic factors of the sea, including hypoxia,
Carrier, T. J., Reitzel, A. M., and Heyland, A., Section 3 Summary—Larval transport, settlement, and metamorphosis.
In. Evolutionary Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland:
Oxford University Press (2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0015
food limitation, salinity, and ocean acidification, af-
fect life after metamorphosis.
These contributions uniquely combine ecological,
physiological, and developmental mechanisms, and
emphasize their evolutionary implications. For exam-
ple, to understand the dispersal potential of embryos
and larvae in the plankton, one has to consider both
physical and biological factors as well as the specific
benthic habitat requirements of these species. Mod-
els for larval transport, therefore, need to integrate
divergent issues, such as larval sensory, feeding, and
swimming ecology along with simluations of ocean
currents, gyres, and eddies. The plankton-to-benthos
transition is also frequently accompanied by a drastic
metamorphosis, where larvae become juveniles. This
transition can occur within a remarkably short period
of time and needs to be timed properly within the life
cycle of an organism. Post-settlement performance
and, by extension, fitness, is therefore linked to vari-
ous aspects of metamorphosis and development. Due
to the small size of most marine invertebrate larvae, it
is difficult to track individuals from the plankton to
the benthos. Genetic population analyses provide an
excellent tool to link planktonic and benthic popula-
tions, and much progress has been made as sequenc-
ing costs of large number of samples have become
more affordable.
 C h a pt er 16
Ecology and Evolution of Larval
Dispersal in the Deep Sea
Craig M. Young, Shawn M. Arellano, Jean-François Hamel, and Annie Mercier
16.1 Introduction
The deep ocean contains more unexplored habi-
tats than anywhere else on Earth, and probably
more undiscovered organisms as well (Grassle and
Maciole­k, 1992), yet most of the nearly 26,000 de-
scribed species of deep-sea benthic invertebrates
(Glover et  al., 2016) are known only from the ini-
tial descriptions of their morphology. Among the
relatively well-studied species from continental
slopes and chemosynthetic environments, we still
have only a rudimentary understanding of life his-
tory biology, including the basics of gametogenesis,
spawning, reproductive timing, and embryonic and
larval development in a small but growing subset
of species (Young 2003; Mercier and Hamel, 2009a;
Watling et al., 2011). Because deep-sea sampling is
expensive, there have been relatively few opportu-
nities to sample populations repeatedly and obtain
viable gametes for laboratory study. A limited num-
ber of larvae, probably less than 50 species world-
wide, have ever been collected from the deep sea
(Bouche­t and Warén, 1994; Hilário et al., 2015), and
fewer than 20 species have been cultured to meta-
morphosis from these field collections or from gam-
etes obtained in the laboratory (e.g. Mercie­r and
Hamel, 2008; 2009b; Sun et al., 2010; 2011; Mercier
et  al., 2014; 2015; 2016; Montgomery et  al., 2017a).
Information about habitat selection at settlement re-
mains fragmentary (Sun et al., 2010; Mercier et al.,
2016). Overall, we know almost nothing about
larval swimming, feeding, or nutrition (Mercier
et al., 2014; Beaulieu et al., 2015; Montgomery et al.,
2017a), and we still have no idea where the larvae of
Young, C. M., Arellano, S. M., Hamel, J.-F., and Mercier, A., Ecology and evolution of larval dispersal in the deep sea.
In. Evolutionary Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland:
Oxford University Press (2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0016
most species spend their entire developmental peri-
ods. Our sample sizes of species within phyla are so
small that generalizations are nearly always unwar-
ranted. The deep sea therefore remains a frontier
not only for exploration, but also for the biology of
early life history stages, including larvae.
This paper is not intended as a comprehensive
synthesis of deep-sea reproduction, which has last
been reviewed some 15–20 years ago (Young and
Eckelbarger, 1994; Young, 2003). Several reviews
discuss larval biology in vent systems (Mullineau­x
and France, 1995; Tyler and Young, 1999; Adams
et al. 2012), and a pair of reviews discuss the emerg-
ing fields of deep-sea genetic connectivity (Baco
et  al., 2016) and its underlying biological mecha-
nisms (Hilário et  al., 2015). The present paper fo-
cuses narrowly and somewhat speculatively on
adaptive mechanisms that might impact the dis-
tances, times, and trajectories of larval dispersal in
the deep ocean. Conditions in the deep sea appear
extreme, so it is easy to imagine that strong selec-
tive pressures might yield bizarre and unique de-
velopmental and larval adaptations through natural
selection. Our goal is to ground truth these various
imagined possibilities with the limited empirical in-
formation available.
16.2  Why Deep-Sea Dispersal Matters
Dispersal and genetic connectivity are topics of con-
siderable current interest because of concerns about
a deep ocean that is increasingly impacted by hu-
mans through fishing, mining, dumping, acidifica-
tion, and climatic changes (Priede et al., 2010; Van
230   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r vae
Dover, 2011; Barbier et al., 2014; Hilário et al., 2015).
The long-term maintenance of species and popula-
tions probably depends on biological mechanisms
that exchange genes among metapopulations and
that spread the risk of mortality in the face of chang-
ing ocean conditions. From a practical standpoint,
knowledge of dispersal is critical for making deci-
sions about the siting and sizes of marine protected
areas, and a recent review (Hilário et al., 2015) ad-
dressed the topic of dispersal from this perspective.
Cowen et  al. (2006, p.  522) provided a succinct
justification for studying connectivity and its un-
derlying biological mechanisms.
Defining the scale of connectivity, or exchange,
among marine populations and determining the
factors driving this exchange are pivotal to our
understanding of the population dynamics, ge-
netic structure, and biogeography of many coastal
species.
This is equally true in the deep sea, where popu-
lation dynamics have been studied for relatively
few species (Gage, 1982) because of the difficulty of
repeated sampling. Moreover, the mechanisms un-
derlying major patterns of biogeography and phy-
logeography remain poorly understood. Several
examples of patterns from the literature highlight this
gap: (1) gastropods in Atlantic abyssal basins show a
highly significant gradient of decreasing alpha diver-
sity from low to high latitudes (Rex and Etter, 2010);
(2) strong evidence for genetic connectivity among
methane seeps on the two sides of the Atlantic Ocean
(Cordes et al., 2007; Olu-Le Roy et al., 2007; Olu et al.,
2010); (3) presence of biogeographic barriers in some
species among western Pacific back-arc basins (Thaler
et  al., 2011); (4) endemism and genetic isolation in
hadal trenches (Jamieson, 2015); (5) species shifts of
protobranch bivalves along surprisingly short dis-
tances near the western boundary current off New
England (Etter and Bower, 2015); and (6) size distribu-
tions of mussels (Van Dover et al., 2001) and genetic
fingerprints of tube worms (Shank and Halanych,
2007) not easily explained by simple advection/diffu-
sion models (Okubo, 1971). The patterns themselves
have been discovered through a combination of care-
ful taxonomy and genetics, yet in every case, the eco-
logical and phylogeographic mechanisms remain in
the realm of speculation because of an inadequate
understanding of dispersal trajectories.
Finally, it is likely that the larvae themselves have
important roles in the food webs of the deep ocean.
A thoughtful paper by Levin et al. (2016) considers
the possibility that dispersing gametes and larvae
transfer organic material of chemosynthetic origin
through pelagic food webs all the way from the
sea floor to the euphotic zone (Pond et  al., 2000;
Arellan­o et al., 2014) and horizontally on the scale
of hundreds or thousands of kilometers (Adams
et al., 2011; Young et al., 2012; Hilário et al., 2015).
The quantitative importance of these transfers re-
mains completely unexplored.
16.3  Is Dispersal Advantageous
to Deep-Sea Species?
The adaptive significance of larval dispersal has
been debated for decades (Strathmann, 1974; 1987;
Pechenik, 1999; Shanks, 2009; Mercier et  al., 2013;
Burgess et al., 2015). Numerous advantages for dis-
persal have been proposed, including (1) reducing
competition between adults and their offspring; (2)
minimizing competition among siblings; (3) coloni-
zation of new habitats; (4) decreasing predation by
benthic predators; (5) reducing negative effects of
inbreeding; and (6) spreading the risk of mortality
in spatially and temporally variable environments.
However, Shields (1982) and Strathmann (1987)
have both discussed the counterintuitive possibility
that expression and elimination of deleterious traits
in isolated populations may create a situation where
inbreeding depression is insignificant and selection
favors retention rather than wide dispersal of off-
spring. Indeed, many clades, particularly of sessile
animals, have evolved life cycles with larvae that
disperse only short distances (Shanks, 2009; Burges­s
et  al., 2015). Examples in the deep sea include the
sea anemone Urticina sp. (Mercier et al., 2016) and
the cup coral Flabellum angulare (Mercier et al., 2011),
which both free spawn gametes, yet have embryos
and larvae that remain close to the parents (<1 m)
for the duration of their development.
There has also been much discussion about
whether dispersal per se has a significant evolu-
tionary advantage, or whether dispersal is a conse-
quence of evolutionary pressures that favor other
traits such as feeding and pre-metamorphic growth

(Strathmann, 1974; Havenhand, 1995; Pechenik,
1999). Palmer and Strathmann (1981) tested the
relative advantages and risks of dispersal for in-
tertidal barnacles, concluding that the advantages
of dispersal decrease with length of larval life and
dispersal distance. Etter and Caswel­l (1994) and
Caswell and Etter (1999) used a cellular automa-
ton model to consider the potential advantages of
dispersal in the deep sea as a function of distur-
bance frequency. In their model, in which distur-
bance frequency was assumed to vary with depth,
the advantages of dispersal did not decrease with
distance as Palmer and Strathmann (1981) sug-
gested, but were highest in systems with interme-
diate frequencies of disturbance. This finding fits
the observed distribution of developmental mode
in gastropods along a bathymetric gradient (Rex
and Warén, 1982).
Recent compilations of dispersal times (pelagic
propagule durations—PPDs; or planktonic larval
durations—PLDs) for a wide assortment of spe-
cies (Grantham et  al., 2003; Shanks 2009; Mercier
et  al., 2013) show that the proportions of species
with various dispersal strategies shift with climate
zones and habitat type in nearshore ecosystems.
For example, a high proportion of species living
on sandy bottoms have long-distance dispersal
mechanisms, whereas rocky-shore species that de-
pend on a narrow strip of habitat are more likely
Probability of Settlement Success
Retention in
Natal Habitats
Planktonic Larval Duration
La r va l D i s p e r sa l i n t he D ee p S ea    231
to disperse short distances, presumably to reduce
offshore loss of larvae (Shanks, 2009). The vast
abyssal basins of the deep ocean are analogous to
sandy continental shelves in that they should pro-
vide abundant and easily located settlement sites in
appropriate habitats. Insular habitats such as drop-
stones (Meyer et al., 2016) and seamounts (Rogers,
1994) are somewhat analogous to shorelines; they
are probably difficult to find, being spatially scat-
tered over expanses of unsuitable muddy bottoms.
Hydrothermal vents and methane seeps are also
insular habitats for the symbiotic species that de-
pend on them. One might argue that such insular
habitats could select for a bimodal structure of dis-
persal strategies favoring either very short disper-
sal (retention), or a long enough larval life to assure
that new habitats will be encountered (Figure 16.1).
It is also important to consider the stability of sites.
Hydrothermal vents are known to be stable only
on decadal scales (Van Dover, 2000), so a retention
strategy might be less likely to evolve at vents (but
see Adams and Mullineaux, 2008) than at methane
seeps, which may be stable over century-long tem-
poral scales (Bergquist et al., 2000).
Jackson and Strathmann (1981) noted that for
coastal species whose habitat are located along the
shore, species with long precompetent dispersal
periods must compensate by having long post-
competent periods to return to suitable settlement
Figure 16.1 Hypothetical bimodal distri-
bution of dispersal strategies evolving in
species living in isolated insular habitats in
the deep ocean. In a homogenous habitat
such as the abyssal plain, the distribution
of planktonic larval durations (PLDs) would
be represented by a horizontal straight line
Discovery of instead of a parabola because the probability
Dispersed Habitats of finding a suitable habitat should not
depend on the distance traveled or the
duration of the search.
232   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r vae
sites. Grantham et  al. (2003) and Shanks (2009)
refined this idea by considering the directions of
currents and the frequency of onshore transport
and retention mechanisms. In deep ocean species
living on the vast, sedimented abyssal plains, one
might assume that shorter postcompetent periods
might be possible, since mud is available almost
everywhere. On the other hand, where deep-sea
species depend on highly patchy habitats such as
vents or seeps, dispersal might increase the proba-
bility of locating a suitable settlement site, but only
if larvae have postcompetent periods long enough
to drift near the bottom and encounter such habi-
tats. Thus, both the advantages and disadvantages
of dispersal and the duration of the competent pe-
riod might be very different in the deep sea than in
shallow coastal habitats that have been considered
in earlier studies.
16.4  How Common is Brooding
in the Deep Sea?
Thorson (1950) suggested that larval development
should be absent or nearly so in the deep sea, since
dispersal should not be required in such a uniform
muddy environment, and because food should be
inadequate to support planktotrophic larvae. The
fact that the first two deep-sea echinoderm larvae
to be cultured (Prouho, 1888; Mortensen, 1921) were
both planktotrophic did little to sway the prevailing
opinion over several decades (Young, 1994).
The speciose peracarid crustaceans (about
4,600 species of isopods, amphipods, cumaceans,
mysids, and tanaids) and nematodes (about
600 species) are constrained phylogenetically to
brood embryos at all depths (Young, 2003), as
are pycnogonids (Mercier et  al., 2015). Among
polychaetes, a large proportion of infaunal spe-
cies with known reproductive modes brood in
their tubes (observations by L. Levin, tabulated
in Young, 2003), while a dorvilleid (Ophryotrocha
sp.) was discovered to rely on mucous chambers
(Mercier et  al., 2014). Egg capsules of gastropods
have often been observed on hard deep-sea sub-
strata, but only in rare instances have they been
cultured long enough to determine whether em-
bryos hatch as juveniles or larvae (Metaxas, 2011;
Montgomery et al., 2017a; Van Gaest, 2006). A re-
markable gastropod (Ifremeria nautili) was discov-
ered at vents in the western Pacific to brood its
larvae within a modified mucus gland in the sole
of its foot (Reynold­s et  al., 2010). Many deep-sea
cnidarians, including octocorals (Sun et  al., 2010,
2011; Mercie­r and Hamel, 2011) and sea anemones
(Mercier et al., 2016) have been found to be brood-
ers or demersal developers.
Charles Wyville Thomson’s (1876) discovery of
Antarctic echinoderms brooding large embryos
during the Challenger Expedition established the
longstanding view that large eggs are indicative
of brooding in the deep sea, and also, therefore, of
short dispersal distances. Thus, when the eggs of
echinothuriid echinoids, elasipod holothuroids,
and various asteroids were found to be large and
yolky (Tyler and Gage, 1984), these species were
often assumed to be brooders rather than plank-
tonic lecithotrophs (Young, 1994). The evolution
of brooding in two clades of Antarctic echinoids
has been discussed by Poulin and Féral (1994) and
by Pearse et al. (2009), who conclude that the evo-
lution of Antarctic brooding can probably be ex-
plained by vicariant speciation events rather than
adaptation to environmental conditions. Thus,
one would not necessarily expect the evolution
of brooding in the deep sea at large. A few eury-
bathic and deep-water sea stars from the North
Atlantic—for example, genera Pteraster, Henricia,
and Leptasterias (Boolootian, 1966), including Hen-
ricia lisa (Mercier and Hamel, 2008)—are known
to brood. In contrast, only two species of deep-
sea holothuroid are known to brood outside of
the Antarctic (Hansen, 1968; Pawson et al., 2003),
with the possible exception of a newly discovered
species (Gebruk et  al.  2013). This contrasts dra-
matically with the number of echinoderms that
brood in shallow water; Boolootian (1966) listed
30 species of shallow-water brooding holothu-
rians and 30 brooding asteroids, and several ad-
ditional species have now been added. Although
deep-sea animals with benthic or brood-protected
development have since been reported in most
taxonomic groups, overall, it still appears likely
that planktonic embryonic/larval development is
more common than protected modes of develop-
ment in the deep sea.

16.5  Do Deep-Sea Larvae Demonstrate
Distinctive Developmental Strategies
that Either Enhance or Reduce Dispersal
Distance?
Planktonic larval durations (PLDs) or the more in-
clusive pelagic propagule durations (PPDs, includ-
ing the embryonic phase) of deep-sea species can
impact dispersal distances significantly. Slow meta-
bolic rates imposed by cold temperatures extend
larval durations (O’Connor et  al., 2007b) in deep-
sea species, and some species show some notable
unique developmental strategies that can lead to
longer PLDs and hence time spent dispersing.
It has been widely assumed that unusual devel-
opmental modes and strange larval forms would be
common in the deep sea. As larvae have been cul-
tured, this has not generally proven to be true. With
very few exceptions, the larvae of most deep-water
species are morphologically similar to those of their
shallow-water relatives, presumably because of
phylogenetic constraints (Eckelbarger and Watlin­g,
1995). Thus, deep-sea crustaceans have nauplii, gas-
tropod molluscs have veligers, cnidarians have plan-
ulae, polychaetes have trocophore-like larvae, and
the various echinoderm classes produce larvae simi-
lar in most respects to those of their shallow-­water
relatives. One notable exception is Warén’s larva,
an unusual ciliated larval form of the aforemen-
tioned hydrothermal-vent gastropod Ifremeria nautili
(Reynold­s et al., 2010). These unusual larvae emerge
from their pedal brood chamber as fully ciliated lar-
vae with cilia protruding through a protective cu-
ticle. Eventually they metamorphose into veligers,
but neither their PLD nor their dispersal potential is
known. However, haplotype analysis of this species
(Thaler et al., 2011) indicates that populations within
back-arc basins show no genetic differentiation over
a scale of 1,000 km, suggesting that the unusual lar-
vae of I. nautili are capable of long-distance dispersal.
Peculiar reproductive adaptations reported from
the deep waters of the northwest Atlantic include
the dual brooding and broadcasting strategy of the
sea star Henricia lisa (Mercier and Hamel, 2008),
the negatively buoyant (benthic) lecithotrophic lar-
vae of the broadcasting sea anemone Urticina sp.
(Mercie­r et al., 2016), and the tethered oocytes of the
cup coral Flabellum angulare that remain bundled
La r va l D i s p e r sa l i n t he D ee p S ea    233
and attached to the female only until they get ferti-
lized (Mercier et al., 2011).
Other larvae thought to be unique to the deep sea
include the enormous Planktosphaera pelagica larvae
of enteropneusts (Spengel, 1932; Hart et  al., 1994),
which have long been hypothesized to have deep-
sea adults (Hadfield, 2001; Osborn et al., 2011), and
the large (15 mm long) “Auricularia nudibranchia”
larva (Chun, 1888), which has now been identified
as the larva of a deep-sea synaptid holothurian,
Protankyra brychia (Pawson et  al., 2003). Both spe-
cies are planktotrophic (Hart et  al., 1994; Sewell
and McEuen, 2001) and are remarkable not only for
their elaborate forms and large sizes, but also for
their presence in surface waters.
Developmental modes of many deep-sea animals
have been inferred from egg sizes and other life
history observations, but using assumptions devel-
oped for shallow-water fauna can be problematic.
This difficulty is well illustrated by the long “spa-
ghetti worm” enteropneust, Saxipendium coronatum,
that drapes over rocks near vents on the East Pacifi­c
Rise. On the basis of its broadly conical sperm
and small observed egg size, Franzén et  al. (1985)
predicted that this species should be a broadcast
spawner with planktotrophic larvae. When this spe-
cies was reexamined two decades later, however,
the eggs were found to be yolky and up to 1.3 mm
in diameter (Young, unpublished), suggestive of the
kind of demersal development known for others
in the same family (Hadfield, 2001). Franzé­n et al.
(1985) apparently based their conclusion on im-
mature specimens. A promising avenue in the con-
firmation of lecithotrophic development revolves
around the combination of egg size and pigmenta-
tion (Montgomery et al., 2017b).
16.5.1  Increasing Dispersal Through Parental
Investment
Several observations suggest that deep-sea eggs are
invested with a larger amount of yolk or yolk with
higher energy content than those of shallow species,
although a clear exception exists to this generaliza-
tion in the northwest Atlantic, where both shallow-
water and deep-sea species are represented by a large
percentage of lecithotrophic species. Bivalve larvae
found off Barbados at depths as great as 5,000  m
234   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r vae
(S. Maslakova and C.M. Young, unpublished) were
obviously planktotrophic (they had 40–60 growth
increments in a large prodissoconch II), yet the pro-
dissoconch I was more than 300 µm in diameter,
suggesting a huge egg that would be expected of a
lecithotrophic brooder in Ockelmann’s (1965) tradi-
tional scheme (Scheltema, 1994). In this species, it ap-
pears that a large egg with a significant yolk reserve
must develop into a long-lived planktotrophic larva.
The eggs of another bivalve, the cold-seep mussel
“Bathymodiolus” childressi, are similar in size to those
of shallow-water mytilids, but are about twice as
energetic (Arellano, 2008). In all of these deep-sea
bivalves, extra energy stores might play a role in ex-
tended larval life, perhaps by permitting extra time
to migrate to the euphotic zone.
Another example of significant yolk storage in a
planktotroph comes from the bathyal echinoid Aspi-
dodiadema jacobyi, which produces a ten-armed echi-
nopluteus with a posterior spine that superficially
resembles that of an irregular echinoid (Young and
George, 2000). While otherwise unremarkable in
form, this large larva comes from an unusually
small (70 µm) egg. During the blastula stage, large
mesenchyme cells migrate into the blastocoel, fill-
ing virtually all of the space. The resulting plutei are
able to develop without feeding for more than 50
days before becoming obligate planktotrophs. This
extended period of self-sufficiency is more than an
order of magnitude longer than the pre-feeding
stage of typical shallow-water plutei developing at
a similar temperature (Young et al., 1989).
16.5.2  Increasing Dispersal Through Arrested
Development
Pechenik et  al. (1984) demonstrated that some tel-
eplanic larvae arrest their growth and develop-
ment while drifting over the Atlantic, presumably
to conserve energy. Arrested development also
plays a role in extending dispersal time of two
­hydrothermal-vent animals on the East Pacific Rise.
The giant siboglinid tube worm Riftia pachyptila fer-
tilizes its eggs internally with the aid of a spermath-
ecal sperm-storage compartment (Hilário et  al.,
2005). During the time the zygotes are stored inter-
nally, no cleavage occurs; the zygotes are arrested at
the germinal vesicle stage (first meiotic prophase)
despite being already inseminated. Meiosis is com-
pleted and embryogenesis begins only after the
eggs are released into seawater. Because of very
slow cell cycles (described later), most dispersal
takes place during the embryonic period before the
larvae develop ciliation (Brooke and Young, 2009).
The alvinellid polychaete Alvinella pompeiana has an
analogous mechanism. In this case, development
is dependent on the warm seawater emitted from
a vent. While the embryos are adrift through the
cold, deep water between vent fields, their devel-
opment is arrested, presumably allowing very long
dispersal with little energy expenditure. Cleavage
begins anew when the embryos encounter warmer
temperatures indicative of the very warm habitats
occupied by the adults on the sides of hydrothermal
chimneys (Pradillon et al., 2005; 2001).
16.5.3  Controlling Dispersal Depth with Egg
Density
In shallow water, large yolky eggs such as those of
many echinoderms and cnidarians are generally
buoyant, and buoyant yolky eggs are also found
commonly in the deep sea (Young and Cameron,
1987). Egg densities for deep-sea species have sel-
dom been measured empirically, but buoyancy
measurements of large (1.1mm diameter) yolky eggs
of the echinothuriid (soft-body) echinoid Phormo-
soma placenta have been measured in the laboratory
and within chambers at several depths (Young and
Cameron, 1987). Given an average flotation rate of
0.42 cm/s, it was estimated that eggs could ascend
to the surface from 800 m in about two days. In dra-
matic contrast, the bathyal sea anemone Urticina sp.,
maintained over several years in the laboratory, pro-
vided direct evidence of benthic development in a
broadcast-spawning lecithotrophic species (Mercie­r
et al., 2016). The eggs were negatively buoyant and
remained near the parents throughout develop-
ment. Such a combination of free-living, nonfeed-
ing, and negatively buoyant larvae is unique among
the known lecithotrophic developers of the North
Atlantic (Montgomery et al., 2017b).
The siboglinid tube worm Riftia pachyptila has
lipid-filled eggs only 110µm in diameter that float at
a rate of about 2–3 m per day under ambient pres-
sures and temperatures, a rate that would retain

them in the deep ocean for the entire embryonic
period (Brooke and Young, 2009). Eggs of virtually
identical size from the methane seep tube worm
Lamellibrachia luymesi float at an average rate of 720
m per day (Young, unpublished), a rate that would
propel them to the surface in one day. These large
differences among eggs of similar size probably re-
flect differences in wax ester composition (Marsh
et al., 2001) interacting with factors such as temper-
ature and pressure that influence the density and
viscosity of the water. Because buoyancy is an es-
sential parameter for dispersal models, much more
information is needed.
16.5.4  Increasing Dispersal through Slow
Development and Long Planktonic Larval
Durations
Conventional wisdom dictates that dispersal dis-
tances are shortest among demersal developers or
brooders and that planktotrophic larvae disperse
farther than lecithotrophic larvae. Even in shallow
water, these relationships do not always hold true
(Mercier et  al., 2013). As an extreme example, the
shallow subtidal sea star Mediaster aequalis, which
has lecithotrophic larvae originating from large
yolky eggs, has been maintained in culture for
more than a year (Birkeland et al., 1971). Slow me-
tabolism coupled with trans-epidermal transport of
dissolved organics can extend the duration of lec-
ithotrophs substantially in cold water (Jaeckle and
Manahan, 1989; Shilling and Manahan, 1994). Bosch
(1989) showed that an Antarctic lecithotrophic aster-
oid in the genus Porania has a longer larval life than
its planktotrophic congener developing at the same
temperature. These and other examples (Mercie­r
et  al., 2013) suggest that lecithotrophy should not
necessarily equate with short dispersal times or
distances in the cold water of the deep ocean. This
has significance because the vast majority of deep-
sea species with planktonic development appear
to have lecithotrophic larvae (Young, 2003; Rex
and Ette­r, 2010), contrasting dramatically with the
famous calculations of Gunnar Thorson (1961) for
shallow-water species: he estimated that about 80%
of the 195 species he studied had planktotrophic de-
velopment, whereas only 10% were lecithotrophs.
This is not to say that planktotrophy is uncommon
La r va l D i s p e r sa l i n t he D ee p S ea    235
in the deep sea. Rex and Etter (2010) demonstrate
a clear increase in planktotrophic development of
deep-sea gastropods with depth; thus, in stark con-
trast to the predictions of Thorson’s rule (Thorson,
1950; Mileikovsky, 1971), planktotrophic gastro-
pods are more common on the abyssal plain of the
North Atlantic than on the continental slope where
feeding larvae would presumably have a shorter
distance to migrate for food. A high incidence of
planktotrophy is also known among bathyal echi-
noderms from tropical latitudes (Young 2003), and
10% of the 30 gastropod species identified by Mills
et al. (2007) from vents on the East Pacific Rise were
planktotrophic.
When larvae and embryos of the hydrothermal-
vent worm Riftia pachyptila were first cultured, in-
vestigators were amazed to discover that, under
normal conditions of temperature (4 °C) and pres-
sure (25 MPa), the embryos developed at a rate
of only one cleavage per day (Marsh et  al., 2001;
Brooke and Young, 2009). To investigate whether
slow developmental rates are common in the deep
sea, we assembled a large set of developmental data
for various major clades of invertebrates, then made
comparisons among species classified by depth dis-
tributions. As a starting point, we used the database
available as supplemental material in Hilário et al.
(2015), adding additional species that were missed
as well as a few unpublished data points from our
own recent work. The spreadsheets are available
from the authors.
Our comparison of cell cycle duration was lim-
ited to echinoids, the only group for which details of
developmental timing are available for a number of
species. Fortuitously, it is possible to separate depth
effects from temperature effects in echinoids because
some deep-sea species at low latitudes develop over
the same range of temperatures as ­shallow-water
species at higher latitudes. We compared bathyal
species in the Bahamas to shallow-water species in
the Northeast Pacific. Strictly speaking, depth is con-
founded with geography in these comparisons, so
conclusions should be treated cautiously; neverthe-
less, the patterns were intriguing. Mean cell cycle du-
ration was significantly longer in the deep than the
shallow species in both temperature ranges tested
(Figure 16.2A). This general trend also applied dra-
matically when echinoids from other regions were
236   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r vae

(A)

Coelopleurus floridanus

Archaeopneustes hystrix

Paleobrissus hilgardi

Linopneustes longispinus

Cidaris blakei

Stylocidaris lineata
Bathyal Bahamas

Shallow Eastern Pacific

Dendraster excentricus

Strongylocentrotus droebachiensis
10°C
Strongylocentrotus franciscanus 14-15°C

Strongylocentrotus purpuratus

0 1 2 3 4
Cell Cycle Duration (h)
(B)
8
Shallow
12

deep 8 5

0 1 2 3 4
Cell Cycle Duration (h)
(C)
8
Shallow
12

deep 9 4

0 10 20 30 40 50 60 70
Time to Hatching (h)

Figure  16.2 Cell cycle durations and time to hatching for deep-dwelling and shallow-water echinoderms at two different temperatures. (A)
Comparison of cell cycle duration between bathyal echinoids from the Bahamian Slope and shallow echinoids from the Northeast Pacific. Species from
both faunas develop over similar temperature ranges. (B) Mean cell cycle durations for a larger dataset including echinoids and asteroids worldwide
that develop at similar temperatures as those in A. Error bars are standard errors and sample sizes are indicated near each bar. (C) Comparisons
of mean times to hatching for deep and shallow echinoderms developing at two temperatures. Standard errors and sample sizes are shown as in B.

added to increase the sample size (Figure  16.2B). When we classified the echinoderm data by en-
Time from fertilization to hatching differed in the vironment (shallow polar, shallow temperate, shal-
same manner (Figure 16.2C), though the difference low tropical, and bathyal), and analyzed asteroids
at 10°C was not significant. and echinoids separately, we found similar patterns

for both echinoderm classes (Table 16.1). Deep-sea
larvae disperse for a much longer time than shal-
low-water species from either temperate or tropical
faunas (Table 16.1).
To determine the effect of bathymetric range on
dispersal distance, we classified the entire dataset by
vertical distribution, grouping species within major
taxa (phyla or classes) as shallow stenobathic, eury-
bathic (extending from shallow water into at least
bathyal depths), or deep (>200 m). With the excep-
tion of polychaetes, all major taxa for which data on
deep-sea species with planktonic development are
available showed significantly longer PLDs than
either shallow-water stenotherms or eurybathic
species (Figure 16.3). Dramatic examples of species
with long planktonic larval lives are known from
several phyla. For example, anecdotal informa-
tion indicates that larva of the ­hydrothermal-vent
crab Bythograea thermodon has the ability to survive
even at the pressures of the upper water column for
more than 10 months (Epifani­o and Dittle, personal
communication cited in Mullineaux and Manaha­n,
1999). Larvae of Sclerasterias tanneri, an asteroid
found at bathyal depths in the Gulf of Mexico
and the western Atlantic, were maintained in cul-
ture by Richard Emlet (University of Oregon) for
nearly two years (Young et  al., 2012), and disper-
sal times of more than a year have been estimated
for the gastropod Bathynerita naticoidea (Arellano
et  al., 2014), the bivalve “Bathymodiolus” childressi
Table 16.1  Mean Planktonic Larval Durations (PLD) for Echinoids and
Asteroids from Shallow Polar, Shallow Temperate, Shallow Tropical, and
Bathyal (Slope) Environments.
Environment Mean PLD S.E.
Echinoidea
polar 105.0 — 1
temperate 40.8 3.6
tropical 25.4 3.5
bathyal 114.8 15.0
Asteroidea
polar 117.4 21.0
temperate 38.8 5.6
tropical 17.5 2.4
bathyal 270.0 — 1
La r va l D i s p e r sa l i n t he D ee p S ea    237
(Arellano and Young, 2009), and the sipunculan
Phascolosoma turnerae (Rice et al., 2012), all inhabit-
ants of bathyal cold seeps.
Long planktonic larval lives are not found ex-
clusively in the deep sea. The shallow-water
ranellid gastropod Fusitriton oregonensis, for exam-
ple, can spend at least 4.5 years in the plankton
(Strathman­n and Strathmann, 2007), and several
species including architectonicid gastropods and
sipunculans from shallow water are known to drift
for long distances, possibly across ocean basins,
suggesting very long larval lives. Nevertheless,
of those species whose PLDs have been measured
empirically or estimated from indirect methods,
long larval life appears to be the rule rather than
the exception in deep water.
16.5.5  Does Release from Predation Permit
Long Dispersal Times in the Deep Sea?
One possible explanation for the evolution of slow
development and extended periods of larval dis-
persal is that embryos and larvae experience lower
mortality in the plankton than their shallow-water
relatives. Strathmann et al. (2002) have argued that
embryos at low risk evolve slower developmental
rates. In a large experiment with multiple evolution-
ary contrasts, Staver and Strathmann (2002) showed
that embryos developing in the water column had
shorter cell cycles than protected embryos living
in a less perilous benthic environment. A similar
process might explain the dramatic differences in
developmental rate and larval life between shallow
and deep water. Figure 16.4 shows the vertical dis-
tributions of three types of potential larval preda-
N tors (Young and Chia, 1987) from our own replicate
MOCNESS (multiple opening-closing net) plankton
samples in the Gulf of Mexico and the Bahama­s.
Fish, euphausiids, and chaetognaths were all more
13 abundant in the euphotic zone above the perma-
22 nent thermocline (~200 m) than at bathyal depths
3 as deep as 900 m. In the most dramatic example,
chaetognaths represent a potential predation pres-
5
sure about 200–300 times higher above the thermo-
cline than anywhere in the deep sea. Euphausiids
16
are also hundreds of times more abundant in the
22
euphotic zone. Visual predators including chae-
tognaths, myctophids, and euphausiids might
238   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r vae

800 160 50 50
Asteroidea 3 Echinoidea Holothuroidea Ophiuroidea
1 140 9
10
40 40
Maximum PLD (days) 600 120

100 30 30
400 80
19
60 20 20
16
6
200 40 43
10 10
16 20
41
0 0
0 0 0 0
D E S D E S D E S D E S
350 200 120 100
1 Brachyura Anomura Caridea Cirripedia
300 2 3
100
80
Maximum PLD (days)

250 150
80
200 60
2
100 60
150 5
6 13 40
40 9
100 4
19 50
20 20
50
0 0
0 0 0 0
D E S D E S D E S D E S
250 500 50
140 Polychaeta
1 Gastropoda Bivalvia Cnidaria
1
200 400 120 40
Maximum PLD (days)

5
100
150 300 30 5
20
80
6
100 200 60 4 20
13
40
50 2 100 10
12 20
0 0
0 0 0 0
D E S D E S D E S D E S

Figure  16.3 Means and standard errors of maximum PLD (planktonic larval duration) values for 298 invertebrate species with planktonic
development and belonging to major deep-sea taxa. Each species is classified by its bathymetric range as shallow (S; limited to shallow seas within
the euphotic zone), shallow eurybathic (E; extending from shallow water to at least upper bathyal depths, and deep (D; limited to bathyal and abyssal
depths). Sample sizes are indicated above each bar. The gastropod data do not include shallow species with teleplanic larvae such as ranellids and
architectonicids. These have very long larval lives but PLDs have been documented for only a few species.

also be unable to prey upon transparent larvae in
the darkness of the deep sea. Although ecological
release might explain why long larval durations are
possible in the deep sea, there must also be a selec-
tive advantage for evolving slower development
since certain deep-sea clades have been proposed to
originate from shallow-water ancestors.
16.6  Ontogenetic Vertical Migration
and Its Evolutionary Significance
Long PLDs alone do not necessarily equate to long
larval dispersal distances (Shanks, 2009). In shallow
water, horizontal larval transport is often medi-
ated by behaviorally regulated vertical migrations
 La r va l D i s p e r sa l i n t he D ee p S ea    239

(A)
25000
Fish
Euphausiids
2000 Chaetognaths

600
Mean Abundance

500

400

300

200

100

0
0-150 150-250 250-350 350-450 450-550 550-650 650-750 750-900

(B) Depth (meters)

Fish
Euphausiids
2000 Chaetognaths

600
Mean Abundance

500

400

300

200

100

0
0-50 50-100 100-200 200-300 300-400 400-500 500-600
Depth (meters)

Figure 16.4 Bathymetric distributions of three planktonic taxa known to be important predators on larvae. Samples were collected by MOCNESS
opening-closing plankton nets in the Bahamas (A) and the Gulf of Mexico (B). Error bars are standard deviations. The fishes were mostly small
myctophids (lantern fish).
240   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r vae
by larvae among water strata that may result in ei-
ther dispersal or retention (reviewed by Young and
Chia, 1987; Shanks, 1995; Pineda et  al., 2007). Evi-
dence for ontogenetic vertical migration, defined
as a change in vertical distribution with larval age,
in the deep sea has accumulated over the past two
decades, but details are lacking. We do not fully un-
derstand the timing of migrations for any deep-sea
species, nor do we know how prevalent migrations
are. Evidence that they occur, mostly consisting of
larval capture far off the bottom, has been summa-
rized elsewhere (Bouchet and Warén, 1994; Adams
et al., 2012). Here we discuss potential causes and
evolutionary advantages of ontogenetic migration.
16.6.1  Ontogenetic Vertical Migration May
Be a Phylogenetically Constrained Character
Fossil records and phylogenetic analyses of cer-
tain clades suggest invasion and diversification
of the deep ocean by multiple shallow-water taxa
over different timescales (reviewed in Brown and
Thatje, 2014). Limited data for some deep-water
crustaceans, molluscs, and echinoderms suggest
that reproductive patterns and early life history
strategies, including development mode and larval
feeding strategies, are phylogenetically constrained
characters (Bouchet and Warén, 1994; Eckelbarger
and Watling, 1995; Tyler and Young, 1999; Young
2003). Likewise, ontogenetic vertical migrations are
common behaviors in shallow-water taxa and, thus,
may simply be a remnant of evolutionary history
for deep-sea species.
The strongest evidence of larval migrations to the
euphotic zone comes from two taxa, crustaceans
and molluscs, with many examples of larval migra-
tory behaviors in shallow water. Larvae of shallow-
water brachyuran crabs change their behavioral
responses to light, gravity, and hydrostatic pressure
with age to match specific demands for horizontal
transport (reviewed by Epifanio and Cohen, 2016).
Some of the best evidence for long ontogenetic ver-
tical migrations is for deep-sea crabs (Kelly et  al.,
1982; Dittel et  al., 2008; Pochelon et  al., 2014). For
example, the hydrothermal-vent brachyuran crab
Bythograea thermydron maintains eye pigments sen-
sitive to the light spectra characteristic of shallow
water (Dittel et  al., 2008), and is able to tolerate
surface pressures despite the fact that larvae are re-
leased at 25 MPa.
Among shallow mytilid mussel larvae, swim-
ming responses to light, gravity, and pressure
change through ontogeny and result in upward
swimming as early veligers and bottom swim-
ming in competent veligers (Bayne, 1963; 1964).
Deep-sea hydrothermal-vent and cold-seep mussels
(bathymodiolinae) are estimated to have radiated
from shallow-water mytilid mussels as organic-fall
specialists (Distel, 2000; Lorion et  al., 2010) just 21
million years ago (Miyazaki et al., 2010). It is not sur-
prising, then, that bathymodiolin veligers have been
found in surface tows in the Gulf of Mexico, more
than 600 m above their adult habitat (Arellano et al.,
2014; Maslakova and Young, unpublished data), de-
spite a paucity of evidence that these planktotrophic
larvae require the algal food found in surface waters
(Salerno et al., 2005; Arellano and Young, 2009). The
morphology of the protoconch suggests that many
abyssal gastropods are planktotrophic (Bouchet and
Warén, 1994), and data on oxygen isotopes indicate
that some of these veligers migrate into warmer wa-
ters to feed (Bouchet and Fontes, 1981; Killingley
and Rex, 1985), observations that have been borne
out by the collection of abyssal larvae in the surface
plankton (Bouchet and Warén, 1994).
16.6.2 Planktotrophic Larvae of Deep-Sea
Animals May Require Migration to Reach
Productive Surface Waters
Moseley (1880) first suggested that larvae of deep-
sea animals might migrate into the euphotic zone to
feed and develop. His hypothesis, though, was over-
shadowed by Thorson’s (1950) assertion that such
long migrations would be energetically impossible.
Long migrations will require substantial energy and
those costs should be even greater for those organ-
isms that remain negatively buoyant throughout
their development (e.g. crabs; Queiroga and Blan-
ton, 2004). Migrating larvae of deep-sea organisms
must, therefore, either pack their eggs with the re-
quired energy to reach the surface before feeding or
use deep-water food sources along the migration.
Young et al. (1996) proposed an energetics model
that predicts the cumulative energy consump-
tion of an upward-migrating larva as a function of
 La r va l D i s p e r sa l i n t he D ee p S ea    241

swimming speed and temperature-dependent met-
abolic rate. The cumulative energy consumed can
be compared to the vertical distance the larva could
swim in a given time and the initial energy avail-
able in the egg, in order to determine the maximum
vertical distance possible for a larva to migrate
without feeding. In simulated migrations of bath-
yal sea urchin larvae, Young et al. (1996) used meta-
bolic and swimming rates taken from the literature
on shallow-water urchin larvae to make predictions
on whether the bathyal larvae could migrate to a
depth at which they could obtain enough energy to
balance their metabolic needs. In all cases, larvae
with a high (though potentially realistic) metabo-
lism swimming at a low (yet potentially realistic)
speed exhausted the energy in the egg long before
reaching the compensation depth, but could reach
the compensation depth even at high metabolism
if they swam as fast as some of their shallow-water
relatives (Young et al., 1996).
However, the boundaries of these model results
are sufficiently wide to accommodate virtually
any conclusion, because none of the model param-
eters were measured empirically. Using empirically
derived metabolic rates of cultured trochophores
of the deep-sea, cold-seep mussel “Bathymodiolus”
childressi, Arellano (2008) predicted that a “B.” chil-
dressi larva swimming upward from 650 m depth
could reach the surface within 10 to 13 days, assum-
ing “B.” childressi veligers swim at mean swimming
velocity for Mytilus edulis veligers, before consum-
ing the energy available in the egg (Figure  16.5).
This conclusion is largely possible due to the fact
that preliminary estimates suggested that the eggs
of “B.” childressi are more than twice as energetic
as those of their shallow-water mytilid relatives
(Arellan­o, 2008), providing sufficient energy to
make a long vertical migration at wide range of
metabolic and swimming rates even without feed-
ing along the way. So far the parameters important
to deep-sea feeding, metabolism, and swimming
have not all been measured empirically for any spe-
cies of deep-sea larva throughout its entire larval
development. Recent advances in precise sampling
of larvae (e.g., the SyPrid sampler on the Sentry
AUV; Billings et al., 2017) should make it possible to
collect viable larvae in sufficient numbers to meas-
ure these metabolic and swimming rates.

Trochophore velocities Veliger velocities

0
–1
0.105 m h 3.96 m h –1

100 0.510 m h –1 7.20 m h–1

1.06 m h–1
200
Depth (m)

300

400

500

600

0 1 2 3 0 1 2 3
Cumulative energy consumed (mJ)

Figure 16.5 Predicted cumulative energy consumption (mJ) as a function of depth for “Bathymodiolus” childressi based on measured temperature-
dependent respiration rates and swimming speeds in cultured trochophores (Arellano, 2008). We used a starting depth of 650 m and temperature of
7 °C, and ranges of swimming speeds measured for “B.” childressi trochophores (left; Arellano, 2008) and mean (Konstantinova, 1966) and maximum
(Sprung, 1984) swimming speeds reported for Mytilus edulis veligers (right). For veligers (right), we used a starting depth of ~575 m (the depth after
six days of swimming as trochophores; Arellano and Young, 2009). Points on the curves are 24 hours apart. The solid vertical lines represent the mean
total energy measured in the “B.” childressi egg with standard deviations represented by dotted lines.
242   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r vae

Planktotrophic larvae of deep-sea organisms,
however, probably use diverse food sources rather
than relying solely on photosynthetic food. We
showed that cultured larvae of the deep-sea, cold-
seep snail Bathynerita naticoidea, which migrates
to the surface (Arellano et  al., 2014), can feed on
phytoplankton (Figure 16.6A; Van Gaest, 2006), but
similar attempts to feed algae to “Bathmodiolus”
childressi veligers were unsuccessful (Arellano and
Young, 2009).
Heterotrophic bacteria also represent a poten-
tially important food source for planktotrophic lar-
vae, and these prokaryotes are abundant in cooler
waters below the deep chlorophyll maximum, as
well as in the surface layer. In fact, the abundance
of free-living heterotrophic bacteria in the mesope-
lagic zone is nearly as high as in the euphotic zone
(Cho and Azam, 1988; DeLong and Pace, 2001).
Larval bacterivory was first reported in Antarctic
starfish larvae (Rivkin et  al., 1986) and was inter-
preted as an adaptation to very oligotrophic waters.
Subsequently, the larvae of a common fouling poly-
chaete, Hydroides elegans, has been shown to be able
to complete metamorphosis with an exclusive bac-
terial diet during portions of its extended breeding
season when phytoplankton are rare (Gosselin and
Qian, 1997), and some larval bivalves can obtain sig-
nificant amounts of energy from Synechococcus spp.
(Baldwin and Newell, 1991; Gallager et  al., 1994).
We have shown that some deep-sea echinoderm and
polychaete larvae consume species of Synechococcus,
Prochlorococcus, and heterotrophic bacteria (Pile and
Young, 2006; Young, Bosch, and Cameron, unpub-
lished) and that two species of bathyal gastropod
veligers feed opportunistically on cultured Synecho-
coccus sp. (Figure 16.6; Arellano and Krodel, unpub-
lished data for Shinkailepas n. sp.; Cziko and Young,
unpublished data for a naticid), suggesting that
members of the microbial loop in the mesopelagic
zone might provide an important source of food for
bathyal larvae. Thus, ciliated invertebrate larvae
may play a similar role as tintinnids and other mid-
water protists in planktonic food chains by grazing
directly on the microbes of the microbial loop.
16.6.3  Ontogenetic Vertical Migrations Impact
Biogeography by Mediating Transport
or Retention of Larvae
Larval dispersal influences gene flow among popu-
lations, which, on evolutionary timescales, affects
biogeographic patterns and ultimately speciation.
Ontogenetic vertical migrations by deep-water
species may mediate dispersal distances and pop-
ulation connectivity, with very long-distance dis-
persal becoming possible through access to faster
surface currents, while larvae that remain in slow
near-bottom flows presumably have more limited

(A) (B)

50 μm

Figure 16.6  Epifluoresence migrographs indicating larval feeding in two deep-sea gastropods. (A) Bathynerita naticoides veliger larva cultured from
egg capsules collected at Brine Pool NR1 cold seep in the Gulf of Mexico, fed a mixed diet of Thalassiosira pseudonana and Isochrysis galbana (Van
Gaest, 2006). (B) Veliger larva of Shinkailepas n. sp. cultured from egg capsules collected from NW Rota-1 hydrothermal vent on the Mariana Arc.
Bright spot in the gut is the autofluroescence of Synechococcus Strain CC9311 (Arellano and Krodel, unpublished data) (see Plate 14).

dispersal (reviewed in Adams et  al., 2012; Hilário
et al., 2015). Whether long-distance dispersal is ad-
vantageous for deep-sea species continues to be de-
bated, though, particularly for species that inhabit
spatially fragmented and specialized habitats, like
chemosynthesis-based ecosystems (whale falls, hy-
drothermal vents, and cold seeps). On one hand, ex-
tensive larval transport may lead to larval wastage
as larval propagules are swept away from their spe-
cialized habitats, a hypothesis that initially drove
the thinking about larval dispersal at hydrothermal
vents (Lutz et al., 1980; Turner et al., 1985). On the
other hand, intermittent long-distance dispersal
can explain the broad biogeographic ranges of most
deep-sea taxa (McClain and Hardy, 2010; Young
et al., 2012; Arellano et al., 2014; Mitarai et al., 2016).
Model simulations of larval transport that in-
corporate vertical larval migrations into the upper
water columns layers have been used to suggest
that long vertical migrations of larvae may result
in infrequent connectivity of deep-sea populations
that could play an important role over evolution-
ary timescales (McClain and Hardy, 2010; Young
et  al., 2012; Arellano et  al., 2014; Mitarai et  al.,
2016). Arellano et al. (2014) suggests that the broad
amphi-Atlantic distribution of a member of the
­ near the seafloor. Citing earlier work with infaunal
“Bathymodiolus” species complex, “B.” mauritanicus
(Cordes et al., 2007; Olu-Le Roy et al., 2007; Génio
et al., 2008; Olu et al., 2010) may have resulted from
historical connectivity aided by intermittent wide-
spread dispersal of long-lived larvae that migrate
into surface waters. Indeed, model simulations us-
ing empirically derived PLDs and depth distribu-
tions of “B.” childressi larvae suggest that dispersal
within the top  100 m of the water column can re-
sult in larval transport up to 1,000 km (Young et al.,
2012; McVeigh, 2016). Model simulations by Mitarai
et al. (2016) suggest that ontogenetic migrations by
vent larvae from the otherwise isolated Okinawa
Trough to above 600 m depth would allow the
Kuroshio Current to connect them with vents in
the Izu-Bonin Arc, which is 1,200 km away. These
connections would be rare, however—once every
tens to thousands of years—which is not frequent
enough to prevent genetic differentiation between
barnacle populations in both basins (Mitarai et al.,
2016). Conversely, hydrothermal-vent larvae within
other western Pacific basins (Manus, Lau, and
La r va l D i s p e r sa l i n t he D ee p S ea    243
North Fiji) do not need to undergo long vertical
migrations to enable connections between basins.
Regardless of dispersal depth, model simulations
suggest rare unidirectional connections between
basins that agree with the genetic data for popula-
tions of the vent gastropod Ifremeria nautili within
these basins (Thaler et al., 2011; Mitarai et al., 2016).
Less evidence of larval behaviors enhancing lo-
cal retention is available for deep-sea species. Etter
and Bower (2015) suggest that larval behaviors may
explain the strong genetic divergence of bathyal
protobranch bivalves populations along depth gra-
dients in the western North Atlantic because this di-
vergence cannot be explained by modeled transport
of passive larvae, but they cannot rule out pre- and
post-settlement selection processes either.
16.6.4  Demersal Drift May Either Retain
Larvae or Increase Encounter Rates for Isolated
Substrata
Demersal development, either for the entire lar-
val period or part of it, may limit dispersal, but
field evidence of demersal development is rare
because plankton samples are difficult to collect
polychaetes and polar echinoderms, Mileikovsky
(1971) identified demersal development (a term
coined by Pearse, 1969) as one of the main catego-
ries of larval development. Demersal development
has been proposed for various Antarctic larvae
(Pearse and Giese, 1966; Pearse, 1969; Picken, 1980;
Clarke, 1983; 1987; Bosch, 1989; Marsh and Mana-
han, 2000), though the hypothesis has now been
rejected for some (Pearse and Bosch, 1986) and the
data remain equivocal for others. Scheltema (1994)
proposed that deep-sea planktotrophic larvae
might remain near the bottom, feeding on hetero-
trophic microbes, protists, or detritus rather than
migrating into a food-poor, deep-water column.
Shimek’s (1986) in vitro observations of feeding in
the planktotrophic veligers of a eurybathic turrid
gastropod provided strong evidence for this idea
(the turrids are the most abundant family of gas-
tropods in the deep sea), and Bouchet and Warén
(1994) suggested that many deep-sea lecithotrophs
are likely to be demersal as well. Citing these and
other bits of early evidence, Rex and Etter (2010)
244   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e La r vae
speculated that demersal drift of lecithotrophic lar-
vae is probably the dominant form of dispersal in
the deep ocean.
Whether demersal development would aid in
retention is unclear. At low temperatures near the
bottom, decreased metabolic rates would lead
to longer PLDs (O’Connor et  al., 2007a) and con-
sequently longer dispersal times. Few studies
of near-bottom dispersal of deep-sea species are
available, and those that are take place within the
complex bottom flow systems present around hy-
drothermal vents. Studies at hydrothermal vents
on the East Pacific Rise (EPR) show that vent larvae
are more abundant near the bottom than higher in
the water column (Mullineaux et al., 2005). Within
the valleys of the EPR, where near-bottom current
velocities are weak (Adams et al., 2012), larval sup-
ply data provide evidence of local retentions of
hydrothermal-vent gastropods and very limited
­ appear to be pelagic lecithotrophs.
dispersal (<1 km; Adams and Mullineaux, 2008).
Conversely, strong currents along ridge flanks can
transport bottom-dwelling larvae over hundreds
of kilometers, whereas larvae that migrate to the
height of the plume are more likely to be retained
near the ridge crest (McGillicuddy et  al., 2010).
Thus, in the case of fauna that inhabits vents along
mid-ocean ridges, demersal swimming behaviors
may actually favor long-distance connectivity
among vent sites along the ridge.
We suspect that for many species, the optimal
dispersal strategy is mixed, involving both vertical
migration and demersal drift. Larvae nearing com-
petency should spend extended periods dispersing
near the bottom (Figure  16.1), where long larval
lives mediated by low temperatures and minimal
metabolic costs could greatly increase the likeli-
hood of finding suitable settlement sites.
The influence of long planktonic larval durations
and ontogenetic vertical migration behaviors on
gene flow of deep-sea species cannot be understood
until we have a better grasp on where the larvae of
deep-sea species spend their entire larval periods.
Despite the rapidly expanding interest in modeling
larval dispersal using biophysical models and sen-
sitivity analysis (reviewed by Hilário et  al., 2015),
larval trajectories will remain within the realm of
speculation until we have empirical data on the
timing of migratory behaviors in deep-sea larvae.
16.7 Summary
1. Genetics have given important insights into
connectivity and phylogeography of deep-sea
populations, but the underlying mechanisms
remain poorly understood.
2. Egg laying and brooding have been docu-
mented in most major deep-water taxa but re-
main proportionally less frequent than reports
of planktonic development. A major taxon of
deep-sea animals, the peracarid crustaceans,
are all brooders because of phylogenetic con-
straints. Brooding is also common in poly-
chaetes, but appears relatively infrequent in
echinoderms and molluscs.
3. Planktotrophic larval development is not un-
common in the deep sea, though the vast major-
ity of species with free-swimming development
4. We hypothesize a bimodal distribution of dis-
persal strategies among animals living in spe-
cialized, isolated habitats such as methane
seeps and dropstones. Such animals should
have either very short dispersal for retention or
very long larval lives to increase the probability
of finding settlement sites.
5. Development is significantly longer in the deep
sea than in shallow water, as measured by cell
cycle duration, time to hatching, and plankton-
ic larval duration (PLD).
6. We hypothesize that low predation pressure in
the deep sea has permitted slow development
and long larval lives.
7. The planktotrophic larvae of several deep-sea
molluscs and echinoderms develop from eggs
with a surprisingly high maternal energy in-
vestment.
8. Arrested development extends dispersal time
in some deep-sea species.
9. Several deep-sea larvae have been found in the
upper water column, suggesting that larvae
may migrate vertically over ontogeny.
10. Demersal development also appears to be a via-
ble dispersal strategy in the deep ocean, though
research has been hampered by the difficulty of
collecting plankton near the sea floor. Labora-
tory work has recently provided evidence in
support of this hypothesis.

11. Biophysical models have become popular for es-
timating dispersal and connectivity, but the bio-
logical parameters of the models are generally
inadequate. Good models will require empirical
information on the movements and depth distri-
butions of larvae through ontogeny. Such infor-
mation is not available for any deep-sea species.
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 C h a pt er 17
Larval Ecology in the Face of Changing
Climate—Impacts of Ocean Warming
and Ocean Acidification
Maria Byrne, Pauline M. Ross, Symon A. Dworjanyn, and Laura Parker
17.1 Introduction
The Earth’s marine ecosystems are being altered by
anthropogenic carbon emissions, which are driving
climate change and ocean acidification (OA) (IPCC,
2014; Gattuso et al., 2015). Climate change is causing
warming of the air and ocean, altering ocean circu-
lation and increasing heat waves (Coma et al., 2009;
Garrabou et al., 2009; Smale and Wernberg, 2013). Ac-
cording to the RCP2 (stringent scenario to keep global
temperature increase below 2 °C in the twenty-first
century) and RCP8.5 (business as usual) CO2 emis-
sions scenarios, ocean warming (OW) of 1.2 °C and
3.2 °C, and OA of 0.14 pH and 0.4 pH units, will occur,
respectively (IPCC, 2014; Gattuso et al., 2015).
Marine invertebrates are exhibiting a range of
responses to OW and OA, including local popula-
tion declines and extirpations, altered planktonic
dispersal pathways leading to changes in species
distribution, changes in phenology (e.g., time of
spawning), declines in calcification, and decreased
body size (Philippart et  al., 2003; Burrow­s et  al.,
2011; Sheridan and Bickford, 2011; Byrne et  al.,
2013a; Smale and Wernberg, 2013). As many spe-
cies have a benthic-pelagic life cycle, climate
change impacts on larvae can have major impli-
cations for the persistence of populations and
marine ecosystem function (Byrne, 2011; Harley,
2011; Kroeker et  al., 2013). Thus far, most climate
change–driven alterations to marine biodiversity
observed in nature are ascribed to increased tem-
perature (Poloczanska et al., 2016).
Byrne, M., Ross, P. M., Dworjanyn, S. A., and Parker, L., Larval ecology in the face of changing climate—impacts of ocean warming and
ocean acidification. In. Evolutionary Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas
Heyland: Oxford University Press (2018). © Oxford University Press.DOI: 10.1093/oso/9780198786962.003.0017
As the coasts and oceans warm and the veloc-
ity of climate change increases, marine ecosystems
are changing in unpredictable ways. Some species
are moving poleward or to deeper water, follow-
ing their thermal niche (the range of temperatures
where they function optimally), while others re-
main, many unable to migrate (e.g., polar species
at a thermal “dead end”) (Burrows et al., 2011; Sun-
day et al., 2012; 2015; Poloczanska et al., 2013; 2016).
­Species distributions are also contracting from lower
latitudes (Sunday et al., 2012; Smale and Wernberg,
2013), as seen in the decline of bivalves attributed
to climate-driven declines in recruitment (Philip-
part et  al., 2003). Marine larvae are following, or
having greater success in more favorable poleward
isotherms (Figure  17.1). Neo-expander­ s (natural
invasions not directly mediated by humans; Sorte
et al., 2010), colonizing through new larval migra-
tion pathways, can be highly successful in their
new location with negative impacts on receiving
ecosystems, as seen in the invasion of barrens form-
ing sea urchins to Tasmania (Ling et al., 2008; Banks
et  al., 2010). As coastal regions warm, a transition
to thermotolerant species is predicted, along with
local ­extinctions of less tolerant species (Poloczan-
ska et al., 2013). These trends are seen worldwide in
the tropicalization of temperate faunas, with tropi-
cal species such as corals invading kelp ecosystems
(Yamano et al., 2011; Baird et al., 2012; Vergés et al.,
2014). C ­ limate-driven change in propagule pres-
sure is creating novel communities, novel species
interactions, and ecological mismatches (Philippart
252   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert ebrat e L ar vae

et al., 2003; Harley, 2011; Burrows et al., 2011; Sun-
day et al., 2012; 2015; Poloczansk­a et al., 2013; García
Molinos et al., 2016).
Studies suggest that decreasing body size may
be a general ecological response to global warming
due to the inverse relationship between tempera-
ture and body size in ectotherms (temperature-size
rule; Huey and Stevenson, 1979; Sheridan and
Bickfor­d 2011). This counterintuitive relationship
holds for many marine ectotherms living well
within their thermal tolerance, with conspecifics
from cooler climes or larvae from cooler seasons be-
ing larger than those from warmer climes/seasons
(Irie and Fischer, 2009; Sheridan and Bickford, 2011).

–10 Acanthaster planci

72h

–15

Arachnoides placenta
60h
27

–20

Centrostephanus rodgersii
23 72h
% Normal Development

–25
Latitude

Heliocidaris tuberculata
–30 72h
19

Heliocidaris erythrogramma
–35
72h

17

–40 15
Asterias amurensis
72h
13

11

–45
142 144 146 148 150 152 154 156 158 160

Longitude
Temperature (°C)

Figure 17.1 Thermal envelopes for development to the early larval stage (60–72 hours post-fertilization) in tropical and temperate echinoderms
that occur along the east coast Australia, and mean isotherms (adapted, with permission, from data in Pecorino et al., 2013; Hardy and Byrne, 2014;
Hardy et al., 2014; Lamare et al., 2014).
 Cha n g i n g Cl i mat e — O c ea n W arm i n g a n d A c i d i f i c at i o n    253
For larvae, stressful temperatures nearing ther-
mal limits also stunt growth (Nguyen et  al., 2012;
Kamy­a et al., 2014; Karelitz et al., 2016). Decreased
larval body size also results from OA, especially for
calcifying larvae where the energetic constraints to
produce a skeleton under CO2-driven hypercapnia
and reduced calcium carbonate mineral saturation
state (Ω) and teratogenic effects impair calcification
and growth (Byrne et al., 2013a; Parker et al., 2013;
Stumpp et al., 2013; Whittman and Pörtner, 2013).
Marine invertebrate larvae live in a multistressor
world and it is important to understand the inter-
active effects of climate change–related stressors
(Przeslawski et  al., 2015). Priority stressors differ
regionally. For instance, acidification is the major
concern for upwelling and polar regions, while
warming is the major stressor for western bound-
ary current regions experiencing incursions of
warm tropical water, and hypoxia is associated
with warming in eutrophic areas (Ridgway, 2007;
Yamano et al., 2011; Wu et al., 2012; Melzner et al.,
2013; Waldbusser and Salisbury, 2014). Thus, it is
essential to have a regional and habitat level un-
derstanding of the present-day environmental con-
ditions that larvae experience and how these are
changing, and will continue to change, as the cli-
mate warms.
This chapter considers the impacts of two major
ocean change stressors, warming and acidification,
on marine invertebrate larvae. The burgeoning lit-
erature on the responses of marine invertebrates to
OW and OA has been the subject of many reviews
(e.g., Byrne, 2011; Ross et  al., 2011; 2016; Kroeker
et al., 2013; Byrne and Przeslawski, 2013; Whittma­n
and Pörtner, 2013; Parker et  al., 2013). A recent
meta-analysis on the stress ecology of invertebrate
development quantified the effects of climate-­
related stressors on early life stages (embryo to lar-
vae) based on studies where more than one stressor
was used (Przeslawski et  al., 2015). This analysis
showed that there are differences among the life
history stages in their sensitivity to OW and OA and
that molluscs and echinoderms are more vulnerable
than arthropods and cnidarians (Przeslawski et al.,
2015). As the Echinodermata and the Mollusca are
ecologically and commercially important phyla, we
focus here on the impacts of climate change and
ocean acidification on their larvae and implications
for larval ecology. In many regions, marine species
with pelagic propagules are on the move, exhibit-
ing a poleward trend as sea surface temperatures
rise and ocean currents change (Poloczanska et al.,
2013; García Molinos et  al., 2016). This emerging
phenomenon is also a focus of this review.
17.2  Global Warming—Larvae
on the Move
Temperature is the major driver of biological pro-
cesses and is significant to marine invertebrate de-
velopment in controlling developmental timing, the
length of the dispersive stage, larval energetics, and
survival, and it also plays a key role in recruitment
dynamics (Pechenik, 1987; Hart and Scheiblin­ g,
1988; O’Connor et  al., 2007; Byrne et  al., 2011a;
Hardy et al., 2014; Kapsenberg and Hofmann, 2014).
Within thermal tolerance limits, developmental rate
increases with warming due to enhanced physi-
ological rates, and so planktonic durations shorten
at higher temperature (O’Connor et al., 2007; Byrn­e,
2011; Kendall et  al., 2016). This, however, may re-
quire the nutritive regime to be able to support
growth. The sensitivity of development to increased
temperature varies among developmental stages
and species (Byrne and Przeslawski, 2013). This
differential sensitivity influences survival, invasive
potential, faunal shifts, and community function in
a changing ocean.
There is a tight relationship between tempera-
ture and spawning for many marine invertebrates,
the timing of which often corresponds to optimal
temperatures for larval development (Johnson and
Babcock, 1994; Reitzel et  al., 2004; Byrne, 2011).
For species with planktotrophic larvae, spawn-
ing also often coincides with the period of opti-
mal food availability. Changes in spawning due
to ocean warming and the resultant altered tim-
ing of larvae in the plankton can lead to a trophic
mismatch, with planktotrophic larvae appearing
earlier than the plankton blooms they depend on,
as seen for echinoid, gastropod, and bivalve larvae
(Philippart et  al., 2003; Edwards and Richardson,
2004; Kirby et al., 2007; Moore et al., 2011). Change
to phytoplankton community structure is a major
outcome of changing climate (Montes-Hugo et al.,
2009). Thus, species with feeding larvae may be
254   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert ebrat e L ar vae
particularly vulnerable unless spawning times can
be adjusted to match phytoplankton availability.
As species with nonfeeding lecithotrophic lar-
vae do not depend on the vagaries of a planktonic
food supply, they may have an advantage in a cli-
mate change ocean (Hardy and Byrne, 2014). For
echinoderms, this advantage is indicated by line-
age survival through past climate change–driven
extinction events (see Uthicke et al., 2009). Overall,
it appears that nonfeeding, noncalcifying larvae
such as the lecithotrophic larvae of the asteroids
and echinoids and coral planula larvae are more
resilient to OA (Dupont et al., 2010; Nguyen et al.,
2012; Putnam et  al., 2013; Hardy and Byrne, 2014;
Przeslawsk­i et al., 2015).
Parental conditioning, the temperature at which
gametes develop, influences the optimal tempera-
ture for embryonic and larval development (Mita
et  al., 1984; Fujisawa, 1989; 1995; Hamdoun and
Epel, 2007; Rahman et al., 2009; Byrne et al., 2011a).
Gametic acclimatization, particularly that of the
egg, can dramatically shift the thermotolerance of
progeny, and so can shape habitat and regional dif-
ferences in optimal temperatures for development,
even within species (Fujisawa, 1989; 1995; Johnson
and Babcock, 1994; Bingham et  al., 1997; Rahman
et al., 2009; Zippay and Hofmann, 2010; Byrne et al.,
2011a). With respect to OW, the offspring of moth-
ers that experience warm conditions during egg de-
velopment perform better in these same conditions
(Bingham et  al., 1997; Burgess and Marshall, 2011).
Thus, maternal thermal history influences the phe-
notype of offspring to enhance their performance
under thermal stress. Whether this is due to overt
manipulation by the mother to protect offspring
(Burgess and Marshall, 2011) or due to a hardwired
genetic acclimatization (and potentially epigenetic)
response (Hamdoun and Epel, 2007) is not known.
For species with broad latitudinal distributions, the
concept of physiological differences among popula-
tions, with metabolic temperature compensation to
different thermal regimes, suggests the potential that
developmental stages may have substantial thermal
plasticity to cope with warming (Vernberg, 1962; By-
rne et al., 2011a).
The strong relationship between fitness and
growth/performance traits in ectotherms in re-
sponse to temperature can be represented in
thermal reaction norms (Figure 17.1) and in the bio-
energetics framework based on changes in aerobic
scope with temperature (Pörtner and Farrell, 2008:
Pörtner, 2012; Sokolova, 2013). Determination of the
thermal envelope (cool to warm tolerance) of devel-
opment with respect to present-day and projected
future marine isotherms during the planktonic
phase across distribution ranges is crucial to predict
the distribution of larvae under projected future
marine isotherms. Most studies have focused on
upper thermal thresholds for development (Byrn­e,
2011). As a result, the data on cool tolerance re-
quired to model potential for poleward range shifts
mediated by larval transport are limited (Bates
et al., 2013). Upper thermal limits for development
provide insights into the risk of local extinction/
range contraction as OW increases, while lower
limits provide insights into potential for poleward
range extension through dispersal of planktonic
larvae. The latitudinal distribution of many marine
ectotherms is strongly influenced by larval ther-
mal tolerance limits (Andronikov, 1975; Jones et al.,
2009; Sunday et al., 2012), and so this information is
important to assess how adult population distribu-
tion may be altered by OW (Byrne et al., 2016).
There is a predicted relationship between physi-
ological tolerance and latitudinal range such that
species with extensive latitudinal distributions are
expected to have broader thermal tolerance limits
than smaller-range species (Stevens, 1989; Somero,
2012). Broadly distributed species that have a wide
developmental thermotolerance appear best able to
avail themselves of the new opportunities provided
by OW (Bates et  al., 2013; Gianguzza et  al., 2014;
García Molinos et al., 2016), as shown for a suite of
tropical and temperate echinoderms that occur along
the east coast of Australia (Figure 17.1). This region
is a warming hotspot, where increasing tempera-
tures are exacerbated by increased flow of a tropical
western boundary current, and where marine heat
waves generated by strong warming of the continent
(5 °C by 2070) are also a risk (Hobda­y and Lough,
2011). The thermal tolerance of embryos and lar-
vae were characterized across a broad temperature
range (12 °C + temperatures) (Pecorino et al., 2013;
Hardy et al., 2014; Lamare et al., 2014; Karelitz et al.,
2016). For all six species studied, the reaction norms
for successful larval development approximated a
 Cha n g i n g Cl i mat e — O c ea n W arm i n g a n d A c i d i f i c at i o n    255
flat normal distribution encompassing a breadth of
temperatures where development rate and larval
growth are optimal, with slower or no development
in the cool range and high abnormality or mortal-
ity in the warm range. A similar thermal envelope
profile is characteristic of development in other as-
teroids and echinoids, including tropical, temper-
ate, and polar species (Hoegh-Guldberg and Pearse,
1995; Stanwell-Smith and Peck, 1998; Delorme and
Sewell, 2013). Due to decreased development time
with warming, Karelitz et  al. (2016) suggest that
near-future OW may even improve prospects for
some species by reducing the time in the plankton,
thereby enhancing larval survival.
Comparison of the thermal envelope for larval de-
velopment of the tropical sand dollar Arachnoides pla-
centa and the temperate sea urchin Centrostephanus
rodgersii shows that they have optimal ranges
17–31 °C and 13–22 °C, respectively (Figure  17.1).
Thus, as expected, from the very broad pan-tropical/
subtropical distribution of adult A. placenta, the lar-
vae of this species are robust to a broader tempera-
ture range than those of C. rodgersii, which has a less
extensive distribution. However, while the larvae of
A. placenta have the potential to be transported to,
and survive in, cooler water, this species has not
expanded its range poleward. The colonization po-
tential of A. placenta is likely limited by the lack of
suitable tropical sandy beach habitat at higher lati-
tudes, and perhaps also by the lack of a larval settle-
ment cue. Sand dollar larvae are known to settle in
response to chemical (as well as physical) cues pro-
vided by adult populations (Highsmith, 1982; Burke,
1984; Pearce and Scheibling, 1990; Hodin et al., this
volume). In contrast, C. rodgersii, has extended its
range by thousands of kilometers through poleward
transport of larvae (Ling et  al., 2008; Banks et  al.,
2010). Success of this species was facilitated by ex-
tensive availability of its temperate rocky reef habi-
tat poleward and associated larval settlement cues
(e.g., coralline algae). For the crown-of-thorns sea
star Acanthaster planci, the developmental thermal
envelope (24–31°C) includes temperatures typical of
high latitude coral habitats (Lamare et al., 2014), in-
dicating that the poleward expansion of corals seen
in the northern and southern Pacific (Yamano et al.,
2011; Baird et al., 2012) may also be accompanied by
a spread of their major predator.
Although larval thermal envelope studies rarely
encompass more advanced larval and juvenile
stages, species distribution models (SDM) deter-
mined from adult distribution data can provide
an integrative assessment indicating where the
planktonic life phase is successfully completed. For
instance, data from larval thermal reaction norms
coupled with adult SDM indicated that the prog-
eny of the invasive population of the Arctic sea star
Asterias amurensis in Tasmania have the potential to
invade the Southern Ocean and Antarctica (Byrne
et al., 2016). This type of comparison is also useful
in showing where adult distribution does not corre-
late with larval thermal tolerance and where other
biological and ecological processes are more impor-
tant (Sunday et al., 2015).
For molluscs, most studies of the impacts of tem-
perature on development have investigated the
optimal temperatures for aquaculture production
and so have focused on the impacts of warming
(O’Connor and Lawler, 2004; Dove and O’Connor,
2007; O’Connor et al., 2015). The complete thermal
envelope for development has only been deter-
mined for four species: Mytilus edulis, Ostrea edulis,
Argopecten irradians, and Mulinia lateralis (Calabrese,
1969; Davis and Calabrese, 1969; Tettelbach and
Rhodes, 1981; Mestre et  al., 2009). For the mussel
M. edulis, development to the D-stage veliger larvae
is successful between 10–20 °C (± 5°C with respect
to ambient) (Mestre et al., 2009). This suggests that
local populations would be at risk of local extinc-
tion or range contraction due to OW. Similar sen-
sitivity to warming was found in the scallop A.
irradians and the clam M. lateralis, with successful
development to the D-stage veliger larvae occur-
ring between 20–25 °C (ambient temperature 25 °C)
and 12.5–25 °C (ambient temperature 20 °C), re-
spectively (Calabrese, 1969; Tettelbach and Rhode­s,
1981). In contrast, for the oyster O. edulis, where am-
bient temperature was 15 °C, the thermal envelope
for larval development was 17.5–30 °C, with an up-
per temperature 10 °C above ambient (Davis and
Calabrese, 1969). This result indicates that O. edulis
would be robust to OW. High tolerance to warm-
ing is a common trait of oyster species; for instance,
the larvae of the pearl oyster Pinctada imbricata and
the commercial oysters Saccostrea glomerata, Crassos-
trea gigas, and Ostrea angasi are robust to warming
256   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert ebrat e L ar vae
4–5 °C above ambient (O’Connor and Lawler, 2004;
Dove and O’Connor, 2007; Parker et al., 2010).
Of the oyster species investigated, P. imbricata had
a greater capacity to withstand cooler temperatures,
with a lower thermal limit for the development to
the D-stage veliger larvae being 10 °C below their
thermal optimum (O’Connor and Lawler 2004),
while for O. angasi and S. glomerata, the thermal
limits below optimum were 7.5 °C (larval growth
and development) and 5 °C (embryonic develop-
ment to D-stage veliger), respectively (Dove and
O’Connor, 2007; O’Connor et al., 2015). There was
an improvement in the thermotolerance of S. glom-
erata with advancing developmental stage, with
adequate survival of later larvae at the lowest tem-
perature tested (9 °C lower than optimal) (Dove and
O’Connor, 2007).
Climate change–related studies of molluscan
development have focused on warming (Parker
et al., 2010; Talmage and Gobler, 2011; Byrne et al.,
2011b; Ko et  al., 2014). Byrne et  al. (2011b), for
example, found that a +4 °C warming from the
optimum temperature of 20 °C breached the ther-
motolerance for larval development in the abalone
Haliotis coccoradiata. Further, Talmage and Gobler
(2011) found that a +4 °C warming led to a 50%
reduction in the survival of larvae of the clam Mer-
cenaria mercenaria and the scallop A. irradians. In
contrast, Ko et al. (2014) found that overall growth
rate of larvae of the oyster C. gigas was increased
following a +4 °C warming. There was, however,
a significant reduction in the total lipid index of C.
gigas larvae when compared with larvae from the
ambient controls. These studies suggest that these
molluscs may be living close to their upper ther-
mal limit for development. However, cool tem-
perature tolerance and the potential for poleward
range shifts was not determined.
As the ocean warms, the developmental dynamics
of many species will change. The thermal envelope
of many of the species investigated thus far exceeds
their current temperature ranges (both warm and
cold), but we only have detailed developmental ther-
mal windows for a few species, mostly echinoderms.
The adult stage of many marine species currently fill
their latitudinal thermal niche (Sunday et al., 2012).
Evidence thus far indicates that a +4 °C warming
approximates the upper tolerance threshold for
development in many mollusc and echinoderm spe-
cies (Byrne, 2010; 2011). This level of warming (pejus
temperature; Pörtner, 2012) may occur for many
species under the worst-case scenario (RCP8.5) and
during heat waves. Successful development at tem-
peratures below ambient suggests that cooler water
is not a barrier to poleward migration through lar-
val transport corridors, with the caveat that larval
settlement and postlarval performance also have to
be successful. Overall, while species ranges are in-
fluenced by larval thermal tolerance, the capacity
for OW-driven poleward colonization varies among
species. Understanding the relationships between
the thermal tolerance of the benthic adult stage as
indicated by SDMs together with larval thermal
windows are important in predicting range change,
both contraction and extension.
17.3  Ocean Acidification—Multiple
Drivers
Ocean uptake of anthropogenic CO2 is reducing
seawater pH and Ω while increasing organism pCO2
(hypercapnia) (Gattuso et al., 2015). Thus, OA incor-
porates three covarying stressors, each with their
own mode of action on larval physiology (Byrne
et  al., 2013a; Waldbusser et  al., 2014, 2015; Jaeckle,
this volume). Decreased pH may erode skeletons
and impair cellular functions such as pH-depend-
ent enzyme activity; reduced Ω compromises the
ability of larvae to build skeletons and shells; while
hypercapnia alters metabolism. These responses
have raised questions as to which variables of the
inorganic carbonate system altered by OA are re-
sponsible for the negative effects on larvae. Is this
a pH, pCO2, or Ω problem—or due to a combina-
tion of these factors? Disentangling the mechanisms
of action of these highly correlated parameters on
larval performance is a challenge. Recent research
shows that their impact differs among taxa, with
pCO2 being the key stressor for sea urchin calcifica-
tion, while mineral saturation is more important for
bivalves (cf. Byrne et  al., 2013a; Waldbusser et  al.,
2014; 2015).
Ocean acidification will have wide-ranging im-
pacts on larval survival, calcification, and physi-
ological processes (Pörtner, 2010). The skeletons and
shells of marine larvae are the most fragile calcium
 Cha n g i n g Cl i mat e — O c ea n W arm i n g a n d A c i d i f i c at i o n    257

carbonate structures produced by marine organ-
isms, and so OA research has focused on calcifying
larvae (Kroeker et al., 2013; Parker et al., 2013; Prze-
slawski et al., 2015). Projected near-future OA has a
­general miniaturizing effect resulting in smaller lar-
vae with less skeleton and smaller shells ­(Figure 17.2;
Figure  17.3), often combined with abnormal de-
velopment indicating teratogenic effects (Byrne,
2012; Byrne et al., 2013a; Parker et al., 2013). Ocean
acidification is a greater stressor for calcifying than
noncalcifying embryos and larvae of sea urchins.
Echinoplutei and bivalve veligers have been used
as model systems to investigate the effects of OA on
calcification (see Byrne et al., 2013; Parker et al., 2013;
Ross et al., 2016), and so the impact of OA is com-
paratively well understood for echinoderms and
molluscs (Figure 17.3). The stunting effect of near-
future OA (~pH 7.7–7.9 vs. control ~pH 8.0–8.2) on

Arachnoides placenta

pH 8.2 7.8 7.6

Echinometra mathaei
Tropical

pH 8.1 7.8 7.7 7.5

Tripneustes gratilla

pH 8.15 7.8 7.6

Heliocidaris tuberculata
Temperate

pH 8.2 7.8 7.6 7.4

Centrostephanus rodgersii

Figure 17.2  Echinoplutei from tropical, tem-

pH 8.2 7.8 7.6 perate, and polar sea urchins reared in control
Strechinus neumayeri and OA treatments. Larvae reared in low pH
treatments are smaller. (Echinometra mathaei,
Antarctic

Centrostephanus rodgersii, and Sterechinus

neumayeri adapted, with permission, from
images in Byrne et  al., 2013; Tripneustes
pH8.1 7.8 7.6 gratilla adapted, with permission, from images
in Sheppard Brennand et al., 2010.)
258   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert ebrat e L ar vae

(A)
1.2 Arachnoides placenta

0.8

0.6
pH 8.2
0.4 pH 7.8
pH 7.6
0.2

0
0 1 2 3 4
(B) 1.2 Odontaster validus

1
Proportion of surviving larvae

0.8

0.6

pH 8.2
0.4
pH 7.8
pH 7.6
0.2

0
0 1 3 6 9 14 21 23 30 36 48 58
(C)
1.2 Saccostrea glomerata

0.8

0.6

0.4
pH 8.2

0.2 pH 7.9 Figure  17.3 Proportion of the larvae of (A) Arachnoides

placenta, (B) Odontaster validus, and (C) Saccostrea glomerata
0 remaining when reared at ambient and OA treatments (adapted,
1 3 5 7 9 11 13 15 17 19 with permission, from data in Gonzalez-Bernat et  al., 2013a;
DAYS 2013b; Parker et al., 2012).

sea urchin echinoplutei is evident in the smaller larval growth and calcification in near-future OA
larvae with shorter skeletal rods compared with is also widely documented in the veliger larvae of
larvae reared in ambient conditions (Figure  17.2). bivalves and gastropods (Byrne et al., 2011b; Crim
This trend has been documented for ~20 species of et al., 2011; Gazeau et al., 2013; Parker et al., 2013;
echinoplutei from across world (Kurihara 2008; By- 2017). In a review by Parker et al. (2013), 26 out of
rne et al., 2013a; Wangensteen et al., 2013; Karelitz 30 species of mollusc larvae showed a reduction in
et al., 2016), and is also reported for the calcifying shell length, area, or thickness following exposure
ophiopluteus larvae (Dupont et al., 2008). Reduced to elevated pCO2.
 Cha n g i n g Cl i mat e — O c ea n W arm i n g a n d A c i d i f i c at i o n    259
The presence of smaller echinoplutei of Stron-
gylocentrotus droebachiensis and Paracentrotus livi-
dus in OA treatments is suggested to be due to
delayed development resulting in larvae that are
at a “younger” ontogenetic stage (Martin et  al.,
2011; Stumpp et al., 2011; Zhan et al., 2016). There
is a similar report for an asteroid (Gonzalez-Bernat
et al., 2013a). For molluscs, delayed development to
the first veliger stage due to OA is widely reported
(Gazeau et al., 2013; Parker et al., 2013). In Sterechi-
nus neumayeri and Strongylocentrotus purpuratus, the
timing of early events, cleavage, and blastulation,
however, do not differ in embryos reared in OA
and ambient conditions, although there was a slight
delay in hatching for the former species (Place and
Smith, 2012; Yu et  al., 2013). For Strongylocentrotus
intermedius, a marked delay in cleavage and hatch-
ing occurs in embryos reared from fertilization in
near-future OA conditions (Zhan et al., 2016). In the
mussel Mytilus galloprovincialis, there was no effect
of OA on the timing of embryogenesis until the tro-
chophore stage, when shell formation first began
(Kurihara et  al., 2008). In one of the few studies
where echinoplutei were reared in OA conditions
through to metamorphosis, the larvae of Arbacia lix-
ula were smaller but development was not delayed.
The larvae reared at low pH had the same plank-
tonic duration and metamorphosed at the same
time as controls, albeit producing smaller juveniles
(Wangensteen et al., 2013). A similar result was evi-
dent for nonfeeding sea urchin larvae (Byrne et al.,
2011b). In contrast, in the bivalves Mercenaria mer-
cenaria, A. irradians, and Crassostrea virginica, lar-
vae exposed to reduced pH exhibited significantly
delayed metamorphosis compared with larvae ex-
posed to ambient pH (Talmage and Gobler, 2011).
In A. irradians, 76% of larvae had metamorphosed
into juveniles after 16 days at reduced pH, whereas
100% of larvae had metamorphosed in the ambient
conditions at this time.
To understand the impacts of pCO2-driven de-
velopmental heterochrony, details of the appear-
ance of developmental milestones (e.g., cell cycle,
blastulation, coelomogenesis, skeletogenesis) need
to be documented. The apparent delay may also
be due to mortality of faster developing genotypes
(see Pechenik, this volume). To understand the
developmental implications of OA with respect to
the questions of delayed development in younger
stages or just a general stunting of growth may re-
quire transcriptome assessment of gene expression
and a better understanding of cellular processes.
Echinoplutei of ~7 species reared in OA condi-
tions exhibited altered body morphology, including
increased abnormality, increased left-right asym-
metry, and a change in the allometric relationships
between arm and body size (Sheppard Brennand
et al., 2010; Chan et al., 2011; Uthicke et al., 2013a;
Karelitz et al., 2016; Zhan et al., 2016), while some
species did not exhibit altered larval allometry
(Martin et  al., 2011; Stumpp et  al., 2011; Karelitz
et al., 2016). Chan et al. (2011) suggest that changes
in the shape of Dendraster excentricus larvae reared
in OA conditions may be a plastic response to com-
pensate for reduced skeletal mass and loss of sta-
bility. Mollusc larvae also experience alterations in
their body morphology following exposure to el-
evated CO2. Abnormal shell development, includ-
ing convex hinge, protrusions of the mantle, and
shell alteration and malformation has been found in
most species studied to date (Kurihara et al., 2008;
Gazeau et al., 2013; Parker et al., 2013). These sub-
lethal abnormalities which appear early in larval
development will likely have lethal consequences
for larvae later in their development (Pechenik, this
volume).
Some echinoid and ophiuroid plutei exhibit high
survival in moderately low pH (~pH  7.8) condi-
tions (Dupont et  al., 2008; Byrne 2011; Stumpp
et  al., 2011; 2012a; Gonzalez-Bernat et  al., 2013a).
However, with greater acidification (~pH  7.6),
­
there is a 10–20% increase in mortality for echino-
derm and mollusc larvae, indicating that this level
of OA may approximate a tipping point for de-
creased survival (Dupont et al., 2008; Gazeau et al.,
2013; Parker et  al., 2012; 2013; Byrne et  al., 2011b;
2013a; Gonzalez-Bernat et al., 2013a). In long-term
rearing experiments, larval mortality in OA con-
ditions increases with time (Figure  17.3) (Dupont
et  al., 2008; Stumpp et  al., 2013). Larvae of Stron-
gylocentrotus purpuratus reared in lower pH had
significantly slower feeding rates (Stumpp et  al.,
2011; 2012b), indicating a reduction in feeding ca-
pacity. A lower feeding efficiency is also indicated
by the smaller stomachs in Dendraster excentricus
larvae reared in OA conditions (Chan et al., 2011).
260   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert ebrat e L ar vae
For Strongylocentrotus droebachiensis, OA interferes
with gastric pH homeostasis, causing a drop in pH
thereby decreasing digestive efficiency (Stumpp
et al., 2013). Decreased growth of echinoplutei un-
der OA conditions is likely associated with changes
to larval energy budgets and the cost of maintain-
ing gastric pH homeostasis (Stumpp et al., 2013).
For molluscs, larvae of the mussel Mytilus califor-
nianus showed a delay in the onset of feeding when
reared at elevated pCO2. At 44 h post-fertilization,
fewer veligers had initiated feeding in the elevated
pCO2 treatment compared with those in the ambient
control (Waldbusser et  al., 2015). In contrast, there
was no effect of reduced pH on the feeding rate of
larvae of the mussel Mytilus edulis or the limpet Pa-
tella vulgata (Marchant et al., 2010; Bechmann et al.,
2011). For mollusc larvae, reductions in the lipid
content and tissue mass, and changes in the im-
mune response and acid-base balance, along with
increased larval metabolism have also been reported
as responses to OA, indicating impacts on larval en-
ergy budgets (Gazeau et al., 2013; Parker et al., 2013).
Other responses of echinoplutei to OA include
altered gene and protein expression (Todgham and
Hofmann, 2009; O’Donnell et al., 2010; Martin et al.,
2011; Stumpp et  al., 2011; 2012a; Dineshram et  al.,
2013; Gazeau et al., 2013; Padilla-Gamiño et al., 2013;
Parker et al., 2013). A meta-analysis of gene expres-
sion data from four echinoid species indicated that
this change involved down-regulation of genes as-
sociated with energy production pathways, an indi-
cation of metabolic depression, as well as changes in
expression of genes involved in biomineralization
and ion transport (Evans and Watson-Wynn, 2014). In
general, there was a down-regulation of genes within
energy pathways (Evans and Watson-Wynn, 2014).
A meta-analysis of data on the impacts of OA on
growth of 15 species of echinoplutei from tropical to
polar regions used multiple linear regression mod-
els to analyse the relative influence of decreased
Ω, increased pH, and increased pCO2 to assess the
effects of each of these stressors on skeletogene-
sis (Byrne et  al., 2013a). Only data for pre-feeding
echinoplutei were used because after food is intro-
duced, comparisons between studies are likely to
be confounded by arm growth plasticity. Well-fed
echinoplutei often develop small arms, a strategy
to prioritize growth of the juvenile rudiment (Soars
et al., 2009; McAlister and Miner, this volume). This
plasticity trait differs among species (Soars et  al.,
2009). The analysis indicated that pH and pCO2
are stressors of similar strength, and that the best-
fitting models had a high weight for pCO2 (Byrn­e
et al., 2013). Thus, for the global dataset, pCO2 was
the most important stressor in reducing sea urchin
larval growth. This is consistent with what is known
for the calcification processes in sea urchin larvae
(echinoderms transport bicarbonate, not carbonate
at the site of calcification) and use respiratory CO2
as a source of carbon (Sikes et  al., 1981). Stunted
echinoplutei in OA conditions appear largely to be
a pCO2-hypercapnia problem—indicatin­g energetic
constraints and altered metabolism. This is likely
associated with metabolic depression (Pörtne­ r,
2010; Evans and Watson-Wynn, 2014) and impaired
feeding (Stumpp et  al., 2013). The combined data-
set revealed that a significant reduction (~20%) in
larval growth occurs at ~0.3–0.4 pH units below
ambient (near-future OA), with a sharp decrease in
growth in far-future OA (Byrne et  al., 2013a). That
near-future OA also has a stunting effect on aster-
oid bipinnaria larvae (Patiriella regularis, Acanthaster
planci) that do not need to calcify also points to the
importance of hypercapnia and pH as major stress-
ors (Byrne et al., 2013d; Kamy­a et al., 2014; Karelitz
et al., 2016). H
­ owever, the opposite occurred for the
bipinnaria of the Antarcti­c species Odontaster validus
­(Gonzalez-Bernat et al., 2013b; Karelitz et al., 2016).
The negative effects of OA on echinoplutei are
suggested to be due to the following: (1) delayed de-
velopment (i.e., the larvae are normal and will catch
up with time); (2) reduced Ω causing physiological
stress on calcification systems; (3) a reduced scope
for growth resulting from higher larval metabolism
and the diversion of energy to acid/base regulation;
and (4) teratogenic effects on developmental mecha-
nisms at the genetic and cellular level with respect to
differentiation and body patterning (e.g., asymmetry
and allometry), causing developmental malforma-
tions (O’Donnell et  al., 2010; Sheppard Brennand
et  al., 2010; Martin et  al., 2011; Stumpp et  al., 2011;
2012a; 2012b; 2013; Byrne, 2012; Uthicke et al., 2013b;
Yu et al., 2013). While the meta-analysis showed the
overarching importance of pCO2-hypercapnia, it
seems that many of these mechanisms may be im-
portant, underlying the broad-ranging effects of OA
 Cha n g i n g Cl i mat e — O c ea n W arm i n g a n d A c i d i f i c at i o n    261
from gene expression to growth, abnormality, and
mortality.
For mollusc development, studies investigate the
effects of pH, pCO2, Ω, and carbonate ion concentra-
tion by chemical manipulations of seawater where
each stressor was independently altered (Gazeau
et  al., 2011; Waldbusser et  al., 2014; 2015). Devel-
opmental success to the D-stage veliger and shell
growth was affected by Ω (Crassostrea gigas, Mytilus
galloprovincialis, M. californianus; Waldbusser et al.,
2014; 2015) and calcium carbonate concentratio­n
(C. gigas; Gazeau et  al., 2011) but not by pH or
pCO2. For M. californianus, respiration rates were
elevated under low pH (~7.4) and initiation of lar-
val feeding was delayed when exposed to elevated
pCO2 (Waldbusser et al., 2015). These results show
that components of the seawater carbonate system
impact different physiological processes in larvae,
and that OA is a multistressor in itself. Waldbusser
et al. (2014; 2015) concluded that nonlinear changes
to the seawater carbonate system indicate that Ω
will be crossed decades or even centuries earlier
than pH thresholds are reached.
It appears that the major stressor associated with
OA causing impaired growth of echinoderm and
mollusc larvae differ, with pCO2 being more impor-
tant for echinoplutei while Ω is more important for
veliger larvae. This may also be due to marked differ-
ences in skeletogenesis in these two major phyla. In
echinoid larvae, the skeleton has an epithelial cover,
with calcification generated by ion pumping into an
intracellular space, stressing the importance of ener-
getic costs (Byrne et al., 2011b; Stumpp et al., 2012b;
Dubois, 2014). In contrast, mollusc larval shells are di-
rectly exposed to water chemistry, and so reduced Ω,
and perhaps decreased pH, have a more direct impact
on the shell (Byrne et al., 2011b; Parker et al., 2013).
Reduced larval size and a potentially longer
pelagic larval duration in a high pCO2 ocean will
have a negative impact on echinoderm and mollusc
larvae, such as making them more susceptible to
predation and decreasing chances of survival and
recruitment (Allen, 2008; Soars et  al., 2009; Chan
et  al., 2011; Parker et  al., 2013), although this may
be tempered by OW (Przeslawski and Byrne, 2013).
For echinoderm plutei, the skeletal rods are essen-
tial for feeding, swimming, and protection from
predation (Hart and Strathmann, 1994; Soars et al.,
2009; Chan et  al., 2011). Misshapen echinoplutei
with an altered body profile and smaller and ab-
normal mollusc larvae, as seen in OA treatments,
are poor swimmers and eaters (Chan et  al., 2011).
For mollusc larvae, normal shell formation is im-
portant for protection from predators. In addition,
smaller larvae may have lower feeding efficiency,
making them susceptible to starvation (Hart and
Strathmann, 1994). Overall, the negative impact of
OA on the size, shape, and survivorship of echino-
plutei, veligers, and other larvae will lower their fit-
ness, compromising success of the pelagic life stage.
In addition to direct effect on larvae, OA alters the
substrate cues (from coralline algae) required for
settlement and metamorphosis (Doropoulos and
Diaz-Pulido, 2013; Uthicke et al., 2013b).
Parental conditioning to environmental pCO2/
pH during gamete development may influence
the resilience of progeny to OA. Acclimation of ur-
chins and oysters to moderately elevated pCO2 can
resul­t in intergenerational enhancement of larval
resilience to OA, indicating phenotypic plasticity
in gamete development (Parker et al., 2012; Dupon­t
et al., 2013). The progeny of selectively bred oysters
acclimated in OA conditions were more resilient
than those of wild-type oysters, a potential genetic
(adaptive) or epigenetic effect (Parker et al., 2012).
Although this is maladaptive with regards to other
stressors (e.g. temperature, Parker et. al., 2017). Pa-
rental acclimation of Strongylocentrotus purpuratus
in OA conditions conveyed resilience to larval de-
velopment, a response dominated by maternal ef-
fects with potential for a genetic influence (Sunda­y
et  al., 2011). However, acclimation of Echinometra
sp. adults did not result in larvae that were more
resilient to OA (Uthicke et al., 2013a).
The larvae of Strongylocentrotus purpuratus found
in the California Current upwelling system are com-
paratively tolerant to OA, with only a modest de-
crease in arm growth at pH  7.7, potentially due to
the wide seasonal flux in pH that they experience
in nature (Yu et al., 2011; Kelly et al., 2013). Popula-
tions of S. purpuratus from this region have experi-
enced upwelling-low pH conditions for thousands
of years, and so their reproductive biology may be
acclimatized/adapted to low pH water (Pespeni
et al., 2013a; 2013b). As the adults also occur in inter-
tidal rock pools that experience regular hypercapnic
conditions, the comparative resilience of the larvae
to OA may also have been influenced by the tide
262   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert ebrat e L ar vae
pool conditions in the adult habitat. For Paracentro-
tus lividus, Moulin et al. (2011) found that the larvae
were more robust to reduced seawater pH if their
parents inhabited tide pools with variable pH. Simi-
larly, larvae of some bivalves are resilient to the nega-
tive effects of ocean acidification, possibly owing to
adaptation to naturally acidic environments (e.g.,
low pH sediments) or naturally high local alkalinity
(Talmag­e and Gobler, 2011), emphasizing the impor-
tance of environmental history on stress tolerance.
For barnacle larvae, variation due to maternal pro-
visioning and parental environmental history influ-
ences the tolerance to OA (Pansch et al., 2014). More
data are needed to explore potential trends in the dif-
ferences in response of the larvae generated from in-
tertidal and subtidal parents and those from regions
differing in pH, and the potential influence of genetic
and epigenetic effects (Foo and Byrne, 2016).
17.4  Multistressor Effects—Interactive
Effects of Ocean Warming
and Acidification on Larvae
As OW and OA are increasing simultaneously, with
potential for interactive effects, it is critical to inves-
tigate the effects of simultaneous exposure of these
stressors on the developmental processes of ma-
rine invertebrates (Byrne and Przeslawski, 2013).
Increased temperature and acidification can have
additive (total effect = sum of individual effects),
synergistic (total effect > sum of individual effects),
or antagonistic (total effect < sum of individual ef-
fects) effects (Przeslawski et  al., 2015). Larval fit-
ness is modified in their physiological responses to
multiple stressors, and so we need to understand
how multiple stressors affect larval performance
(Pörtne­r, 2012). A recent meta-analysis of the in-
teractions of OW and OA on the early life history
stages of marine invertebrates across four phyla
from multistressor studies revealed that synergistic
interactions (76% of individual tests) are more com-
mon than additive (9%) or antagonistic (16%) inter-
actions, reinforcing the importance of multistressor
studies (Przeslawski et al., 2015). The nature of the
responses to combined OW and OA is influenced
by developmental stage, phylum, and stressor lev-
els used (Przeslawski et al., 2015).
The responses of larvae to increased temperature
and acidification as individual stressors (as above)
prompt predictions as to the combined effects of
OW and OA. Prediction 1: the faster pace of devel-
opment and faster growth with modest warming
in response to stimulation of metabolism and the
suppressive effects of OA predict an antagonistic
interaction between these stressors with increased
temperature (within limits), ameliorating some of
the negative effects of acidification. Prediction 2:
additive negative effects or negative synergistic in-
teractions would be predicted as thermal and pH/
pCO2 tolerance thresholds are exceeded, leading
to developmental failure. The stressor combina-
tion thus makes matters worse. As predicted by the
bioenergetics–thermal window hypothesis (Pörtner
and Farrell, 2008; Pörtner, 2012), thermal tolerance
should reduce under hypercapnia. Conforming to
Prediction 1, OA and OW (~3 × 3 stressor levels) an-
tagonistically interacted to affect growth of echino-
plutei (Figure 17.4), with the stunting effects of OA
ameliorated by near-future warming (~+ 3–4  °C)
(Sheppard Brennand et al., 2010; Byrne et al., 2013b;
2013c). This interactive effect is not evident for mol-
luscs (Parker et al., 2010; Byrne et al., 2011b).
A recent study Karelitz et  al. (2016) addressed
the bioenergetics–thermal window hypothesis with
five echinoderm species (asteroids and echinoids)
from tropical to polar regions to determine if OW
and OA interact to alter developmental thermal
envelopes. Development in all five species was af-
fected by temperature and pH, in line with expecta-
tions (i.e., OW—faster development; OA—stunted
growth of echinoplutei), but developmental ther-
mal envelopes, the temperature range at which
normal development can occur, were not altered by
OA. When the entire developmental thermal enve-
lope is considered, warming and acidification ex-
erted additive negative effects (Prediction 2), with
OA-driven abnormality and growth reductions
consistent across the thermal gradient used. This
suggests a partitioning of mechanisms involved in
coping with OW and OA (Karelitz et al., 2016).
The results of Karelitz et al. (2016) are consistent
with the paradigm that temperature is the primary
environmental factor influencing the physiology
of marine invertebrates (Vernberg, 1962; Hoegh-
Guldber­g and Pearse, 1995; Byrne, 2011; Przeslawski
 Cha n g i n g Cl i mat e — O c ea n W arm i n g a n d A c i d i f i c at i o n    263

(A) Tripneustes gratilla

200
180
Mean Arm Length (μm)

160
140 8.2
120
7.8
100
7.6
80
60
40
20
0
24°C 27°C
(B) Heliocidaris tuberculata
500
450
Mean Arm Length (μm)

400
350
300
8.2
250 7.8
200 7.6
150 7.4
100
50
0
20°C 24°C

(C) 350 Sterechins neumayeri

300
Mean Arm Length (μm)

250 Figure  17.4 Mean arm length of the echinoplutei of three

echinoid species reared in multiple OW-OA treatments—fully
200 orthogonal independent replicate design. (A) For Tripneustes
8.0
150 7.8 gratilla, there was an antagonistic interaction between
7.6 warming and acidification. (B,C) For Heliocidaris tuberculata
100
and Sterechinus neumayeri, there was no significant interactive
50 effect, but the larvae reared in warmer conditions were larger
across all pH treatments (adapted, with permission, from data
0
–1°C 1°C in Sheppard Brennand et al., 2010; Byrne et al., 2013b; 2013c).

et al., 2015). For the early life stages of marine inver-
tebrates, it appears that thermal stress is more likely
to cause mortality, while pH is more likely to result
in sublethal effects (hypercapnia, impaired metabo-
lism, reduced calcification, altered gene expression)
(Przeslawski, 2004; Todgham and Hofmann, 2009;
Beniash et al., 2010; Stumpp et al., 2012b; Evans and
Watson-Wynn, 2014). Thus, temperature is likely to
be more important than OA in influencing marine
invertebrate development and thus species lati-
tudinal distributions in a changing ocean. Further
research on the relationships between fitness and
growth/performance of larvae in a multistressor
context is needed, and the bioenergetics framework
(e.g., dynamic energy budget) is likely to be an in-
formative approach. In addition, other stressors
(e.g., salinity, UV, hypoxia) are also very important
to consider, but the data available on their impact
on marine larvae in a multistressor context are lim-
ited (Przeslawski et al., 2015).
17.5  Adaptation and Acclimation
of Marine Larval Stages in a Changing
Climate
As for considerations of OW and OA individually,
acclimatization of parents to both stressors in the
environment and adaptation to local conditions is
264   E v o l u t i o n ary E c o l o g y o f M ar i n e I n v ert ebrat e L ar vae
important to consider (Sanford and Kelly, 2011).
Parental conditioning to combined OW and OA in-
fluences the stress tolerance of sea urchin progeny
(Suckling et al., 2014). In this study, longer-term (6 vs.
17 months) acclimation of adult Sterechinus neumayeri
to near-future warming and acidification improved
the performance of larvae, and this may have been
influenced by production of bigger eggs by urchins
in climate change treatments (Suckling et al., 2014).
Improved performance has also been shown in
coral larvae following acclimatization of parents to
OW and OA (Putnam and Gates, 2015). Exposure
of adult corals Pocillopora damicornis to OW and OA
led to a higher size-dependent metabolic rate (meta-
bolic acclimation) at elevated temperature and CO2
compared with larvae from non-exposed parents
(Putnam and Gates, 2015). This type of phenotypic
plasticity potentially associated with maternal and/
or paternal effects may help mitigate the effects of
changing climate on fitness over generations, poten-
tially providing a temporal window for evolutionary
(genetic) adaptation to occur. We need more infor-
mation across the life cycle of marine invertebrates
to understand the potential for transgenerational
adaptive capacity in response to multiple stressors
(Sunda­y et al., 2011; 2014; Kelly et al., 2013; Munday
et al., 2014; Foo and Byrne, 2016; Ross et al., 2016).
Several studies note the presence of a subset of
progeny (the survivors) that are more resilient to
near-future OW and/or OA indicating the exist-
ing phenotypic plasticity or genetic variation that
might provide species’ resilience in a changing
ocean. This will, however, lead to a reduction in
genetic variation in populations if these stress-­
tolerant genotypes are selected for (see Pechenik,
this volume). Quantitative genetics studies indicate
the presence of genetic traits to facilitate adaptation
to climate change for echinoid but not for bivalve
development (Sunday et  al., 2011; Foo et  al., 2012;
2014; 2016; Kelly et al., 2013; Foo and Byrne, 2016).
As the ocean will change over coming decades
more gradually than in climate change experiments
reviewed here, acclimatization of gametogenesis,
particularly during egg development (maternal
imprinting), in a warmer and lower-pH ocean may
facilitate production of more resistant offspring and
buy time to facilitate genetic adaptation. We also
need better data on the temperature, pH, CO2, and
carbonate chemistry of adult and larval habitats
to achieve a better understanding of the threat of
global change to marine invertebrates.
17.6 Conclusion
Due to the “developmental domino effect,” expo-
sure to stress early in development can result in del-
eterious downstream effects, because performance
of later ontogeny depends on the success of early
stages (Pechenik, 1987; this volume; Byrne, 2011).
Developmental failure, regardless of stage, will
have a negative effect for the persistence of marine
populations and the ecosystems in which they re-
side (Harley et  al., 2006; Brierley and Kingsford,
2009; Hofmann et al., 2010). The impaired ability of
calcifying larvae to produce a larval skeleton due
to OA in a timely manner may be a weak link for
persistence of a broad suite of invertebrates.
We need whole development studies from ferti-
lization to settlement and postlarval development
in multistressor experiments to assess potential out-
comes for species. For instance, while barnacle larvae
are resilient to OA, negative effects of a weak skel-
eton are evident for the juvenile stage (McDonal­d
et al., 2009). Similarly, in a study of the impacts of
combined OW and OA in an echinoid from fertiliza-
tion to the juvenile stage, the subset of larvae that
survived the bottleneck of early mortality due to
warming were able to develop to the juvenile stage,
but their skeleton was compromised, depending
on stressor levels (Byrne et  al., 2011a). In the oys-
ter, Ostrea lurida, exposure to elevated pCO2 during
larval development had strong carryover effects for
the growth of juveniles (Hettinger et al., 2012). One
week after metamorphosis, juveniles of O. lurida
that had been reared at elevated pCO2 during larval
development had a 41% reduction in shell growth
rate. This negative effect of pCO2 exposure during
the larval phase on juvenile shell growth persisted
for 1.5 months after the juveniles had been trans-
ferred to a common garden of ambient seawater.
Differences in the survival of benthic marine in-
vertebrates through past climate change are likely
to have been influenced by life history traits, with
species that had nonfeeding larvae being favored
(Valentine and Jablonski, 1986; Uthicke et al., 2009).
These species have a buffered life history free of
 Cha n g i n g Cl i mat e — O c ea n W arm i n g a n d A c i d i f i c at i o n    265
the vagaries of a planktonic food supply. If they do
not need to calcify, they will also be less vulner-
able to OA. Thus, species that produce large eggs
and have noncalcifying/nonfeeding larvae (e.g.,
lecithotrophic echinoderm larvae, coral planulae,
barnacle cyprids) may have a competitive advan-
tage in a future ocean (Dupont et al., 2010; Anlauf
et al., 2011; Byrne et al., 2011a; 2011b; Nguyen et al.,
2012; Chua et  al., 2013; Hardy and Byrne, 2014),
although they may have a mortality bottleneck at
the early juvenile stage (Anlauf et al., 2011; Byrne
et  al., 2011a; McDonald et  al., 2009; Wolfe et  al.,
2013). For echinoderms, the increase in diversity of
lecithotrophic developers over evolutionary time
indicates that this trait is associated with higher
speciation rates, as seen in several extant clades
(Jeffery et  al., 2003; Uthicke et  al., 2009; Byrne,
2013). If possession of nonfeeding/noncalcifying
larvae conveys a competitive advantage for adults
in a future ocean, flow-on alterations to benthic as-
semblages would be expected.
The larval and early juvenile stages of marine
invertebrates experience naturally high mortality
rates, and relatively small perturbations in recruit-
ment can translate to large alterations of adult pop-
ulations (Eckman, 1996; Lamare and Barker, 1999;
Uthicke et  al., 2009). The responses of the pelagic
life phase, adverse or otherwise, to OW and/or
OA will alter marine communities and populations
(Harley et  al., 2006; Brierley and Kingsford, 2009).
There may be some redundancy, with more resilient
species continuing to fulfill important functional
roles. Understanding the level of species specific-
ity, adaptive capacity, and the degree of response
to changing ocean conditions is important in order
to predict the future composition and dynamics of
marine communities under a “winners and losers”
scenario (see Somero, 2010; Fabricius et al., 2011).
17.7 Summary
1. Marine invertebrate larvae live in a multistressor
world, and it is important to understand the inter-
active effects of climate change–related stressors
in a regionally and habitat-relevant context. Prior-
ity stressors differ regionally, with some regions,
for instance, exhibiting strong warming, while in
other regions acidification is more important.
2. Species are on the move as their larvae are fol-
lowing more favorable poleward isotherms, an
emerging global trend seen in the tropicalization
of temperate faunas. The sensitivity of larvae to
increased temperature varies among species, and
this influences their invasive potential. Broadly
distributed species that have a wide develop-
mental thermotolerance appear best able to avail
themselves of the new opportunities provided
by ocean warming.
3. Ocean acidification is a multistressor in itself,
and the impacts of its covarying stressors differ
among taxa. Increased pCO2 is the key stressor
impairing calcification in sea urchin larvae, while
decreased mineral saturation is more important
for calcification in bivalve larvae.
4. It appears that nonfeeding, noncalcifying larvae
are more resilient to climate change stressors.
5. As the ocean continues to change, some species
may be able to persist through acclimatization or
adaptation and produce more resilient offspring.
Understanding the capacity for adaptation/ac-
climatization, and the response to climate change
stressors in trans- and multi-generational studies
is important for predicting the future species com-
position and dynamics of marine communities.
Acknowledgments
Thanks to Andreas Heyland, Tyler Carrier, and
Adam Reitzel for inviting this contribution. Re-
search was funded by grants from the Australian
Research Council. Thanks to Dr. Miles Lamare for
providing the images for Figure 17.3. We acknowl-
edge the assistance of Selma Klanten and Paula
Cisterna­s, University of Sydney. This is supported
by an ARC Discovery Grant (M.B., S.D.). Thanks to
the comments from reviewers that improved the
manuscript. Sydney Institute of Marine Science
Contribution #212.
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 C h a pt er 18
Ecotoxicology in Marine
Environments: The Protective Role
of ABC Transporters in Sea Urchin
Embryos and Larvae
Ilaria Corsi and Luis Fernando Marques-Santos
This chapter is dedicated to the memory of Prof. Valeria
Matranga. Part of the work presented on the effects of na-
noparticles was conducted in collaboration with Valeria
Matranga, a mentor, a dear friend, and a distinguished
scientist.
18.1 Introduction
Contaminants with the potential to disrupt bio-
logical functions enter aquatic ecosystems from a
diversity of sources (industry, agriculture, sewage,
landfill sites, etc.). Such anthropogenic impacts pose
a potential risk to the long-term sustainability of
these ecosystems by disrupting food webs through
extirpation of species, reductions in the health and
reproduction of resident species, and changes in the
overall quality of the environment. While a broad
diversity of organisms is being used in ecotoxico-
logical bioassays to assess the risks of particular
pollutants, comparatively little work has been done
in marine ecosystems, especially for larvae of ma-
rine invertebrates. Understanding the sensitivity of
these developmental stages to anthropogenic con-
taminants is needed to predict how the biodiversity
of organisms that live in the plankton respond to
current and predicted future levels of contamina-
tion. This need is increasingly urgent because the
diversity of pollutants continues to increase in both
number and type. For example, point-source re-
lease of contaminants, including polychlorinated
Corsi, I. and Marques-Santos, L. F., Ecotoxicology in marine environments: the protective role of ABC transporters in sea urchin embryos
and larvae. In. Evolutionary Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland:
Oxford University Press (2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0018
biphenyls (PCBs) and toxic metals, from freshwa-
ter sources due to industrial processing remain a
consistent local stress for species in estuaries and
nearshore habitats. Release of polycyclic aromatic
hydrocarbons (PAHs) from oil spills can now re-
sult in the release of toxic compounds over broad
geographic ranges. Finally, new types of pollutants,
such as micro- and nanoplastics, are growing at an
increasing rate in marine ecosystems, with signifi-
cant impacts on pelagic species.
There are estimated to be upwards of 2.2 million
species of animals in the world’s oceans, most of
which are likely invertebrates (Mora et  al., 2011),
and this, at times, makes it difficult to select the
most appropriate model species to assess risks as-
sociated with the exposure to anthropogenic con-
taminants reaching the marine environment. In
particular, several adaptations across different
taxa, such as mechanisms of protection, robust-
ness, resistance, and plasticity, make such choices
very complex. Still, embryo-larval bioassays using
marine invertebrate species have been recently in-
troduced as a promising tool in ecotoxicology, as
they are reliable in assessing the biological effects of
exposure to contaminants. Ecotoxicology bioassays
are currently used in environmental risk assessment
protocols in water pollution monitoring, and direct
toxicity of both legacy and emerging contaminants
provide sensitive tools to assess both exposure and
effects of toxic compounds. Up to now, a number
274   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e
of standardized ecotoxicity test protocols using em-
bryo or larvae of several taxonomic groups have
been developed and successfully recommended
for determining the toxicity of chemicals to aquatic
species by regulatory bodies, such as the Organiza-
tion for Economic Co-operation and Development
(OECD), the American Society for Testing and Mate-
rials (ASTM), the Environmental Protection Agency
(EPA), and the International Organization for Stand-
ardization (ISO). For example, embryos and larvae
of echinoids and bivalves have been proposed as
appropriate models for ecotoxicological studies, as
many species in this group are abundant and have
early planktonic life stages which are directly ex-
posed to environmental change. More importantly,
several of these species are easy to work with in the
lab and have completely sequenced genomes (e.g.,
Sea Urchin Genome Sequencing Consortium, 2006).
Based on the assessment of different bioassay
protocols on species sensitivity to a wide variety
of toxicants, bioassays currently used are based on
the most available (worldwide) and ecologically
relevant marine invertebrate species, including the
purple sea urchin Strongylocentrotus purpuratus, red
abalone Haliotis rufescens, and bay mussel Mytilus
galloprovincialis; species used more recently include
the Pacific oyster Crassostrea gigas and sand dollar
Dendraster excentricus (U.S. EPA, 1995). Moreover,
other marine echinoids have recently been pro-
posed, mainly due to the need to find an ecologically
relevant model that is also abundant and easily ac-
cessible in nature; these include the Mediterranean
sea urchin Paracentrotus lividus and the red urchin
Arbacia sp. Ecotoxicology tests are commonly based
on short-term (<96 h) exposure conditions and sub-
lethal endpoints as embryo development but also
fertilization. For instance, echinoids are the most
sensitive test species to certain chemicals, where
it is easy to perform and screen a large number of
chemicals/environmental matrices in a short time
(Environment Canada, 2011; ASTM, 2012).
Sea urchin embryos have been used as models in
developmental biology for more than a century. As
deuterostomes they share many genes with verte-
brates and specifically mammals. More importantly,
they produce large numbers of transparent embryos
that can be studied in the lab in a short period of time;
cellular processes can be directly linked to molecular
changes thanks to a completely sequenced genome
(Sea Urchin Genome Sequencing Consortium, 2006).
In addition, sea urchin embryos’ sensitivity toward
environmental perturbations was investigated for
sensitivity toward several stressors, including UV-B
radiation and anthropogenic contaminants, the
latter being at low environmental concentrations
(Kobayash­i, 1980; Morroni et al., 2016).
Much attention has been recently devoted
towar­ d the sea urchin “chemical defensome,”
or genes predicte­ d to be involved in chemical
defense to confe­ r resilience and survival to de-
veloping embryos. These genes include stress-
activated receptors (aryl hydrocarbon receptor,
pregnane-X-recepto­ r, constitutive androstane re-
ceptor, peroxisome p ­roliferator-activated recep-
tors, liver-X-receptor, and farnesoid-X-receptor),
oxidizers (cytochrome P-450 enzymes and flavo-
protein monooxygenase), reducers and conjugated
enzymes (aldo-keto reductase­s, NAD(P)H-quinone
oxidoreductases, glutathione-S-transferases, UDP-
glucuronosyl transferases, N-acetyl transferases),
oxidative detoxification proteins (superoxide dis-
mutases, catalase, glutathione peroxidases), sol-
ute carrier family and efflux pumps (ATP-binding
cassette-ABC-transporters) (Goldstone et  al., 2006;
William­s and Carrier, this volume). Among these,
the ABC transporters, which have been and are
highly conserved over evolutionary time, have
received a lot of attention from the scientific com-
munity due to their involvement in the cellular
detoxification, including in sea urchin embryonic
development (see Table 18.1 for references).
18.2  Sea Urchin Development
Preceding the formation of the juvenile sea urchin,
development occurs freely in the water column
and serves as a key component of the zooplank-
ton. According to the species, the first cell divi-
sion occurs between 90 minutes (tropical species)
and 12 hours (Antarctic species) post-fertilization
(Hoegh-Guldber­g and Pearse, 1995). The embryos
then undergo a series of cell divisions leading to the
blastula stage, at which time hatching occurs. Sea
urchin embryos gastrulate in the plankton, while
swimming. The time needed to reach the larval
stage depends on the species, climate conditions
Table 18.1  Anthropogenic Contaminants and Their Interactions with MXR Phenotype and Larval Toxicity in Marine Invertebrate Embryos and Larvae.

Class of Compound Use MXR interaction Larval Toxicity Test

Contaminant (Model Organism) Model Organism (LC50)

Antifouling agent Sea-Nine 211 Marine industry Substrate* Mytilus edulis (38–372 nM)9
(Sea urchin embryos and larvae1) Artemia sp. (0.318 mg/L)10
Antimicrobials Triclosan Biocide, personal care products, plastic materials Circumvention** Palaemonetes pugio (154.0 μg/L)11
(Sea urchin larvae2) Strongylocentrotus nudus (0.987
μM)12
Fluorosurfactants Heptadecafluorooctane sulfonic acid Emulsion polymerization in metal plating, fire-fighting Circumvention (Sea urchin larvae2) -
foams and other materials
Fungicide Dichlofluanid Agriculture Substrate (Sea urchin embryos Mytilus edulis (224–4311 nM)9
and larvae1) Artemia sp. (154.94 mg/L)10
Cadmium (CdCl2) Photography, fabric printing, electronics component Substrate (Mussel embryos and Haliotis rubra (4,515 mg/L)13
manufacture, analytical chemistry and others larvae3)
Mercury (inorganic) Industrial processes, personal care products and medical Substrate (Sea urchin embryos4) Haliotis rubra (21 mg/L)13
applications
Metals
Copper (CuSO4) Agricultural insecticide, fungicide and herbicide, catalyst Substrate/Circumvention Haliotis rubra (7 mg/L)13
in pharmaceutical industry and antiseptics or disinfectants (Sea urchin embryos5)
Zinc (ZnSO4) Fertilizer, nutritional supplement, astringent Substrate/Circumvention -
in pharmaceutical industry (Sea urchin embryos5)
Copper oxide nanoparticles Catalyst in several industrial process, chemical sensor, Circumvention -
semiconductor (Sea urchin embryos5)
Zinc oxide nanoparticles Catalyst in several industrial process, optical and electrical Circumvention (Sea urchin Artemia salina (>100 mg/L)14
Nanomaterials
devices, personal care products, drug delivery embryos5)
Carboxylated polystyrene nanoparticles Plastic materials, personal care products Up-regulation of ABCB1 gene -
(Sea urchin embryos6)
Nonionic surfactants Pluronic P-85 (nanoparticles) Drug delivery systems Circumvention (Sea urchin larvae2) -
Oil hydrocarbons - Petroleum industry Substrate (Fat innkeeper worm -
larvae7)

(continued)
E c oto x ic o l o g y i n M a r i n e E n v i r o n me n t s    275
Table 18.1  (Continued )

Class of Compound Use MXR interaction Larval Toxicity Test

Contaminant (Model Organism) Model Organism (LC50)

Dichlorodiphenyltrichloroethane (DDT) Insecticide Substrate/Circumvention Litopenaeus stylirostris (10.79

(Sea urchin embryos and larvae2) mg/L)15

Organochlorines Pentachlorophenol Insecticide and disinfectant Circumvention (Sea urchin larvae2) Artemia salina (0.30 mg/L)16
Crangon crangon (0.11 ppm)17
Palaemon elegans (0.08 ppm)17
Palaemonetes varians (0.36 ppm)17
Phenols Bisphenol A (BPA) Plastic industry Circumvention (Sea urchin larvae2) -
Up-regulation of ABCB1 gene -
(Sea urchin embryos8)

*Based on toxicity assays in the presence of ABC transporter blockers.

**Based on ABC transporter substrate accumulation assays or toxicity assays in the presence of toxic ABC transporter substrate.
1Xu,
X. et al., Ecotoxicology (2011) 20:419–428
2Anselmo,
H.M.R. et al., Ecotoxicology (2012) 21:2276–2287
3McFadzen,
I. et al., Marine Environm Res (2000) 50:319–323
276   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e

4Bošnjak, I. et al., Environ Sci Technol (2009) 43:8374–8380

5Wu, B. et al., Environ Sci Technol (2015) 49:5760–5770
6Della Torre, C. et al., Environ Sci Technol, (2014) 20:12302–12,311
7Hamdoun, A.M. et al., Aquat Toxicol (2002) 61:127–140
8Bošnjak, I. et al., Aquat Toxicol (2014) 156: 21–29
9Bellas, J., Sci Total Environ (2006) 367:2–3
10Jung, S.M. et al., Mar Pollut Bull (2016) 16:30969–9
11Delorenzo, M.E. et al., Environ Toxicol (2008) 2:224–32
12Hwang et al., Ecotoxicol Environ Saf (2014) 100:148–52
13Groski, J. et al., Environ Toxicol Chem (2006) 25:1360–7
14Ates, M. et al., Environ Sci Process Impacts (2013) 15:225–33
15Galindo Reys et al., Ecotoxicol Environ Saf (2002) 53:191–5
16Barahona, M.V. et al., Bull Environ Contam Toxicol (1996) 56:271–8;
17van Dijk, J.J. Bull Environ Contam Toxicol (1977) 17:622–30.

such as temperature and sunlight, and the composi-
tion and availability of nutrients. The larval stage
lasts for several weeks (four to six) and ends with
the metamorphosis into a benthic juvenile.
Pluteus larvae are bilaterally symmetrical and
develop a series of arms, supported by an endog-
enous skeleton. These arms also create scaffold for
an extensive ciliated band used for feeding. Several
morphological features as well as morphometric
measurements of the pluteus larvae have been used
in the evaluation of sea urchin larval development
in ecotoxicological studies. These include abnormal
or delayed development; number and distribu-
tion of pigment cells; number and size of the arms;
length of larval axes; formation and size of larva ru-
diment; swimming behavior; and absent, incorrect,
or incomplete localization of the larval skeletal rods
(Carballeira et al., 2012).
18.3  ABC Transporters: Brief History
and General Overview
The ABC transporters constitute a large family of
transmembrane proteins linked to the MDR/MXR
phenotype (multidrug resistance and multixenobi-
otic resistance, respectively). The first description of
the MDR phenomenon in eukaryotic cells dates from
the beginning of 1970s. Biedler and Riehm (1970),
studying leukemic cell lineages and Chinese ham-
ster lung cells exposed to increasing concentrations
of actinomycin D, observed that cells acquired re-
sistance not only to actinomycin D but also to drugs
chemically and pharmacologically unrelated. Subse-
quently, Danø (1973) suggested that an active efflux
transport was responsible for the cross-resistance
to daunomycin and vinca alkaloids in the murine
Ehrlich ascites tumor cells. Three years later, the first
ABC transporter family member—P-glycoprotein
(currently also known as ABCB1)—was described in
Chinese hamster ovary cells with a pleiotropic cross-
resistance (Juliano and Ling, 1976).
The cloning and sequencing of three mdr mu-
rine genes was firstly reported in 1986 (Gros et al.,
1986), and soon studies demonstrated that mdr1
gene transfection confers the MDR phenotype to
nonresistant cells (Gros et  al., 1986). At the end of
1980s, the development of monoclonal antibod-
ies against intracellular and extracellular epitopes
E c oto x ic o l o g y i n M a r i n e E n v i r o n me n t s    277
of P-­ glycoprotein revealed the physiological ex-
pression of the transporter in several mammalian
tissues (Thiebaut et  al., 1987). Nevertheless, only
with the seminal work of Schinkel et  al. (1994),
who demonstrated the P-glycoprotein expression
in the blood-brain barrier and its involvement in
drug bioavailability, was the physiological role of
the ABC transporters highlighted. Since then, ABC
transporters have been studied in several organ-
isms, from bacteria to humans.
P-glycoprotein was first described in marine or-
ganisms by Kurelec (1992), who coined the term
multixenobiotic resistance (MXR). The first descrip-
tions were of the Mediterranean mussel Mytilus gal-
loprovincialis, the swan mussel Anodonta cygnea, and
the sponges Tethya aurantium, Verongia aerophoba,
and Geodia cydonium. The main result of the MXR
mechanism is to decrease the intracellular concen-
tration of chemical hazardous compounds, both as
parent and as metabolites. However, ABC trans-
porters are not only involved in the efflux of xeno-
biotics but also of several endogenous compounds
and ions, such as hormones, sterols, lipids, phos-
pholipids, oligopeptides, nucleotides, glutathione
S-conjugates, and chloride ion (Gottesman and
Pastan, 1993). Despite their nonspecificity in bind-
ing chemical substrates, common features—such as
moderate hydrophobicity, small size, and positively
charged domains of their substrates—can be used
to better understand their role in cell protection and
the ability to respond to toxic chemical exposure.
The ABC transporters are found in the plasma
membrane (e.g., ABCB1/P-gp, ABCC1/MRP,
ABCC7/CFRT, and ABCC8/SUR) and the endomem-
brane system of eukaryotic cells (e.g., ABCB2/TAP1,
ABCB3/TAP2, PGH1, ABCD2/ALDP, ABCD3/
PMP70, and MTABC) (Higgins, 1992), and exhibit
two highly conserved nucleotide-binding domains
which hydrolyze ATP (Walker et al., 1982). The ATP-
binding domains were the bases to coin the name of
this large family of proteins in the early 1990s (Hyde
et  al., 1990), and their highly conserved sequences
across animal taxa have allowed the identification of
ABC transporter genes in various organisms.
The eukaryotic ABC superfamily is divided into
eight subfamilies (ABCA through ABCH) and com-
prises proteins involved in efflux of endo- and xeno-
biotics, but also in the regulation of protein synthesis.
278   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e
Members associated with the MDR/MXR pheno-
type—and involved in the efflux of endo- and xeno-
biotics—are distributed into the subfamilies ABCB,
ABCC, and ABCG, notably the transporters ABCB1,
ABCC1, and ABCG2. Subfamily A comprises pro-
teins associated with the transport of lipids (e.g.,
cholesterol). Subfamily D is represented by genes
that encode proteins involved in the uptake of fatty
acid by peroxisomes; the genes of subfamilies E and
F encode proteins that lack the transmembrane do-
main and are not involved in transport but in the
regulation of protein synthesis. Finally, subfamily
H is closely related to subfamily G, and has been
described in invertebrates, such as insects and sea
urchins, and also in the zebrafish Danio rerio.
The ABC transporters can be expressed in the
cell membranes as a single polypeptide chain (full-
transporters; subfamilies B and C) or as hetero- or
homodimers (half-transporters; subfamilies B
and G). Nonetheless, independently of the struc-
tural configuration of the protein in the cell mem-
branes, a functional ABC transporter comprises two
­nucleotide-binding domains and two large trans-
membrane domains, each one composed by several
α-helices transmembrane segments. Several genes
coding for ABC transporters have been described in
eukaryotic cells. The human genome has 48 genes
encoding functional ABC proteins; this number
increases to 52 genes in Danio rerio and reaches 65
genes in the sea urchin Strongylocentrotus purpuratus
(Annilo et al., 2006; Goldstone et al., 2006). Never-
theless, only 33 ABC transporter genes were found
in the sea squirt Ciona intestinalis and in the sea lam-
prey Petromyzon marinus (Annilo et  al., 2006; Ren
et  al., 2015). In spite of the great variability in the
number of ABC transporter genes according to the
organisms, the three most representative subfami-
lies are ABCB, ABCC, and ABCG. In the sea urchin
Strongylocentrotus purpuratus, the ABCC subfamily
represents almost 50% of all ABC genes.
Several compounds compromise the efflux me-
diated by ABC transporters and have been used as
pharmacological tools in investigations of the MDR/
MXR phenotype in a wide range of biological models
(Robert and Jarry, 2003). From verapamil to MK571,
dozens of compounds have been shown to modulate
the activity of ABC transporters; they contributed to
the understanding of the role of the transporters in
several biological systems. These compounds can
act as competitive or non-­competitive inhibitors and
disrupt the protective effect of the pumps against xe-
nobiotics or endogenous compounds.
Studies conducted with mammalian ABC
transporters revealed that many pesticides—­
halophenoxies (tetrachlorohydroquinone), or-
ganochlorines (chlordecone, endosulfan, and
heptachlor), and organophosphates (dicapthon
and parathion)—affect ABCB1 transporters and
act as MXR chemosensitizers (Bain and Leblanc,
1996). Some of them, like the organochlorine en-
dosulfan, are also embryotoxic to the oyster Cras-
sostrea gigas (Wessel et  al., 2007). However, their
effects on the MXR phenotype and on cell defense
against anthropogenic or natural toxins are still un-
known, and more studies should address if their
toxicity might be linked to the circumvention of
the MXR phenotype in marine invertebrate em-
bryos and larvae. Studies have shown that marine
natural compounds—such as those derived from
algae (caulerpin), tunicates (ipatellamides and
amellarins), sponges (agosterol A, sipholenol A),
bryozoans (bryostatin 1), red algae (parguerenes),
and cyanobacteria (alkaloids and the porphyrin
tolyporphi­n)—are able to disrupt ABC transporters
by blocking ABCB1 efflux activity and/or circum-
vent the MDR/MXR phenotype (Schroeder et  al.,
1998; Lopez and Martinez-Luis, 2014).
Recently, studies have linked the exposure to ma-
rine natural toxins with the expression level of ABC
transporters in marine invertebrates. The co-culture
of the mussel Perna viridis and the bivalves Ruditapes
philippinarum, Scapharca subcrenata, and Tegillarca
granosa with the toxic dinoflagellate Prorocentrum
lima has been shown to up-regulate the expres-
sion of ABC transporters in directly affected tissues
(Huang et al., 2015a; 2015b). Moreover, Tanaka et al.
(2002) showed that the expression of ABCC1 trans-
porter confers resistance to polyoxygenated steroids
that are derived from corals. These data encourage
the investigation about the effects of marine natural
toxins on ABC transporters’ expression and activity
in marine invertebrate embryos and larvae, which
could increase current knowledge of the biology of
marine invertebrates not only in terms of develop-
ment, ecology, and evolution, but also in terms of
the overall ecosystem’s dynamics.

18.4  ABC Transporters in Sea Urchin
Embryos and Larvae
ABC transporters are present in sea urchin gam-
etes, embryos, and the pluteus. The first report of
the expression of an ABC transporter in sea ur-
chin cells was from the early 2000s. In studying
the acrosome reaction, Mengerink and Vacquier
(2002) observed that the major glycoprotein of the
sea urchin sperm membranes was closely related
to the human ABCA3 transporter. Two years later,
Hamdoun et al. (2004) showed that the activity of
ABCB and ABCC-like subfamily transporters is
up-regulated just following fertilization in the sea
urchin Strongylocentrotus purpuratus. Since then,
several works have mapped the expression, activ-
ity, and role of ABC transporters in the sea urchin
development (De Souza et al., 2010; Torrezan et al.,
2012; Shipp and Hamdoun, 2012; Whalen et  al.,
2012; Shipp et al., 2015).
Twenty ABCB, ABCC, and ABCG transporter
subfamily genes are expressed during sea urchin
embryonic development—from unfertilized egg
to early prism stage—with ABCB1a being the most
abundant. Nevertheless, the expression of ABC
transporter genes is not constant during sea urchin
development. ABCB subfamily genes are present in
the egg but decrease in expression until the hatch-
ing blastula stage, when expression is restored
until the early prism. ABCC5a and three ABCG tran-
scripts are absent until the hatching blastula, when
expression increases, particularly to the ABCC5a.
The ABCG2b transcript is expressed only in the
early prism stage. Finally, three ABCC transcripts
(ABCC1, ABCC4b, and ABCC4c) increase during
hatching blastula and early gastrula stages (Shipp
and Hamdoun, 2012).
Expression of ABC transporters in sea urchin
gametes, embryos, and larvae is crucial to the estab-
lishment of a biochemical barrier that protects gam-
etes as well as embryonic and larval development
against toxicants from both natural and anthropo-
genic sources. When exposed to the ABC substrate
calcein-AM, sea urchin embryos exhibit a low ac-
cumulation of the dye; however, when embryos are
exposed to calcein-AM in the presence of the ABCB1
blocker reversin 205 or the ABCC1 blocker MK571,
there is a notable increase in dye accumulation in
E c oto x ic o l o g y i n M a r i n e E n v i r o n me n t s    279
the embryonic cells (Figure 18.1). In spite of embry-
onic cells exhibiting a higher activity of ABC trans-
porters than gametes, eggs and spermatozoa also
exhibit significant activity of the transporters (De
Souza et al., 2010). The increase in ABC transporter
activity, and consequently in the MXR phenotype
in sea urchin embryonic cells, can be attributed to
the exocytosis of the cortical granules and the dis-
posal of the ABC transporters in the microvilli of
the cellular membrane of the zygote (Whalen et al.,
2012). In fact, sea urchin embryonic development is
more sensitive to toxicants when eggs are exposed
to them prior to fertilization and zygote formation
(De Souza et  al., 2010). A similar pattern of ABC
transporter expression and activity was observed
in freshwater invertebrates, such as in the zebra
mussel Dreissena polymorpha (Navarro et  al., 2012),
suggesting that the regulation of efflux pumps is a
conserved biological process during the embryonic
and larval development in aquatic organisms.
Apart from their protective role against xeno-
biotics, ABC transporters are also involved in the
transport of endogenous signaling molecules that
regulate embryonic development, and are involved
in several cellular processes (Shipp and Hamdoun,
2012). It has been demonstrated that the exposure of
sea urchin spermatozoa to ABC transporter blockers
decreases the fertilization rate, suggesting that these
proteins may be involved in cell membrane homeo-
stasis and perhaps gamete fusion (Silva-Neta et al.,
2012). It has also been reported that the inhibition
of ABC transporter activity in sea urchin embryos
disturbs the segregation of small micromeres to the
coelomic pouches, and because it is the left coelomic
pouch gives rise to the adult rudiment, this may
subsequently impact metamorphosis. Members of
the ABCC subfamily are also important to the sea
urchin’s embryonic development: Sp-ABCC5a is
highly expressed in pigment cells and aboral non-
skeletogenic mesoderm cells during and immedi-
ately following gastrulation. Studies have shown
that this protein is required for gut formation and
hindgut orientation, and its ablation causes gut pro-
lapse in developing embryos (Shipp et al., 2015).
The activity of ABC transporters has also been
reported in the sea urchin pluteus larvae. The use
of the intracellular calcein-AM accumulation assay
and pharmacological tools has shown a noteworthy
280   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e

control Reversin 205 MK571

ll

lll

lV

Figure 18.1  Effect of ABC transporter blockers in the MXR phenotype. Echinometra lucunter two-cell embryos (panels I and II) and gastrula (panels
III and IV) were pre-treated with the ABC transporter blockers reversin 250 or MK571 (both 10 μM) and incubated with calcein-AM (250 nM) for 30
minutes. The two-cell embryos treated with the ABCC1 transporter blocker MK571 exhibited a stronger calcein fluorescence signal than two-cell embryos
treated with the ABCB1 transporter blocker reversin 205 (panel II); the treatment with the ABCB1 blocker reversin 205 revealed a more intense calcein
fluorescence signal in the gastrula ectoderm and archenteron (panel IV). Representative photomicrography. Panels I and III = optical microscopy; panels
II and IV = fluorescence microscopy.

difference in the activity of ABCB1 and ABCC1
transporter in Echinometra lucunter pluteus and
calcein distribution. While ABCB1-like transporter
activity is remarkably found in the larval gut,
ABCC1-like transporter activity is present in the
whole larva (Figure 18.2) (Torrezan et al., 2012). This
configuration is closely related to that found in ver-
tebrates, where ABCB1 transporter is located in sev-
eral blood barriers, such as the blood-brain barrier,
and also in the apical membrane of intestinal cells.
 E c oto x ic o l o g y i n M a r i n e E n v i r o n me n t s    281

arms arms

gut

Figure 18.2  Calcein disposition in the sea urchin pluteus larvae. Echinometra lucunter pluteus larvae were pre-treated with the ABC transporter
blockers reversin 250 or MK571 (both 10 μM), and incubated with calcein-AM (250 nM) for 30 minutes. The treatment with the ABCB1 transporter
blocker reversin 250 revealed activity of the ABCB1-like transporter in the larval gut (left image); the treatment with the ABCC1 transporter blocker
MK571 revealed activity of the ABCC1-like transporter in the whole pluteus larvae (right image); untreated pluteus larvae show no fluorescence
signal (not shown). Images are three-dimensional reconstructions from serial sections obtained under confocal laser scanning microscopy.

18.5  ABC Transporters, Echinoderms,
and Ecotoxicology
The ABC transporters are responsible for the MXR
phenotype and act as the first line of cell defense
against both natural and anthropogenic toxicants.
These efflux proteins have been well documented
in marine invertebrates, such as mussels, oysters,
and echinoderms. While members of the ABCB and
ABCG subfamilies are involved in the pumping of
xenobiotics out of the cells (Phase 0 or uptake), mem-
bers of the ABCC subfamily are enrolled in the efflux
of metabolites (Phase 3 or elimination) (Figure 18.3).
Not directly involved in the metabolism of the toxic
chemicals, they play an important role in their cel-
lular uptake and disposition. Thus, the commitment
of ABC transporters activity can seriously affect the
biotransformation/detoxification system. Such a de-
fense system is able to play a protective role in pol-
luted environments, and therefore this mechanism
is highly conserved by marine organisms, including
embryos and larvae, which allows them to respond
to different types of stressors (physical, chemical,
and biological) (Epel et al., 2008).
Since the first description of ABC transporter ex-
pression in sea urchin embryos (Hamdoun et  al.,
2004), the echinoderms have emerged as a suitable
model for ecotoxicological studies and the investiga-
tion of the MXR phenomenon in marine organisms.
The ABC transporters are crucial to sea urchin devel-
opment because they protect the embryonic and larval
stages against chemical (potential toxic compounds;
De Souza et al., 2010) and physical (ultraviolet radia-
tion; De Araújo Leite et  al., 2014) stressors. Several
works have shown that the occurrence of toxicants
affects sea urchin embryos and pluteus larvae when
ABC transporters are blocked. Sea urchin embryos
normally develop when exposed to low concentra-
tions (100 nm) of the anticancer agent vinblastine;
however, when embryos are co-exposed with ABCB1
transporter blockers, such as verapamil, trifluopera-
zine, chlorpromazine, or reversin 205, development
is arrested (De Souza et al. 2010; unpublished data).
ABC transporter inhibitors also induce chemosensiti-
zation to UV radiation. Abnormal and delayed larval
development occurred when gametes were exposed
to UVB in the presence of the ABC transporter block-
ers reversin 205 and MK571 (Figure 18.4).
282   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e

1 ABCB1/ABCG2
Transporter MXR Phenotype
ABCC1 Transporter

5 Phase-ll detoxification enzymes

3 anthropogenic contaminant
natural toxicant
4 ABC transporter blocker (natural or anthropogenic source)
conjugation functional group

Figure 18.3 ABC transporters and the MXR phenotype. A natural toxicant or anthropogenic contaminant may be pumped out of the cell by the
ABCB1/ABCG2 transporters (Phase 0, uptake) or (1) enter the cell (2) and—after Phase I biotransformation—(3) be conjugated with other molecules
(Phase II biotransformation) such as glutathione, and (4) pumped out of the cells by the ABCC1 transporter (Phase III, elimination). Nevertheless, a
natural toxicant or anthropogenic contaminant may also, directly or competitively, block an ABC transporter and circumvent the MXR phenotype, thus
(5) making cells susceptible to the toxic effects of many chemicals.

control reversin 205 MK571

unexposed

exposed sptz

exposed eggs

Figure 18.4 ABC transporters and ultraviolet B (UVB) radiation. Representative photomicrography of Echinometra lucunter pluteus larvae (48 h post-
fertilization). Gametes were treated previously to UVB exposure with the ABC transporter blockers reversin 205 and MK571 (both10 μM) and submitted
to fertilization procedure. Spermatozoa and eggs were exposed to 0.45 kJ m-2 and 14.4 kJ m-2 UVB, respectively. Control represents pluteus larvae derived
from treated or untreated unexposed gametes.

18.6  ABC Transporters and Bioassays
In the absence of specific antibodies against ABC pro-
teins for most marine invertebrates, investigations of
ABC transporters have focused on gene expression
and efflux activity. In spite of the higher specificity of
gene expression assays, efflux assays are crucial, since
the presence of the MXR mechanism is a relevant fea-
ture in ecotoxicological studies, and changes in gene
expression levels may not alter MXR phenotype and
have no ecotoxicological impact. On the other hand,
gene expression monitoring can reveal molecular
targets to be used to understand ABC transporters
involvement in the response to toxicants.
Several studies have investigated the effects of
exposure to toxic compounds on the expression of
ABC transporters in marine invertebrates, including
embryos and larvae. The up-regulation of several
genes encoding for efflux pumps has been shown
to confer resistance against toxic chemicals (e.g.,
cadmium) (Ivanina and Sokolova, 2008; Della Torre
et al., 2014b). Despite such studies clearly elucidat-
ing the regulation of specific ABC efflux pumps,
the connection between mRNA abundance, protein
expression, and activity has not yet been clearly
elucidated; thus linking these remains a major chal-
lenge. The lack of antibodies to detect specific efflux
pumps in marine species is also a major obstacle in
understanding the regulation of ABC transporter ef-
flux pumps, as well as functional characterization.
We also lack an understanding of the mechanism(s)
whereby the expression of ABC transporters is in-
duced, and how this varies between tissues and
species. A cAMP/PKA-mediated regulation of the
expression of ABCB in marine mussels has been
shown by Franzellitti (2013); drugs as well as metals
and other environmental contaminants have been
shown to induce cAMP, so affecting ABCB tran-
scription, which could act as a defense strategy.
Several ABC transporter substrates have been
used both for in vitro and in vivo assays as a means
to monitor the efflux activity of the pumps. Moreo-
ver, fluorescent dyes, such as rhodamine 123, rho-
damine B, and calcein-AM, are among the most
employed fluorophores that are used to study ABC
transporter activity and the MXR phenotype. In
spite this, these fluorescent substrates are not spe-
cific to each type of ABC transporter, as the use of
E c oto x ic o l o g y i n M a r i n e E n v i r o n me n t s    283
these dyes specifically predicts the presence of the
MXR phenotype. Since ABC transporters reduce the
accumulation of the substrate inside the cells, a low
fluorescent signal correlates with a higher activity
of the pumps, and, at the same time, an increased
fluorescent signal reflects a lower activity of the
transporters.
It is important to emphasize that certain fluores-
cent substrates are not suitable for use in the inves-
tigation of ABC transporter activity of cells from
marine invertebrates (e.g., rhodamine 123). Thus,
calcein-AM and rhodamine B are the widely used
in efflux bioassays in marine invertebrate embryos
and larvae, with most attention given to the former.
Calcein-AM has an advantage over rhodamine B, as
the dye emits fluorescence only after the removal
of the acetomethoxy group by intracellular ester-
ases. So, no further washing steps are needed for
its use under fluorescence or confocal microscopy.
Nonetheless, the use of dyes conjugated with ace-
tomethoxy group requires the use of positive con-
trols (ABC transporter blockers), since fluorescence
emission depends on the activity of intracellular
esterase to convert the non-fluorescent conjugated
form into its free fluorescent form. This is impor-
tant, especially when comparing dye accumulation
between samples from environment with different
level of contamination. Therefore, the data obtained
can be attributed to ABC transporter efflux activity
and not to a lower activity of intracellular esterases.
In this regard, MK571 and reversin 205 (ABCB1 and
ABCC1 transporter blockers, respectively) are the
most used positive controls.
The choice of the culture medium to be employed
in the evaluation of the ABC transporter activity is
another important aspect that must be considered
when setting an efflux assay protocol. According
to the cell type, the use of artificial culture medium
may induce artifacts in the use of ABC transporter
blocker as have been observed with sea urchin coe-
lomocytes (unpublished data). So, preferentially,
the culture medium must be the original body fluid,
or, depending on the biological model, natural sea-
water instead of artificial seawater as in the case of
marine echinoid embryos and larvae.
Short-term assays (30 min up to 1h), known as
competitive binding assays, provide good informa-
tion regarding the involvement of ABC transporter
284   E v o l u t i o n a ry E c o l o g y o f M a r i n e I n v e rt e b r at e L a r va e
activity in its cellular disposition as well as to test
the effect on efflux functionality. On the other end,
long-term assays may elucidate a more realistic ef-
fect of a contaminant on the MXR phenotype since
gene expression can also be addressed. So, it is
highly recommended that both approaches are ap-
plied in order to understand the impact of a contam-
inant on the ABC transporter activity and its related
MXR phenomenon. ABC transporter’s fluorescent
substrates can also be used in in vivo assays to moni-
tor the distribution of the dyes in several tissues.
This allows the evaluation of the integrity of the bio-
chemical barriers supported by the ABC transport-
ers and predicts the effect of contaminants in the
disposition of other toxicants in the whole organism.
18.7  Current Knowledge and Future Gaps
Current knowledge of how marine embryos and
larvae deal with several toxicological stressors dur-
ing development focuses on the defensive role of
the ABC transporters (Hamdoun and Epel, 2007).
The identification of substrates or inhibitors among
toxic compounds, as well as properties of the ef-
flux transporters in terms of tissue distribution,
regulation, and physiological function, represents
the main target of research on ABC transporters
in ecotoxicology. A concern that has emerged in
the field is the circumvention of the MXR pheno-
type in realistic exposure scenarios where mixture
of toxic compounds is present (Kurth et  al., 2015).
These proteins have been proposed as potential bi-
omarkers of toxic chemical exposure, despite clear
evidence of their ability to be involved in cell pro-
tection against mixtures of chemicals, as observed
in natural environmental scenarios. Some toxic
chemicals can bind to the transporters acting as
substrates or inhibitors, thus affecting subsequent
activity of chemosensors (Smital et al., 2004).
Because ABC transporters can be blocked by doz-
ens of compounds that are not chemically related,
there is a demand for a broad knowledge regard-
ing not only the direct effect of contaminants on the
activity of the pumps, but also their effects on the
expression of ABC transporters and the MXR phe-
notype. Moreover, the effect of seasonal variations
in the expression of ABC transporters represent an-
other relevant issue with very few investigated.
Another point to be considered is that species
belonging to commonly used taxa exhibit different
levels of tolerance to a same toxic compound. For
example, Martino et  al. (2016) showed substantial
differences in the sensitivity to the lanthanide metal
gadolinium in larvae from four echinoid species
from different geographical regions. Studies com-
paring the sensitivity of different marine inver-
tebrate larvae to several toxicants, as insecticides
(lindane and chlorpyrifos), herbicides (diuron),
and organometallic antifoulants (tributyltin), have
shown a different pattern of resistance among spe-
cies. Moreover, the sea urchin (Paracentrotus lividus)
and ascidians (Ciona intestinalis) larvae were more
resistant to lindane and chlorpyrifos than larvae
from the decapods Maja squinado and Palaemon ser-
ratus (Bellas et  al., 2005). These data highlight the
need of broader and deeper studies regarding ABC
transporters in embryonic and larval development,
addressing both the level of gene expression as well
as the activity of the transporters in geographically
and phylogenetically distant species and their in-
teractions with different ecosystems. Such knowl-
edge will increase the current understanding of the
biological role of ABC transporters and their con-
tributions to each taxon towards protection against
natural and anthropogenic toxicants. The design
of in situ experimental protocols and in vitro and
in vivo laboratorial studies with natural seawater
samples, and their effects on MXR phenotype, may
bring new insights about the relevance of the ABC
transporters to the protection against toxicants in
the marine ecosystem.
The biological relevance of the ABC transporters
in the protection against chemical or physical stress-
ors is clearly evidenced by the strong evolutionary
conservation of these proteins in the biological sys-
tems, including the aquatic invertebrates (Jeong
et al., 2017). However, further studies on embryos
and larvae of marine invertebrates are needed in the
ecotoxicological research field. The biochemical and
cellular factors driving the toxicokinetic and toxico-
dynamic properties of a toxic compound need to be
further elucidate. Major progresses have been made
in identifying ABC transporter genes in marine or-
ganisms but a general lack of knowledge concern
both embryos and larvae. Many gaps regarding
both the presence and physiological function of

ABC transporters as for instance their role in chemi-
cal uptake and their inhibition (chemosensitization)
in marine embryos and larvae need to be filled in
order to better understand the ecotoxicological rel-
evance of ABC transporters as defense mechanisms
towards toxic compounds.
18.8 Summary
1. Anthropogenic contaminants with the poten-
tial to disrupt biological functions enter aquatic
ecosystems from a diversity of sources. Their
impacts pose a potential risk to the long-term
sustainability of these ecosystems. Larvae and
other developmental stages can be used as sys-
tems to determine mechanisms for ecotoxicity of
these compounds.
2. ABC transporters, as a component of the “chemi-
cal defensome,” are responsible for the MXR
phenotype and act as first line of defense against
both natural and anthropogenic toxicants. They
are present in sea urchin gametes, embryos, and
the pluteus. Apart from their protective role
against xenobiotics, they are involved in the
transport of endogenous signaling molecules
that regulate embryonic development and sev-
eral cellular processes.
3. Several studies have investigated the effects of
exposure to toxic compounds on the expression
and activity of ABC transporters in marine in-
vertebrates, including embryos and larvae. The
up-regulation of several genes encoding for ef-
flux pumps has been shown to confer resistance
against toxic chemicals.
4. The identification of substrates or inhibitors
among toxic compounds, as well as properties
of the efflux transporters in terms of tissue dis-
tribution, regulation, and physiological function,
represents the main target of research on ABC
transporters in ecotoxicology. A concern that has
emerged is the circumvention of the MXR phe-
notype in realistic exposure scenarios where a
mixture of toxic compounds is present.
5. The involvement of ABC transporters in larval
lethality and the reversal of the MXR pheno-
type have not been properly addressed to date.
Further studies are thus recommended that link
E c oto x ic o l o g y i n M a r i n e E n v i r o n me n t s    285
ABC transporter gene expression and efflux
pump activity with toxicity, including mixtures
of compounds and chemical resistance among
different taxa.
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 Ch a pt er 19
An -Omics Perspective on Marine
Invertebrate Larvae
Elizabeth A. Williams and Tyler J. Carrier
19.1 Introduction
As larval biologists, we strive to understand how
evolution and ecology shape larval development,
morphology, behavior, and physiology. Larvae are
particularly fascinating subjects for studying evolu-
tionary pressures and environmental interactions,
primarily because this life stage of marine inverte-
brate is under strong selective pressures (Thorson,
1950; Pedersen et al., 2008), and larvae must inter-
act closely with their environment to successfully
complete development and ultimately transition
into juveniles (see Hodin et  al., this volume). Sev-
eral pioneers of larval biology, including Gunnar
Thorson, Fu-Shiang Chia, and Richard Strathmann,
were integral in emphasizing the importance of the
environment on larval evolution. Using a suite of
techniques spanning microscopy and high-speed
video combined with morphological descriptions,
behavior, and physiology, as well as records of bi-
otic and abiotic environmental parameters, they
provided fundamental insights into how the envi-
ronment influences development, life history mode,
metamorphic induction, and adult reproductive
biology (Thorson, 1950; Strathmann, 1987; Chia,
1989). What these and other seminal researchers
were unable to address, however, are the molecular
mechanisms underlying the translation of environ-
mental pressures into the evolution of phenotypic
traits (Figure 19.1).
With the advent of molecular biology, the
field began defining the mechanisms mediating
the interaction between the environment, larval
genotype, and the resulting larval phenotype(s).
Williams, E. A. and Carrier, T. J., An -omics perspective on marine invertebrate larvae. In. Evolutionary Ecology of Marine
Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland: Oxford University Press (2018).
© Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0019
The translation from genotype to phenotype is a
complex process that integrates sets of external (en-
vironmental) signals with larval physiology and the
regulation of gene expression during development.
Genotype is not related to phenotype in a direct
one-to-one ratio, rather, it is possible for one geno-
type to generate multiple phenotypes, depending
on environmental stimuli (see McAlister and Miner,
this volume), and conversely, multiple genotypes
can generate the same phenotype (genetic canaliza-
tion; see Flatt, 2005).
In recent years, larval biologists have turned to
molecular biological techniques, such as qRT-PCR,
subtractive screens of cDNA libraries, and in situ
hybridization, to explore the molecular mecha-
nisms underlying larval phenotypes. These ap-
proaches have allowed for the identification of key
genes involved in the regulation of larval develop-
ment and metamorphosis. For example, the Manx
gene is responsible for generating the brain sensory
organ, notochord, and tail in tailed (urodele) ascid-
ian larvae (Swalla and Jeffery, 1996); morphoge-
netic and anticipatory developmental pathways in
abalone metamorphosis (Degnan and Morse, 1995);
and ß-catenin, a key regulator of animal-vegetal axis
patterning in embryos of the sea urchin Lytechinus
variegatus (Wikramanayake et al., 1998). Molecular
approaches have also facilitated the identification of
potential molecular mechanisms underlying larval-
juvenile carryover effects (Williams and Degnan,
2009; Pechenik, this volume). While we are now
developing a better understanding of the molecu-
lar mechanisms underlying larval development, we
can also begin to address how these mechanisms
 A n - o m i c s P e r s pect i v e    289

genomics

genotype

environment developmental
(internal,
external) gene expression

transcriptomics
metabolomics
peptidomics

microbiomics

connectomics

phenotype

phenomics

Figure 19.1  -Omics approaches to the larval genotype-phenotype map. Genotype does not directly translate to phenotype; the process is influenced by
developmental gene expression and both internal and external environmental factors. External and internal environment can influence gene expression,
and gene expression can influence internal environment. Genomics allows large-scale mapping of genotype, while transcriptomics and peptidomics can
be used to explore developmental gene expression, as well as the effects of environmental changes on gene and peptide expression. Metabolomics
and microbiomics can be used to investigate the internal larval environment by large-scale measurements of metabolites and microbiota, respectively.
Connectomics can be used to explore the larval nervous system, which is responsible for sensing the external environment and generating initial
behavioral responses to the environment. Connectomics may also be considered a way to investigate larval phenotype, specifically, nervous system
connectivity. Phenomics can be used for large-scale mapping of larval phenotype, including behavioral and morphological parameters.

interact with internal and external environmental
changes to bring about the evolution of phenotypes.
The nature of these investigations requires integra-
tive studies in which data on larval genotype, mo-
lecular expression, physiology, and phenotype can
be combined with environmental monitoring in a
systems-level approach.
Numerous technical advances in DNA sequenc-
ing, computational analyses, and microscopy have
allowed for faster and larger-scale assaying of
molecules. These high-throughput approaches are
collectively known as “-omics,” and they enable
a transition from studying single or small groups
of genes, proteins, or metabolites to simultane-
ously profiling all molecules within a single larva
or group of larvae. Similarly, -omics studies with a
morphological and/or behavioral focus enable si-
multaneous profiling of multiple phenotypes within
an organism or across several groups of organisms.
These -omics approaches can facilitate larval biolo-
gists to study the interaction of genotype and en-
vironment to generate unique larval phenotypes,
ultimately providing a complementary platform for
advancing our understanding of larval evolution-
ary ecology (Figure  19.1). The main fields of -om-
ics are described in Box 19.1. The level of biological
organization and type of biological variation that
can currently be measured in marine invertebrate
larvae varies with each technique (Table 19.1).
Applying -omics techniques to larval ecology
and evolution has the potential to provide fruit-
ful insights into many aspects of the field that are
highlighted in this edited volume; namely, larval
development and life history transitions (Collin
and Moran, this volume), larval-microbe symbioses
(see later), coping with environmental variation
290   E v o l u t i o n a ry E c o l o g y o f Ma r i n e I n v e rte b r ate La r vae

Box 19.1  Types of -omics approaches.

Genomics—Study of the genome (entire DNA comple-
ment) of an organism (Del Giacco and Cattaneo, 2012).
Subfields of genomics include metagenomics, the study of
genetic material from organisms collected from environ-
mental samples (Riesenfeld et  al., 2004), microbiomics,
the study of the genomes of the microbial communities
inhabiting the living tissue of metazoans (Rosenberg and
Zilber-Rosenberg, 2011), and epigenomics, the study of
genome-wide epigenetic modifications to an organism’s
DNA (Biemont, 2010).
Transcriptomics—Detection and quantification of all
expressed transcripts within a cell, tissue or organism, in-
cluding mRNAs, non-coding RNAs, and small RNAs (Wang
et al., 2009).
Peptidomics—Study of the peptide complement produced
by an organism or system (Ivanov and Yatskin, 2005).
Metabolomics—Study of the entire complement of me-
tabolites produced by an organism, cell, tissue, or organ,
including endogenous (amino acids, organic acids, fatty ac-
ids, amines, sugars, vitamins, co-factors, pigments, antibiot-
ics) and endogenous (drugs, environmental contaminants,
food additives, toxins, xenobiotics) metabolites (Kell, 2004).
A subfield of metabolomics is lipidomics, study of the lipid
complement in a cell, tissue, or organism (Shevchenko and
Simons, 2010).
Connectomics—Reconstruction of the synaptic connec-
tions between the neurons of an animal’s nervous system,
to generate neuronal circuits underlying different behaviors
(Helmstaedter, 2013).
Phenomics—Large-scale measurements of the physical
and biochemical traits of organisms in response to genetic
and/or environmental change (Houle et al., 2010).

Table 19.1  Examples of -omics Approaches That Can Be Used to Study Marine Invertebrate Larvae, the Minimal Level of Biological Organization at
Which They Can Currently Be Performed, and the Related Molecular Biology Methods Used for Generating Data.

Field of study Minimal level of biological Related techniques

organization

Genomics Single cell DNA sequencing: Sanger/shotgun sequencing, which is rapidly being replaced by next-
generation sequencing (Illumina Solexa-HiSeq, MiSeq) and assembly from short reads
Metagenomics Single organism DNA sequencing, as above
Microbiomics Single organism DNA sequencing, as above
Epigenomics Pooled larval samples Chromatin immunoprecipitation-DNA microarray (ChIP-chip), chromatin
immunoprecipitation-next-generation DNA sequencing (ChIP-Seq)
Transcriptomics Single cell Microarray, quantitative real-time PCR, next-generation sequencing (RNAseq)
Peptidomics Pooled larval samples 1- or 2-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (1D/2D
SDS-PAGE), high-performance liquid chromatography (HPLC), mass spectrometry,
peptide sequencing, enzyme-linked immunoabsorbent assay (ELISA), mass spectrometric
immunoassay (MSIA)
Lipidomics Pooled larval samples Nuclear magnetic resonance (NMR), mass spectrometry, electrospray ionization (ESI), matrix-
assisted laser desorption/ionization (MALDI)
Metabolomics Pooled larval samples Separation: gas chromatography (GC), liquid chromatography (LC), HPLC, capillary
electrophoresis (CE). Ionization: electron ionization (EI), chemical ionization (CI), electrospray
ionization (ESI). Detection: mass spectrometry (MS), nuclear magnetic resonance (NMR)
Connectomics Single cell Transmission electron micrography (TEM), image analysis
Phenomics Single cell to single larva, Various imaging techniques, imaging analysis
depending on phenotypic trait

Note. Further description of methods can be found in Online Resource 1: Description of Techniques for the Acquirement of -omics Data.

(Byrne et al., this volume), and physiology of feed-
ing, growth, and nutrition (Pernet, this volume;
McAliste­r and Miner, this volume; Jaeckle, this vol-
ume). In this chapter, we aim to highlight the role
of -omics in our understanding of the following: (1)
larval development and life cycle evolution; (2) how
larvae cope with environmental variation; (3) inter-
actions between larvae and their symbiotic microbi-
ota; (4) larval physiology and feeding; and (5) larval
behavior. We also discuss current potential biases
and limitations of -omics approaches and provide
suggestions for future research directions.  -omics
studies can serve as tools for the generation of novel
hypotheses on gene function, however, it is impor-
tant that functional analyses are also carried out to
holistically test these hypotheses. Designing -omics
studies in a way that captures the extent of natural
variation within larval individuals and populations
is also crucial to improving our understanding of
microevolutionary processes at the molecular level.
With organismal-level profiling, -omics approaches
can provide us with the capacity to study the in-
teraction of genetic and environmental factors in
shaping the genotype-phenotype map in a holistic
manner (Figure 19.1).
19.2  Role of -Omics Approaches
in Deciphering Molecular Mechanisms
of Larval Development and Life History
Evolution
To study the influence that the environment has
on the genome, requires an understanding of the
gene expression pathways regulating larval de-
velopment. This understanding, in turn, provides
a substrate to outline the genetic mechanisms un-
derlying development processes, and we can then
begin comparing these between species to look
at how developmental programs have changed
over evolutionary time. The ongoing application
of transcriptomic-scale approaches to study gene
expression patterns across developmental stages
of diverse organisms was initially by use of mi-
croarray and subtractive hybridization methods.
For example, these techniques allowed Heyland
and Moroz (2006) to summarize common signal-
ing mechanisms of metamorphosis in ascidians
A n - o m i c s P e r s pect i v e    291
and molluscs as well as insects and amphibians.
Specifically, transcripts related to stress response,
immunity, and apoptosis, as well as signaling medi-
ated by hormones and nitric oxide, were found to
be associated with metamorphosis in all phyla in-
vestigated (Heyland and Moroz, 2006). Microarrays
and subtractive hybridization were subsequently
replaced with transcriptomics and peptidomics as
well as mass spectrometry in order to provide a
more comprehensive assessment of genes and pep-
tides with putative roles in development (Nomura
et al., 2009; Huan et al., 2012; Brekhman et al., 2015;
Huan et  al., 2015). For example, broad-spectrum
transcriptional profiling (i.e., mRNA sequencing)
that spans developmental stages enabled Levin
et al. (2016) to conduct metazoan-wide comparisons
of developmentally important genes.
Transcriptomic approaches, when complemente­d
by experimental and molecular manipulations,
provide a more comprehensive insights into the
molecular mechanisms of larval development
than when employed alone. For example, larvae
of the polychaete Hydroides elegans are induced to
settle and undergo metamorphosis by the bacte-
rium Pseudoalteromonas luteoviolacea. By combining
genomics and transcriptomics with pharmacologi-
cal treatments and genetic manipulation of the bac-
terial inductive cue, Shikuma et al. (2016) were able
to locate regions of the P. luteoviolacea genome and
the mitogen-activated protein kinase (MAPK) sign-
aling pathways in H. elegans responsible for the de-
velopmental transition. More specifically, H. elegans
larvae are induced to settle and metamorphose by
P. luteoviolacea-produced phage tail-like metamor-
phosis-associated contractile structures (MACs)
(Shikuma et  al., 2014). Moreover, by generating
mutant P. luteoviolacea missing an ~8.2 kb region of
the genome, Shikuma et al. (2016) showed that this
genomic region was important for the morphologi-
cal changes of metamorphosis, but not for inducing
settlement behavior, as larvae exposed to the mu-
tant bacteria undergo settlement, but not metamor-
phosis. Larvae induced by bacteria missing part of
the genome responsible for structural properties of
MACs undergo neither settlement nor metamor-
phosis, suggesting that the physical properties of
MACs are important for initiating the behavioral re-
sponse. Like most transcriptome studies comparing
292   E v o l u t i o n a ry E c o l o g y o f Ma r i n e I n v e rte b r ate La r vae
large-scale gene expression between different larval
developmental stages, this study identified several
candidate genes likely to play a role in the regulation
of metamorphosis, including genes with putative
functions in tissue remodeling, innate immunity,
and MAPK signaling. Pharmacological treatments
of larvae with MAPK pathway inhibitors showed
that this signaling pathway is important for regu-
lating morphological changes after the induction of
metamorphosis. Ideally, the function of candidate
genes identified by transcriptomic studies of devel-
opment could be confirmed by functional analysis
using gene knockdown or knockout. The applica-
tion of functional analyses in -omics experiments is
discussed further below.
Due to accessibility of embryos and diversity of
developmental modes, one marine invertebrate
phylum that has been a focus of larval develop-
mental and life history evolution studies is the
echinoderms. Prior to -omics approaches, molecu-
lar phylogenetics enabled researchers to broadly
survey this taxonomic clade to locate gains, losses,
and transitions in developmental mode. This was
done primarily through sequencing of the mito-
chondrial 16S ribosomal RNA and cytochrome
c oxidase subunit I genes, and was coupled with
larval type (planktotrophy, lecithotrophy, direct
developer) as a character state (e.g., Hart et  al.,
2004). This approach hypothesizes where and
when clades gained or lost a given larval type, as
well as infers the ancestral larval state. Advances
in sequencing technologies in the last decade have
enabled far broader phylogenetic approaches, with
greater sample sizes in terms of species number, to
investigate these questions. These data are collected
in online databases where they can be freely ac-
cessed by the research community for use in a wide
variety of comparative studies, including phylo-
genetics. A recent example is the web-based appli-
cation Echino­ DB, which collates next-generatio­ n skeleton, gut, and nervous system. Comparative
transcriptome data from an initial count of 42
specimens from 24 orders and 37 families for phylo-
genetic analysis (http://echinodb.uncc.edu; Janies
et al., 2016). Using this and other publicly available
databases, it is feasible that a complete mapping of
larval type to echinoderm molecular phylogeny,
building on the efforts of Hart et al. (2004), can be
completed within a reasonable time frame.
In addition to a detailed and broad phylogenomic
profile in echinoderms, comparative transcriptom-
ics can also be used to identify key mechanistic
differences between sister taxa with different life
history strategies. In a study by Israel et al. (2016),
mRNA-seq was used to profile the developmental
transcriptome of the sea urchins Heliocidaris tuber-
culata, a species with planktotrophic larvae, and
H. erythrogramma, a species with lecithotrophic lar-
vae, as well as Lytechinus variegatus, an outgroup
with planktotrophic larvae. In doing so, they
showed that the gene regulatory network (GRN)
between life history strategies was significantly
rewired between sister species, despite having di-
verged a comparatively short five million years ago.
GRN rewiring was more severe in the transition
from planktotrophy (for H. tuberculata) to lecitho-
trophy (H. erythrogramma) than between distantly
related planktotrophs (H. tuberculata vs. L. variega-
tus). This differential magnitude of GRN rewiring
indicates that a combination of subtle shifts in gene
expression patterns and loss of gene expression for
several genes that continue to be expressed in the
development of both planktotrophic species was at
least in part a driver of this major life history transi-
tion (Israe­l et al., 2016).
The comparative transcriptomic approach taken
by Israel et  al. (2016) permits simultaneous detec-
tion of changes in gene expression from several
different functional categories. For example, plank-
totrophic and lecithotrophic echinoderm larvae
display differential expression of genes with puta-
tive roles in skeletogenesis, and gastrointestinal
and neural pathways, suggesting genes and path-
ways that were likely to have been selected for or
against during this transition (Israel et  al., 2016).
These changes in gene expression account at least
in part for the distinctive morphological differences
between planktotrophic and lecithotrophic larval
transcriptomics can be broadly applied to other
interesting systems of life history evolution found
among marine invertebrates, which may enable us
to detect the underlying molecular mechanisms of
developmental transitions. Examples include po-
ecilogony, where planktotrophic and lecithotrophic
larvae are produced within a single species (Chia
et al., 1996), or comparisons between facultative and

obligate planktotrophs (e.g., the echinoid Clypeaste­r
spp., Emlet, 1986). Marine invertebrate larvae also
offer the exciting possibilities of studying hybrid
larvae, such as those produced by crosses between
planktotrophic and lecithotrophic echinoderms
(Raff et  al., 1999) and tailed and tail-less ascidian
larvae (Swalla and Jeffery, 1996).
19.3  -Omics Approaches for Measuring
Larval Response to Environmental
Change and Challenges
While changes in genotype and gene expression
undoubtedly play a role in determining larval phe-
notype, the influence of environmental cues on
phenotype cannot be ignored. The environment
can and does impact expression levels of genes,
proteins, and metabolites, each of which can lead
to downstream changes in larval morphology,
physiology, and behavior. Initial transcriptomic and
peptidomic investigations into the effects of envi-
ronmental change on larvae have been prompted
by concerns over the ability of larvae to cope with
predicted climate change scenarios (e.g., increased
temperature and/or decreased pH; see Byrne et al.,
this volume). At both levels of investigation, a com-
mon theme is the up-regulation of transcripts and
peptides involved in stress response, metabolism,
and calcification (Meyer et  al., 2011; Runcie et  al.,
2012; Evans et al., 2013; Padilla-Gamiño et al., 2013;
Chandramouli et  al., 2015; Harney et  al., 2016).
Moreover, studies using global molecular profiling
tend to use either transcriptomics or peptidomics,
but rarely both. For example, transcriptome studies
of environmental effects on larval gene expression
have mainly been carried out in anthozoans and
echinoids, while peptidomic studies have focused
mainly on bivalves, polychaetes, and barnacles.
Transcriptomic and peptidomic studies alone can
indicate genes and peptides that alter their expres-
sion under changing environmental conditions, and
thereby give an understanding of which molecular
pathways and specific tissue types or organ systems
within larvae are primarily affected and linked or
co-regulated at a molecular level. However, an even
more informative approach to understanding the
effects of the environment on larvae can be found in
A n - o m i c s P e r s pect i v e    293
the combination of transcriptomics and/or peptid-
omics with genomics, metabolomics, or phenomics.
An example of combining different -omics ap-
proaches for understanding how larvae cope with
environmental stress can be seen in an investigation
of the effects of increased temperature on sea urchin
larvae. Runcie et al. (2012) used mRNA-seq to com-
pare global gene expression differences in larvae of
the sea urchin Strongylocentrotus purpuratus reared
at 12 °C, 15 °C, or 18 °C. Larvae reared at higher
temperatures up-regulated expression of stress
response genes, such as Hsp70 and Hsp90, and
showed changes in expression of genes with puta-
tive function in protein folding, RNA processing,
and development. Morphological analyses showed
that larvae at 18 °C were significantly shorter along
the animal-vegetal axis. However, even at higher
temperatures, larvae still successfully completed
gastrulation. By analyzing transcriptomic profiles
in the context of the well-resolved sea urchin GRN,
this study found that the ectoderm/endomesoderm
specification GRN acts as a buffer for suppressing
the effects of increased temperature on larval de-
velopment (Runcie et  al., 2012). That is, the tem-
perature responses of upstream regulatory genes
did not systematically lead to similar responses in
their corresponding downstream genes. Thus, by
combining genomic and transcriptomic elements,
Runcie et al. (2012) could begin to reveal a molecu-
lar mechanism through which larvae can at least
partially maintain their developmental phenotype.
One means to translate the influence of the en-
vironment on larval phenotypes is phenomics,
as this rising branch of -omics may aid in linking
genotype and phenotype (Houle et al., 2010). Video
tracking can provide high-throughput analyses of
larval behavior, while 2D, 3D, and 4D (Hejnol and
Schnabel, 2006) imaging can be used for morpho-
logical analyses. An example of integrating -omics
approaches with the measurement of environmen-
tal parameters is already being set by the Tara ex-
pedition. From 2009 to 2013, Tara Oceans launched
a study of the global ocean ecosystem to sample
the plankton of the world’s oceans. Both genomics
and transcriptomics, in the form of Illumina next-
generation sequencing, were applied to plankton
samples, in combination with oceanographic meas-
urements (e.g., temperature, salinity, nutrients) and
294   E v o l u t i o n a ry E c o l o g y o f Ma r i n e I n v e rte b r ate La r vae
visual monitoring of plankton (Sunagaw­ a et  al.,
2015). High-throughput imaging tools, including
flow cytometry, microscopy, FlowCam, and ZooS-
can, were used to characterize the morphology of
marine invertebrate larvae (Karsenti et  al., 2011).
ZooScan allows automated measurements of zoo-
plankton size, abundance, and mass (Gorsky et al.,
2010). FlowCam (Fluid Imaging Technologies Inc.),
on the other hand, allows automated analysis of the
size and shape of planktonic organisms in seawa-
ter. High-throughput imaging technologies such as
these provide the possibility for researchers to de-
velop large-scale phenomic datasets to complement
genomic, transcriptomic, or peptidomic analyses
and relate genotype to phenotype in population
studies, improving our understanding of microevo-
lutionary processes.
Comparing sequencing data from eight oceanic
provinces at 67 stations in the Tara Oceans project
to measure organism abundance and biodiversity,
Lima-Mendez et al. (2015) were able to construct an
interactome among photic zone plankton groups, in-
cluding viruses, prokaryotes, and eukaryotes. In do-
ing so, they indicated that temperature and dissolved
oxygen were the strongest correlates of community
composition in the surface layer (Lima-Mendez
et  al., 2015). A photosymbiotic interaction between
an acoel flatworm (Symsagittifera sp.) and a green
microalga (Tetraselmis sp.) could also be predicted by
the interactome, which was confirmed by confocal
microscopy (Lima-Mendez et al., 2015). This study is
a further example of the usefulness of combing phe-
notypic measurements from imaging studies with
genomic data in order to discover new biological
phenomena. Analysis of marine invertebrate larval
samples from the Tara expeditions may also be used
in furthering our understanding of environmental
effects on marine invertebrate larvae as holobionts.
19.4  Marine Invertebrate Larvae
and Their Microbiome
Symbioses with microbes span Metazoa, are critical
components in the potential for animals—­including
their early stages (i.e., embryos and larvae)—to ac-
climate and adapt, and are evolutionary agents
(Gilbert et al., 2015). The role of bacteria and other
microbes in the evolutionary ecology of marine
invertebrate larvae ranges from the symbiotic mi-
crobiome to the induction of metamorphosis by
microbial biofilm complexes (see Hodin et al., this
volume). A majority of the literature regarding
larval-microbe interactions focuses on the latter,
­
while the former remains little explored. This comes
as no surprise, because only recently has the im-
portance of symbiotic microbiota been recognized
(­Zilber-Rosenberg and Rosenberg, 2008; Rosenberg
et  al., 2009; Gilbert et  al., 2012; Bordenstein and
Theis, 2015; Gilbert et al., 2015), a trend mediated by
recent advancements in next-generation sequencing
technologies (e.g., MiSeq and 454-pyrosequencing)
and computational techniques (e.g., QIIME and
mothur). These components have enabled the mi-
crobial consortiums associated with marine inver-
tebrate larvae to be characterized and quantified,
providing a substrate to determine their potential
role in larval biology. Here, we briefly discuss how
a dynamic hologenome provides a buffer against
environmental stressors, and the role these symbi-
onts may play in larval evolution.
While in the plankton, marine invertebrate lar-
vae experience acute (e.g., entering a warm-water
mass) and chronic (e.g., ocean warming) periods of
temperature-induced physiological stress (Byrne
et al., this volume). A highly conserved and broadly
recognized mechanism used by larvae to cope with
temperature stress is the expression of heat-shock
proteins. Larvae, however, also expel or down-
regulate (by decreasing abundance) less advanta-
geous associated microbes while either horizontally
acquiring (Figure  19.2) or up-regulating other mi-
crobial taxa. For instance, following temperature
stress (from present, 28 °C, to future, 34 °C), larvae
of the sponge Rhopaloeides odorabile expel Nitros-
pira, Chloroflexi, and Roseobacter bacterial lineages
and are primarily replaced with γ-proteobacteria
(Webster et  al., 2011). In the face of ocean warm-
ing, chronic temperature stress will be common for
planktonic larvae. Given that the composition of
associated microbiota adapts faster than the host
genome, over the course of a few generations it is
suspected that larvae could inherit microbes (i.e.,
acquire as obligate symbionts) to cope with climate-
related temperature stress (Figure  19.2). This type
of hologenomic plasticity may be applied more
broadly to other abiotic or biotic stressors that
 A n - o m i c s P e r s pect i v e    295

(A) Neutral
Horizontially transmitted
Vertically transmitted
Novel environmental symbiont
Novel vertically transmitted
symbiont
Novel horizontally transmitted
symbiont

Epithelial Microbiome Gut Microbiome

Environmental Metagenome

(B) Phase 1 Phase 2 Phase 3

Pre-Environmental Stress Post-Stress (Acute) Post-Stress (Chronic)

Figure 19.2 Larvae are holobionts. Marine invertebrate larvae are classically viewed as “biological individuals” consisting of a eukaryotic genome.
Recent research suggests a symbiotic way of life, where all eukaryotics form intricate partnerships with a diverse community of prokaryotic microbes,
including bacteria, viruses, and Archaea. Moreover, current theory proposes that hosts co-evolve with these symbiotic microbiota and that this
consortium functions together as a unit of natural selection, with variation in that community being derived from environmental as well as genomic
influences. These symbionts are an extension of the host genome (together termed the “hologenome”) and remain distinct from the environmental
metagenome; in that, the host’s microbiome is of microbes that are commonly <0.1% of the environmental metagenome (termed “rare biosphere”).
(A) For most animals, there are two sub-communities among the larval microbiome: the epithelial microbiome lining the larval-seawater interface, and
the gut microbiome that colonizes the digestive track of the larva. The microbes within these communities come in several “flavors”: neutral (dark
and light gray and white), those that appears to provide to functional advantage; vertically transmitted, those maternally or paternally inherited; and
horizontally transmitted, those acquired from the environment. (B) The composition of the hologenome is highly dynamic in the face of environmental
stressors. Whether by stochastic association or selection by the larval host, novel microbes apart from the environmental metagenome may join the
hologenome in response to an abiotic or biotic stressor. If this association provides an advantage to the holobiont, maintaining a partnership is selected
for, and the degree to which the symbionts are required by the host subsequently determines, in part, whether an obligate (commonly horizontally
transmission) or facultative (commonly vertical transmission) partnership is established. (see Plate 15)

larvae commonly face, including but not limited
to diet (i.e., shifts in phytoplankton communities;
see Pernet, this volume), the physical ocean (Pineda
and Reyns, this volume), and toxicants (Corsi and
Marques-Santos, this volume).
A common concern of microbiome-centric stud-
ies, including those of marine invertebrate larvae,
is whether this consortium is symbiotic and spe-
cific to the host. An example supporting a larval-
selected microbiome is Galac et  al. (2016), who
identified teleplanic asteroid larvae from the west-
ern North ­Atlantic, and, using 454-pyrosequencing,
characterized the microbiome associated with those
larvae and that of the corresponding seawater. There
are, as with studies of other systems, marked differ-
ences between the asteroid larval microbiota and
that of the seawater, suggesting that these larvae
recruit specific microbes. Further support for this
claim is that the most abundant bacteria associated
with the larval hologenome were detected at very
low levels in the seawater; thus, larvae were not
colonized by the most abundant bacterial types;
instead, they specifically select for microbial taxa.
A similar pattern is observed between species of
296   E v o l u t i o n a ry E c o l o g y o f Ma r i n e I n v e rte b r ate La r vae
larvae, where, for example, the microbiome of
­Mithrodia clavigera and other Oreasterideans har-
bor distinct microbial taxa (Galac et al., 2016). This
suggests that the microbial community is species-
specific, implying a co-­evolutionary relationship be-
tween host and community. Examples such as these
are sparse in the larval literature, primarily because
the microbiome of invertebrate larvae has only de-
veloped in recent years. Therefore, this branch of the
field is at a critical stage and is encouraged to avoid
a shotgun approach when deciphering the role of
microbes in larval evolutionary ecology, but instead
should ­default to hypothesis-driven investigations.
Later we attempt to connect this rising branch of lar-
val evolutionary ecology to sections of this volume.
Crosstalk between host and symbionts is essential
for the development of key morphological features
(e.g., the light organ of Euprymna spp.). Pernet (this
volume) and many before describe and link feeding
mechanisms with larval morphology. In the diver-
gence and convergence of larval form and function
of key morphological features, may novel acquisi-
tion or loss of microbial taxa have mediated these
transitions? Microbes also play a large role in game-
togenesis (e.g., Wolbachia for many terrestrial arthro-
pods). Marshall et al. (this volume) as well as Collin
and Moran (this volume) both discuss components
of paternal investment and life history transitions,
respectively. Using an extant clade along this life his-
tory continuum, are there smooth or marked tran-
sitions in the microbial consortiums of these larval
types and, if so, may a particular microbe be linked
to gameteogenesis and investment strategy? Such
marked or smooth transitioning may also be evident
in the context of larvae coping with environmental
extremes. More specifically, how can members of
the microbiome ameliorate developmental or physi-
ological effects of toxicants (Corsi and Marques-
Santos, this volume) and buffer the consequences
of warming and acidification (Byrne et al., this vol-
ume)? These and related questions remain largely
unexplored and can serve as a platform for a holis-
tic understanding of larval biology when integrated
with transcriptomics (host expression), proteomics
(host function), and metatranscriptomics (microbi-
ome function), which may be further complemented
with techniques for functional manipulations (mor-
pholinos, RNAi, and CRISPR-Cas9).
As mentioned, 16s metagenomics is the primary
tool for assaying microbial communities, and the
power and value of this technique goes without
question; however, there are biases. There are often
three steps (DNA isolation, PCR amplification and
sequencing, and taxonomic classification) from tis-
sue sample to publication-ready figures, and each
is accompanied with their respective biases. For
DNA isolation, the kit used directly relates to the
extent of extractable genomic material; PCR am-
plification and sequencing are most influenced by
the polymerase and sequencing platform, respec-
tively; and taxonomic classification is most affected
by the analysis platform and corresponding refer-
ence database (Brooks et  al., 2015). Nevertheless,
to elucidate key questions, both old and new, in
larval evolutionary ecology it remains paramount
to complement current primary research endeavors
with 16s metagenomics to holistically study larval
holobionts.
19.5  -Omics Approaches to Further
Understanding of Larval Physiology:
Growth, Feeding, and Nutrition
In terms of generating phenotype from genotype,
larval physiology links the internal and external
environments—primarily through the ingestion
and digestion of nutrients—that influence larval
growth, as well as developmental gene expression
to alter phenotype (see earlier). Larvae may also ex-
hibit physiological responses, whether metabolic or
otherwise, when stressed by environmental varia-
bles (e.g., temperature and food availability), which
further facilitate changes in growth, metabolism,
and feeding during larval development.
Both transcriptomic and proteomic approaches
have been employed to study larval physiology. At
the level of transcriptome, it is possible to predict
the diet of marine invertebrate larvae during devel-
opment and also to assess the effects of nutrition
on global developmental gene expression patterns.
Using mRNA-seq, Wei et al. (2014) revealed diverse
expression patterns of 16 digestive enzymes during
the development of the white shrimp Litopenaeus
vannamei, showing that expression of digestive en-
zymes is directly linked with diet type. This reflects

the capacity of L. vannamei larvae with distinct nu-
tritional requirements to efficiently exploit diverse
diets (Wei et al., 2014). On the other hand, Carrier
et  al. (2015) profiled gene expression patterns by
mRNA-seq of diet-restricted and well-fed Strongy-
locentrotus droebachiensis echinopluteus to identify
key pathways used for withstanding starvation.
In doing so, they showed that diet restriction re-
presses the expression of genes putatively involved
in growth, development, metabolic activity, and
stress, while increasing the expression of putative
lipid transport, environmental sensing, immune de-
fense, and genes shown to slow aging in other taxa.
A complement to transcriptomics is lipidom-
ics, which has been used since the 1980s for the as-
sessment of nutritional state in aquaculture species
(e.g., Langdon and Waldock, 1981). Large-scale lipid
and fatty acid profiles of marine larvae show com-
plex patterns of metabolite expression depending
on developmental stage and diet. A combination
of thin-layer chromatography-mass spectrometry
(TLC-MS) and gas chromatography-mass spectrom-
etry (GC-MS) in larvae of two sympatric crab species
(Carcinus maenas and Necora puber) detected a total
of 98 molecular species across seven phospholipid
classes (Rey et al., 2015). The phospholipid comple-
ment becomes more complex throughout develop-
ment, but low interspecific differences indicated that
the two different species of crab incorporate similar
energetic contents into their eggs (Rey et al., 2015).
Furthermore, Da Costa et  al. (2015) used GC-MS
to profile fatty acids in larvae of the Pacific oyster
Crassostrea gigas as a means to differentiate dietary
and endogenous sources of fatty acids. They found
that on a diet deficient in docosahexaenoic acid (an
omega-3 fatty acid), larvae compensate by convert-
ing precursor molecules from their endogenous li-
pid resources through the up-regulation of a delta-5
fatty acid desaturase enzyme (Da Costa et al., 2015).
Finally, Bassim et  al. (2015) used both transcrip-
tomics and lipidomics to profile fatty acids in larvae
of the blue mussel Mytilus edulis fed a diet deficient in
essential fatty acids. In combination with phenotypic
measurements, this approach allowed the authors
to show that a diet deficient in essential fatty acids
(e.g., arachidonic and eicosapentaenoic) mediates
elevated mortality, reduced shell growth, and im-
paired postlarval performance (Bassim et al., 2015).
This study remains an exceptional example of how
A n - o m i c s P e r s pect i v e    297
combining -omics with details of environmental
parameters and phenomics can enhance our under-
standing of marine invertebrate larvae, and how
larval experience influences juvenile success (see Pe-
chenik, this volume). Future research efforts directed
toward larval physiology can benefit from combin-
ing measurements of metabolic aspects with large-
scale analysis of the gene, peptide, and metabolite
complements of larvae through various -omics-scale
technologies for a detailed overview of physiology
throughout development and under different envi-
ronmental conditions.
19.6  Connectomics: An -Omics Approach
to Shed Light on Larval Behavior
Larval behavior may be viewed as a phenotype that
is generated through response to both the external
and internal environment. Behavior is an output
of the larval nervous system and changes in gene
expression, as well as larval physiology, can result
in behavioral changes. Conversely, changes in be-
havior can also indirectly affect gene expression.
For example, behavioral changes in locomotion
may expose the larvae to unfamiliar habitats, with
new environmental stressors. There have been few
analyses of the genes, peptides, or metabolites that
regulate larval behaviors, with the exception of lar-
val settlement behavior (Hodin et al., this volume).
However, connectomics, an ‘omics approach which
involves mapping neuronal circuitry based on the
synaptic connections between neurons (Box  19.2),
has the power to elucidate larval behavior.
A connectomics approach has been used to inves-
tigate how the nervous system regulates swimming
behavior of larvae of the polychaete Platynereis
dumerilii in response to light (Randel et  al., 2014).
By reconstructing the connectome of a three-day-old
P. dumerilii larva, researchers explained how a four-
eye visual circuit mediates phototactic behavior.
From a transmission electron microscopy dataset
consisting of 1,690 sections, researchers identified
21 photoreceptor cells, 42 interneurons, and eight
motor neurons that were linked by 1,106 chemical
synapses to form an eye circuit (Randel et al., 2014).
By integrating behavioral analyses with eye abla-
tion experiments and connectome analysis, Randel
et al. (2014) showed that the eyes compare light in-
tensities at the left and right sides and generate the
298   E v o l u t i o n a ry E c o l o g y o f Ma r i n e I n v e rte b r ate La r vae

Box 19.2  The methods behind connectomics.

Connectomics requires the fixation of an organism, fol-
lowed by sectioning into ultrathin sections, which are
then scanned with an electron microscope at very high
resolution to reveal the ultrastructural details of cells and
their organelles. Scanned images of each section are then
stitched together, and sections are aligned using image
registration procedures to reconstruct the organism in a
three-dimensional image stack. Axons and neurites of neu-
rons are then traced through the stack, marking synapses
between different cells and labeling pre- and post-synaptic
cells. This allows the reconstruction of distinct neuronal
circuits that can then be subjected to network analyses to
reveal pathways of information flow through the nervous
system (Figure 19.3).

(A) (B) (G)

larval eyespot PRCar PRCal
adult eye
adult eye photoreceptor cells PRCpr PRCpl

IN1pr IN1pl
INintr INintl
IN1ar IN1al
INdcr INdcl
(C) (D) (E) dorsal
interneurons INvcr INvcl
INtonr INtonl

right left INpror INprol

INsnr INsnl
INvncr INvncl

ventral larval eyespot PRCle-r1r PRCle-r1l PRC

le-new1l

photoreceptor PRCle-r3r
(F) PRCle-r3l
cells
INarcr INarcl
dense core vesicles
mitochondria motorneurons
MNr MNl

ciliated celll
ciliated cellr
effectors musclel
muscler
epithelia cellsr epithelia cellsl

Figure 19.3 Neuronal circuit reconstruction in the three-day-old Platynereis dumerilii larva. (A) Scanning electron micrograph of a three-day-
old nektochaete larva, dorsal view. (B) Light micrograph of the larval head, showing the dorsal adult eyes and ventral larval eyespots. (C) Larva
immunostained with an antibody raised against acetylated tubulin (white) to mark neuronal scaffold and cilia, ventral view. Gray boxes indicate
sectioning of the larva for connectome reconstruction. (D) Schematic example of tracing of an axon through layers of a transmission electron
micrography (TEM) stack. (E) A single layer of TEM scans stitched together to reconstruct the entire circumference of the larval head, anterior
view. Neural plexus is marked by white dashed line. (F) Close-up high-resolution TEM scan in the neural plexus of the larva. White asterisk and
arrow mark a synaptic connection between two neurons. (G) Connectivity graph of the reconstructed visual and larval eye circuit. Line thickness
represents average synapse number (neurons) or total synapse number (effectors). Nodes represent groups of cell types. (Adapted from Randel
et al. 2014 additional images provided by Réza Shahidi, Jürgen Berger.) (see Plate 16)

appropriate muscle contractions that lead to bend-
ing of the larval tail, causing the swimming larva
to turn either toward or away from a light source,
thereby mediating spatial vision. Mapping the con-
nectome of the Ciona intestinalis tadpole larvae has
clarified the neuronal pathways that are responsible
for changes in larval swimming (Ryan et al., 2016).
19.7  A Future Perspective
As seen from the previous examples, -omics are a
suite of techniques that can improve our understand-
ing of the molecular and environmental mechanisms
at work in the translation of larval genotype to phe-
notype during development, over evolutionary time,
and in different ecological niches. However, these
large-scale approaches alone cannot provide a holis-
tic understanding of larval biology.
One drawback of -omics approaches is a tendency
for researchers to generate long lists of genes, pep-
tides, or metabolites with differential expression
between larvae of different species, developmental
stages, or environments, without functional investi-
gation of the newly identified candidates to properly
understand their role in regulating the genotype-
phenotype connection. Until recently, there were
limited molecular tools available that allowed for
the investigation of gene function in marine inver-
tebrate larvae, but rapid developments in this field
now allow for this possibility. For example, RNA
interference (RNAi) or morpholino-­mediated gene
knockdown have proven to be effective methods for
decreasing gene expression in marine invertebrate
larvae, and can potentially be applied in a large-
scale study to investigate the function of several can-
didate genes. While these methods cause a transient
knockdown of gene expression, morpholinos may
be injected in later-stage larvae to knockdown gene
expression later in development (Heyland et  al.,
2014). Moreover, vivo-morpholinos, which have
the capacity to cross cell membranes, may also be
used to knockdown expression by direct addition to
the larval culture medium, negating the need for a
time-consuming and technically challenging injec-
tion procedure (Luo and Su, 2012). Additionally, UV
laser-activated photo-morpholinos can be employed
to control timing and spatial location of gene expres-
sion knockdown (Tallafuss et al., 2012).
A n - o m i c s P e r s pect i v e    299
Another recent development in targeted genome
editing that can be employed for the functional anal-
ysis of marine invertebrate genes is CRISPR/Cas9.
This technique employs an aspect of the bacterial
immune system that consists of the Cas9 enzyme
that binds to a guide RNA sequence to generate
double-stranded breaks in virus DNA (Gasiunas
et  al., 2012; Cong et  al., 2013). The guide RNA se-
quence can be designed to target specific sites in the
genome of diverse organisms. Errors occurring dur-
ing the repair of the DNA break by the gDNA repair
machinery can introduce mutations that lead to the
knockout of target genes (Cong et al., 2013). In Ciona
intestinalis larvae, knockout phenotypes generated
with CRISPR/Cas9 can be efficiently identified in
the F0 generation, negating the need to culture ani-
mals for several generations in the lab to establish a
knockout line (Sasaki et al., 2014; Stolfi et al., 2014).
In addition to genome editing, CRISPR/Cas9 can be
used to silence gene expression through the use of a
Cas9 mutant (dCas9) which binds to DNA but has
no endonuclease activity, thus blocking gene tran-
scription without altering genotype (Perez-Pinera
et al., 2013). CRISPR/Cas9 techniques are currently
being established for a number of marine inverte-
brate species that can be kept in the lab (Ikmi et al.,
2014; Perry and Henry, 2015; Lin and Su, 2016). The
CRISPR/Cas9 system also allows for future possi-
bilities of high-throughput screening for functional
genomics, for example, via the generation of large-
scale guide RNA libraries delivered via a lentiviral
system in combination with rapid screening of sim-
ple phenotypes (Zhou et al., 2014).
It is often not feasible to perform genetic manipu-
lations of larvae in their natural environment; nev-
ertheless, studying larvae outside of the lab remains
an important part of understanding how the envi-
ronment shapes larval gene expression and pheno-
type. While large-scale genomic and transcriptomic
studies based on natural larval populations may
be considered descriptive, this kind of data is still
important for the generation and testing of hypoth-
eses about larval ecology and evolution (Dunn and
Munro, 2016). An understanding of how variable
larval genotypes and phenotypes can be in nature
is essential to our understanding of the genotype-
phenotype map. With transcriptomic methods, it
is now possible to survey gene expression variance
300   E v o l u t i o n a ry E c o l o g y o f Ma r i n e I n v e rte b r ate La r vae
between different individuals and populations of
larvae, sourced from different environments. Gene
expression studies at the scale of single cells and
larvae in marine invertebrates have already been
carried out (Achim et  al., 2015; Levin et  al., 2016);
however, to date, these studies have not focused
on investigating natural variation in expression
between individuals or populations, and tend to
report gene expression as means, without indicat-
ing variance. Currently, transcriptomic methods
provide the best possibility for measuring variation
at the level of single cells or larvae; current peptid-
omic and metabolomic methods have not yet been
used in individual marine invertebrate larvae or
cells. However, there is the possibility that peptid-
omics and metabolomics may be applicable in the
future at least on a single-larva scale, as direct pep-
tide profiling of single neurons from adult sea hares
Aplysia californica and Pleurobranchaea californica by
matrix-assisted laser desorption/ionization time-
of-flight mass spectrometry is already being carried
out (Garden et al., 1996; Rubakhin et al., 2011).
To develop a systems-level understanding of
marine invertebrate larvae, a promising approach
for the future is the combination of different -om-
ics approaches with each other, classical and novel
molecular biology techniques, functional analyses
of developmental genes, and large-scale profiling of
phenotype and environmental parameters. For ex-
ample, single-cell transcriptomic analyses in combi-
nation with cell lineage and spatial gene expression
studies could be used to map developmental and
cell differentiation signaling pathways to specific cell
lineages. Genome editing can be combined with me-
tabolomics and morphological measurements to ex-
amine the influence of key developmental genes on
both larval physiology and morphology. Including
transcriptomics in this study would allow the identi-
fication of downstream target genes of the knockout
gene. Including monitoring of multiple environmen-
tal parameters in studies of the microbiome in lar-
vae sourced from different geographic locations will
show the contribution of environment to microbiome
composition. Combining connectomics with spatial
mapping of gene expression, single-larva transcrip-
tomics, and video behavioral analyses will reveal
how variation in the molecular signaling of the nerv-
ous system generates different larval behaviors.
Several marine invertebrates representing multi-
ple phyla have now been developed as lab model
organisms. With published genomes and larval
transcriptome resources, and in most cases, estab-
lished methodologies for functional genetic analy-
ses, these organisms provide a platform for the
commencement of the aforementioned integrative
-omics studies. Examples of established and up-
coming marine invertebrate model species include
the solitary ascidian Ciona intestinalis, the colonial
ascidian Phallusia mammilata, the sea urchin Stron-
gylocentrotus purpuratus, the starfish Patiria miniata,
the snails Ilyanassa obsoleta and Crepidula fornicata,
the crustacean Parhyale hawaiensis, the polychaete
Platynereis dumerilii, the anthozoan Nematostella
vectensis, the hydrozoan Clytia hemisphaerica, and
the sponge Amphimedon queenslandica. Functional
studies in these model organisms can be used to
identify key developmental genes in larvae. The ex-
pression and presence/absence of these genes can
then be studied in natural larval populations in or-
der to understand the contribution of environment
to phenotype. The rise of -omics discussed here
facilitates studies of natural populations; however,
in cases where it is not possible to source appropri-
ate material for transcriptomics, peptidomics, or
metabolomics, traditional molecular biology tech-
niques for studying gene expression, such as qRT-
PCR and in situ hybridization, could be employed
in a high-throughput manner to study natural
variation in gene expression in larval populations.
Although -omics approaches alone will not solve
all remaining questions in larval biology, they cer-
tainly represent a step forward in our study of these
intriguing and complex organisms.
19.8 Summary
1. Molecular methods have been an important ad-
dition to larval biology, as they enable us to be-
gin to understand the mechanisms underlying
the larval genotype-phenotype map. As an ex-
tension of this, -omics are large-scale, often high-
throughput approaches that allow simultaneous
profiling of all molecules within a cell or organ-
ism, phenotypes within a population, or synaptic
connections within a nervous system.

2. In the study of larval development, transcrip-
tomics can be analyzed in conjunction with gene
regulatory networks, or the experimental ma-
nipulation of external environmental cues for
developmental transitions, to identify molecular
regulators of development within a species and
how these evolve to generate different morpho-
logical phenotypes.
3. Large-scale analyses of larval genomics, tran-
scriptome, peptidome, and metabolome provide
an efficient means of exploring the impact of ex-
ternal environmental factors on larval biochem-
istry. Combined with large-scale phenotypic
measurements and monitoring of environmen-
tal parameters, this approach will allow us to
explore how environment shapes larval pheno-
type, including behavior and physiology.
4. Marine invertebrate larvae harbor species-­
specific microbiota, including bacteria, archaea,
and viruses, that can influence their physiology
and morphology and may provide a means for
adapting to environmental change. Large-scale
sequencing of genomes and transcriptomes will
allow rapid identification of larval microbiota
and their influence on larval gene expression and
phenotype.
5. Connectomics, the study of neuronal connectiv-
ity, can provide us with a functional understand-
ing of how and why larvae respond to different
environmental cues with different behaviors,
and insight into the evolution of the larval nerv-
ous system.
6. Important next steps for -omics approaches in
studying marine invertebrate larvae include the
development of tools for large-scale functional
genetic analyses, measurement of natural varia-
tion in molecular expression between individuals
and populations, and a systems-level approach
that combines multiple -omics approaches to
further explore the influence of environment and
genotype on larval phenotype.
Acknowledgments
We would like to thank Sandie Degnan for her
thoughtful comments on earlier versions of the
manuscript, Csaba Verasztó for his constructive
comments on later versions, as well as Réza Shahidi
A n - o m i c s P e r s pect i v e    301
(TEM) and Jürgen Berger (SEM) for providing im-
ages for use in Figure 19.3. Lastly, we thank Adam
Reitzel and Andreas Heyland for their collaboration
in the undertaking of this project, for the invitation
to write this chapter, and for helpful comments on
structuring it.
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 C h a pt er 20
Section 4 Summary—Larval Ecology
at the Extremes
Much of our understanding of marine invertebrate
larval ecology, until much of the last two decades,
was derived from easily accessible intertidal spe-
cies. The study of larvae in species from other en-
vironments, particularly the deep sea, has grown
tremendously in the past decades. In Chapter  16,
Craig Young and colleagues discuss the selective
pressures on deep-sea larvae and how those, in
turn, may have directed the observed frequencies
of feeding modes. Further, they hypothesize that
the dispersal strategies by invertebrates in these
specialized habitats is bimodal, with very short dis-
persal or very long larval lives being favored.
While technological advances have allowed the
field a glimpse of deep-sea larval ecology, intertidal
species have been faced with a multitude of anthro-
pogenic stressors. In Chapter 17, Maria ­Byrne and col-
leagues focus on two: ocean acidification and ocean
warming, along with their interaction. In response
to ocean warming, benthic marine invertebrates and
their larvae are following poleward isotherms, while
coping with an acidifying ocean is far more complex.
On the other hand, in Chapter  18, Ilaria Corsi and
Luis Marques-Santos assess how toxicants affect
embryonic and larval functionality with a focus on
the ABC transporters of echinoids. They provide a
history of ABC transporters, discuss the utility of
Carrier, T. J., Reitzel, A. M., and Heyland, A., Section 4 Summary—Larval ecology at the extremes. In. Evolutionary
Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland:
Oxford University Press (2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0020
echinoids in ecotoxicology research, and emphasize
the need to find complementary models to fully cap-
ture the consequences of aquatic toxicants.
Similar to the technological advances in deep-
sea biology, a trend has occurred with -omics. In
Chapter  19, Williams and Carrier, discuss how
­
next-generation sequencing and other -omics ap-
proaches have enabled the field to globally profile
responses by larvae (and their associated microbi-
ome) to environmental variability and anthropo-
genic stressors. Moreover, they emphasize how and
why -omics may complement many topics in this
volume, from evolutionary history to transitions in
feeding mode to feeding physiology.
Since the publication of Ecology of Marine Inver-
tebrate Larvae in 1995, the study of larval evolution-
ary ecology in these extremes has seen tremendous
growth, so much so that none of the chapters in this
section were featured in McEdward (1995). Never-
theless, these disciplines remain in their infancy: the
deep sea and their fauna remain little explored, and
the rippling effect of climate change–related and
toxicological stressors on larvae is only beginning.
-Omics approaches, however, are developing rap-
idly and are now being applied to the larval field.
We believe this area will take center stage of larval
research going forward.
 Ch a pt er 21
Marine Invertebrate Larvae: Model
Life Histories for Development,
Ecology, and Evolution
Alan C. Love and Richard R. Strathmann
21.1 Introduction
The questions raised for the study of marine inver-
tebrate larvae have implications for the evolution of
development, the life histories of animals, and life
in the sea more generally. These questions began to
coalesce in the nineteenth century around two main
factors. The first was the discovery of marine larvae.
Through careful observation, investigators detected
and confirmed that the development of animals ex-
hibited stages surprisingly different from the previ-
ously known adults and adult-like juveniles. Famous
examples include the demonstration that barnacles
were arthropods because of a shared larval form
(Thompson, 1830), the characterization of bilateral
echinoderm larvae by Johannes Müller (summarized
in Huxley, 1851), and the unexpected chordate af-
finities of tunicates (Kowalevsky, 1867). Subsequent
studies also demonstrated that adults with similar
morphological features could have radically differ-
ent larval forms and life histories (e.g., Giard, 1905).
The second factor crucial to a coalescing of ques-
tions around marine invertebrate larvae was the
emergence of various forms of evolutionary think-
ing. Although this factor has been well documented
(e.g., Bowler, 2003), it is important to highlight how
it articulates with the first factor of discovering
larval forms. For example, the dramatic metamor-
phosis in echinoderms from a pelagic larva with
bilateral symmetry to a benthic adult with radial
symmetry provided a visible demonstration of a
major morphological transformation. Larval forms
Love, A. C. and Strathmann, R. R., Marine invertebrate larvae: model life histories for development, ecology, and evolution.
In. Evolutionary Ecology of Marine Invertebrate Larvae. Edited by Tyler J. Carrier, Adam M. Reitzel, and Andreas Heyland:
Oxford University Press (2018). © Oxford University Press. DOI: 10.1093/oso/9780198786962.003.0021
served as indicators of developmental variability
that were relevant to conceptualizing substantial
evolutionary change. “Embryologists are famil-
iar with many other illustrations of this kind of
development.  .  .  . It shows by easy steps how the
dramatic metamorphosis of Pilidium into a Nemer-
tine may have arisen” (Garstang, 1928, p. 79–80). A
number of these illustrations are captured memora-
bly in Walter Garstang’s playful poetry, such as the
plausibility of an evolutionary transition between
the cyphonautes larva of bryozoans and actinotroch
larva of phoronids (Garstang, 1951, p. 51):
The Cyphonautes larva wears a cocked hat for a shell,
With involuted sucker and a vestibule as well:
His mask, a very thin disguise, may be a playful joker’s,
But whose the visage underneath if not the Actinotrocha’s?
These morphological transformations not only
contributed to phylogenetic speculation (Bowler,
1996), but also suggested particular mechanisms
of evolutionary change, such as the origin of nov-
elties from larval forms via heterochrony, with the
adult morphology being subsequently abandoned:
“Qualitative evolutionary novelties can and do ap-
pear at all stages of ontogeny, and not solely in the
adult. . . . A big change in evolution is more likely to
occur if ancestral youthful characters become those
of adult descendants” (de Beer, 1941). Evidence in
support of this mechanistic scenario for marine lar-
vae has not surfaced, but tracking morphological
changes across a life history has provided a fertile
source for testable evolutionary hypotheses.

Additionally, interest in the evolutionary ecology
of marine larvae manifested naturally in studies of
how larval behavior and morphology were adapted
to the environment, and what effects various larval
stages had on the ecology of populations, such as
through dispersal and varying survival (Hjort, 1914;
Garstang, 1928; Thorson, 1950; reviewed in Young,
1990). Although some evolutionists overemphasized
particular ecological dimensions (“the primary bio-
logical significance of free-swimming larvae is that
of dispersal”; Mayr, 1963, p. 607), questions about re-
cruitment, speciation, adaptation, and biogeography
clearly differed from those about phylogeny, develop-
ment, and the origins of evolutionary novelty. Over-
all, a wide range of diverse but interrelated questions
arose from a combination of evolutionary thinking
and discoveries that the life histories of marine inver-
tebrates displayed sexually immature, mobile, and
(often) feeding larval stages that differed morpholog-
ically from corresponding adults and were adapted
to the special conditions of marine environments.
In the present chapter, our aim is to provide a
broad conceptual framework for studies of the de-
velopmental and evolutionary ecology of marine
invertebrate larvae and illustrate how the contribu-
tions to this volume demonstrate both past achieve-
ments and the future fecundity of this research
program. Our conceptual framework is anchored in
the idea of model life histories, which is a category
of investigation similar to but distinct from model
organisms and model clades. Although it may not
appear that marine invertebrate larvae constitute a
coherent domain of research, we argue precisely the
reverse. Not only is it a coherent domain of research,
but it also affords novel investigative pathways una-
vailable in more standard investigative frameworks,
especially with regard to emerging issues surround-
ing the impact of climate change on the world’s
oceans. As a consequence, there are good reasons to
invest and participate in research on the model life
histories of marine invertebrate larvae.
21.2  Model Life Histories
We live in the age of model organisms. Much of
modern biological research is focused on a small
set of organisms—Caenorhabditis elegans, Drosophila
melanogaster, Danio rerio—that permit researchers
M o d e l L i f e H i s to r i e s    307
to experimentally investigate biological processes
in great depth, and facilitate a precise dissection
of causal relationships (Davis, 2004; Ankeny and
Leonelli, 2011). A basic presumption about model
organisms is that they are representative of other
organisms with respect to the properties under
investigation (e.g., “We study flies and frogs as ex-
amples for the development of animals in general”;
Nüsslein-Volhard, 2006, p.  138). This presumption
has been anchored by the finding that many core
genetic and cellular mechanisms are evolutionarily
conserved (Kirschner and Gerhart, 1998; 2005). This
means a model organism (“source”) can represent
or exemplify other unstudied species (“targets”).
Critics have questioned whether model organ-
isms are good exemplars of other species because
of inherent biases involved in their selection, such
as rapid development and short generation time
(Bolker, 1995). Preparing model organisms for ex-
perimental analysis also involves intentionally mis-
representing mechanisms or phenomena, such as
through standardized temporal stages that ignore
the degree and kinds of variation present (Love,
2010). Model organisms are sometimes utilized
as surrogates to represent one particular genetic
mechanism or phenotypic trait; they provide in-
direct experimental access to otherwise inaccessi-
ble phenomena, such as mouse models of human
psychological disorders (Bolker, 2009). Although
evolutionary conservation undergirds using model
organisms as exemplars, a more recent common
ancestor can undergird viewing one as a surrogate,
even if it is not a good representative generally.
In addition to criteria of representation, model or-
ganism research is also justified in terms of capaci-
ties for experimental manipulation: “Model species
have each been selected because they have some
particular experimental advantages” (Slack, 2006,
p. 61). These advantages can dominate the rationale
for model organism research and trump objections
about how well they represent other species. As-
pects of the capacity for manipulation include how
easy it is to initiate a regime of experiments (e.g.,
availability, cost, and infrastructural prerequisites)
or employ different techniques (e.g., the kinds of
manipulation possible and how quickly they can
be done). Criteria of representation and manipula-
bility are both modulated by the types of research
308   E v o l u t i o n ary Ec o l o g y o f M ar i n e I n v e rt e b rat e L ar va e
questions in view, including but not limited to
questions about adaptation, cellular mechanisms,
diversity, inheritance and variation, development,
the origins of novelty, classification and phylogeny,
biogeography, coevolution, and speciation. Model
organisms might be well suited for representing
and manipulating phenomena or mechanisms perti-
nent to some domains of biological problems rather
than others. Marine invertebrate larvae prompt par-
ticular kinds of inquiry about the evolution of mor-
phology, and it is therefore unsurprising that their
absence as models from key research trajectories of
mid-twentieth century biology was accompanied by
a neglect of these scientific questions (Love, 2009).
Model organisms have been less attractive in
evolutionary research because of questions about
representativeness that stem from both the taxa
chosen (e.g., short generation time is a derived trait
in many lineages) and the reduction of naturally
occurring variation due to laboratory domestica-
tion (e.g., purified genetic lines and standardized
environmental regimes) (Bolker, 1995; Frankino
and Raff, 2004). Evolutionary investigations of the
origin of traits, like the vertebrate head, require the
simultaneous study of multiple taxa that represent
variation relevant to a particular phylogenetic junc-
ture (Hanken, 1993). Instead of one species (e.g.,
C. elegans) being scrutinized intensely in laboratory
experiments (though this is sometimes informa-
tive for evolutionary questions; see, e.g., Rose et al.,
2005), it is crucial to make comparisons across many
species using an empirically supported phylogeny
(Harvey and Pagel, 1991). We can speak of a model
taxon or clade (Griesemer, 2013; 2015) in circum-
stances where phylogenetic relationships are well
established, detailed morphological and behavioral
knowledge about the different species exists, and
extant representatives can be studied in more con-
trolled settings to achieve different degrees of ex-
perimental manipulation.
Just as model organisms are intended to be rep-
resentative of other organisms, so also model clades
are intended to be representative of evolutionary
processes occurring in other clades. David Wake
and colleagues assembled an array of morphologi-
cal, phylogenetic, and biomechanical approaches to
answer suites of interrelated questions pertaining
to lungless salamanders (Griesemer, 2013; 2015).
Over time, an evolutionary morphological research
platform emerged that combined a variety of mate-
rial and conceptual resources organized into stand-
ardized routines of investigation (Wake and Larson,
1987). This platform yielded key findings about the
evolution of anatomical features (e.g., the tongue
or body size) and associated functional systems
(e.g., feeding, locomotion), especially in terms of
the mechanisms contributing to these patterns (e.g.,
miniaturization due to changes in developmental
timing; large cell size resulting from constraints
of genome size) (Wake, 2009). These patterns ap-
peared across the model clade and helped in the
formulation of evolutionary generalizations, such
as the significance of homoplasy for processes of
morphological evolution (Wake, 2015).
Another example is the genus Anolis—­represented
by species found in diverse habitats within the Car-
ibbean—which has been studied intensively by the
Losos group (Losos, 2011). A manageable clade made
up of a large number of species with good phyloge-
netic resolution, while also being regionally delim-
ited and accessible, has generated a fruitful platform
for the investigation of ecological and evolutionary
questions with a constellation of different methods.
Across various islands, similar morphology and be-
havior are observed for lizards in different micro-
environments. Applying the comparative method
facilitates discriminating between the independent,
convergent evolution of these “ecomorphs” and
a single origin and subsequent dispersal (Mahler
et  al., 2013). Laboratory investigations and field
studies of the functional capabilities of these lizards
help to decipher how distinctive traits—correlated
with specific physical or social habitats—are adap-
tive. Generalizations about the dynamics of adap-
tive radiations have been gleaned from comparisons
and contrasts across members of the model clade
(Stroud and Losos, 2016).
Model clades can be scored on the three dimen-
sions highlighted for model organisms: repre-
sentation, manipulation, and research questions.
Unlike the focus on a small set of exemplar spe-
cies in model organisms, model clades represent
multiple species comparatively across a span of
evolutionary history. This is essential for discern-
ing patterns of homoplasy or adaptive radiation,
which are not properties of particular species.

Although model organism features can be com-
pared, any conclusions drawn must be calibrated
with additional phylogenetic information. Studies
of model clades result in generalizations that typi-
cally do not have as wide a scope, but are robust
across nearby spaces in the tree of life (e.g., tetra-
pods). Manipulations relevant to model clades are
less focused on genetics, cell biology, or ontogeny,
and instead concentrate on morphology and be-
havior using different methodological approaches
(e.g., biomechanics and field experiments). Al-
though diverse manipulations are possible, many
are time- and cost-intensive. The significance of
research questions applies here because the types
of relevant manipulations align with the ecologi-
cal and evolutionary problems in view (e.g., adap-
tation, biogeography, and speciation). Studies of
model clades also take advantage of material avail-
ability in terms of natural abundance or compre-
hensive museum specimen collections (e.g., cost
and infrastructure prerequisites). Just as many or-
ganisms would make poor model organisms (e.g.,
due to long gestation times), so too many clades
would not be good models (e.g., due to too few or
too many species without a consensus phylogeny
or inadequate knowledge of relevant traits).
Marine invertebrate larvae clearly do not fit into
the category of model organisms. Although a few
model organisms, such as the purple sea urchin
(Strongylocentrotus purpuratus), exhibit indirect de-
velopment with a pelagic larval form (echinoplutei),
these are typically studied with little or no atten-
tion to larval features because they manifest later
in time. (Practical reasons, such as ease of embryo
maintenance or manipulation, undergirded their
selection as models.) Most model organisms are di-
rect developing without distinctive larval forms, a
characteristic of brief life histories, which served as
a basis for selecting many (though not all) model or-
ganisms. Additionally, many of the more advanced
experimental tools that permit a dissection of pre-
cise causal relationships either do not work or have
not been worked out for marine invertebrate larvae.
Despite decreasing costs for sequencing and the
standardization of many experimental protocols
(Crotty and Gann, 2009), the full breadth and depth
of manipulative tools from genetics and molecular
biology cannot be applied.
M o d e l L i f e H i s to r i e s    309
Marine invertebrate larvae do not fit into the
category of a model clade either. Studies of ma-
rine clades have revealed much about evolution of
modes of larval development, including changes in
developmental processes that accompany the loss
of larval feeding (the sea urchin genus Heliocidaris;
Wray and Raff, 1991), transitions and homoplasy
(asterinid sea stars; Hart et  al., 2006), and conse-
quences for speciation and extinction (sacoglossan
molluscs; Krug et  al., 2015). However, although
studies of larval forms are frequently executed
within explicit phylogenetic contexts, the category
itself (marine invertebrates) is not a monophyletic
group. Many of the comparisons central to studies
of marine larvae involve cross-clade evaluations
of functional requirements for specific ecological
settings that are not confined to a clade but rather
exhibit a broad taxonomic distribution. Resulting
generalizations about larval forms transgress phy-
logenetic boundaries and facilitate a larger number
of insightful comparisons and contrasts both within
and across clades. Some generalizations revolve
around particular instantiations of larvae that ex-
emplify a broader type (e.g., trochophore). In this
sense, marine invertebrate larvae play a represen-
tational role as exemplars. Some comparisons have
wider scope across the tree of life. Similar patterns in
feeding vs. nonfeeding or planktonic vs. benthic de-
velopment are discernible in disparate taxa. Broad
generalizations from the marine environment per-
mit contrasts between dispersal as a byproduct of
other adaptations in many marine life histories and
dispersal that is itself more commonly adaptive in
terrestrial life histories (Strathmann, 1990; Vermeij
and Grosberg, 2010; Burgess et al., 2016).
The study of marine invertebrate larvae can be
understood in terms of a category different from
both model organisms and model clades: model
life histories. In addition to the differences already
described, a crucial distinguishing feature of the
category is marked by the name: life history. Con-
centrating on a life history rather than an organ-
ism means that the temporal sequence within
each generation is foregrounded. Concentrating
on a life history rather than a clade means that the
comparisons and contrasts can be made across this
temporal sequence within an individual species,
in addition to those made within or across clades.
310   E v o l u t i o n ary Ec o l o g y o f M ar i n e I n v e rt e b rat e L ar va e
And because the temporal sequence of a life his-
tory ranges over different biological problems,
studies of model life histories do not exhibit the
typical divide of questions clustered around ge-
netics, cell biology, development, and physiology,
as compared to those clustered around ecology,
evolution, adaptation, and phylogeny. As a conse-
quence, model life histories demand an integration
of disciplinary approaches and diverse methodol-
ogies, something that is explicitly highlighted by
practitioners:
The conditions of breeding and larval development con-
stitute a very essential point for all studies of the repro-
duction and fundamental ecology of marine bottom
invertebrates. . . . The fact that a mode of reproduction
and development, well fit for an arctic area, is unfit in a
temperate or tropical area of the sea is probably one of
the main reasons for the restricted distribution of spe-
cies. (Thorson, 1950, p. 3, 37)
As developmental stages that interact behaviorally with
their environments, [larvae] play important roles in de-
velopment, ecology, and evolution and are therefore of
interest to a wide community of biologists. (Emlet et al.,
2009, p. 201)
Investigations of model life histories require mul-
tidisciplinary research configurations: “The study
of marine invertebrate larvae is necessarily inter-
disciplinary” (McEdward, 1995, Preface). This re-
quirement has even survived the explicit severing
of disciplinary connections, such as the previously
central role of comparative embryology in phyloge-
netic reconstruction (Hand, 1959; Anderson, 1973).
Many of these research configurations are mani-
fested in marine stations where the availability,
cost, and infrastructural prerequisites for manipula-
tion are in place to support the relevant array of ap-
proaches simultaneously. The original motivations
for marine stations bear this imprint.
Marine stations are necessary for biological science, since
nowhere but in the sea can there be found a host of types
whose study is indispensable if one desires to form a clear
and concise idea of the ensemble of the organic world. . . .
The sea is really the source of organic life in its ensem-
ble; researches on the relationships of different animals,
on their origin, on their individual development, from the
first visible germ to the completion of their life cycle, are
continually and necessarily leading us back to marine or-
ganisms. (Carl Vogt, in Whitman, 1893, p. 11–12)
Model organisms, clades, and life histories offer
different investigative strategies in biological re-
search that can be evaluated in terms of criteria for
representation, manipulation, and research ques-
tions. Importantly, not all of biological research falls
into one of these three categories. However, the con-
cept of a model life history provides a wider concep-
tual framework for understanding the coherence of
research into marine invertebrate larvae. It accounts
for the unique juxtaposition of exemplar and com-
parative representational dimensions, the diversity
of manipulative approaches utilized (as well as lim-
itations on the application of particular techniques),
and the combination of diverse biological problems
not seen in research programs on model organisms
or model clades. This arises, in part, because studies
of model life histories focus on multiple aspects of a
temporal sequence rather than particular segments
of that sequence (e.g., early embryonic patterning
or adult feeding) and different dimensions within
those aspects of the sequence (e.g., embryonic de-
velopment, larval feeding, metamorphosis, disper-
sal, and reproduction). The category of a model life
history also provides for the recognition of novel
kinds of representation, such as marine invertebrate
larvae acting as surrogate models of ocean environ-
ments under changing climatic conditions.
21.3  Evolution: Diversity, Origins,
and Adaptation
Studies of the evolution and ecology of marine lar-
vae are interdisciplinary; they use a wide range of
tools from systematics, life history theory, biome-
chanics, oceanography, population genetics, and
paleontology to address different questions, such
as how and why larval stages originated, how and
why they persist (or are lost), and what their eco-
logical and genetic consequences are within the tra-
jectory of a life history. Answering these questions
requires expanding observations to all stages in life
histories, as well as analyzing marine environments
on all spatial scales, and extending the historical
scale scrutinized for animal evolution back through
the Phanerozoic and into the Proterozoic. Although
questions about developmental genetic mecha-
nisms and the experimental tools of contemporary
molecular biology have been (until recently) less

prominent in communities concentrating on marine
invertebrate larvae, new methods for studying the
molecular genetics of development have facilitated
broader comparisons among animals and promoted
interest in the role of developmental processes in
evolution (Hinman et  al., 2003). Additionally, old
questions about whether larval forms are ancestral
or derived in particular lineages have been revis-
ited (Sly et  al., 2003), and speculative hypotheses
about their origination refuted (Hart and Grosberg,
2009). A better understanding of the mechanisms
that produce animal morphology illuminates how
developmental processes have constrained or fa-
cilitated evolutionary change. A renewed interest in
the evolution of development generally, including
substantial advances in phylogenetic reconstruc-
tion, assisted in reuniting studies of developmental
mechanisms and the evolutionary ecology of lar-
vae. A number of the preceding chapters illustrate
the ways in which constellations of new methods
have enabled answers to old questions (Love and
Raff, 2003), as well as created new questions.
Nielsen (this volume, Chapter  1) and Marlow
(this volume, Chapter  2) review hypotheses and
evidence for the origins of ciliated bilaterian larvae
(all subsequent chapter references will be to the pre-
sent volume). Molecular genetic evidence has con-
tinued to improve phylogenetic inferences about
relationships among classes and phyla (Hart 2005;
see Collin and Moran, Chapter  4). The individual
expression of and interaction among genes during
morphogenesis has revealed deep similarities un-
derlying the development of disparate morpholo-
gies and pointed to genetic changes responsible for
the divergence of those forms (Hinman et al., 2007).
For bilaterians generally, a more extended pattern
of Hox gene expression in adults compared with
larvae indicates that ancestral adults had more to
their bodies than present larvae display (Arenas-
Mena et al., 2000; Kulakova et al., 2007; Hiebert and
Maslakova, 2015). However, the development and
morphology of some features implies that several
larval structures antedate the divergence of par-
ticular phyla. A plausible hypothesis, supported
by different studies from developmental genetics,
is that larvae and adults diverged as distinct stages
from more directly developing ancestral bilateri-
ans (Marlow, Chapter 2). Feeding mechanisms and
M o d e l L i f e H i s to r i e s    311
associated forms of larvae differ greatly among ma-
jor clades of animals (Nielsen, Chapter  1; Pernet,
Chapter 7), which implies that there were multiple
origins of larval feeding and multiple divergence
events for larval and adult forms. There is no evi-
dence for a single, primary form of feeding larva in
metazoans or, more specifically, for bilaterians.
In bilaterian development, Hox genes tend to be
expressed sequentially through time from anterior
regions in the embryo to more posterior regions
(Mallo et al., 2010). This patterning of body devel-
opment is commonly accompanied by a differentia-
tion of structures that also proceeds from anterior to
posterior. This sequence of body patterning and dif-
ferentiation in bilaterians prompts two related ques-
tions: (1) Did this sequence influence the evolution
of larval forms and their functional capabilities? (2)
Did this sequence influence the evolutionary origins
of larval forms (e.g., by what differentiated first and
most anteriorly in the ancestor)? An answer to these
questions can be formulated through a combination
of evidence from mechanistic studies of develop-
mental processes (Marlow, Chapte­r 2), biomechani-
cal studies of larval performance in swimming
and feeding (Pernet, Chapter 7), and adaptationist
studies of trade-offs in the evolution of life histories
(Marshall et al., Chapter 3; Pernet, Chapter 7; Young
et al., Chapter 16).
With development proceeding from front to back,
selection for mobility and feeding should favor
early functioning in anterior structures. On this hy-
pothesis, the mobility and feeding functions based
on anterior structures permit development from a
smaller egg and thus production of more eggs, as-
suming there is growth before patterning and dif-
ferentiation (and, therefore, functionality) in more
posterior structures. Earlier motility reduces mor-
tality by facilitating, for example, escape from pred-
ators, and is a key feature of life history adaptations
(Pennington and Chia, 1984). The limited structures
at the anterior end of the developing animal ne-
cessitate a form and function different from what
is found in the adult. This helps to account for the
recurring origin of head larvae that have been docu-
mented in the three major branches of bilaterians:
deuterostomes, lophotrochozoans, and ecdysozo-
ans (Gonzalez et al., 2016; see Marlow, Chapter 2).
Does this hypothesis match observations of egg
312   E v o l u t i o n ary Ec o l o g y o f M ar i n e I n v e rt e b rat e L ar va e
sizes and developmental rates for structures asso-
ciated with feeding and mobility? There is a large
amount of overlap in the sizes of eggs for animals
with head larvae and those that initiate mobility
and feeding at later stages, and (unfortunately) few
extensive comparisons. However, head larvae are
associated with the smallest eggs within annelids
(Schroeder and Hermans, 1975) and with the small-
est eggs and briefest times to first feeding in crusta-
ceans (Strathmann, 2017).
Although life histories involving transitions be-
tween planktonic and benthic habitats are docu-
mented throughout this volume, the origins of
head larvae do not necessarily depend on larvae
and adults occupying different habitats. The trilo-
bite protaspis—an early stage prior to the develop-
ment of articulated segments—appears to have been
a head larva, and many protaspids are inferred to
have been benthic (Chatterton and Speyer, 1997).
Today, pycnogonids are benthic as both head larva
(protonymphon) and adult. Most benthic copepods
have benthic head larvae (nauplii). Many plank-
tonic and nektonic crustaceans, such as copepods,
euphausiids, and sergestid shrimp, have planktonic
head larvae. These observations suggest that the
evolution of distinctive larval stages was not neces-
sarily associated with a benthic-planktonic life his-
tory. Overall, studies of development, function, and
life history indicate that the differentiation of ante-
rior structures before posterior structures did have
an influence on the origins of bilaterian larval forms.
Importantly, the evolution of larvae in other ani-
mal clades differs. For example, the modifications
of adult ctenophores appear to have resulted in the
earlier stage being distinctively different and hence
a larva. Adults in the paraphyletic Cydippida pre-
sumably resemble the ancestral adult of extant cten-
ophores (Podar et  al., 2001; Haddock, 2007). If so,
development from a cydippid larva into strikingly
different adults in other ctenophores represents the
evolution of diverse, novel adult forms, whereas the
larvae retain features of a more direct development.
Several kinds of larvae have a broad taxonomic
distribution and similar features that suggest a sin-
gle, early evolutionary origin. Examples include
trochophores of spiralians, dipleurulas of hemi-
chordates and echinoderms, and nauplii of crus-
taceans (Nielsen, Chapter  1; Pernet, Chapter  7).
These forms are either head larvae or appear to be
modified head larvae (Marlow, Chapter  2). As far
as we know, head larvae originated early in the
diversification of animals and not later. The taxo-
nomic distribution of head larvae and their inferred
homologies (indicating early origins) prompt the
question: Why did the kinds of larvae observed to-
day so often originate early in the evolution of the
clade in which they occur? Development and ecol-
ogy suggest nonexclusive hypotheses.
On the one hand, a developmental hypothesis is
that subsequent evolutionary changes in how ante-
rior adult structures develop may have precluded
a later evolution of head larvae (Collin and Moran,
Chapter 4; Pernet, Chapter 7). As an extreme exam-
ple, echinoderm larvae may have originated as head
larvae, as indicated by inferred homologies with the
head larvae of hemichordates, but present-day echi-
noderms lack heads in post-metamorphic stages.
When an echinoderm loses a feeding larva, there re-
mains no basis of phenotypic variation for selection
to operate on and produce a new kind of head larva.
However, their relatives the enteropneust hemi-
chordates have kept their heads. Given that head
larvae did originate at one or more points early in
the history of metazoans, it is less clear why a head
larva could not originate again. On the other hand,
an ecological hypothesis is based on an evolution-
ary escalation in the interactions among organisms.
In this scenario, the first steps in the evolution of a
feeding larva were advantageous early in animal
evolution but then selected against as the biologi-
cal environment became more demanding, perhaps
from increased predation. A combination of factors
from development, functional morphology, and
ecology are likely needed for an adequate explana-
tion. Notably, early origins of head larvae are a dis-
tinctively marine phenomenon. A few feeding larval
forms evolved much later in marine, freshwater, and
terrestrial environments, but these larvae use more
posterior structures for mobility and feeding, which
requires the prior development of most of the body
axis (e.g., the zoea stage of decapod crustaceans and
larvae of stomatopod crustaceans, as well as larvae
of teleost fish, amphibians, and insects).
Across marine invertebrates, species differ in
whether the larva feeds and whether there is a free
larval stage or none at all. How is larval feeding

lost? How do larvae get deleted from a life his-
tory? Under what conditions, if any, can losses be
reversed? Attempts to answer these questions have
required diverse kinds of methods and observations
(Collin and Moran, Chapter  4). Inference of losses
and reversal depend on a phylogenetic hypothesis
detailing the taxonomic distribution of modes of
development in existing species, but computational
methods for inferring ancestral traits are based on
models of evolutionary change and reversal (Joy
et  al., 2016). Constraints on evolutionary transi-
tions differ among taxa. The extent and frequency
of modifications in larval development depend
on properties of processes of ontogeny (e.g., the
sequence of regional differentiation), feeding and
swimming (e.g., controlled vs. passive transport),
maternal protection of young (e.g., encapsulation
and brooding), and ecological challenges (e.g.,
mortality and dispersal). All of these properties
differ among clades of marine invertebrates. Both
functional morphology and developmental genet-
ics help in identifying and characterizing what fa-
cilitates and inhibits evolutionary transitions. Some
indications of irreversible change are simple and
direct, such as mutations in unexpressed genes
(Huber et  al., 2000) or the loss of a gut and allied
structures for capturing food (Love et  al., 2008).
Barriers to loss include an inability to synthesize es-
sential compounds acquired from food or the need
for feeding structures to consume nurse eggs. The
potential influences on evolutionary transitions
are numerous, disparate, and different among taxa
(Collin and Moran, Chapter 4).
The ancient origins of larvae alongside a bias
in evolutionary transitions away from feeding or
mobility raise the question of how a larval stage
persists. For some species, dietary requirements or
nurse egg feeding may limit loss of larval feeding
(Collin and Moran, Chapter  4). Species selection
may be a more general influence on persistence
and loss of feeding larvae. Insofar as modes of lar-
val development affect dispersal, gene flow, and
geographic range (Marko and Hart, Chapter  12;
Pineda and Reyns, Chapter 11; Young et al., Chap-
ter 16), modes of development are expected to affect
rates of speciation and extinction, which combine
to affect rates of species diversification (Collin
and Mora­n, Chapter  4). The proportion of species
M o d e l L i f e H i s to r i e s    313
without larval feeding increased in several fami-
lies of gastropods from the Early Tertiary, which is
consistent with the hypothesis that speciation rates
are greater under conditions of less larval dispersal
(Hansen, 1982). A study of sacoglossan gastropods
points to the reverse, consistent with the hypoth-
esis that extinction rates are greater with less larval
dispersal (Krug et  al., 2015). Similarly, a study of
molgulid and styelid tunicates found lower diversi-
fication rates for species without a swimming larva
(Malisk­a et al., 2013). However, extinction rates can
also depend on differences in geographic range that
are independent of modes of larval development
(Jablonski and Hunt, 2006).
21.4  Ecology: Dispersal, Feeding,
and Metamorphosis
Descriptions of evolutionary changes in larval de-
velopment do not explain the adaptive advantages
of different modes of development or their eco-
logical consequences. Other kinds of observations
and investigative methods are needed to compre-
hend how larvae help animals meet the challenges
of completing a life cycle in marine environments.
Two adaptive advantages are commonly suggested
(McEdward, 2000). One is that larval feeding per-
mits development and growth from a smaller egg to
a larger adult-like juvenile. The other is that a mobile
larva enables dispersal of animals that are sessile or
sedentary as juveniles and adults. Developing and
testing these hypotheses requires the use of math-
ematical and physical models, molecular genetics,
and laboratory experiments to establish functional
mechanisms and responses. Concepts and methods
from different disciplinary lines of inquiry must be
combined. Some conclusions from these interdisci-
plinary inquiries are counterintuitive.
Larval feeding does permit greater growth than
reliance entirely on dissolved organic materials.
However, the gain comes at the expense of a longer
larval period during which ephemeral larval struc-
tures are developed and food is gathered (Jaeckle,
Chapter 9; Marshall et al., Chapter 3; McAlister and
Miner, Chapter  8; Pernet, Chapter  7; Young et  al.,
Chapter  16). There is a trade-off between number
and size. A mother can produce more offspring if
314   E v o l u t i o n ary Ec o l o g y o f M ar i n e I n v e rt e b rat e L ar va e
they are smaller, but the smaller offspring experi-
ence greater mortality during a longer period of
growth. The modeled trade-offs are based on neces-
sary limits to what mothers can devote to reproduc-
tion and the division of that effort among all of her
offspring, but those limits are shaped in complex
ways. Initial mathematical models for this trade-off
confined the effects of offspring size to growth and
mortality in the larval period. Subsequently, it has
been shown that egg size and larval experience can
affect development and growth after metamorpho-
sis, and parental experience can affect growth and
survival of larvae (Byrne et al., Chapter 17; Marshall
et al., Chapter 3; Pechenik, Chapter 14).
Although more information is needed, there are
now studies that relate developing larval structures
to larval growth quantitatively via the larva’s capac-
ity to capture food, digest it, and then translocate
materials to growing tissues (Jaeckle, Chapter  9;
Pernet, Chapter 7; McAlister and Miner, Chapter 8).
The trajectory of development through a series of
larval forms is not fixed. Plasticity in many aspects
of reproduction and development makes it possi-
ble to adjust for differences in the food or predators
encountered. The effectiveness of the plasticity de-
pends on the availability of a stimulus that serves as
a reliable predictor of future conditions. The plastic-
ity observed in the development of structures for
feeding, storage of nutrients, and development of
juvenile structures is particularly striking. There
are presumably advantages from shifting growth
among structures for feeding, digestion, food stor-
age, and postlarval life depending on food supply,
but the advantages are difficult to quantify. One
potential strategy for demonstrating the costs of a
fixed development and gains from plasticity is ma-
nipulating the development of a larval form while
holding the food supply constant (McAlister and
Miner, Chapter  8). Stimuli from predators also in-
duce changes in larval form. A response in echino-
derms is larval cloning, which occurs with stimuli
from predators and also under conditions favorable
for growth (Vaughn and Allen, 2010; Allen et  al.,
Chapter 5).
The costs of a longer period of planktonic devel-
opment are difficult to assess. Mortality results from
both predation and transport away from favorable
habitats. Losses from predation and from transport
are often difficult to distinguish, although estimates
of mortality are improving for some kinds of lar-
vae (White et al., 2014). Risks do not necessarily de-
crease as larval size increases. Some predators, such
as small fish, prey more readily on larger larvae,
perhaps because they are easier to detect (Vaughn
and Allen, 2010). Risks may often depend more on
larval behavior than size (Eiane and Ohman, 2004).
Uncertainty about these and other risks limits infer-
ences about trade-offs of size vs. number (Marshall
et al., Chapter 3; Young et al., Chapter 16). Popula-
tion density in the adult stage also can affect size-
number trade-offs in several ways. For example, in
animals that shed eggs and sperm into the water
column (i.e., broadcast spawning), the size of eggs
affects encounters with sperm and thereby modu-
lates losses from either few sperm-egg contacts or
polyspermy (Marshall et al., Chapter 3).
A planktonic larval stage can account for much
of the dispersal in an otherwise sedentary animal
(Marko and Hart, Chapter  12). Because a plank-
tonic larval stage leads to spectacular dispersal in
some circumstances, it is tempting to infer that this
stage is an adaptation for dispersal. However, the
costs and benefits depend on the stage of develop-
ment. Many invertebrates are sessile or sedentary as
juveniles and adults. A mobile larva can find suit-
able habitat at a distance from parents and siblings,
but it is a challenge for planktonic larvae of benthic
adults to encounter suitable habitat on the seafloor
after days, weeks, or months in the plankton (Hodi­n
et  al., Chapter  13; Pineda and Reyns, Chapte­r  11;
Young et al., Chapter 16). The benefits of larval dis-
persal are more apparent when the larva is com-
petent for settlement and metamorphosis. During
a long period of larval feeding and growth, before
metamorphic competence, dispersal is not halted
when a favorable benthic habitat is encountered.
As a consequence, the risk of transport away from
a suitable habitat might outweigh any advantages
of dispersal. Many larvae disperse less than if they
were neutrally buoyant and passive in the water
column. Larvae enhance retention near suitable ben-
thic habitats or enhance returns to these habitats by
adjusting vertical position in currents that vary with
time or depth (Hodin et al., Chapter 13; Pineda and
Reyns; Chapter 11; Young et al., Chapter 16). Even
during competence for metamorphosis, habitat

selection is a formidable challenge because the sea is
large, much of the seafloor inhospitable, and speeds
of swimming larvae are commonly less than the
speeds of ocean currents.
The challenges do not cease when a larva ar-
rives at a favorable site on the seafloor. A larval
form adapted for life in the plankton differs from
a juvenile form adapted for a benthic life. Most
transitions from a planktonic to a benthic form are
rapid, so that the duration of less capable interme-
diate stages is brief. Some aspects of the internal
signaling that initiates rapid metamorphosis are
widespread (Marlow, Chapter  2), but the external
stimuli differ greatly across different species, can
be numerous within a given species or of different
types (e.g., stimulatory or inhibitory), and operate
over a range of scales near suitable benthic habitats
(Hodin et al., Chapter 13). Since larvae must be pre-
pared for a rapid metamorphosis when the time is
right, they must develop competence for settlement
and metamorphosis in advance of encountering the
inducing stimulus. The length of time that a species’
larvae can maintain metamorphic competence is ex-
pected to depend on their chances of encountering
a suitable habitat (Young et al., Chapter 16); extend-
ing habitat search during competence can decrease
size or growth after metamorphosis (Pecheni­ k,
Chapter 14). There also can be stimuli that induce
an earlier competence, as in the response of sea ur-
chin larvae to turbulence (Hodin et al., Chapter 13).
Anatomically, rapid metamorphosis is facilitated by
rudiments of juvenile structures. These rudiments
develop internally where they interfere less with
larval functions, but then can be deployed rapidly
when needed. Although juvenile rudiments and
rapid metamorphoses evolved independently in
many clades, the evolution of distinctive larvae ap-
pears to have preceded the evolution of more radi-
cal metamorphoses. Not all larvae change habitat,
and some of these larvae transform gradually into
an adult-like juvenile.
The risks of planktonic development raise the
question: how does a larval stage help an animal
cope with its environment in a way that direct
development does not? As we have already seen,
a feeding larva reduces maternal investment per
offspring by obtaining its own food for growth.
For sedentary animals, a mobile larva provides
M o d e l L i f e H i s to r i e s    315
dispersal and habitat selection. In addition, a mo-
bile larval stage reduces costs of parental protec-
tion. Larvae can be analyzed as adaptations that
achieve these ends (Hodin et  al., Chapter  13; Jae-
ckle, Chapter 9; Marshall et al., Chapter 3; McAlis-
ter and Miner, Chapter 8; Pernet, Chapter 7; Pineda
and Reyns, Chapter 11; Young et al., Chapter 16), yet
many puzzles remain (and are detailed throughout
this volume). For example, many marine inverte-
brates protect offspring in broods or capsules until
the larvae are capable of feeding or settling. This
protection can reduce risk during vulnerable early
stages of development. Protection at stages that
cannot feed and are far from metamorphic com-
petence seems like an adaptive strategy, but many
invertebrates nevertheless engage in broadcast
spawning. The reality is that protection entails costs
and limitations, which are varied and incompletely
quantified (Marshall et al., Chapter 3).
Although the preceding chapters concentrate on
invertebrates with planktonic larvae and benthic
adults, some invertebrates are benthic as both larva
and adult, and others swim far from the seafloor as
both larva and adult. Why do larva and adult so
often (but not always) live in different habitats?
The question is bolstered by similar patterns in ter-
restrial species, but the answers are uncertain. For
larvae in general, including those of amphibians
and insects, different habitats provide differences
in food and safety, as well as different opportuni-
ties for dispersal, habitat selection, and mating. A
small mobile animal can disperse to a favorable
habitat more effectively in the ocean than on land
or in ponds and streams. But it is less clear why the
precompetent feeding stages of marine invertebrate
larvae are commonly planktonic. Head larvae are
conspicuously less developed than adult-like juve-
niles, but that does not necessarily preclude benthic
life. Several kinds of marine ecdysozoan larvae are
benthic. It is uncertain whether ciliated larvae are
somehow less readily adapted for benthic feeding.
Possibly more larvae develop near the seafloor
than is currently recognized simply because it is
difficult to sample larvae there. However, many
larvae swim upward, some even from great ocean
depths to the surface (Young et al., Chapter 16). For
larvae that begin life at great depths, faster develop-
ment and growth occur near the surface at higher
316   E v o l u t i o n ary Ec o l o g y o f M ar i n e I n v e rt e b rat e L ar va e
temperatures and with more food, but that ad-
vantage may not apply generally in more shallow
waters. One explanation for why larvae enter the
plankton (in addition to advantages from dispersal
and habitat selection) is the existence of greater pre-
dation near the seafloor. In the few studies made,
planktonic larvae are at greater risk from predators
near the seafloor, even at stages adapted for settle-
ment (Acosta and Butler, 1999; Allen and McAlister,
2007). Departing the seafloor may decrease risk for
larvae. Although diverse kinds of tiny animals live
at the seafloor, the plankton may be safer at earlier
stages of development for many animals.
21.5  Emerging Issues: Environmental
Hazards and Extreme Environments
Human activity is changing the environment of ma-
rine life through global warming, ocean acidifica-
tion, hypoxia, toxins, habitat loss, and harvesting.
Attempts to learn what kinds of changes are occur-
ring, as well as how we might anticipate and cope
with them, necessarily include studies of marine in-
vertebrate larvae. How do larvae adapt in real time
to novel environmental conditions? What evolution-
ary trajectories might be expected (apart from extinc-
tion)? Does prior knowledge derived from ecological
and evolutionary studies provide answers?
One of the methods for managing, harvesting,
and maintaining habitats is the establishment of
marine protected areas. The sizes and locations of
protected areas that will be most effective in main-
taining populations depends on the nature of lar-
val dispersal, which differs greatly among species
(Marko and Hart, Chapter  12). And since larval
behavior also influences the transport of larvae
(Pineda and Reyns, Chapter  11), marine protected
areas are expected to impose selection on those be-
haviors (Baskett et al., 2007). However, the direction
and magnitude of responses that might result un-
der these selection regimes are unknown.
There has been more progress in studies of ac-
climation and the potential for adaptation of lar-
vae to acidification and warming (Byrne et  al.,
Chapte­r  17). Observations of gene expression aid
the interpretation of the differing effects of acidifi-
cation and warming on development and growth.
When there is sufficient dispersal, the present
range of a species may provide some indication,
if not a prediction, of its resilience. Such disper-
sal is common with long planktonic larval dura-
tions (Marko and Hart, Chapter  12; Young et  al.,
Chapte­r 16). The fossil record also indicates effects
of geographic range on survival of mass extinctions
(Jablonsk­i, 1986). Thus, one question is whether
particular properties of larvae indicate resilience
in the face of environmental stress. Another ques-
tion is whether extreme environments favor larval
forms or not. Biogeographic patterns in modes of
larval development aid prediction of responses to
environmental change. Species with feeding larvae
are more prevalent in warmer waters. The propor-
tion of species with planktonic-feeding larvae and
the size of offspring in this group are not correlated
with the mean abundance of food but its seasonal-
ity and predictability (Marshall et  al., Chapter  3).
Larvae of species that occupy a broader range of en-
vironments may be more resilient to environmental
changes (Byrne et al., Chapter 17).
Paleontological studies can indicate effects of
ecological perturbations on the persistence or loss
of larval forms. The mass extinction of marine ani-
mals at the end of the Permian was extraordinarily
severe (Erwin, 2006). Environmental changes as-
sociated with this extinction include ones that we
are causing now: increased warming, acidification,
and hypoxia (Knoll et al., 2007). An irreversible loss
of feeding larval forms during this extinction was
suggested for crinoids because today’s crinoids
lack feeding larvae and few lineages of crinoids
survived the extinction. However, the discovery
of a nonfeeding but auricularia-like larval stage in
a stalked crinoid (Nakano et  al., 2003) now casts
doubt on this hypothesis. Echinoderms eventu-
ally lose structures associated with larval feeding
after a long history of nonfeeding larval develop-
ment (Wray, 1996; Collin and Moran, Chapter  4).
The hypothesis that a nonfeeding, auricularia form
was retained through more than 200 million years
is less plausible than the alternative hypothesis of
a more recent loss of larval feeding in this lineage.
Few lineages of sea urchins survived the Permian
extinction, but some clearly had feeding larvae, as
their descendants show. Although this global en-
vironmental change caused many extinctions, it is

difficult to prove that there was selection against
feeding larval forms. However, tracking patterns
of species survival with and without feeding larval
forms under today’s environmental regimes may
both yield insight into these earlier evolutionary
processes and prompt more urgent action to miti-
gate these climatic changes.
Despite a wealth of developmental, ecological,
and evolutionary knowledge about marine inver-
tebrate larval forms, it is critical that we reflect on
where we might be handicapped in approaching
these new problems. One obstacle is the dispari-
ties among taxa in larval forms. Some differences
between clades arise from the presence or absence
of a calcium carbonate skeleton, whose deposition
is affected by carbonate undersaturation. Other dif-
ferences are not so visible (Byrne et al., Chapter 17).
Many potential influences of environmental change
are unknown but amenable to study. One example
is the possible shifts in the microbiomes associated
with different larvae and their effects on the sur-
vival and growth of larvae (Williams and Carrier,
Chapter  19). Microbial populations clearly change
in response to ocean acidification (Witt et al., 2011),
but linking this to effects on the growth and devel-
opment of larval forms contributes to understand-
ing the nature and extent of these animal-bacteria
interactions (McFall-Ngai et al., 2013). Another ex-
ample is the effects of anthropogenic stressors on
the removal of toxic compounds by transporters
and their effectiveness (Corsi and Marques-Santos,
Chapter  18). In these cases, marine invertebrate
larvae have the potential to be surrogate models of
the ocean environment, and the associated research
questions should spur novel manipulative experi-
mentation to enhance the information that can be
gleaned from them.
A perennial challenge in studies of marine in-
vertebrate larvae is observing very small animals
that live in a very large environment; within this
environment, one habitat (the seafloor) is nearly
stationary and another (the overlying water) is
in constant motion. The mechanics of processes
related to swimming, feeding, and the internal
transport of materials are vastly different from the
mechanics of processes related to the transport of
larvae in currents. Because observations of larvae in
situ are difficult, laboratory studies have provided
M o d e l L i f e H i s to r i e s    317
much of the information we have, through both ex-
periments and observations of individuals. But the
environment in containers of seawater differs from
the environment in the sea. The moment seawater
is placed in a container, changes in its chemistry
and populations of microorganisms begin to occur
(Hovanec and DeLong, 1996), some of which are
wholly unknown to us. Additionally, larvae live
in a world with turbulence, density layering, and
heterogeneity on many scales (Vogel, 2003; 2009).
These features are imperfectly reproduced in the
laboratory—extrapolation requires care—though
important insights have been secured from meticu-
lous experimentation (e.g., Emlet, 1991). Molecula­r
methods may aid comparisons between larval
physiology in the lab and measurements taken in
the field. Extreme environments, such as the deep
ocean, exacerbate these challenges but also could
yield the most enlightening data (i.e., high-risk,
high-yield science).
Another challenge is the timescales for the rel-
evant processes in tracking a life history. Devel-
opmental sequences for many larval forms are on
the order of several weeks, and observations of
feeding mechanisms often require recordings with
hundreds of images per second. Given that their
development (and evolution) can be influenced by
stimuli lasting seconds to days, there is a real con-
cern that we will miss the causes that make a dif-
ference. Extreme environments can impose extreme
difficulties in accomplishing scientific work within
them. Technically challenging scientific investiga-
tion, especially when there is a need to draw on
multiple lines of disciplinary expertise, requires in-
creased time, coordination, and money. Many past
studies of marine invertebrate larvae have been
done by small teams, but these considerations point
in the direction of larger teams and more substan-
tial financial investments. In a world experiencing
significant environmental upheaval, and with the
oceans a specific locus of concern, the need for these
kinds of endeavors increases; research on the future
of larvae as they do (or do not) adjust to environ-
mental changes may reveal broader impacts.
These challenges are formidable but many are
surmountable. The influence of technical advances
is evident when comparing the contributions to this
book with those from Ecology of Marine Invertebrate
318   E v o l u t i o n ary Ec o l o g y o f M ar i n e I n v e rt e b rat e L ar va e
Larvae (McEdward, 1995). An expansion of compu-
tational capacity has enhanced observations over
all scales of space and time in numerous domains:
phylogeny, genetics of populations, gene expres-
sion, gene regulatory networks, larval behaviors
(e.g., from analyses of confocal images and re-
cordings), and the motion of water and its varia-
tion. Some molecular methods should help bridge
gaps between observations in laboratory and
field. For example, sequencing techniques enable
new ways of identifying what larvae and associ-
ated organisms are present; -omics studies make
it possible to identify and track larval physiologi-
cal and developmental responses to their environ-
ment (Williams and Carrier, Chapter  19). Studies
of symbioses between larvae and microorganisms,
previously impractical, are increasingly feasible
with combinations of molecular methods, and may
provide unanticipated explanations of larval struc-
tures and behaviors (McFall-Ngai, 2014). Technical
advances have enhanced sampling, observation,
and even experiments at great ocean depths, tak-
ing larval biology from the shallows into the
world’s largest and most unknown habitat (Young
et al., Chapter 16).
21.6 Summary
Marine invertebrate larvae constitute a coherent,
structured research program as model life histories
that represent developmental, ecological, and evo-
lutionary processes in different ways. They facili-
tate the investigation of diverse research questions
about larval ontogeny, feeding, dispersal, adapta-
tion, and origination, as well as providing a window
on emerging issues where alterations of the marine
environment can be tracked in an anthropogenic,
global natural experiment. While originally emerg-
ing from a confluence of evolutionary thinking and
the discovery of larvae that differed radically from
adults, this research program has expanded to in-
clude a broad range of questions that have nurtured
interdisciplinary research across distinct questions
and theoretical stances. “Larval forms have been
central to the study of evolutionary and develop-
mental biology since the inception of these fields
in the mid-nineteenth century, yet they still offer
contemporary biologists tremendous opportunities
for deciphering fundamental biological principles”
(Emlet et al., 2009, p. 201). The expanded questions
and answers speak to central problems in the his-
tory of life and urgent issues confronting human
society. Success in studies of model life histories
provides a strong case for sustained professional,
institutional, and financial support to carry these
endeavors forward.
Many prospects and challenges for studying
marine invertebrate larvae are described herein. In
conclusion, one more benefit from investigations of
model life histories can be described. A great chal-
lenge in contemporary biology is the integration of
approaches across diverse disciplines (NRC, 2008).
Work on model clades has shown how fruitful this
integration can be, though the range of integration
has tended not to bridge the divide of ecology, evo-
lution, and behavior on the one side, and genetics,
cell, and developmental biology on the other. We
have observed throughout this chapter that stud-
ies of marine invertebrate larvae are an exemplar
of interdisciplinary research that encourages and
nurtures the integration of different fields of study,
each of which have been dominated by different
types of question: (1) Mechanism: what are the pro-
cesses that produce this pattern? (2) History: what
is the sequence of changes that led to these patterns
and processes? (3) Adaptation: how do these pat-
terns and processes help the organism meet life’s
challenges? The past achievements and future
promise of interdisciplinary investigations of these
questions in the context of marine invertebrate lar-
vae testify to enduring and ongoing contributions
from studies of model life histories found in ocean
environments to the challenges facing today’s bio-
logical sciences.
Acknowledgments
We would like to thank Tyler Carrier, Andreas
Heylan­d, and Adam Reitzel for the invitation to con-
tribute to this volume and for helpful comments on
an earlier version of the chapter. Alan Love is grateful
for feedback from participants at the Marine Biologi-
cal Laboratories—Arizona State University History
of Biology Seminar on “Why Marine Studies” (May
2016), where some of these ideas were advanced in
larval form. Alan Love is supported in part by a grant

from the John Templeton Foundation (“Integrating
Generic and Genetic Approaches to Biological Phe-
nomena”; grant no. 46919). Richard Strathmann’s
studies were supported by the Friday Harbor Labo-
ratories of the University of Washington, where he
has been educated by students and other colleagues.
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 Index

A biotic induction  73–4 Branchiostoma japonicum 13

ABC transporters  281–4 budding 69–70 bryozoans 10
Acanthaster planci  107, 252, 255, 260 cloning  69, 71, 74–6 feeding mechanisms  90–1
Achelia assimilis 11 feeding larvae  69 phylogenetic studies  52
Acropora tenuis  209, 214 future directions  76–8 Buccinium undatum 3
actinotroch 8 induction 72–4 budding 68–9
active response particle capture  89 other taxa  74 adaptive nature  75
adaptive behavior in larval flow  148 paratomy 70–1 echinoderms 69–70
adaptive sampling  154 types 68–9 Bugula
advection 149 Aspidodiadema jacobyi 234 B. californica 42
mechanisms 149–50 Asterias B. neritina  42, 218
Alderia willowi 8 A. amurensis  131, 252, 255 Bythograea
Alvinella pompeiana 234 A. forbesi  72, 132 B. thermodo 237
Ambulacraria 10 A. pectinifera  76, 131 B. thermydron 240
Amphimedon queenslandica 300 attachment 194
annelids  3, 9, 92 Aurelia labiata 131 C
Anodonta cygnea 277 Auricularia nudibranchia 233 cadmium 211
anuran larvae  8 Australostichopus mollis 108 Calyptreae lichen 8
apical plate  20–1 automated larval species cAMP response element-binding
Aplysia californica 300 recognition 155 protein (CREB)  115
Apostichopus japonicus 108 autonomous remote sampling  155 Capitella teleta  42, 210, 212, 214, 216
approximate Bayesian autonomous underwater vehicles carbon dioxide (CO2)  259–61, 264
computation 170 (AUVs) 155 anthropogenic 256
approximation population autotomy in echinoderms  71–2 Carcinus maenas 210
methods 170 Carinaria japonica 5
Arachnoides placenta  252, 255, 258 B catastrophic metamorphosis  3, 17, 21
Arbacia lixula 259 Balanus Celleporaria sp. 42
Archaeopneustes hystrix 236 B. amphitrite  210, 214 cell-type trees  22
archenteron 70–1 B. glandula  42, 210, 212, 217 Centrostephanus rodgersii  107, 252,
Argopecten irradians  216, 255 Bathymodiolus childressi  234, 237, 241 255, 257
arm length  109, 111, 113 Bathynerita naticoidea  237, 242 Cephalochordata 13
relative to body length  112 benthic life stage  3, 19 cephalopods 7
arrested development and biphasic life cycle  3, 6 CERVUS method  172–3, 178
dispersal 234 Bittium alternatum 130 Chaetoceros gracilis 214
Artemia franciscana  126, 130, 131 bivalves 52 chemical defensome  274
Arthropoda  10, 92–3 body length  112–13 chordates 13
ascidians 7–8 relative to arm length  112 Cidaris blakei 236
phylogeny 52 body plan  20–1 Ciona
Ascidiella aspersa 210 Boltenia villosa 42 C. intestinalis  13, 210, 278, 284, 300
asexual reproduction  67–8, 78 Bostrycapulus odites 55 C. savigni 42
abiotic induction  72–3 Botrylloides violaceus  13, 42 clearance rate  94
adaptive nature of larval boundary layer  192 cloning  69, 71
cloning 74–6 Brachipodia 10 adaptive nature  74–6
autotomy 71–2 feeding mechanisms  91 Clypeaster subdepressus 107
324   i n d e x

Clytia hemisphaerica 300 parental investments  233–34 Echinarachnius parma  68, 73

Cnidaria  8–9, 90 predation 237–8 echinoderms 10–11
coalescent population genetics  prevalence of brooding  232 ABC transporters  279–82
169–70 demersal drift  243–44 asexual reproduction by feeding
coastal boundary layer  193 Dendraster excentricus  72, 73, 74, 76, larvae 69
coelom formation  71 107, 129, 236, 259, 274 autotomy 71–2
Coelopleurus floridanus 236 depth-dependent larval flow  148 budding 69–70
Cominella developmental mode feeding mechanisms  93–94
C. maculosa 42 feeding  87–8, 99 future research  76–8
C. virgata 42 Diadema life cycle  3
competence 26 D. antillarum 107 paratomy 70–1
connectivity, genetic vs. D. mexicanum 107 phylogenetic studies  52
demographic 180–2 diel vertical migrations (DVMs)  196 size spectrum of catchable
connectomics  289, 290, 297–9 Diplosoma particles 96–7
continuous settlement–relocation, D. listerianum 218 toxicant exposure  274–7
definition 194 D. macdonaldi 42 echinoids 61–2
Cowen, R.K.  230 direct life cycle  3, 17 Echinometra
Corella inflate 42 dispersal  229, 244–5 E. lucunter  107, 280, 281
Crambe crambe 211 see also larval dispersal E. mathaei 257
Crandall, E.D.  177 advantages 230–2 E. vanbrunti 107
Crassostrea arrested development  234 E. viridis 107
C. gigas  108, 124, 130, 131, 255, bathymetric distribution  239 echinoplutei 259
261, 274 bimodal distribution of dispersal eco-evolutionary dynamics of
C. virginica  126, 130, 259 strategies 231 parental investment
Crepipatella peruviana 210 cumulative energy consumption  offspring quality  44–5
Crepidula 8 241 offspring size  43
C. coquimbensis 55 dispersal distance consequences  offspring size drivers  43–4
C. fornicata  73, 108, 130, 209, 212, 233–38 within-brood variation  45
216, 300 egg density  234–5 ecotoxicology 273–4
C. onyx  209, 212, 217 importance 229–30 bioassays for ABC
Crepipatella long PLDs  235–7 transporters 283–4
C. dilatata  55, 210, 212, 214 ontogenetic vertical migration  echinoderm ABC transporters 
C. peruviana 210 238–44 281–2
CRISPR technique  299 parental investments  233–34 future directions  284–5
Crustacea 10 predation 237–8 latent effects following
cryptic complexity  22 prevalence of brooding  232 metamorphosis 210–11
Cryptosula pallasiana 42 dispersal distance estimation  176 sea urchin ABC transporters 
Ctenophora 9 dispersal kernels  26, 178–80 279–80
isolation-by-distance method  sea urchin development  274–7
D 176–8
dissolved organic materials  125–6,
eddy diffusion  148
Edwardsiella lineata 76
D'Aloia, C.C.  178
131, 133–35, 218 egg density  234–5
deep-sea dispersal of larvae  229,
Distaplia occidentalis 42 egg size  37–8, 109–11
244–5
Distolasterias nipon 72 energy content  38
see also larval dispersal
dithiothreitol 73 per-offspring investment  39
advantages 230–2
diversity of marine larvae  8–9 Elminius modestus 11
arrested development  234
Ambulacraria 10 embryo 51
bathymetric distribution  239
Chordata 13 asexual reproduction  69
bimodal distribution of dispersal
Ecdysozoa 10 lecithotrophic modifications  55
strategies 231
Spiralia (Lophotrochozoa)  9 Emlet, R.B.  34, 67, 237, 310
cumulative energy consumption 
DNA methylation  219 Encope michelini 107
241
Dollo's law  51 energy content of eggs  38
dispersal distance consequences 
Doratopsis 7 constraints on maternal
233–38
Dreissena polymorpha 279 investment 39
egg density  234–5
per-offspring investment  39
importance 229–30
long PLDs  235–7 E Enteropneusta 10
Entopoctia 10
ontogenetic vertical migration  East Pacific Rise (EPR)  244
feeding mechanisms  92
238–44 ecdysozoans 10

environmental changes, responses
to 135–6
epigenetics
epigenomics 290
episodic transport  153, 154
ETS domain-containing protein
(Elk) 115
Eucidaris
E. thouarsii 107
E. tribuloides  68, 73, 107
Evechinus chloroticus 107
evolutionary development of marine
larvae  16–17, 29
ancestrality 18
body plan  20–1
cell type comparison  22–3
developmental networks  24–6
gene regulation  21–2
gene regulatory network  25, 292
heterochrony 26–7
homologous larval features  17–18
homology assessment  23
molecular approaches to
comparison 18–20
ontogenetic process
assessment 23–4
pelago-benthic metamorphosis  effects of  212–13
27–8
terminology 17
transcriptional programs  24–6
transcriptional signals  23–4
excitatory amino acid transporter  115
extracellular yolk  53–4
F hypoxia, latent effects of  207–8
FST -based metrics  175
facultative planktotrophs  36
feeding  87–8, 99
diverse larval forms and feeding
mechanisms 89–94
inferences from nature  98–9
maximum clearance rate  95–6
methods of feeding  88–94
particle encounter and capture  genetic analysis  164–5, 182–3
88–9
predictions from forms and feeding
mechanisms 94–5
relative clearance rates  97
size spectrum of catchable
particles 96–8
feeding performance  94–5
inferences from nature  98–9
feeding physiology  124, 136
absorption 129
challenges 124–5
digestion 129
dissolved organic materials
(DOM) 133–5
extraction efficiency  130–1
internal transport systems  131–2
material acquisition  125–6
material movement  126–7
responses to environmental
change 135–6
ventilation of digestive
system 127–9
feeding structures  103, 118–9
marine invertebrate larvae  105–6
phenotypic plasticity  103–5
planktotrophic larvae phenotypic
plasticity 106–18
fertilization 40
fibroblast growth factors  115
filter feeding  88–9
finite volume community ocean
model 156
Flabellum angulare  230, 233
fluid dynamics  190–1, 203–4
environmental parameters sensed
by larvae  194–9
navigation techniques of
larvae 199–203
sensory aspects of larvae  191–9
food/nutrient limitation, latent
Forkhead Box
FoxA 26
FoxG 115
FoxO 116
front propagation  151–2
Fu-Shiang Chia  288
Fusitriton oregonensis 237
G
Gaelodea 3
Garstang, W.  306
Gastropods, phylogenetic
studies 52
gastrulation 70
gene regulation  21–2
gene regulatory network  25, 292
biological correlates of larval
dispersal 173–6
consequences of larval
dispersal 180–2
dispersal distance estimation  Isarachnanthus nocturnus
176–80
individual-based methods  170–3
larval dispersal  165–6
population-based methods  166–70
genomics  289, 290
genome sequencing  18–20
genome-wide sequencing  22
Geodia cydonium 277
GLWamide 28
i n d e x    325
G-protein-coupled family
of receptors  27
gut passage time  126–7
Gyrodactylus elegans 68
H
Haeckel, E.  18
Haliotis
H. asinine 218
H. coccoradiata 256
H. diversicolor  210, 214
H. rufescens 274
Hardy–Weinberg
equilibrium 167, 171
hatching 3
head patterning network  21
heat stress, latent effects following
metamorphosis 215
Heliocidaris
H. erythrogramma  34, 252
H. tuberculata  34, 107, 252, 263
Hemichordata feeding
mechanisms 94
heterochrony 26–7
horizontal larval flow
velocity 147
hox gene  21
hydrodynamic variability  149
Hydroides
H. dianthus 210
H. diramphus  42, 212
H. elegans 242
hydrosol filtration  88, 89
hydrothermal vents  231, 234, 244
I
Ifremeria nautili  3, 232, 233
Ilyanassa obsoleta  300, 130
indirect life cycle  3, 17
individual-based genetic
analysis 170–1
assigning individuals to
families 172–3
assigning individuals to source
populations 172
clustering individuals from
different samples  171–2
manii 4
island model, Wright's  168
Isochrysis galbana  126, 212
isolation-by-distance method
dispersal distance estimation 
176–8
dispersal kernels  178–80
planktonic larval stage
duration 174
326   i n d e x

J caution in interpretations  216 Mercenaria mercenaria 259

Janua pagenstecheri 42 consequences not always metabolomics  289, 290
negative 216–17 metagenomics 290
delayed metamorphosis  214–15 metamorphosis 3
K food/nutrient limitation  212–10 see also latent effects following
kernels  26, 178–80
hypoxia 211–12 metamorphosis
impact and implications  219–20 catastrophic  3, 17, 21
L mechanisms 217–19 definition 194
laminar flow  192 ocean acidification  215 pelago-benthic 27–8
larvae  3–6, 13 salinity stress  213–14 metamorphosis-associated contractile
adaptive nature of cloning  74–6 thermal stress  215 structures 291–2
anuran larvae  8 toxicant exposure  210–11 microbiomics 289
asexual reproduction  68–9 lecithitrophy  8, 17, 36, 50, 51 invertebrate larvae  294–6
ciliated larvae  7, 17 embryo modifications  55 Microcosmus squamiger 42
origin 6–7 lipids 61 Micrura alaskens 9
variation in types  7–8 legacy effects  209 minimally indirect
larval accumulation  151–2 Leodia sexiesperforata 107 development 17
larval behavior  147–8 Leptasterias aequalis 42 Mithrodia clavigera 296
larval dispersal  165–6, 182–3 life histories  306–7, 318 mitogen-activated protein
see also deep-sea dispersal of larvae ecology 313–16 kinase  291, 292
biological correlates  173–6 emerging issues  316–18 Molgula socialis 210
consequences 180–2 evolution 310–13 molluscs 9
dispersal distance estimation  model life histories  307–10 feeding mechanisms  91
176–80 Lingula anatina 4 life cycle  3
individual-based methods  170–73 linkage disequilibrium  171 Montastraea faveolata  209, 217
population-based methods  166–70 Linopneustes longispinus 236 morpholino-mediated gene
larval diversity  8–13, 34–5 lipidomics  290, 297 knockdown 299
larval physiology  296–7 Litopenaeus vannamei 296 Mortensen, T.  34, 70
larval size  151 Littorina obtusata 3 Mulinia lateralis 255
larval transport in coastal zones  lophophore evolution  26 multixenobiotic resistance  277,
145–6, 158 Loricifera 10 278, 291
accumulation 151–2 Luidia Mytilus
advection mechanisms  149–50 L. foliolata  76, 107 M. californianus  5, 260, 261
autonomous remote sampling  155 L. senegalensis 77 M. edulis  131, 255, 260, 297
challenges 153–55 Lytechinus M. galloprovincialis  126, 259, 261,
components 147–9 L. pictus  126, 129, 131 274, 277
future directions  158 L. variegatus  68, 73, 76, 106, 107
hydrodynamic variability  149 L. variegatus carolinus 113
imaging in species recognition  155 N
larval behavior  147–84 nauplii  92–3, 97
larval duration  151 M navigation techniques of larvae 
numerical-modeling techniques  Macrophiothrix 199–200
156–8 M. koehleri 108 larval scale (millimetres–
other first-order phenomena  151–3 M. longipeda 108 centimetres) 202–3
patchiness and episodic M. lorioli 108 local scale (centimetres–
transport 153 M. rhabdoti 108 metres) 202
physical transport mechanisms  Maculaura cerebrosa 4 meso scale (0.1–1 km)  201–2
148–9 Maja squinado 284 macro scale (1–1000 km)  200–1
recent study approaches 155 Markov chain Monte Carlo Nematostella vectensis  4, 300
robotic techniques in in species methods  169, 170, 171 Nemertea 9
recognition 156 maximally indirect development  17 feeding mechanisms  90
scales 146 maximum clearance rate  94–5 neurohormones 28
spatial variability  153 predicting 95–6 neutral genetic markers  167
swimming proficiency and McEdward, L.R.  34, 37, 38, 41, 59, 62, nitric oxide/cyclic guanosine
size 151 67, 191, 310 monophosphate signaling  197
last common bilaterian ancestor  20–1 Mediaster aequalis 235 nonylphenol 211
latent effects following Melitta tenuis 107 numerical-modeling techniques for
metamorphosis  208–10, 220–21 Membranipora membranacea 109 larval transport  156–8

O paralarval stage  7
ocean acidification (OA)  251–3, 264–5
adaptation and acclimation  263–4
drivers 256–62
feeding physiology responses
to 135–6
latent effects following
metamorphosis 215
multistressor effects  262–3
ocean warming  251–3, 264–5
adaptation and acclimation  263–4
larval migration  253–6
multistressor effects  262–3
Odontaster
O. meridionalis 134
O. validus  258, 260
offspring quality  44–5
offspring size  43
drivers 43–4
within-brood variation  45
offspring size–fitness functions  39–40
fertilization success  40
planktonic period  40–1
post-metamorphic
performance 41–3
omics perspective  288–91, 300–1
connectomics 297–99
future directions  299–300
larval physiology  296–7
larval response to environmental
change 293–4
microbiome of invertebrate
larvae 294–6
molecular mechanisms  291–3
ontogenetic vertical migration  phenotypic plasticity in feeding
238–40
cumulative energy
consumption 241
demersal drift  243–4
migration to surface  240–2
phylogenetic constraint  240
transport of retention of
larvae 242–3
operational taxonomic units  58
Ophiopholis aculeata 12
Ophiura albida 12
Ostrea
O. angas 255
O. edulis  131, 255
O. lurida 215
P planktonic larval stage duration  Riftia pachyptila  234, 235
Palaemon serratus 284
Paleobrissus hilgardi 236
Pantinonemertes californiensis 4
Panulirus japonicus 133
Paracentrotus lividus  107, 274, 284
paratomy in echinoderms  70–1
parental conditioning  254, 261
parental investment  34–5, 46–7
biogeography 35–6
eco-evolutionary dynamics  43–5
egg size and size–number trade-
off 37–9
future directions  45–6
increasing dispersal  233–4
offspring size–fitness functions  platyhelminths  7, 10
39–43
theory 36–7
parent–offspring analysis  172–3
Parhyale hawaiensis 300
particle encounter and capture in
feeding 88–9
patchiness  153, 154
Patella vulgata 260
Patiria miniata  76, 126, 129, 131,
134, 300
Patiriella regularis 260
pelagic lifestyle  3, 19
pelagic propagule durations  population-based genetic
231, 233
Penaeus
P. monodon 126
P. setiferus 126
peptidomics  289, 290, 293
Perna viridis 278
Petromyzon marinus 278
Phallusia mammilata 300
Phascolosoma turnerae  3, 237
phenomics  289, 290, 293
structures  103–5, 118–19
marine invertebrates  105–6
planktotrophic larvae  106–18
Phormosoma placenta 234
Phoronida 10
feeding mechanisms  91
Phoronis
P. architecta  128, 129
P. muelleri  3, 8
Phragmatopoma lapidosa californica  R
3, 108
phylotypic stage  23
Pinctada imbricata 255
Pisaster ochraceus  12, 72, 73, 74,
76, 107
Planktomya henseni 151
planktonic larvae  40–1, 50–1
173–6, 231, 233
increasing dispersal  235–7
Planktosphaera pelagica 233
planktotrophic lifestyle  3, 7, 8, 17,
50, 51
i n d e x    327
energetic trade-offs  106–11
experimental designs and
analyses 116–18
expression patterns and
environmental cues  111–14
facultative planktotrophs  36
feeding structure developmental
mechanisms 114–16
food limitation  106
resource acquisition  106
feeding mechanisms  90
Platynereis dumerilii  297, 298, 300
Pleurobranchaea californica 300
Pocillopora damicornis 264
poecilogony 8
point of no return  135
polychlorinated biphenyls  273
polycyclic aromatic
hydrocarbons 273
Polydora ciliata 9
polyembryony 68
Polygorius 3
analysis 166–7
approximation methods  170
classical population genetics  167–8
coalescent population
genetics 169–70
neutral genetic markers  167
Porifera 8
Porites astreoides  209, 215
Priapula 10
primary larvae  7, 17
principal component analysis  117–18
Prionocidaris baculosa  68, 73
Prorocentrum lima 278
Protankyra brychia 233
Pseudochinus huttoni 107
Pteraster tessulatus 60
Pterobranchia 11
Pygospio elegans 8
recruitment, definition  194
regional oceanic modeling
system 156
relative clearance rates  97
retention time of ingested food  130
Reynold's number  191–2
Rhopaloeides odorabile 294
RNA interference  299
RNA sequencing  22
robotic techniques in species
recognition 156
Ruditapes philippinarum 278
328   i n d e x

S Spongia ceylonensis 4 toxicant exposure  273–4

Saccostrea glomerata  255, 258 Squilla mantis 11 bioassays for ABC
Sagmariasus verreauxi 11 steady-state clearance rate  95 transporters 283–4
salinity stress, latent effects of  213–14 Sterechinus neumayeri  134, 257, echinoderm ABC transporters 
Saxipendium coronatum 233 263, 264 281–2
scan-and-trap particle capture  89 Stokes drift  148 future directions  284–5
Scapharca subcrenata 278 Stone, C.J.  93, 97 latent effects following
Schizocardium brasiliense 132 Strathmann, R.R.  34, 36, 39, 87, 92, metamorphosis 210–11
schmoo 10–11 95, 113, 288 sea urchin ABC transporters 
Sclerasterias Strongylocentrotus 279–80
S. mollis 107 S. droebachiensis  68, 73, 108, 129, sea urchin development  274–7
S. tanneri 237 216, 236 transcriptomes  25–6, 289, 290, 293,
Scopalina lophyropoda 211 S. franciscanus  12, 108, 236 296–7
Scrupocellaria scruposa 4 S. purpuratus  73, 131, 134, 236, 259, Tripneustes gratilla  108, 257, 263
Scylla serrata 126 261, 274, 278, 300 Tubularia 9
secondary larvae  7, 17, 18 STRUCTURE genetic program  turbulent flow  192
sensory aspects of larvae  190–1, 203 171–2
environmental parameters Styela U
sensed 194–9 S. gibbsii 42 urochordates 13
navigation techniques  199–203 S. plicata  42, 218 Urticina felina 42
responses 191–9 Stylocidaris lineata 236
scales of sensory input  198 suspension sieving in feeding  88–9
sensory cilia  17 sweepstakes reproductive success V
(SRS) 201 Vance, R.  36–41
settlement, definition  194
swimming proficiency of larvae  151 model of offspring size  36–7
simulation techniques for larval
velum  3, 62
transport 156–8
Verongia aerophoba 277
single nucleotide polymorphisms  170 T vitellogenesis 44–5
site frequency spectrum  170 Tachypleus tridentatus 11
size spectrum of catchable Tegillarca granosa 278
particles 95 Terebratalia transversa 4 W
predicting 96–8 Tethya aurantium 277 Wake, D.  308
Skeletonema costatum 212 thermal stress, latent effects following Wald test  38
Solaster stimpsonii  12, 38 metamorphosis 215 Watersipora subtorquata  42, 43, 218
species distribution models  255 Thomson, C.W.  232 Wright, S.  168, 169
species recognition Thorson, G.  34, 35, 50, 232, 235, 240, island model  168
imaging 155 288, 310
robots 156 Thorson's rule  35 Z
spiralians 9 totipotent cells  68 zoeae 93