ПАРАЗИТОЛОГИЯ, 2022, том 56, № 6, с. 443-459.
УДК 595.122.2:594.32
GUIDE TO NOTOCOTYLIDAE (DIGENEA)
PARASITIZING COASTAL GASTROPODS
OF THE WHITE AND BARENTS SEAS
© 2022 A. Gonchara, b, *, K. V. Galaktionovb, a
aSt Petersburg University, Universitetskaya emb., 7/9, St Petersburg, 199034 Russia
bZoological Institute of the Russian Academy of Sciences,
Universitetskaya emb., 1, St Petersburg, 199034 Russia
*e-mail: anya.gonchar@gmail.com
Received 28.10.2022
Received in revised form 24.11.2022
Accepted 01.12.2022
Recent studies on the digenean fauna at the White and Barents Seas have shown that there are
more species than previously thought. These data are emerging in a series of publications, and we
suggest to summarize them in a form that is convenient for practical use. Here we provide a guide
that covers the 11 species from the family Notocotylidae that we have recorded in the intertidal
gastropods Ecrobia ventrosa, Peringia ulvae, Littorina spp. and Onoba aculeus. We recap brief de-
scriptions of rediae and cercariae, documented host and geographic range, though for several species
the information is incomplete. We also refer to the DNA barcodes from GenBank, including the new
ones. For the better usability, we include hints on the mollusc identification and explain how to deal
with the parasites (field and lab procedures).
Keywords: Notocotylidae, Digenea, cercariae, rediae, intermediate hosts, DNA barcodes
DOI: 10.31857/S0031184722060011; EDN: FHLCKE
Gastropods at the White and Barents Seas serve as the first intermediate hosts for
a variety of digeneans. Many of these parasites have been subject to at least some research,
but the actual set of species within each family of Digenea in this region is still unclear.
Recent integrative taxonomy studies have revealed diversity that had been hidden before,
for example, in Brachycladiidae (Kremnev et al., 2020), Fellodistomidae (Krupenko et al.,
2020), Himasthlidae (Galaktionov et al., 2021), Derogenidae (Krupenko et al., 2022), Reni-
colidae (Galaktionov et al., 2022), and a few other families. These data are emerging in
a number of publications that are not so easy to follow. At the same time, the correct spe-
cies identification of the digeneans’ intramolluscan stages is a key to any type of research
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on this material. That is why we take up a task to summarize the available data and compile
a series of guides. This paper deals with the representatives of the family Notocotylidae.
Notocotylidae (Pronocephaloidea) are digeneans with a two-host life cycle. Their
maritae (sexual adults) infect birds and, to a lesser extent, mammals. The eggs get into
the environment, but there is no free-swimming miracidium. The first intermediate host,
a gastropod, gets infected after accidental ingestion of eggs. In the gastropod host, rediae
develop and reproduce, and eventually cercariae get formed. Cercariae of most notocotylids
have a tail and leave the mollusc. There is no second intermediate host, and the definitive
host gets infected by consuming metacercariae that had encysted on some object in the
water.
Until recently, life cycles of two species of notocotylids were known at the White and
Barents Seas. For Parapronocephalum symmetricum Belopolskaja, 1952 the life cycle was
part of the original species description; for Paramonostomum alveatum (Mehlis in Creplin,
1846) Lühe, 1909 the intermediate host was discovered later (Kulachkova, 1954). However,
more species were actually recorded (Chubrik, 1966; Podlipaev, 1979), at least in Littorina
spp. and Onoba aculeus (A. Gould, 1841). Employing the molecular genetic data, we have
preliminary estimated that at least 11 species of Notocotylidae occur in Littorina spp.,
Peringia ulvae (Pennant, 1777), Ecrobia ventrosa (Montagu, 1803) and O. aculeus at the
White and Barents Seas.
This guide includes the overview of the mollusc identification, sampling and research
procedures; and the information on all the notocotylid species that we found.
MATERIALS AND METHODS
The guide was compiled from the results of our own research conducted in 2002-2022, both
published and unpublished. We collected most of our samples at the White Sea (various locations in
the vicinity of Keret Archipelago, Chupa Inlet, Kandalaksha Bay); Barents Sea (Western and Eastern
Murman, Pechora Sea); and in Iceland. We screened at least 20 thousands of molluscs (E. ventrosa,
P. ulvae, Littorina spp. and O. aculeus) for infection with the Notocotylidae. We also acknowledge
relevant earlier research on the Notocotylidae at the White and Barents Seas, as well as in the North
Atlantic, commenting on its validation.
We open the guide with some tips on the correct identification of the gastropod intermediate
host species. Next, we provide the description of the typical procedures to collect and identify noto-
cotylid intramolluscan stages (according to our research experience). It is followed by the summary
of identification characters that allow distinction of the Notocotylidae from other digeneans that are
common in the same region and the same snail hosts. Then, the guide is structured by the gastropod
intermediate host taxa. Taxonomy follows the WoRMS (WoRMS editorial board, 2022).
Some species of Notocotylidae are properly named and characterized, while other are just tem-
porarily denoted, with most details lacking. We call these “Notocotylidae gen. sp.” followed by
a number that happened to be in use in our laboratory, “WS” standing for their origin from the
White Sea and a letter addressing the cercaria morphotype (see below); for example, Notocotylidae
gen. sp. 2 WSM (White Sea Monostomi). We include information on size and structure of cercariae
and rediae, occurrence, and the definitive hosts. Size in micrometers is given as range with mean
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in parenthesis (if more than five measurements were made), or simply as mean (less than five
measurements); length and width are separated with “×” sign.
We refer to the GenBank IDs of the reference sequences, both previously published and the
new ones.
The photographs were taken at different times between 2002 and 2022, using a variety of com-
pound microscopes and cameras. The appropriate image modifications, drawings and figure layout
were made in Corel Photo-Paint 24.0.0.301 and CorelDRAW 24.0.0.301.
RESULTS AND DISCUSSION
Highlights on gastropod identification
Before further work with any parasite, its host must be correctly identified. We empha-
size this for Notocotylidae and their molluscan hosts, because their specificity is not yet
clear. Still, this guide is structured by molluscan taxa, and we start with a brief comment
on their specific differentiation. All of the molluscs we are dealing with are now classified
in Caenogastropoda, Littorinimorpha.
Hydrobiidae
Two members of Hydrobiidae (Truncatelloidea) are found at the White Sea: E. ventrosa
and P. ulvae; these mudsnails are also widely known under their old generic name, Hyd-
robia W. Hartmann, 1821. No distinction between the two species had been made in this
region until their co-existence was highlighted (Gorbushin, 1992). So, in earlier records
of digeneans from “Hydrobia spp.”, information on the host species is not quite accurate:
it could have been either E. ventrosa or P. ulvae, or a mixture of two.
Hydrobiidae belong to an abundant superfamily Truncatelloidea where many representa-
tives are morphologically and ecologically uniform (Falniowski, 2018). Its systematics has
recently been revised, so we call for attention when addressing the old data on digeneans
from “hydrobiids” - some of the species are now in different genera and families (Wilke
et al., 2001). For example, Hydrobia salsa (Pilsbry, 1905) from North America is now
classified as Spurwinkia salsa (Cochliopidae) (Davis et al., 1982).
At the White Sea, E. ventrosa and P. ulvae occur in sympatric populations and inhabit
mudflats, often in the estuaries. However, they differ ecologically (Gorbushin, 1995) and in
their response to harsh environmental conditions (Berger, Gorbushin, 2001). Morphological
distinction is as obvious as it is ambiguous (Gorbushin, 1992). The size range of the two
species overlaps, though P. ulvae are generally larger. The shell morphology is not a reliable
character, especially in infected snails and when the surface is eroded. Most of the times,
however, the periostracum differs in two species. In P. ulvae it is strong and coloured in
copper to dark brown. In E. ventrosa it is thin and transparent, often faded, resulting in
the pale bluish appearance of the shell. The oval aperture always has a narrowed tip in
P. ulvae and just sometimes in young E. ventrosa. The umbilicus is slitlike and covered
by the fold of the inner lip almost completely (P. ulvae) or just in half (E. ventrosa). The
distinct external character that allows to distinguish species is the dark spots of pigment
near the tip of the tentacles of P. ulvae (absent in E. ventrosa). The penis shape is a good
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discriminating character: it is “small and pointed” in E. ventrosa and “big and stout” in
P. ulvae (Muus, 1963, p. 135). Although this is not directly relevant for practical species
discrimination, an important difference is that P. ulvae has a free-swimming larva, while
E. ventrosa has direct development (Gorbushin, 1992).
Distribution of E. ventrosa is validated in the Mediterranean, as well as in northern
Europe: Iceland, North and Baltic Seas (Vandendorpe et al., 2019). P. ulvae has a slightly
different range: it is absent from Iceland, and may spread further north along the Euro-
pean coast (Wilke, Davis, 2000). At the Barents Sea both species probably have patchy
distribution: they are not found in the Pechora Sea and around Dalniye Zelentsy (Eastern
Murman), but present in the south-western Barents Sea in Varangerfjord and in Sommarøy
(personal observations).
Rissoidae
O. aculeus is a member of Rissooidea, Rissoidae. It is distributed on both sides of
the northern Atlantic, and apparently is the only species of this genus in the White Sea
(Matveeva, 1974; Golikov, 1987; Loskutova, Granovitch, 2006). Onoba aculeus inhabits
the intertidal and the upper subtidal (Matveeva, 1974; Golikov, 1987). At the Barents Sea,
three other species of Onoba occur, but they can be differentiated from O. aculeus mor-
phologically (Nekhaev et al., 2014).
Littorinidae
The common periwinkle Littorina littorea (Linnaeus, 1758) is not considered here be-
cause in our samples these snails were never infected with Notocotylidae.
Other periwinkles in the European north Atlantic are two groups of cryptic species
from the subgenus Neritrema (Littorinoidea, Littorinidae): “saxatilis” (L. saxatilis (Olivi,
1792), L. arcana Hannaford-Ellis, 1978, L. compressa Jeffreys, 1865) and “obtusata”
(L. obtusata (Linnaeus, 1758), L. fabalis (W. Turton, 1825)). Conchiological characters are
used to distinguish between these two groups. The shell is more conical in the “saxatilis”
and more spherical in the “obtusata” group; in the “obtusata” group, the periostracum bears
fine longitudinal striation; the sutures are deeper in “saxatilis” group (Reid, 1996; Grano-
vitch et al., 2004). Species identification within each group is possible only based on the
reproductive anatomy. Littorina saxatilis are special in being ovoviviparous.
Both at the White and Barents Seas, L. obtusata and L. fabalis co-occur. Discrimination
between them is unambiguous following dissection (Reid, 1996; Granovitch et al., 2004;
Maltseva et al., 2021b). Males of L. fabalis bear few (below six) penial glands arranged
in one row, and a long thin filament. In L. obtusata they have small glands arranged in
several rows, and a short filament. In L. obtusata females, the bursa copulatrix is almost
as long as the jelly gland, and in L. fabalis it is less than ½ of the jelly gland length.
At the Barents Sea, all three species of the “saxatilis” group co-occur. Their identi-
fication also requires dissection, but may remain inconclusive (Reid, 1996; Granovitch
et al., 2004; Maltseva et al., 2021b). Females differ in the relative size of the glands in the
reproductive system; also, bursa copulatrix is broad and long in L. arcana and short and
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slim in L. compressa. Males differ in penis morphology: two or more rows of relatively
small penial glands and triangular filament (L. arcana); one row of large penial glands
(not more than six) and an indistinct filament (L. compressa); one row with more than six
(up to 45) small penial glands and a triangular filament (L. saxatilis). At the White Sea,
only L. saxatilis has been recorded.
There is some ecological niche differentiation between the Neritrema species (Reid,
1996; Granovitch et al., 2013; Maltseva et al., 2021a). Littorina arcana are more frequent
in the upper intertidal, L. compressa - in the lower intertidal, and L. fabalis - in the upper
subtidal. Littorina saxatilis and L. obtusata can be found from the lower to the upper zones.
Littorina obtusata and L. fabalis inhabit the macrophytes, with the first species preferring
Fucus vesiculosus Linnaeus, 1753 and Ascophyllum nodosum (Linnaeus) Le Jolis, 1863,
and the second species - F. serratus Linnaeus, 1753. Species of the “saxatilis” group prefer
to live on stones and gravel.
Hybrids occur within both “obtusata” and “saxatilis” groups (Mikhailova et al., 2009;
Costa et al., 2020). Identification problems also arise when the snails are immature or
castrated following digenean infection. Genotyping is possible for the “obtusata” group
(Reid et al., 2012; Costa et al., 2020), but DNA barcoding is problematic for members of
the “saxatilis” group.
Summing up the story of distinction between the Neritrema species, the best practice for
collecting notocotylids from these snails is along with the host species identification based
on the anatomy features. If this has not been done, we recommend stating explicitly that
the hosts were identified only as “obtusata” or “saxatilis” group members, and the actual
species is unknown. This should be, however, taken more liberally for sampling in areas
where only L. saxatilis and/or L. obtusata are known to occur. If in future it is confirmed
that for species within each group (“saxatilis” and “obtusata”) the spectrum of notocotylid
parasites is the same, the laborious task to distinguish periwinkles will be fine to cease.
Field and laboratory procedures for identification of intramolluscan Notocotylidae
Snails that serve as the first intermediate hosts of Notocotylidae at the White and
Barents Seas inhabit the intertidal. They are usually sampled at low tide, by hand (peri-
winkles) or by sieving sediment through a sieve with 1-mm2 mesh size (mud snails and
O. aculeus). Smaller snails usually have dramatically lower prevalence of infection and
are not sampled, unless there is a specific goal to estimate this prevalence. In the lab the
snails are kept in natural or artificial sea water at 4-12°C, depending on the available
equipment. While the fridges with 4°C temperature are more common, higher temperature
is more favourable for activity of snails and development of cercariae, and subsequently
for their shedding (see below).
One method to detect infected snails is to induce cercariae shedding. To do so, snails
are placed individually in the wells of a 24-welled cell culture plate (for mudsnails and
O. aculeus), or similar larger vessels (for periwinkles), containing sea water. Cercariae usu-
ally leave the snails after exposure to bright artificial light or, more effectively, sunlight. The
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favourable period to try this procedure is usually between 10 am and 2 pm. It is important
to observe wells with the snails regularly because cercariae of Notocotylidae often form
cysts soon after release. The rough plan is look through the water in the wells under the
stereomicroscope after 30 min of illumination, and then 2-4 times more at 15 min intervals.
Another method is to dissect snails. This allows to detect early infection, when no
cercariae are yet ready for shedding; also it is a way to observe the rediae. Additionally,
the specific identification of snails may require dissection (see above).
The best way to observe diagnostic features of cercariae is to prepare temporary mounts
and study them under the compound microscope. Any sharp movements may trigger cer-
cariae encystment, so pipetting should be smooth and coverslip should be placed slowly. In
a thick layer of water, cercariae can swim and no details of the structure are visible. When
excess water is removed, cercariae start to contract and stretch their body; this way they
can crawl. Such a medium coverslip pressure is best for identifying cercariae, with special
emphasis to their morphotype (see below; Fig. 1F-1H). When water is deficient, cercariae
stop moving and get over-flattened. The ideal moment to take photographs is before this
moment, when cercarial activity is already low.
For measurements, the standard procedure is to kill the cercariae by heating over the
flame of a spirit lamp in a drop of water on a glass slide; to make a temporary mount with
a medium coverslip pressure; and to use an ocular micrometer or an image produced by the
camera. Using photographs for measurements is also possible (e.g. in ImageJ; Schneider
et al., 2012).
To verify identification with DNA sequencing, usually one redia or even one cercaria
is enough to extract DNA with any protocol. In this guide, we offer two rDNA barcodes:
ITS1 (800-100 b.p.) and D2 domain of the 28S rDNA (590 b.p.). The primers for their
amplification are BD1 (GTCGTAACAAGGTTTCCGTA) and 4S (TCTAGATGCGTTC-
GAARTGTCGATG) for the ITS1 (Luton et al., 1992); and C2’B (GAAAAGTACTTT-
GRARAGAGA, Bayssade-Dufour et al., 2000) and D2 (TCCGTGTTTCAAGACGGG,
Vân Le et al., 1993) for the D2. Amplification protocols are in our published articles on
Notocotylidae (e.g. Gonchar, Galaktionov, 2021). The D2 domain of the 28S rDNA am-
plifies very robustly, but it may lack variations to distinguish some of the close species
(one such example is mentioned below). ITS1 also amplifies well and is distinct in all
the species we discovered, but it contains a varying number of repeats, and this should
be accounted for during alignment. Neither of the two fragments is powerful enough for
phylogenetic reconstructions.
The new sequences generated in this study and submitted to GenBank are OP942346-
OP942361.
Recognition of Notocotylidae in the molluscs
Identification of the intramolluscan stages of Notocotylidae to the family level is sim-
ple and reliable. Stereomicroscope is usually sufficient. In a dissected mollusc, one clear
feature is the presence of rediae, they often take up much space in the snail’s haemocoel
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(Fig. 1A). The rediae have prominent pharynx and caecum (as opposed to the daughter
sporocysts in some other digeneans), germinal mass in the posterior region and some de-
veloping cercariae (Fig. 1B). Rediae of different species are similar and mostly differ in
size and the number of embryos. Cercariae have body length of about 250-450 μm and
a set of typical characters: two lateral and one median (usually less prominent) eyespots; oral
sucker but no ventral sucker; dorsal adhesive pockets at the posterolateral edges of the body
(Fig. 1C). The cercariae are pigmented and have excretory ducts filled with refractive
granules. Notocotylid cercariae encyst in the external environment (Fig. 1D-1E); this also
happens in the lab dishes where no natural substrates are available. Observations on the
behaviour of notocotylid cercariae are summarized by Krupenko and Gonchar (2017).
Immature cercariae are often seen inside and outside of the rediae within a mollusc
(Fig. 1A). They may lack median eyespot and have pigment unevenly distributed, but
otherwise have typical appearance of notocotylid cercariae.
Figure 1. Helpful features to identity infection of a mollusc by Notocotylidae and distinguish between
the species. A - hepatopancreas of a periwinkle infected with Tristriata anatis, showing multiple
rediae and cercariae, the arrows are pointing at their eyespots. B - appearance of a notocotylid redia
(exemplified by Paramonostomum alveatum); ph - pharynx, ca - caecum, ce - developing cercariae,
em - embryos, gm - germinal mass; scale bar 100 μm. C - general structure of a notocotylid cer-
caria, modified from Krupenko, Gonchar, 2017; os - oral sucker, me - median eyespot, le - lateral
eyespot, dap - dorsal adhesive pocket. D - microphotograph of a metacercarial cyst (Paramonosto-
mum alveatum) with eyespots and excretory granules still visible; scale bar 100 μm. E - cysts on the
surface of the molluscan shell. F-H - schemes of the anterior part of the main collecting ducts of the
excretory system in notocotylid cercariae that correspond to three cercarial morphotypes: Monostomi
(F), Imbricata (G) and Yenchingensis (H), modified from Rotschild, 1938.
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The minority of notocotylid cercariae lack eyespots, have a knob-like tail and encyst
inside the molluscan first intermediate host. In our study region, these belong to a single
species P. symmetricum (see below).
A couple of other digeneans occur in the similar set of coastal snails and have rediae
as an intramolluscan developmental stage, but they can be readily distinguished from Noto-
cotylidae. Cercariae of Cryptocotyle spp. (Opisthorchiidae) also lack ventral sucker and
have eyespots (two), but they have fin folds on the tail and peculiar intermittent swimming;
they also are smaller (body length below 200 μm) (Stunkard, 1930). Cercariae of Himasthla
spp. (Himasthlidae, Echinostomatoidea) are about as large as notocotylid cercariae, but
they have a collar with spines and a ventral sucker, and no eyes (Galaktionov et al., 2021).
Within Notocotylidae, cercariae are classified into three groups, or morphotypes, de-
pending on the appearance of the main collecting ducts (MCD) of their excretory system
(Rothschild, 1938). The MCD in notocotylid cercariae merge at the front, forming a circle.
In the Monostomi group, the anterior part of the MCD is behind the median eyespot
(Fig. 1F). In the Imbricata group, the anterior part of the MCD forms a loop at/in front of
the median eyespot (Fig. 1G). In the Yenchingensis group, the anterior part of the MCD
forms a diverticulum reaching the median eyespot or further forward (Fig. 1H). Some vari-
ations to the Monostomi morphotype are possible, in particular - one, two or three shorter
inconspicuous diverticula (not to be confused with Yenchingensis).
Summary of Notocotylidae species from different hosts1
Hydrobiidae
Notocotylus atlanticus Stunkard, 1966 (Fig. 2A)
(From Gonchar et al., 2019) Living rediae vary in size: mature rediae are 770-1570 ×
290-370 (1185 × 330); young rediae are 270-600 × 114-257 (418 × 176). Cercariae are
of Yenchingensis morphotype. Heat-killed cercariae are 257-371 (300) × 114-229 (153);
the tail is 243-400 (305) × 29-43 (34); the oral sucker is 29-47 (35). Excretory granules
are located 4-7 in rows across the main excretory ducts, their size is 0.88-1.84 (1.32).
Cystogenous glands cells contain uniform rod-shaped secrete. The cysts are 170-200 (186)
in diameter.
The species was originally described in North America, with S. salsa as the first in-
termediate hosts (Stunkard, 1966). We found E. ventrosa infected with this parasite at
the White Sea and in Iceland. There has yet been no ultimate genetic validation of the
conspecificity of the European and American material. Indirect genetic evidence suggests
that N. atlanticus is also found in Japan (Gonchar, Galaktionov, 2022).
The type host is Somateria mollissima (Linnaeus, 1758). We found the maritae of
N. atlanticus in Anas platyrhynchos Linnaeus, 1758, Mareca penelope (Linnaeus, 1758)
and A. acuta Linnaeus, 1758. Maritae are difficult to distinguish from several other species:
N. intestinalis Tubangui, 1932, N. magniovatus Yamaguti, 1934, N. imbricatus (Looss,
1893) Szidat, 1935, N. attenuatus (Rudolphi, 1809) Kossack, 1911.
Molecular references MH818008 (28S rDNA) and MH818012 (ITS1).
1 The actual range of molluscan hosts for each species of Notocotylids may be wider; this possibility has to
be tested in all future studies.
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Paramonostomum anatis Garkavi, 1965 (Fig. 2B)
Mother rediae reach up to 150-450 × 45-90, and daughter rediae measuring 540 × 180
and more start producing cercariae (Garkavi, 1968). We found that cercariae belong to the
Imbricata morphotype; their body is 270 × 151, the tail is 300 × 30, the oral sucker is 32
(our unpublished data). Cercariae are equally willing to encyst on both vegetate substrate
and the shell of the mollusc which they have left (Fig. 1E) (Gonchar, Galaktionov, 2016).
We found this species only in E. ventrosa, at the White Sea and in Iceland.
The species was originally described at the coast of the Sea of Azov; its life cycle was
also elucidated there, with E. ventrosa as the first intermediate hosts (Garkavi, 1968). There
were no further records of its intramolluscan stages, but Cercaria Notocotylidae sp. no. 12
Deblock, 1980 from Peringia ulvae have similar appearance, particularly the morphotype.
Definitive hosts are A. platyrhynchos (type host, also our unpublished data) and prob-
ably other Anas spp. (Filimonova, 1985).
Molecular references OP942354 (28S rDNA) and OP942347 (ITS1).
Paramonostomum alveatum (Mehlis in Creplin, 1846) Lühe, 1909 (Fig. 2C)
Rediae (measured from the photograph) are 1120 × 283 and contain about four devel-
oping cercariae (Fig. 1B). The cercarial body is 240 × 135, the tail is 300 × 30, the oral
sucker is 32 (our unpublished data). Cercarial morphotype is Monostomi, but some varia-
tions occur: one, two or three short extensions in the anterior part of the main collecting
ducts. Cercariae tend to encyst on the vegetate substrate (Gonchar, Galaktionov, 2016).
We found P. alveatum in both E. ventrosa and Peringia ulvae at the Keret Archipelago,
the White Sea; and in E. ventrosa in Iceland.
Earlier records of the geographic and host range are not clear: it is impossible to
confidently interpret them because we might deal with a group of morphologically simi-
lar species. At the White Sea, the intermediate hosts of P. alveatum were identified as
Peringia ulvae (Kulachkova, 1954; Zelikman, 1966; Chubrik, 1966). In North America,
this species is supposedly hosted by S. salsa (Stunkard, 1967). Cercaria Notocotylidae sp.
no. 11 Deblock, 1980 from E. ventrosa in the Mediterranean may represent this species,
too (Deblock, 1980).
Definitive hosts are Somateria mollissima (our data) and many other species of anatids
(Filimonova, 1985); type host is unclear.
Molecular references OP942355 (28S rDNA) and OP942346 (ITS1). Sequence of the
D2 domain of the 28S rDNA does not ensure reliable distinction from the species Noto-
cotylidae gen. sp. 2 WSM (see below).
Notocotylidae gen. sp. 3 WSI (Fig. 2D)
We found cercariae of Imbricata morphotype measuring 350-409 × 168-220 (body),
380-415 × 38-50 (tail); 31-35 × 33-37 (oral sucker) uniquely in Peringia ulvae (our
unpublished data). We matched them by molecular genetic data to (1) maritae from
A. platyrhynchos that comply with the diagnosis of P. anatis; and (2) a GenBank sequence
for the marita from a wader Tringa erythropus Pallas, 1764 (Charadriiformes) from Kherson
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Region, Ukraine (Tkach et al., 2001) that is also named P. anatis. However, there is genetic
divergence between the P. anatis from E. ventrosa (described above) and this species.
The morphological distinction between Notocotylidae gen. sp. 3 WSI and P. anatis is
scarce, but it appears that the former might have larger cercariae.
Molecular references OP942358 (28S rDNA) and OP942350 (ITS1).
Notocotylidae gen. sp. 2 WSM (no image available)
According to our unpublished data, the body of cercariae is 265 × 152; their tail is
344 × 36; and the oral sucker is 36 × 35. Cercariae are of Monostomi morphotype and
very similar to those of P. alveatum, but appear genetically distinct from them. Both
E. ventrosa and P. ulvae serve as the first intermediate hosts.
We found maritae in S. mollissima from the White Sea that matched these cercariae
genetically; this material was not sufficient for species identification or morphological
description.
Molecular references OP942361 (28S rDNA) and OP942349 (ITS1). Sequence of
the D2 domain of the 28S rDNA does not ensure reliable distinction from the species
P. alveatum (see above).
Notocotylidae gen. sp. 5 WSM (no image available)
One Peringia ulvae mollusc from Krasnyi Island (White Sea, Kandalaksha Bay) was
infected with Notocotylidae that had Monostomi cercariae, but were genetically distinct
from P. alveatum. As for now, this species is delineated only based on molecular data;
no matching maritae have been found. Notocotylidae gen. sp. 5 WS appears as a sister
species to P. alveatum.
Cercariae of Monostomi morphotype from P. ulvae were described in the life cycle of
Catatropis lagunae Bayssade-Dufour et al., 1996 in France. This is the only prior record
that may potentially refer to the same species.
Molecular references OP942360 (28S rDNA) and OP942352 (ITS1).
Notocotylidae gen. sp. 4 WSY (no image available)
In March 2018 and 2019 we collected two P. ulvae molluscs shedding notocotylid
cercariae with Yenchingensis morphotype in the Sukhaya Salma inlet at the White Sea.
The definitive host is unknown. The putative species is suspected based on molecular data.
It appears as a sister species to Catatropis onobae Gonchar, Galaktionov, 2021.
Molecular references OP942359 (28S rDNA) and OP942353 (ITS1).
Littorina spp.
Tristriata anatis Belopolskaja, 1953 (Fig. 2E)
(From Gonchar, Galaktionov, 2017) There may be up to a thousand rediae in one
mollusc (Fig. 1A), they differ in size depending on their age, roughly from 300 to 1800
in length. Cercariae are of Monostomi morphotype, their body size is 315-510 (425) ×
165-270 (230), the tail is 315-488 (408) × 37-83 (62), and the oral sucker is 45-60 (55).
The secrete of the cystogenous glands in rod-shaped.
453
We found T. anatis in three species of Littorina: L. saxatilis, L. obtusata and (in the
Sea of Okhotsk) Littorina sitkana Philippi, 1846.
The species is found across a large geographic range: in the North Pacific (Sea of
Okhotsk) and North Atlantic (Barents Sea, Celtic Sea, Iceland) (Gonchar, Galaktionov,
2020).
Definitive hosts are S. mollissima, Somateria spectabilis (Linnaeus, 1758), Histrionicus
histrionicus (Linnaeus, 1758) and A. platyrhynchos.
Molecular references KX833042 (28S rDNA) and KX833023 (ITS1).
Notocotylidae gen. sp. 6 WSY (Fig. 2F)
We found notocotylid cercariae with Yenchingensis morphotype in L. saxatilis in Kem-
ludy Archipelago (White Sea, Kandalaksha Bay, Chupa Inlet) and in Roscoff (the English
Channel). Few photographs are available, and when measured from a photo, the cercarial
body is 194 × 99, the tail is 146 × 18, and the oral sucker is 29. The corresponding maritae
and the definitive host are unknown. We consider this an independent species based on
the molecular genetic data.
Yenchingensis cercariae from L. obtusata and L. littorea in Roscoff were called Cer-
caria lebouri (Stunkard, 1932). Later accounts of supposedly the same cercariae were from
L. littorea (e.g. Werding, 1969) and L. saxatilis (e.g. James, 1969). The experimental
infection study revealed that Cercaria lebouri from L. littorea correspond to the maritae
identified as Paramonostomum chabaudi van Strydonck 1965 (Evans et al., 1997). This
leaves a question on a specific identity of Yenchingensis cercariae from the representa-
tives of the subgenus Neritrema - both L. saxatilis and L. obtusata: do they also belong to
P. chabaudi or to some different species? And are our Yenchingensis isolates from L. saxa-
tilis at the White Sea conspecific to those from the British Isles (James, 1969)?
Molecular references OP942356 (28S rDNA) and OP942351 (ITS1).
Parapronocephalum symmetricum Belopolskaja, 1952 (Fig. 2G)
(Based on the description from James, 1969). Rediae and cercariae are found in the
visceral haemocoel. There is a single first (410-750 × 120-350) and six to ten second
(600-900 × 290-580) generation rediae, their pharynx is 50-70 in diameter. The body
of the fully-developed cercariae is 710-720 × 280-300, their oral sucker is 140-145, the
tail is stumpy and measures just 100. They have a collar that is 300-305 wide. Cercariae
migrate towards the stomach and encycst there in the haemocoel lining. The cysts are oval,
380-400 × 250-280.
We found L. obtusata infected with P. symmetricum in Kem-ludy Archipelago, Chupa
Inlet, White Sea; and also detected it in the Eastern Murman. The species was originally
described from L. saxatilis in the Seven Islands Archipelago, Barents Sea (Belopolskaja,
1952). It was later recorded, also in L. saxatilis, on the British Isles (Celtic Sea - Bristol
Channel (James, 1969) and Isles of Scilly (Newell, 1986); the North Channel - St Mary’s
Portavogie harbour (Matthews et al., 1985) and Belfast Lough (Irwin et al., 1989)). Ac-
counts in Iceland are from both L. saxatilis and L. obtusata (Skírnisson, Galaktionov, 2002).
454
In a study covering the extended region in the north of Europe, P. symmetricum was found
in both L. saxatilis and L. obtusata in the west (Trøms, Finnmark and Western Murman),
only in L. saxatilis on the Eastern Murman coast and only in L. obtusata in the White Sea
(Galaktionov, Bustnes, 1996).
Type hosts are Calidris maritima (Brünnich, 1764). Adult worms also bear a collar and
resemble the metacercariae from the periwinkles, but reach larger size.
Molecular references OP942357 (28S rDNA) and OP942348 (ITS1).
Onoba aculeus
Catatropis onobae Gonchar, Galaktionov, 2021 (Fig. 2H)
(From Gonchar, Galaktionov, 2021) The rediae measure 218-800 (492) × 106-234
(174), the pharynx is 31-49 (43) × 31-44 (37). In ethanol-fixed cercariae, body size is
243-361 (300) × 118-180 (145); tail is 322-588 (463) × 33-48 (41); oral sucker is 27-40
× 30-43. In living cercariae, body size is 248-379 (297) × 163-215 (181). The morphotype
is Yenchingensis, MCD contain excretory granules 1.45-2.28 (1.86, n = 38) in diameter;
1-2 granules are in a row across main excretory ducts. Secretory granules in cystogenous
glands uniform.
O. aculeus infected with C. onobae occur in Kem-ludy Archipelago, Chupa Bay, White
Sea; Dalniye Zelentsy, Barents Sea; and Grόtta, Grindavik (Iceland). In the same regions,
notocotylids (probably also belonging to C. onobae) had been registered in O. aculeus
before the species was described (Chubrik, 1966; Gorbushin, Levakin, 1999; Galaktionov,
Skírnisson, 2000; Skírnisson, Galaktionov, 2002).
Definitive (and type) hosts are common eiders S. mollissima, maritae are morphologi-
cally indistinguishable from those of at least several other species: C. verrucosa (Frölich,
1789) Odhner, 1905, Pseudocatatropis joyeuxi Kanev and Vasiliev, 1986, and P. dvoryadkini
Izrailskaia, Besprozvannykh, Tatonova et al., 2019.
Molecular references MN963021 (28S rDNA) and MN962974 (ITS1).
CONCLUSIONS
We have shown that the diversity of Notocotylidae infecting intertidal gastropods at
the White and Barents Seas was highly underestimated. The current total of eleven species
may also not be the final number, but probably is close to reality, considering the sampling
effort. The non-genetic discriminating features for these species are limited, but future re-
search may discover more of these. Particularly intriguing is the specificity of Notocotylidae
to their intermediate host. Now there seem to be examples of both strict (one species of
gastropods) and relaxed (members of several families within one superfamily) specificity.
This issue also requires further studies. Moreover, the definitive hosts and maritae are yet
well-defined for just five species out of 11.
Overall, if similar trends are found in other regions, the family Notocotylidae will likely
grow following integrative taxonomy research. This will also lead to better understanding
their evolution, and contribute to the development of evolutionary concepts for the whole
Digenea.
455
ACKNOWLEDGEMENTS
The sampling during the cruises to the Pechora Sea and the molecular genetic stud-
ies were funded by the Russian Science Foundation project no. 18-14-00170. Fieldwork
at the White Sea Biological Station “Kartesh” of the Zoological Institute of the Russian
Academy of Sciences (ZIN RAS) was funded by the research program of ZIN RAS (project
no. 122031100260-0). Part of the material was collected at the Educational and Research
Station “Belomorskaia” of St Petersburg University (SPbU).
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ВИДОВОЙ СОСТАВ NOTOCOTYLIDAE (DIGENEA)
В ЛИТОРАЛЬНЫХ ГАСТРОПОДАХ НА БЕЛОМ И БАРЕНЦЕВОМ МОРЯХ
А. Г. Гончар, К. В. Галактионов
Ключевые слова: нотокотилиды, трематоды, церкарии, редии, промежуточные хозяева,
ДНК-баркоды
РЕЗЮМЕ
Как показали недавние исследования, фауна трематод в брюхоногих моллюсках на Белом и
Баренцевом морях характеризуется бóльшим видовым богатством, чем предполагалось ранее.
Эти данные, опубликованные в сериях статей, мы предлагаем обобщить в форме, удобной для
практического использования. В данной работе мы объединили сведения об 11 видах из сем.
Notocotylidae, которых мы обнаружили в литоральных моллюсках Ecrobia ventrosa, Peringia
ulvae, Littorina spp. и Onoba aculeus. Мы приводим размеры и краткие описания редий и цер-
карий, известный спектр хозяев и географическое распространение, хотя для некоторых видов
информация пока неполная. Мы также ссылаемся на последовательности ДНК из базы данных
GenBank, которые могут послужить для идентификации видов - включая несколько новых
последовательностей. Для удобства использования мы предваряем список видов нотокотилид
советами по идентификации моллюсков-хозяев и проведению основных полевых и лаборатор-
ных процедур.
459