Life Cycle, Cytology, and Morphology of Polypodium hydriforme, a Coelenterate Parasite of

the Eggs of Acipenseriform Fishes

Author(s): Ekaterina V. Raikova
Source: The Journal of Parasitology, Vol. 80, No. 1 (Feb., 1994), pp. 1-22
Published by: The American Society of Parasitologists
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 J. Parasitol, 80(1), 1994, p. 1-22
? American Society of Parasitologists 1994

INVITEDREVIEW

LIFECYCLE,CYTOLOGY,AND MORPHOLOGY OF
POLYPODIUM A COELENTERATE
HYDRIFORME, PARASITE
OF THEEGGS OF ACIPENSERIFORM
FISHES
EkaterinaV. Raikova
Instituteof Cytology,4 Tichoretsky
Ave., 194064
St. Petersburg,Russia

ABSTRACT: Polypodiumhydriformeis the only coelenterateadapted to intracellularparasitismin oocytes of

acipenseridand polyodontidfishes. It occursin both the Old and the New worlds,being parasiticin 12 species
of Acipenseridaeand in 1 species of Polyodontidae.Its earliestparasiticstages are binucleatecells that occur
in previtellogenicoocytes. All embryonicand postembryonicdevelopment(which seems to be parthenogenetic)
up to the budding stolon stage takes place inside fish oocytes and lasts several years. The planula and stolon
have inverted germ layers. All parasiticstages are encircledwith a highly polyploid unicellulartrophamnion
that is homologous to the second polar body. Before spawning,eversion of the stolon takes place inside the
oocyte. At spawning,the everted stolons get into water and the free-livingphase of the life cycle begins. The
stolon fragmentsinto individual specimens that can move and feed. They multiply by longitudinalfission
(paratomy).In mid-summerthey form 2 kinds of endodermalgonads.The so-called"female"gonads(2 ovaria,
each with a gonoduct encircledwith a common envelope) producediploid cells that display no meiotic phe-
nomena.The so-called"male"gonadshave no gonoducts,but theirsex cells undergo2 meiotic divisions, giving
rise to binucleatecells with unequalnuclei. The entire gonad becomes a gametophorewith an ectodermallid
carryingnematocystsand containingmany binucleatecells. Gametophorescan be deposited onto the skin of
prelarvaeof fishes. How the parasitegets into young fish oocytes is not known.

Polypodium hydriforme is the only coelenter- HISTORY

ate adapted to intracellular parasitism in oocytes
of acipenserid and polyodontid fishes. Investi-
gations of its life cycle reveal a unique system of
adaptations of a multicellular organism to pro-
longed (several years) parasitism inside a single
though highly specialized cell, the fish oocyte.
Polypodium hydriforme has serious economic
significance because it affects the most valuable
product of acipenserids, black caviar, and may
severely compromise the reproductive potential
of infected fishes. To control this parasite, a de-
tailed study of all stages of its life cycle and its
parasitic adaptations is necessary. This is es-
pecially urgent for the countries of the former
USSR, because its share in the world catch of
acipenserid fishes approached 90%, and 95% of
this catch came from the Volga-Caspian basin
(Sokolov, 1983) where P. hydriforme seems to
have the highest prevalence. This review sum-
marizes our knowledge regarding the life cycle
of P. hydriforme, highlights new research prob-
lems, and draws this most interesting parasite to
the attention of zoologists, cytologists, and par-
asitologists.
Received27 March 1991;revised 23 June 1993; ac-
cepted 28 August 1993.
1
OF THESTUDYOF POLYPODIUM
A disease of sterlet (Acipenser ruthenus) eggs
was first reported in the Volga River by Ows-
jannikow (1871). Grimm (1873) called the caus-
ative organism "Owsjannikow's parasitic larva."
The animal was more thoroughly described by
Ussov (1885, 1887) and named P. hydriforme.
Lipin (1909, 1911a, 1911b, 1915, 1922a,
1922b, 1925) conducted most of the early studies
on the life cycle. He first discovered the inversion
of the germ layers in the parasitic stolon (Lipin,
1911la) and interpreted it as an adaptation to
parasitism; since then P. hydriforme has attracted
much attention as an "enantiozoan." However,
many questions remained unsolved even after
Lipin's thorough studies: the entire life cycle in-
cluding the mode of infection offish oocytes, and
its systematic position, among others. Studies on
the life cycle were resumed with the work of
Raikova (1958a) and still are continuing.
HOSTANDGEOGRAPHICAL
DISTRIBUTION
During the first 70 yr (1870-1940) after its
discovery P. hydriforme was known as a parasite
of only the Volga sterlet (A. ruthenus). Subse-
quently, Dogiel (1940) discovered its distribu-
tion in new hosts and rivers, reporting it in the

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2 THEJOURNALOF PARASITOLOGY,
VOL. 80, NO. 1, FEBRUARY1994

TABLEI. Distribution of Polypodium hydriforme in various hosts and bodies of water.

Prevalence
(% of hosts Number of infected
infected when eggs per infected
Host River counted) female when counted Reference

Acipenser ruthenus Volga Owsjannikow, 1871; Grimm, 1873;

(sterlet) Ussov, 1885; Lipin, 1911la; Dogiel,
1940
Kama 78 10-100% Persov, 1957; Raikova, 1958a
Don Raikova, 1964a, 1964b
Danube Rasin, 1930; Bogatu, 1961
North Dvina Persov, 1957
Acipenser giildenstiidti Volga Dogiel, 1940
(Russian sturgeon) 12.5-29.0 1-96 Raikova, 1959
7.6-44.8 Markov et al., 1963, 1964, 1965
Kama Raikova, 1959
Don Dogiel, 1940
Ural 100 10-25 Smirnova and Michenko, 1966
Sulak Dogiel, 1940
Acipenser stellatus Volga Dogiel, 1940
(sevrjuga) 0.8-3.4 Markov et al., 1963, 1964, 1965
Don Detlaf and Ginsburg, 1954
Kuban Kamenev and Sachnina, 1956
Sulak Dogiel, 1940
Dnestr 1 specimen 36 Shumilo, 1959
sampled
Ural 12.5 Up to 25% Smirnova and Mishenko, 1966
Sefidrud 1.4 Nechat and Mokhayer, 1974
Acipenser nudiventris Ural Raikova, 1986
(schip) Syr-Darya Trussov, 1947
Lake Balkhash Tlenbekova, 1980
Acipenser schrenki Amur 23.3 Average 290 Svirsky, 1984
(Amur sturgeon) Lake Hanka Yukhimenko, 1985

Acipenser baikalensis Selenga 16.6 Pronin, 1975

(Baikal sturgeon)
Acipenser medirostris Tumnina River (Sa- 25 10-15% Artuchin and Andronov, 1989
(green sturgeon) khalin)
Acipenserfulvescens Black River (Michi- 20 0.07% Hoffman et al., 1974
(lake sturgeon) gan)
Nelson River 2-176 Dick et al., 1991
Saskatchewan River
(Canada)
Acipenser brevirostrum St. John River (Can- 50 Ca. 1% Dadswell et al., 1984
(shortnose sturgeon) ada)
Acipenser transmontanus Davis River (Cali- Collected by M. Kent, pers. comm.
(white sturgeon) fornia)
Huso huso Volga Markov and Trussov, 1966
(beluga) Ural Raikova, 1986
Huso dauricus Amur 42.8 Average 1,636 Svirsky, 1984
(kaluga)
H. huso x A. ruthenus Ponds near Don I. A. Burtsev, pers. comm.
(bester)
Polyodon spathula Osage (Mississippi) 0.01; 88 at Suppes and Meyer, 1975
(paddlefish) rearing

eggs of the Russian sturgeon (Acipenser giilden-
stiidti) in the Volga and Sulak rivers and in the
eggs of the sevrjuga (Acipenser stellatus) in the
Sulak and the Don rivers. It seemed probable
that P. hydriforme already existed there but re-
mained undiscovered. However, the parasite may
have been introduced into some rivers with the
transfer of acipenserids. Dogiel's work was fol-
lowed by numerous other findings of P. hydri-
forme in Eurasia and most recently in North
America in Acipenser fulvescens and Polyodon
spathula (Table I), indicating that this parasite
is not enzootic to the Ponto-Caspian basin and
is distributed in both families of extant acipen-
seriforms, Acipenseridae and Polyodontidae.
Polypodium hydriforme has not been found in

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 RAIKOVA-LIFECYCLEOF P. HYDRIFORME 3

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FIGURE 1. Life cycle of Polypodiumhydriforme.Parasiticstagesof the cycle are within the circle;free-living
ones are outside. I-XII are months of the year. A. Maturestolon with internaltentaclesinside the egg before
spawning.B. Stolon with externaltentaclesemergingfrom egg at the time of spawning.C. Free stolon in water.
D. Fragmentof a stolon. E. Twelve-tentacledspecimen. F. Twenty-four-tentacledspecimen. G. Six-tentacled
specimen.H. Twenty-four-tentacled specimenwith 4 "female"sexualcomplexes.I. Twelve-tentacledspecimen
with 4 "male"gonads. Question markat H indicates that fate of the animals with female gonads is unknown;
questionmark between I and J indicatesthat mode of oocyte infection is unknown.J. Binuclearparasiticcell
inside a young oocyte. K. Morulaencircledby a trophamnion.L. Planula(trophamnionnot shown here and at
any furtherparasiticstage).M. Buddingplanula.N. Stolon without tentacles.O. Stolon with internaltentacles.

the river Ob' where acipenserids are likely not
infected (Svirsky, 1984). Other Siberian rivers
have not been sampled in this respect. In western
European rivers, acipenserids are rare and not
commercially fished (Magnin, 1959); conse-
quently, no data on P. hydriforme are available.
The prevalence ofP. hydriforme in roe-bearing
females varies (Table I). The sterlet seems to be
the most extensively infected species, at a prev-
alence of 78% of mature females in the Volga
and the Kama rivers (Raikova, 1958a) and with
a maximum percentage of infected eggs per fe-
male of 100% (Persov, 1957). Anadromous stur-
geons investigated in this respect (Russian stur-
geon, sevrjuga, ship, and beluga) are appreciably
less infected.
Free-living specimens of P. hydriforme have
been observed in the Volga (Derzhavin, 1910;
Behning, 1913; Smolyanov and Raikova, 1961),
the Oka (Nalivkina and Neizvestnova, 1921;
Behning, 1922), the Dnieper (Jadin, 1940), and
the Don (Skljarova and Raikova, 1967) rivers.
LIFECYCLE
The life cycle of Polypodium hydriforme con-
sists of a parasitic and a free-living phase (Fig.
1). The parasitic stages occur in host oocytes
throughout oogenesis, beginning with early pre-
vitellogenesis up to spawning.
Parasiticstages
Unicellular and "cell-in-a-cell" stages: The
youngest stages of parasitic development, found
in previtellogenic oocytes ofsterlet (stage II, after

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4 THEJOURNALOF PARASITOLOGY,
VOL. 80, NO. 1, FEBRUARY1994

Nedoshivin [1928]), are single cells, 15-30 tm
in diameter (Fig. 2), with 2 unequally sized, Feul-
gen-positive nuclei measuring 9-23 tm and 2.5-
5 tm in diameter (Fig. 3) and cytoplasm filled
with round mucopolysaccharide protein gran-
ules. The volume of the larger nucleus exceeds
that of the other by at least a factor of 50. There-
after, the small nucleus becomes surrounded by
a portion of cytoplasm, and a small new cell is
formed with a clear halo around it that separates
it within the cytoplasm of the original cell (Fig.
4). Then, the large nucleus forms a depression
into which the small cell appears to "sink" (Figs.
1J, 5). Subsequently the pit of the large nucleus
(Fig. 6) closes (Fig. 7), securing the smaller cell
in a hollow space within it and giving rise to a
"cell-in-a-cell" stage.
Segmentation stages: Mitotic divisions of the
small cell, now enclosed in the hollow of the large
cell nucleus (Fig. 8), give rise to blastomeres,
indicating that the small cell is equivalent to the
egg or the generative cell. Its small size and lack
of reserve nutrients ensure that its division is not
hindered. The first blastomeres are ameboid, have
prominent nucleoli and grow in size in compar-
ison with the generative cell (Fig. 9). The large
cell, termed the trophamnion (Raikova, 1980),
has the form of a hollow sphere, and likely it
performs trophic and protective functions. At
early segmentation stages, its cytoplasm still
shows spherical mucoprotein inclusions (Fig. 9),
as during the unicellular stages (Figs. 5, 6). The
entire parasite lies in a parasitophorous vacuole
inside the host oocyte (Figs. 7, 10, 11).
As early as at the 8-celled stage, the blasto-
meres often form a dense aggregate (a morula)
without a blastocoel (Fig. 10). The embryo later
elongates, reaching 80 tm, and its outer cells
epithelialize (Fig. 11). At stages with more than
150 cells, the embryos form lacunae and clefts
within the mass of cells (Fig. 12). There seem to
be signs of gastrulation, similar to the model of
secondary (morular) delamination (Tardent,
1978). The inversion of the germ layers, typical
of the later parasitic stages of P. hydriforme, is
likely to occur during gastrulation of this type.
As the trophamnion grows, its nucleus becomes
a reticulate hollow sphere, appearing in tangen-
tial sections like fragments of a reticulum (Fig.
13). Its cytoplasm becomes differentiated into
loose endoplasm containing, as before, the mu-
coprotein inclusions of various sizes (Figs. 9, 10)
and a dense ectoplasm free of inclusions.
Photometry of Feulgen-stained nuclei has
shown that the small nucleus of the binucleate
cell stage (Fig. 3) is haploid (Raikova, 1965, 1987).
The large nucleus of the same cell undergoes
polyploidization and may contain 18-56C DNA
at this stage. At the 16-cell stage, the blastomeres
contain a near-haploid (1C) DNA quantity and
the trophamnion nucleus, about 400C of DNA
(Raikova, 1965, 1987). Later stages of embryo-
genesis (morula and gastrulation stages) proved
unsuitable for photometry because of a dense
packing of embryonic cells and the reticulate
trophamnion nucleus. Therefore, the moment
and manner of restoring diploidy (characteristic
of free-living P. hydriforme [Raikova, 1965]) re-
main unknown. It is possible that blastomere
fusion or endoreplication is involved.
Planuliform larva: The next stage of devel-
opment, the planuliform larva, with its epithelial
bilayer (Figs. 1L, M; 14), was found in oocytes
already accumulating yolk (stages II-III and III
after Nedoshivin's [1928] scale, see Table II).
This stage has been observed from late May to
late July (of the year prior to spawning). From
this stage onward, infected oocytes can be vi-
sually distinguished from healthy ones by their
darker pigmentation and larger size (in the ster-
let, 1.5-2.0 mm versus 1.3 mm [Raikova, 1958a,
1958b]). The darker color of infected eggs results
from the presence of relatively more yolk and
pigment beneath the envelope than in uninfected
oocytes. Beginning with this stage, the nucleus
of the infected oocyte becomes damaged due to
the parasite's presence (Raikova, 1963a).
In May, these larvae are spherical or ellipsoid;

FIGURES 2-7. Unicellularparasiticstages of the life cycle of Polypodiumhydriformein sterlet oocytes. 2.

Sterletoocyte showingnucleusand infection of unicellularstage of P. hydriforme.Bouin's,hemalum;bar = 10
Lm.3. Binucleateparasiticcell showinga largeand a smallnucleusin common cytoplasm.Bouin's,azanstaining;
bar = 10 pm. 4. Formation of small cell around the small nucleus (arrow).Bouin's, azan; bar = 10 am. 5.
Invaginationof the largenucleusclose to the small cell (arrow).Bouin's,hemalum;bar = 10 Lm.6. Migration
of the small cell into a cavity formed by the largenucleus. Navashin's, Feulgen-lightgreen;bar = 10 7.
/m.
"Cell-in-a-cell"stage;the egg is at arrow.Navashin's,Feulgen-lightgreen;bar = 10 Lm.OC, oocyte cytoplasm;
ON, oocyte nucleus;P, parasite;PV, parasitophorousvacuole; SN, small nucleus;T, trophamnion.

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 RAIKOVA-LIFECYCLEOF P. HYDRIFORME 5

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6 THEJOURNALOF PARASITOLOGY,
VOL. 80, NO. 1, FEBRUARY1994

in late July, they grow and start to form the first
buds (Fig. 1M).
Sections through infected oocytes show that
the planuliform larva lies in a cavity lined by the
trophamnion, which is in close contact with the
oocyte cytoplasm (Fig. 14) and forms outgrowths
that penetrate the latter (Raikova, 1980). There-
fore the parasitophorous vacuole is no longer
visible.
The outer cell layer of the larva may be con-
sidered as the endoderm, because its cells possess
flagella like the gastrodermis of free-living ani-
mals. The inner layer corresponds to the ecto-
derm because of its specific marker, the apical
acid mucopolysaccharide granules typical of epi-
dermal cells of the free-living stages. Therefore,
the planuliform larva already displays the in-
version of the germ layers characteristic of the
subsequent parasitic stages (Raikova, 1958b).
During the planuliform stage, the parasite grows
appreciably (from 0.5 mm to 2.5 mm in length).
It seems to feed via the surrounding tropham-
nion, which phagocytizes and digests yolk, hav-
ing lysosomes in its cytoplasm (Fig. 15). The cells
of the larva do not contain ingested yolk or fat
droplets (Raikova, 1958b, 1980).
Budding stolon: The planuliform larva elon-
gates, forms multiple buds, and becomes the sto-
lon (Fig. IN) in August of the year preceding
spawning, and it is found in older generation
oocytes (stage IY, Nedoshivin's [1928] scale). The
stolon grows and becomes convoluted inside the
fish oocyte. The buds divide longitudinally so
that by September of the same year stolons may
have 30-40 buds in sterlet oocytes and up to 100
buds in those of larger sturgeon.
The trophamnion surrounding the parasite
seems at this time within oocytes in full vitel-
logenesis, to be at the peak of its function (Fig.
15 [Raikova, 1984]). On its inner surface it has
numerous microvilli and is apparently involved
in exocytosis of soluble nutrients to the cells of
the parasite. Large and broad protrusions on the
outer side of the trophamnion ensure endocy-
tosis (phagocytosis) of portions of the host egg
cytoplasm. Primordia of the tentacles appear in
September (Fig. 10); due to germ layer inversion,
they are formed inside the buds. Initially, each
bud forms 2 pairs of short and sturdy supporting
(or walking) tentacles and, later, 4 pairs of longer
and thinner feeling tentacles. The tentacles ac-
quire nematocysts by the end of October (Rai-
kova, 1958a; Raikova et al., 1979).
Studies of winter oocytes from the Ural beluga
collected in January and of early spring oocytes
(from the Kama sterlet, the Ural sevrjuga and
beluga, and the American paddlefish collected in
March) have shown that the parasite apparently
ceases to develop during winter. The state of the
nucleus and the amount of yolk in infected oo-
cytes seemingly remain unchanged. From March
to May of the spawning year, the buds on the
stolon increase in size, the tentacles grow longer,
and the trophamnion is retained. The diameter
of the infected eggs also increases and reaches
4.0-4.5 mm in the sterlet (Raikova, 1958a). The
peripherally located stolon displaces the yolk to-
ward the center of the egg (Fig. 16) and appears
through the egg envelopes as a whitish mass al-
ternating with dark trabeculae of yolk, the in-
fected eggs acquiring the characteristic "marble"
design, which already is visible in autumn but
most clearly seen in spring (Fig. 17).
Everted stolon: Immediately prior to spawn-
ing, the stolon turns inside out, acquiring the
normal position of germ layers. All remaining
yolk accumulated in the egg is drawn into the
forming gastral cavities of the stolon buds, pro-
viding a food reserve for the initial period of life
of the free-living stages (Lipin, 191 la). The ever-
sion apparently is caused by a shift of the osmotic
equilibrium that occurs due to solubilization of
most of the glycogen and especially of the ma-
terial of the apical acid mucopolysaccharide
granules that line the stolon cavity and the inner
tentacles (Raikova, 1960). The strong rise of the
inner pressure, first noted by Lipin (1911 a), likely
is brought about by the extrusion of these gran-
ules from the cells and their swelling. As a result,
the stolon wall ruptures and it turns inside out
through the opening formed by this rupture.
In naturally maturing fish, the stolons are lib-

FIGURES8-11. Segmentationand morula stagesof Polypodiumhydriformeinside the sterlet eggs. Bouin's,

azan. 8. First segmentationmitosis of the egg inside trophamnioncell. Trophamnionnucleusis at arrowhead.
Bar = 10 tm. 9. Segmentingembryo (3 blastomeresare seen). Trophamnionnucleus at arrowhead.Bar = 10
tm.10. Fourteen-celledmorula(arrow)inside the cavity of the trophamnioncell. Bar = 10 Em.11. Multicellular
morula inside the trophamnionwith fragmentsof ramified nucleus (at arrows).Bar = 10 jm. OC, oocyte
cytoplasm;ON, oocyte nucleus;PV, parasitophorousvacuole; T, trophamnion.

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 RAIKOVA-LIFECYCLEOF P. HYDRIFORME 7

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8 VOL. 80, NO. 1, FEBRUARY1994
THEJOURNALOF PARASITOLOGY,
erated from eggs in the oviducts of spawning
females. Thus, normally, free stolons of P. hy-
driforme with outer tentacles, rather than intact
infected eggs, are released into the water along
with healthy eggs (Raikova, 1958a). In hypophy-
sized sterlet and sturgeon females, the everted
stolons usually remain inside the infected eggs,
which do not ovulate in most cases.
Free-living stages
The stolon carrying buds with outer tentacles
is a transition from the parasitic to the free-living
stages of the life cycle (Figs. IB, C; 18). The
reserve of yolk in the gastral cavities of the buds
is sufficient for the first 6-8 days of the free life.
Upon entering the water, the stolon fragments
into pieces with various numbers ofbuds (stellate
colonies with 4 or 8 buds are frequent) (Fig. iD).
On the third or fourth day, this fragmentation
forms single specimens whose numbers of ten-
tacles are multiples of 6 (Fig. 1E-G).
Single free-living animals: P. hydriforme with
24 tentacles initially predominate, whereas in
midsummer, 12-tentacled specimens are com-
mon (at least, in culture conditions). In late sum-
mer, organisms with only 6 tentacles or less are
found (Lipin, 1911 a; Raikova, 1961 a).
After depletion of the yolk within the gastral
cavity, each individual forms a mouth (Fig. 19)
at the place of the former attachment of the bud
to the stolon and begins to feed, capturing with
its feeling tentacles oligochaetes (Tubifex), tur-
bellarians, and rotifers (Lipin, 191 la, 1915; Rai-
kova, 1958a).
Free-living animals reproduce by longitudinal
binary fission beginning at the aboral pole. Be-
fore division, 2 new sets of 6 tentacles each (2
walking and 4 feeling) are laid down near the
center of the aboral body surface, and the divi-
sion furrow forms between them (Fig. 19).
The free-living animals move about with the
aid of walking (supporting) tentacles. They can
be homologized with the medusoid generation
(sexual and settling form) of coelenterates
(Shimkevitch, 1890) but lack an umbrella and
sensory organs (Fig. 19).
Free-living forms with gonads: Two types of
gonads are formed during summer in the free-
living animals (Fig. 1H, I), those formerly con-
sidered as "female" and "male" (Lipin, 1915;
Raikova, 1961b, 1963b).
The female sexual complexes (Figs. 20, 21)
usually form at the base of the tentacles in late
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June. At first, gonoducts are formed, then go-
nads. These gonads may be resorbed, especially
at early stages of their formation and may be laid
down 2 or 3 times in succession in the same
specimen. These are compound endodermal or-
gans, within a common envelope, consisting of
2 "ovaries," each with its own "oviduct" open-
ing into the gastral cavity (Lipin, 1915). Each
ovary contains free round cells emerging into its
lumen from the generative epithelium and poly-
ploid glandular cells (up to 37C of DNA [Rai-
kova, 1965]), producing a mucopolysaccharide
material by apocrine secretion, i.e., by separation
of the apical ends of the cells into the lumen (Figs.
20, 21). Later, generative cells and the muco-
polysaccharide material both seem to be trans-
ported via the oviducts into the gastral cavity.
However, neither Lipin (1915, 1925) nor Rai-
kova (1963b) observed meiotic phenomena in
generative cells. They are diploid according to
their DNA content (Raikova, 1965). Free-living
P. hydriforme captured in nature (from the Don
River) also showed no advanced stage of repro-
ductive cells in these female gonads (Skljarova
and Raikova, 1967). Both in cultures and in na-
ture, degeneration of the entire female sexual
complex begins after the stage with diploid sex
cells is reached. It is therefore unknown whether
the female sex cells mature.
It was observed in cultured P. hydriforme that
male gonads appear after female ones; they are
more stable and once formed rarely degenerate.
These formations also display an invagination of
the endoderm but lack ducts (Fig. 22). They may
appear both in individuals with female gonads
("hermaphroditic" specimens) and in individu-
als without gonads (male specimens). The latter
usually are smaller than the former and cease to
reproduce by binary fission (Raikova, 1961 b).
In August, the male gonads reach maximum
size and often protrude from the animal's body.
Such gonads (gametophores after Raikova [198 5,
1987]) can also separate from the animal (by
rupturing of the body wall), after which the in-
dividual dies (Raikova, 1961 b).
The course of gametogenesis in these gonads
(Lipin, 1915, 1925; Raikova, 1961b) includes
synchronous meiosis in sex cells, which have en-
dodermal origin, unequal cytokinesis of meiosis
I resembling polar body formation, and the lack
of cytokinesis of meiosis II leading to formation
of binucleate cells. There is also an ectodermal
lid with nematocysts that stoppers the mature
gonad filled with binucleate cells (Figs. 23, 24).
 RAIKOVA-LIFECYCLEOF P. HYDRIFORME 9

TABLEII. Correspondence between parasite devel-

opmental stages and stages of sterlet oogenesis.

Infected sterlet oocytes

Stages of
Diameter parasite Season of
(sm) Stage development collection

38-100 Previtellogenesis, Binucleate cell May-August

60-170 slow growth (stage Separation of the
II*) generative cell
85-220 "Sinking" of the
generative cell
120-200 "Cell-in-a-cell"
160-350 Previtellogenesis, 2-8 blastomeres May
270-400 rapid growth 12-56 blasto- May-July
meres, morula
420-690 Early vitellogenesis, 64-576 cells,
rapid growth gastrulation
1,000-1,500 Vitellogenesis, rapid Planula May-July
growth (stage III*)
3,000 Vitellogenesis, rapid Stolon with August
growth (stage IV*) buds; without
tentacles
3,500 Vitellogenesis, rapid Stolon with buds September
growth containing in-
ternal tentacles
4,500 Metaphase II, Stolon with buds May
spawning (stage bearing exter-
V*) nal tentacles

* Stages according to scale of Nedoshivin (1928).

Especially interesting were the collections of

free-living sexual P. hydriforme from the Volga
River (Smolyanov and Raikova, 1961). Individ-
uals were captured in the process of depositing

* -

FIGURES12, 13. Multicellular embryo of Polypo-

dium hydriforme at gastrulation stage. Bouin's, azan.
12. Embryo with cleft (arrow) between the cells. The
parasite is adjacent to the host oocyte nucleus. Bar =
20 tm. 13. Tangential section through the trophamnion
showing reticulate structure of its nucleus (arrowhead).
Bar = 20 tm. OC, oocyte cytoplasm; ON, oocyte nu-
cleus; T, trophamnion nucleus.

Observations made on cultured animals were
confirmed by examination of naturally occurring
free-living P. hydriforme captured in the Don
River (Skljarova and Raikova, 1967).
FIGURE14. Planula stage inside vitellogenic sterlet
oocyte. Bouin's, iron hematoxylin--eosin. Bar = 0.5
mm. OC, oocyte cytoplasm; ON, oocyte nucleus; P,
parasite; PC, second parasitophorous cavity; Y, yolk
of the sterlet egg.

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10 VOL 80, NO 1, FEBRUARY1994
THE JOURNALOF PARASITOLOGY,

;c."~' ...

'4
?np'd 4.i;

~4~,Ivv~' ?

FIGURE15. Trophamnion in infected sterlet oocyte in October preceding spawning. Bar = 2 rm. En, en-
doderm; FI, food inclusions in the trophamnion; Lys, lysosomes; Mv, microvilli; N, nucleus of the trophamnion;
OC, oocyte cytoplasm; OT, outgrowths of the trophamnion into oocyte's cytoplasm; PC, second parasitophorous
cavity; S, stolon; T, trophamnion, Y, yolk of sterlet egg.

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 RAIKOVA-LIFECYCLEOF P. HYDRIFORME 11

ii

~ ~ /1

FIGURE16. Infected beluga oocyte in early March.

Bouin's, azan. Bar = 0.5 mm. ON, oocyte nucleus; S,
stolon, Te, tentacles; Y, yolk of the egg.

their gonads (gametophores) onto the bodies of

prelarvae of A. stellatus (Fig. 25), and these sam-
ples also contained prelarvae with gametophores
of P. hydrnforme already attached to the yolk sac,
head, body, tail, or fin fold. The deposited for-
mations were filled entirely with binucleate cells
similar to those occurring in mature male gonads
of cultured P. hydriforme (Fig. 26). The ecto-
dermal lid of the gonad was disintegrated, and
the nematocysts contained in it were discharged,
helping the gonads attach to the more or less
undamaged skin tissue of the larva (Fig. 26). FIGURE17. Infected sterlet eggs (4 bigger ones)
among normal prespawningeggs. Bar = 5 mm.
The fate of these binucleate cells remains un-
clear. The next known stage is the previously
described binucleate cell starting the parasitic with or without male and/or female gonads, were
phase of the life cycle inside young fish oocytes not successful (Raikova, unpubl. data) as P. hy-
(Figs. 2, 3). This cell differs from the binucleate dnforme individuals did not attach to any part
cells inside the gametophore mainly in its larger of the fish body; neither were they swallowed by
size and highly polyploid large nucleus. Conse- the fish. Following deliberate deposition of P.
quently, in the cytological sense, the life cycle is hydriforme onto sterlets they occasionally at-
complete, from binucleate cells inside the gonads tached to the fins with their tentacles, but for no
of free-living P. hydriforme to those formed in more than 10
min. Polypodium hydriforme spec-
oocytes of the host. imens without gonads do not react on prelarvae
of the sturgeon (Raikova, unpubl. obs.). These
Attempts at artificial infection observations tend to confirm the supposition that
Attempts made to infect 15-17-cm-long ster- infection of acipenserids may be brought about
lets with free-living P. hydriforme from cultures, during early life, when their skin is more delicate

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12 THEJOURNALOF PARASITOLOGY,
VOL.80, NO. 1, FEBRUARY1994

and accessible to the action of nematocysts of P.

hydriforme. However, the reaction of gameto-
phore-carrying specimens to prelarvae of fish re-
mains to be tested experimentally.

GENERALCONSIDERATIONS
Correlationbetween parasite development and
oogenesis in acipenserids
The occurrence of certain stages of develop-
ment of P. hydriforme demonstrates a definite
correlation with the diameter of infected (and
uninfected) fish oocytes (Table II). This has been
shown in the sterlet (Raikova, 1958a, 1987),
Russian sturgeon (Raikova, 1959), American
paddlefish (Raikova et al., 1979), schip, sevrjuga,
and beluga (Raikova, 1986). Binucleate cells oc-
cur in sterlet oocytes in the "slow growth" phase.
Their nuclei clearly display diplotene bivalents,
not yet transformed into typical lampbrush chro-
mosomes, and peripheral nucleoli that have not
yet reached the stage of amphinucleoli. Of the
nutrient reserves, only a few fat droplets and
glycogen are found in the cytoplasm (Fig. 2). Such
cells belong to the junior generation of oocytes
in the sterlet ovary, i.e., to that population of
cells that do not accumulate yolk and, conse-
quently, will not mature for the next spawning.
The cell-in-a-cell stage corresponds to the end of
the slow growth. Segmentation begins when oo-
cytes enter the period of "rapid growth" (average
size 200 jm, lampbrush chromosomes appear),
and, sometimes, segmentation stages were found
in the same sterlet females that contained uni-
cellular parasitic stages in smaller oocytes (Rai-
kova, 1987). Stages (from 64 cells on) undergo
gastrulation and are likely to be of short duration
because the larger oocytes of the same females,
having more yolk, already contain planuliform
larvae (Fig. 14). The planula forms when yolk
fills the oocyte cytoplasm, the stolon when vi-
tellogenesis is at its maximum, and eversion takes
place when host oocytes undergo meiosis. Rai-
kova (1960) demonstrated that the eversion of
the stolon is synchronized with meiotic divisions FIGURES18, 19. Free-living Polypodium hydri-
of maturing uninfected oocytes, that this process forme. 18. Stolon just emerged from an infected egg
can be triggered artificially, e.g., with the help of into water. Bar = 5 mm. 19. Twelve-tentacled speci-
men with 2 sets of 6 tentacles prior to binary fission.
hypophysal injection, and that no meiotic divi- Bar = 2 mm. Arrow points to walking tentacles, ar-
sion occurs in infected oocytes. rowhead points to the mouth.
Thus, the parasitic phase of the life cycle can
last more than a year. If the parasite infects a fish
at the larval stage, this phase of the cycle must but in 2 females 5 oocytes that contained 2 par-
last almost 10 yr (or more), the time of devel- asites, each at identical stages of development,
opment of most acipenserids to maturity. were found (in 4 cases these were binucleate cells
Usually only 1 parasite occurs in an oocyte, and in 1 case they were the cell-in-a-cell stage).

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 RAIKOVA-LIFECYCLEOF P. HYDRIFORME 13

At later stages of parasite development, no dou-

ble infection was encountered (Raikova, 1987).
Although the nucleus of the host oocyte is
damaged (Raikova, 1963a), an infected oocyte
va accumulates yolk (essential for the survival of
the parasite) and is under hormonal control of
the fish (i.e., reacts on hypophysal injection).
Peculiarities of embryogenesis
Binucleate cells probably constitute the longest
stage of development of P. hydriforme (Raikova,
1987). They likely penetrate, in some unknown
way, a very small oocyte (8 tm in diameter; Ta-
ble II) and seem to remain unchanged for several
years, until the respective oocyte enters a critical
, phase, that of rapid growth. Then, the parasitic
:Oo cell undergoes endocytokinesis, and the genera-
tive cell thus formed develops to a morula. An-
other likely critical phase is when it is determined
whether the host oocyte will start vitellogenesis
(and join the senior generation of oocytes due to
be released the following spring) or whether it
will remain blocked in previtellogenesis as part
of the junior generation of oocytes destined for
the next spawning batch of eggs. The parasite
morula would develop further only in the former
case. This suggestion is supported by the occur-
GCNC rence, in the same sterlet female, of oocytes in-
fected with different sized morulas and the em-
bryos undergoing gastrulation. The latter oocytes
contain the first bits of yolk, i.e., they are entering
the senior generation. Differentiation of oocytes
into senior and junior generations occurs in fe-
males both maturing for the first time and re-
maturing after spawning (II and YI-II stages after
Nedoshivin [1928]). In the latter case, the inter-
val between 2 consecutive spawnings is 4-5 yr
in the sterlet (Shilov, 1964). Therefore it is pos-
sible that the morula stage of the parasite also
a long-lasting one. i,
~Oo 4 Since 1-celled and segmentation stages of the
parasite frequently occur in different sized oo-
cytes of the same female, it appears that fish
either are heavily infected or uninfected. This
may mean that infection of the host's oocytes
occurs simultaneously with a rather large num-
ber of parasites (probably binucleate cells). Dif-

" MI1'
.4 -? .,,
'
1
,.-2
FIGURES 20, 21. Sections through the "female" go- Ec, ectodermal layer of the body; En, entodermal layer
nad. 20. "Ovarium" opening into "oviduct." Bouin's, of the animal; GC, gastral cavity; GE, germinal epi-
iron hematoxylin; Bar = 20 jm. 21. Ovarium with its thelium; M, mesogleal space within the gonad; Od,
generative and nurse cells (NC). Zenker's formol, Feul- "oviduct"; Oo, "oogonia"; Ov, "ovarium." Arrow-
gen-light green. Bar = 10 tm. E, envelope of the gonad; head points to the beginning of the oviduct.

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14 VOL 80, NO 1, FEBRUARY1994
THE JOURNALOF PARASITOLOGY,

.,e

i$*

?,

~; .s ?-
M !$
?~

FIGURES22-24. Sections through the "male" gonads. Zenker's formal. 22. Male gonad at the beginning of
gametogenesis. Unna staining; bar = 20 ~m. 23. Male gonad at the end of gametogenesis filled with binucleate
cells and supplied with an ectodermal plate containing nematocysts. Hemalum; bar = 40 ~m. 24. Binucleate
cells with different sized nuclei. Hemalum; bar = 10 jm. BC, binucleate cells; Ec, ectoderm; EcP, ectodermal
plate with nematocysts (at arrow); En, entoderm; G, gonial cells; GC, gastral cavity; GLC, glandular cell; M,
mesogleal space within the gonad.

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.26
..
FIGURES25, 26. Mature "male" gonads in free-
living Polypodium hydriforme. 25. Free-living form de-
positing its male gonads (gametophores) through a body
wall opening (arrow) on the tail of a larva of Acipenser
stellatus (arrowhead); bar = 1 mm. 26. Section through
male gametophore deposited on the surface of the yolk
sac of a larva ofA. stellatus. Arrow points to discharged
nematocysts of the ectodermal operculum. Formol, iron
hematoxylin; bar = 100 gm. BC, binucleate cells; Ec,
ectoderm of the animal body; En, remnants of the en-
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RAIKOVA-LIFECYCLEOF P. HYDRIFORME 15
ferentiation of the most mature oocyte popula-
tion occurs only much later, with the start of
vitellogenesis (stages II-III, Nedoshivin's [1928]
scale) bringing about a differentiation of the par-
asite population into respective groups.
The embryonic development of P. hydriforme
is probably the slowest among all coelenterates.
In contrast, segmentation in free-living coelen-
terates usually takes only hours, and develop-
ment of larva takes only 1 or 2 days (Metschni-
koff, 1886; Van de Vyver, 1967, 1968). The egg
of P. hydrnforme does not undergo splitting as
strictly defined; its blastomeres grow (probably
just because of slow embryogenesis), and the vol-
ume of a multicellular embryo is much greater
than that of the original egg (cf. Figs. 7, 10).
The formation of the small (generative) cell
inside the cytoplasm of the large cell by endo-
cytokinesis corresponds to the postponed cyto-
kinesis of the second meiotic division. In other
words, the initial binucleate cell, which may be
termed "pregamete" or "pre-egg," divides into
the egg (generative small cell) and the vegetative
cell (the trophamnion). The latter, consequently,
corresponds to a hypertrophied second polar body
of meiosis.
The initial segmenting cell in P. hydrnforme,
the homolog of the egg, is 4 times smaller (4-6
tm) than the egg of members of the genus Cunina
(the smallest known eggs among cnidarians [Tar-
dent, 1978]), which measures 24 tm (Metschni-
koff, 1886). This cell lacks elements of deuto-
plasm. For that, it is enclosed within another
amoeboid cell that has functions of a phorocyte
and a nutritive cell. Its trophic function becomes
particularly evident later, when it ingests yolk of
the host oocyte (Raikova, 1958b, 1960, 1980,
1984).
Endocytokinesis in itself is a protozoan fea-
ture. Comparable patterns of endogenous cell di-
vision have been observed during sporulation in
Paramyxa paradoxa, a parasite of polychaetes
of the genus Poecilochaetus (Desportes, 1981).
This division also leads to the position of 1
daughter cell inside the other.
Functioning of a meiotic polar body during
embryogenesis is a rare phenomenon that also
exists in larval-parasitic hymenopterans (Iva-
nova-Kazas, 1961, 1972; Kos~ielski et al., 1978),
todermal envelope of the male gonad; S, skin of the
larva; Y, yolk of larva.
16 VOL.80, NO. 1, FEBRUARY1994
THEJOURNALOF PARASITOLOGY,

although in these it is formed by fusion of 3 polar TABLEIII. Characteristics of type I and II gonads of
bodies (the product of the fusion later undergoes free-living Polypodium hydriforme.
polyploidization, too). The block of cytokinesis I ("female") gonads II ("male") gonads
during meiosis II in P. hydriforme is probably
also an important adaptation to parasitism and Origin Endodermal Endodermal
Organs Ovaria and gonoducts Endodermal fold
is not typical of coelenterates, where meiosis usu- Origin of sex Generative epithelium of Endodermal cells
ally proceeds without interruption (Masui and cells the ovarium
Differentiation Synchronous
Clark, 1979). It would allow the binucleate cells, of sex cells
1-by-1

especially if they infect young sturgeon larvae, to Nurse cells Highly polyploid, with Small cup-shaped, phag-
remain in an arrested stage of development for apocrine secretion ocytized by sex cell
Meiosis I Not observed Unequal division with
years and undergo endocytokinesis 1 or 2 yr be- polar body formation
fore first spawning. This may be at the age of 6- Meiosis II Not observed Binucleate cells with un-
10 yr in sterlet, and not before 16 yr in the beluga equal sized nuclei
Ploidy of sex Diploid Haploid and polyploid
(Berg, 1948a). cells nuclei in binucleate
With the trophamnion existing as atrophic cell cell
Fate of gonads Gametophores, may be
throughout the parasitic phase of the P. hydri- Resorption (?)
attached to prelarvae
forme cycle it seems that inversion of the germ of sturgeon
layers is not an important adaptation for the up- Fate of sex cells May leave gonads via Give rise to future para-
gonoducts entering site generation
take of oocyte nutrients by endodermal cells fac- gastral cavity; further
ing yolk as suggested since Lipin's work (1911 a). fate unknown
More likely it is an adaptation for supplying the
future free-living generation with yolk until the
free individuals are able to capture food.
The establishment of haploidy of both the small and can be resorbed. A new generation of P.
nucleus of the binucleate cell and the blastomere hydriforme develops from cells produced by type
nuclei in early segmentation proves that no fer- II (male) gonads (Table III).
tilization occurs in P. hydriforme (Raikova, 1965) Type II gonads have features indicative of a
and that haploidy may last during the entire uni- male gonad and spermatogenesis, such as sim-
cellular stage and early multicellular stages, i.e., pler organization (absence of gonoducts), pres-
several years until diploidy is restored at, or after, ence of many cells that undergo synchronous
segmentation (Raikova, 1985). This type of hap- meiosis without a period of growth, and lack of
lophase in the life cycle can be compared to that special polyploid nutritive cells, although small
occurring in Myxosporidia (Uspenskaya, 1982). caplike cells occur adjacent to meiocytes, which
are later phagocytized by these meiocytes. The
sex cells themselves lack yolk or other nutritive
Sex of the gonads and parthenogenesis substances. Moreover, these gonads are more
It seems that the small generative cell develops stable and their existence precludes vegetative
parthenogenetically, being surrounded by the reproduction. However, their homology with
trophamnion cell. The binucleate cells produced ovaries and oogenesis is favored because the first
in the male gonad of the free-living P. hydriforme meiotic division of the gonocytes is unequal and
correspond to meiocytes II (Lipin, 1915; Rai- produces 1 functional cell and 1 polar body and
kova, 1961 b). This is a case of natural andro- the second meiotic division of the gonocytes is
genesis because the new generation develops from acytokinetic, so the number of cells does not in-
a male generative cell without fertilization. Sa- crease as a result of meiosis (Table III).
lient features of the 2 types of gonads and the Nematocysts in the egg envelopes, as found in
cytological processes in them are outlined in Ta- gametophores of P. hydriforme, have been de-
ble III. scribed in species of Nausitoe (Metschnikoff,
Type I gonads have been recognized as female 1886) and in overwintering eggs of the antho-
because they have gonoducts (Fig. 20), the gono- medusa Margelopsis haeckeli (which also repro-
cytes are produced there 1-by-1, and the gonads duces parthenogenetically). Nematocysts (gluti-
have nutritive cells with polyploid nuclei (Rai- nants) ensure the attachment of the egg to a
kova, 1963b, 1965) (Fig. 21). These gonocytes, substrate (Werner, 1956) as in the case of the
however, do not undergo meiosis and are ex- "armed egg" of the hydromedusa Bougainvillia
truded as diploid oogonia. The gonads are labile multitentaculata (Sz6llosi, 1969). On the other

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 RAIKOVA-LIFECYCLEOF P. HYDRIFORME 17

and the small nucleus, respectively, in P. hy-

hand, simultaneous arrival of many sex cells at
the same place is brought about by spermato- driforme.
phores that may also have nematocysts, e.g., in
the Cubomedusae (Werner, 1973). Life cycle analysis
Thus, early phenomena in type II gonads re- The life cycle of P. hydriforme is an alternation
semble spermatogenesis, whereas later processes of free-living sexual and parasitic asexual gen-
resemble oogenesis. The organization of the ga- erations (metagenesis). The parasitic generation
metophores has features of both. reproduces by budding followed by longitudinal
Lipin (1925) reevaluated the function of type fission; the free-living one reproduces both sex-
II gonads and termed them "enigmatic forma- ually and asexually (by longitudinal fission or
tions"; he concluded that the male sex in P. hy- paratomy).
driforme was reduced. Based on the fate of the Free-living specimens of P. hydrnforme,being
binucleate cells, type II gonads must be consid- unattached animals, can also be considered as a
ered secondarily female (Raikova, 1985). It may dispersal stage. As they are also sexual, they cor-
be hypothesized that ancestors of P. hydrnforme respond not to polyps but to the medusae of other
were bisexual free-living animals. In the course cnidarians. However, they lack the umbrella and
of evolution, and due to adaptation to parasit- the sense organs. This can be explained by sec-
ism, gonads that were initially male (type II) ondary reduction, perhaps under the effect of the
changed to female. Simultaneously, the female parasitic mode of existence of the previous stage
gonads (type I) ceased to produce mature eggs where these future medusae are formed. Parato-
and became rudimentary, with possibly some my seems to be an adaptation that ensures an
other function (endocrine?). Due to the lack of increase in the number of free-living medusae in
true spermatozoa, development became second- the water basin and thus the probability of in-
arily parthenogenetic, derived from andro- fection of new host individuals. The inverted
genesis, leading to gametophory. This highly stolon with buds then corresponds to the poly-
aberrant feature in the development of P. ploid generation. The budding of the stolon en-
hydrnforme is unknown in other animals in sures that each binucleate cell eventually pro-
natural (not experimental) conditions (Raikova, duces 40-100 free-living medusoid animals.
1985). However, it has been suggested that the free-
Parasitism favored simpler reproductive or- living stages are polyps, rather than medusae,
gans producing more germ cells as well as par- that acquired the ability of larval reproduction
thenogenesis (androgenesis) and gametophory resembling polyembryony (Ivanova-Kazas,
because it eliminated the problem of meeting of 1977).
gametes existing during bisexual reproduction.
The initially male gonads proved more suitable Possible modes of infection of the host oocytes
to follow these tendencies. Although gametophores with binucleate cells
may attach to the body of prelarval fish (Smolya-
nov and Raikova, 1961), it is not known where
Atypical gametes these cells remain in the body of the host before
The gametes of P. hydrnforme displaying nu- oogenesis begins and how they enter young oo-
clear dualism are atypical when compared with cytes. In sections of ovaries of young sterlet fe-
those of other coelenterates. These cells do not males, cells resembling the binucleate ones were
resemble cnidarian eggs or spermatozoa (Tar- repeatedly found. They occurred either in the
dent, 1978). However, they have some pro- germ epithelium of the ovary or in small
tozoan features. Their nuclear dimorphism re- blood vessels, but the second (small) nucleus
sembles that found in ciliates and some could not be distinguished (Raikova, unpub-
foraminiferans, where a generative nucleus lished obs.).
(micronucleus) coexists with a vegetative nu- Several questions regarding the mode of in-
cleus (macronucleus) within the same cell (Rai- fection remain: Can the parasite infect older stur-
kov, 1982). Also, the micronucleus usually re- geon or the same fish more than once? Can the
tains the degree of ploidy, whereas the parasites infect both future females and males?
macronucleus frequently becomes polyploid. Do intermediate hosts (e.g., tubificids) exist? Can
In ciliates, the macronucleus frequently forms gametophores remain dormant in water or even
a depression for the micronucleus, like the large overwinter?

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18 VOL. 80, NO. 1 FEBRUARY1994
THEJOURNALOF PARASITOLOGY,
Systematic position of Polypodium
Presently most authors relate Polypodium to
the suborder Narcomedusae (Hyman, 1940; Ber-
rill, 1950; Bouillon, 1981, 1987; Werner, 1984),
a hypothesis that can be traced to Shimkevitch
(1890). The systematic position is related to the
question of the life cycle being a case of meta-
genesis. Raikova (1988) considered that the life
cycle of P. hydriforme is nearest to that of par-
asitic narcomedusans (order Trachylida, class
Hydrozoa, see Werner [1984]). Developmental
features common to P. hydriforme and the nar-
comedusans are the early differentiation of the
developing egg into a generative cell and phoro-
cyte, which occurs in Narcomedusae during seg-
mentation at the stage of 2 blastomeres (Pegan-
tha [Bigelow, 1909, in Hyman, 1940], Cunina
[Stschelkanowzew, 1906]); the resemblance in
segmentation (leading, in both cases, to a morula)
and gastrulation (by morular delamination
[Bouillon, 1987]); the similar function of the
phorocyte during embryogenesis, in members of
Cunina the phorocyte fixes itself to the endoderm
of the mother organism and has trophic function
(Stschelkanowzew, 1906); the similar change of
the body axes, in Narcomedusae the axis of the
planula does not become the body axis of the
actinula, which is in fact perpendicular to the
former (Bouillon, 1987) like in P. hydriforme
where the axes of the buds (the future medusae)
are at right angles to the primary axis of the
planula; and the similarity in disposition and
structure of the tentacles, narcomedusan tenta-
cles having aboral position and a dense endo-
dermal axis (without a lumen) and carrying a
single type of nematocyst (atrichous isorhiza), as
in P. hydriforme.
There is also evidence against the homologi-
zation of Polypodium with Narcomedusae. First,
Polypodium has unique features distinguishing it
not only from the Narcomedusae but also from
all Hydrozoa. These are the general plan (bilat-
eral symmetry); the occurrence of longitudinal
fission (paratomy), starting at the aboral pole;
the presence of gonoducts; the cytology of ga-
metogenesis; and the inversion of the germ layers
during the parasitic phase of the life cycle. The
nonflagellated (or nonciliated) type of ectoder-
mal cells, the complete separation of epidermal
and muscle cells, as in Cubozoa (see Werner,
1973), and the structure of the cnidocil apparatus
of the nematocytes (see Raikova, 1990) are also
peculiar characteristics. In addition, there are zo-
ogeographical considerations against relating
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Polypodium to the narcomedusans: P. hydri-
forme is clearly a freshwater cnidarian and infects
mostly freshwater acipenserids (especially the
sterlet, the kaluga, the lake sturgeon, and the pad-
dlefish [Raikova et al., 1979]), whereas all nar-
comedusans are marine, avoiding even brackish
water; no representative of the suborder has been
found in the Caspian sea (Naumov, 1960). The
relation to the narcomedusans, if real, must be
very ancient. Based mainly on the unique fea-
tures of Polypodium, it probably should be at-
tributed to a separate class of Cnidaria, Poly-
podiozoa (Raikova, 1973, 1988).
Originof Polypodium
Schulman (1954) noted that the parasites of
Acipenseridae had low host specificity within this
family. In fact, there is no parasite species specific
for even 1 genus (Acipenser or Huso), to say noth-
ing of the different species. This seems to be true
also for Polypodium. The parasitic stages of
members of Polypodium from the eggs of fishes
of the genera Acipenser, Huso, and Polyodon are
morphologically indistinguishable (Raikova et
al., 1979; Raikova, 1986). Probably, 1 species of
Polypodium, P. hydriforme, occurs in hosts of all
3 genera and in both Eurasia and North America.
The possible existence of sibling species needs to
be investigated by modern methods (molecular,
immunological, etc.).
The occurrence of similar forms of Polypo-
dium in Acipenseriformes of both the Old and
the New worlds argues for the great antiquity of
this parasite, at least at the generic level. Ac-
cording to Berg (1948b, 1955), the families Aci-
penseridae and Polyodontidae exist from the up-
per Cretaceous Period. The theory of a common
origin of Acipenseridae and Polyodontidae of the
Old and New worlds has been substantiated both
paleontologically (Yakovlev, 1977), karyologi-
cally (Vassiljev, 1985), and parasitologically with
respect to Monogenea, Amphilina, and Cystoop-
sis (Bauer and Gussev, 1969; Gussev, 1969,
1978). Consequently, Polypodium must have
evolved before the separation of North Ameri-
can from Eurasia, and its members must have
acquired a parasitic mode of life even before that,
probably with the origin of Acipenseriformes.
Afterward, it must have adapted to fresh water,
because presently the main hosts are freshwater
species. Later, there was hardly any divergence
of Polypodium at even the species level. Possibly,
Polypodium has some common ancestor with
Hydrozoa, namely, with Narcomedusae, which

is the most primitive taxon within this class
(Bouillon, 1981).
ACKNOWLEDGMENTS
I am very grateful to Terry Dick for arranging
my visit to Canada in 1990 through a grant from
the Natural Sciences and Engineering Research
Council of Canada. I thank Lu MingChuan for
his patient and tireless help in the use of the
computer and printing of photographs. I also
thank Anindo Choudhury for his generous help
in editing the manuscript and arranging illustra-
tions as well as for other valuable suggestions. I
am thankful to Alexander Karpov and Ilya Sko-
vorodkin for helping me use another computer
program during review of this manuscript in St.
Petersburg.
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