Literature Review: The Evolutionary Influence of Pollinators on Floral Morphology

Introduction

Pollination refers to the process by which pollen, a substance containing a male gamete

emitted from the male anatomy of a flowering plant, is transferred to the stigma of either the

same flower or a different flower for fertilization. A pollinator is an animal that aids in this

process of pollination by transporting pollen between the male anther of a plant and the stigma of

a plant (USDA). Pollinators influence the evolution of floral morphology through a variety of

methods, including selection based on floral attraction and through floral mechanistic adaptation

to pollinators based on increasing the functional fit between pollinator and plant for more

efficient pollen transfer. Our understanding of the relationship between pollinators and floral

evolution is important to our understanding of evolutionary biology as a whole as it elucidates

important concepts such as stabilizing and directional selection as well as providing explanations

for why phenotypic traits not selected for can remain in populations.

Literature Overview

One of the flowering plants in an experiment to test for morphological evolution in

response to pollinator selection was the Alpine sky pilot Polemonium Viscosum (Galen, p. 882)

in the paper “Measuring Pollinator-mediated selection on Morphometric Floral Traits:

Bumbeless and the Alpine Sky Pilot, Polemonium Viscosum”. The pollinator used to pollinate

the flowers was free-foraging queen bumblebees of the species Bombus Kirbyellus (Galen, p.

884). The Polemonium Viscosum is a unique case among the species to be analyzed due it

requiring pollination to reproduce because it is self-incompatible and protandrous, meaning the

male reproductive organs mature before the female organs (Galen, p. 883). The experiment used

two populations of plants: those that were pollinated randomly by hand as controls and
individuals experimentally pollinated by bumblebees (Galen, p. 884). The purpose of the control

in this experiment as it is in most experiments regarding this topic to be presented is to determine

whether the dependent variables tested for are positively correlated with high resource

availability (Galen, p. 884). High resource availability could be a confounding variable affecting

the physical traits of gathered flowers outside the effect of pollinators. 120 flowers were

randomly chosen for the experimental test and 41 bees used as pollinators, with the bees taken

out of the enclosure only after each bee had visited at least 10 plants (Galen p. 884). For each

plant in the control and experimental population the total number of flowers was counted, and

the corolla and inflorescence stalk size measured (Galen, p. 885). To test the fitness for each

plant, the number of seeds per hand-pollinated flower for control plants and the number of seeds

in every flower was counted for the experimental plants (Galen, p. 885). The relative fitness for

each individual was then regressed on the number of times that individual was visited by a bee

(Galen, p. 885). The experiment found seed production of plants was strongly correlated with

how many times that plant was visited by a bumblebee with an F value of 45.58 (Galen, p. 885).

The principal component 1 is highly correlated with stem length, corolla length, and corolla

diameter (Galen, p. 886) while the principal component 2 is highly correlated with flower

number (Galen p. 886). Principal component 1 was found to have a great influence on the

variation in seed number with an F value of 25.40 (Galen p. 886) whereas principal component 2

did not affect seed production (Galen p. 886). These influences along with the correlation

between bumble bee visitations and seed number should point to those morphological traits

associated with principal component 1 also demonstrating a correlation with bumble bee visits.

The paper shows this to be the case with corolla length having correlation score of 0.39 with

bumblebee visits and 0.41 with relative fitness, corolla diameter having a correlation score of
0.31 with bumblebee visits and 0.33 with relative fitness, and inflorescence stalk height having a

correlation score of 0.41 with bumblebee visits and 0.24 with relative fitness (Galen p. 888).

These results show that bumblebees frequented plants with larger flowers on taller stems, thus

displaying a case of directional selection towards plants with these features. An important aspect

of directional selection is heritability, as in order for directional selection to occur the plants with

the morphological traits correlating to pollinator visits and fitness must be able to pass those

traits along to their progeny. An important segment of the paper concerning the control

population shows no fertility advantage inherent to each morphological trait outside of

pollination for hand-pollinated plants (Galen, p. 889). The next paper further demonstrates

evidence of directional selection as a result of pollinator induced influences as well as

demonstrating correlational selection on flower number, an aspect of floral morphology not

found to be correlated with fitness in the first paper.

Cyclopogon elatus is a pollinator dependent orchid used in the next experiment in the

paper “Pollinator-mediated selection on floral traits and size of floral display in Cyclopogon

elatus, a sweat bee-pollinated orchid.” This experiment differs from the previous experiment in

that this paper studies plants found in the field and observed and counted the various pollinators

that visited each chosen plant before capturing the pollinator and harvesting the flowers after

pollination, both for analysis (Benitez-Vieyra et al, p. 949). Two flowers of each plant's nectary

depth, labellum width, and the size of the lower lip were all measured, as well as the number of

flowers on each plant and the time spent on each plant by each pollinator (Benitez-Vieray et al,

p.949). The number of fruit sets and the number of pollinaria exported were used to determine

reproductive success for male and female anatomy, with male corresponding to pollinaria

exported and female corresponding to the number of fruit sets (Benitez-Vieray et al, p. 949). A
variance test conducted in this article that was not conducted in the last article and that would

have strengthened its credibility is comparing the variance among the morphological features of

flowers on an individual plant to the variance of flowers between plants, and only including the

analysis for the traits for which the variance was significantly greater among plants than within

plants (Benitez-Vieyra, p. 951). The results showed higher selection through male function

compared to female function (Benitez-Vierya, p. 951). They also show significant directional

selection acting on smaller-sized lower lips, deeper nectaries, and greater flower number

(Benitez-Vierya, p. 952). The selection on lip size was found to be related to male and female

function, while deeper nectaries are influenced through male fitness and greater flower number

through female fitness (Benitez-Vierya, p. 952). Analysis of the pollinator Augochlora nausicaa

(Benitez-Vierya, p. 948) reveals the causes behind many of these selections. The mean

proboscis length of A. nausicaa and the mean necessary length of flowers are very close

(Benitez-Vierya, p. 954) The close match is important to the function of pollen uptake as the

pollinarium is adhered to the underside of the pollinators labrum (Benitez-Vierya, p. 954). This

is the likely explanation for directional selection related to male reproductive function driving

deeper nectary depth as the the nectary depth needs to be as long as or longer than the pollinators

proboscis in order to optimally export pollinarium through its adherence to the pollinators

labrum (Benitez-Vierya, p. 954). The study also demonstrated how floral morphological traits

could interact synergistically to increase function. This was shown through the positive

correlation between greater flower number and deeper nectaries, likely because flower number

increases the rate of pollinator visitation and deeper nectaries increase pollinarium export so

these effects work multiplicity to increase fitness (Benitez-Vierya, p. 955). While there was

directional selection on flower numbers related to female fitness, this was overwhelmed by the
stabilizing selection detected (Benitez-Vierya, p. 955). This was shown to be caused by more

flowers increasing the attractiveness of a plant to pollinators by a positive correlation between

pollinator visitation and the number of flowers a plant has (Benitez-Vierya, p. 954), yet the more

individual flowers a plant has the lower the fitness of each individual flower above a point due to

there being a limited number of pollinators and a limit to the amount of pollen they can carry

(Benitez-Vierya, p. 955). This stabilizing selection was not found in the first article, likely due

to those plants not being subject to severe pollen limitation. The next article demonstrates

further evidence of the influence of pollinators on directional selection of floral morphological

traits.

The article “Pollinator-Mediated Selection on Floral Display and Flowering Time in the

Perennial Herb Arabidopsis Lyrata,” uses Arabidopsis Lyrata as the flowering plant, and is

significant compared to the other articles for its analysis of how pollinators can affect perennial

time in flowering plants. The experimental setup is similar to the other articles, where 100 plants

are randomly assigned to be pollinated by hand whereas the other plants are subject to open

pollination by pollinators (Sandring, Agren, p. 1293). The morphological traits associated with

pollination: petal size, flower production, and the size of the leaf rosette, are measured for each

plant (Sandring, Agren, p. 1294). Fitness, as in the other experiments, was measured using the

number of seeds produced by each flower (Sandring, Agren, p. 1294). The experiment was

conducted twice, once in 2003, and once in 2005, to give two sets of sample data. In this

experiment correlations are calculated for traits relative to their pollination method, open or

supplemental, rather than between the number of visits by pollinators they receive. This lends

the experiment to better evaluate the effect of pollen limitation on floral evolution, as

presumably there is no pollen limitation in the populations supplemented by hand. The results,
as in the prior experiment, indicate a positive correlation between many flowers, large leaf

rosettes, an early start of flowering, a late end of flowering, and large petals (Sandring, Agren, p.

1295). The number of flowers on each plant and the size of their petals experienced positive

directional selection as well (Sandring, Agren, p. 1296), though the study could not show that the

petal size is influenced by pollinators as the magnitude of selection was not significantly

different between open pollinated plants and plants supplement by hand (Sandring, Agren, p.

1298). This is a deviation from the article about the sky pilot and the future article on

Dactylorhiza lapponica that found a significant influence by pollinators on corolla (petal) size.

The study suggested petal size is correlated with overall increased reproductive output rather

than selected for by pollinators (Sandring, Agren, p. 1298), though due to the findings of the

other papers and the commonly understood function of petals as attractors for pollinators, this

study’s inconclusive results may be due to their research method of comparing only open-

pollinated plants with artificially pollinated plants rather than between open-pollinated plants

based on the number of visitors they receive. This is in contrast to the selection influence on the

number of flowers per plant, which experienced much stronger selection for more flowers among

the open-pollinated flowers as compared to the flowers supplemented by hand (Sandring, Agren,

p. 1298). They also indicate directional influence by pollinators for late start of flowering in one

sample and early end of flowering in the other (Sandring, Agren, p. 1296). The explanation

provided by the article for why pollinators selected for an early end to flowering is pollen

limitation, where pollen was more limited later in the season (Sandring, Agren, p. 1297). An

example of a discrepancy found between the traits found to be significantly correlated with

pollination between the papers can be found in the next article, with their findings showing no

influence by pollinators on flowering start or end.

 The article “Pollinator-mediated selection on floral display, spur length and flowering

phenology in the deceptive orchid Dactylorhiza lapponica” tests pollinators' influence on the

floral morphology of the orchid Dactylorhiza lapponia. The pollinators used in the study were

the bumblebees B. pascorum and B. lucorum (Sletvold et al, p. 387). 85 plants were subject to

pollination by hand and 165 plants subject to open pollination (Sletvold et al, p. 387). The study

found positive directional selection for plants with greater height and longer spurs, while petal

size and start of flowering were not found to be significantly correlated (Sletvold et al, p. 388).

Dactylorhiza lapponica is significant because it is “deceptive.” This means they give no food

reward for their pollinators (Sletvold et al, p. 385). The study’s finding that pollinators did not

influence flowering time is significant because this differs from the previous paper’s finding, and

also differs from another paper cited in this study that tested pollinators influenced on a different

deceptive pollinated plant (Sletvold et al, p. 390). The paper claims this may be due to certain

pollinators adjusting to the lack of reward slowly and thus there being limited selection for early

flowering times as the pollinators will not adjust later in the season soon enough to stop

pollinating the orchids (Sletvold et al, p. 391). However, it does not fully answer why there

would not be significant directional selection for early flowering due to limited pollen

availability later in the season as found in the prior article.

The final paper “Pollinator-mediated selection on flower-tube length in a hawkmoth-

pollinated Gladiolus (Iridaceae)” analyzes the pollinator mediated effects on floral morphology

of Gladiolus by the hawkmoth Agrius convolvuli. The study measured the tube length of 289

flowering plants of the Gladiolus longicollis (Alexandersson, Johnson, p. 631). 15 plants were

chosen randomly to receive pollination by hand (Alexandersson, Johnson, p. 631). The

researchers then captured hawkmoths in the area and checked for the presence of Gladiolus
longicollis pollen as well as measuring the tongue length of each hawkmoth captured

(Alexandersson, Johnson, p. 632). The study found that the mean tube length of plants that

produced a fruit were significantly longer than those that did not produce fruit (Alexandersson,

Johnson, p. 634). This provides evidence for directional selection, as the efficiency for pollen

transfer is increased when the flower tube is longer than the hawkmoth’s tongue (Alexandersson,

Johnson, p. 634). The flowers with longer tubes will be more likely to be pollinated by a

hawkmoth with a tongue shorter than the flower tube, and this is evidenced by the probability for

a flower to produce fruit increasing from about 50% at the shortest to nearly 100% at the longest

(Alexandersson, Johnson, p. 634).

Conclusion

From each article a clear correlation between pollinator visitation and certain

morphological features that either serve to attract pollinators or facilitate the transfer of pollen

was found. For the attraction of pollinators, the number of flowers on a plant was found to be

positively correlated with bee visits, and by the number of flowers on a plant was found to be

positively correlated with that plant’s fitness (Benitez-Vierya, p. 955). An example of

mechanistic directional selection for the transfer of pollen was directional selection for longer

tubes in hawkmoth pollinated Iridaceae as stated earlier. Some minor discrepancies exist among

the traits within the pollinated plants that were found to be selected upon by pollinators. These

were likely due to differences between the plants, such as the Dactylorhiza lapponica being the

only deceptive plant shown and the only plant where floral start or end was not significantly

correlated with the pollinators. Despite these differences, every article demonstrated pollinator-

mediated directional selection on floral morphological traits. Further research could be

conducting these tests over many generations to test the heritability of these traits, as well as
providing further insight into why certain traits, such as long nectary tubes, are not present in

every flower in a population that is influenced by pollinators. Current hypotheses include high

gene flow between populations with polarized morphological traits and selection against certain

traits by herbivores such as herbivores potentially preferring to eat larger flowers

(Alexandersson, Johnson, p. 634).

 Bibliography

Sandring, S., Agren, J. (2008) Pollinator-mediated selection on floral display and flowering time
in the perennial herb arabidopsis lyrata. International Journal of Organic Evolution, 63, 1292-
1300

Alexandersson, R., Johnson, S. (2002) Pollinator-mediated selection on flower-tube length in a

hawkmoth-pollinated Gladiolus (iridaceae). Proceedings of the Royal Society B, 269, 631-636

Sletvold, N., Grindeland, J., Agren, J. (2010) Pollinator-mediated selection on floral display,
spur length, and flowering phenology in the deceptive orchid Dactylorhiza lapponica. New
Phytologist, 188, 385-392

Galen, C. (1988) Measuring pollinator-mediated selection on morphometric floral traits:

Bumblebees and the Alpine sky pilot, Polemonium Viscosum. International Journal of Organic
Evolution, 43, 882-890

Benitez-Vieyra, S., Medina, A., Glinos, E., Cocucci, A. (2006) Pollinator-mediated selection on
floral traits and size of floral display in Cyclopogon elatus, a sweat bee-pollinated orchid.
Functional Ecology, 20, 948-957

What is Pollination? United States Department of Agriculture.

https://www.fs.fed.us/wildflowers/pollinators/What_is_Pollination/