A New Species of Deropristid Trematode from the Sterlet

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Zoological Journal of the Linnean Society, 2020, 190, 448–459. With 4 figures.

A new species of deropristid trematode from the sterlet Acipenser ruthenus (Actinopterygii: Acipenseridae) and revision of superfamily affiliation of the family Deropristidae Downloaded from https://academic.oup.com/zoolinnean/article/190/2/448/5810955 by guest on 02 October 2020

SERGEY SOKOLOV1, EKATERINA VOROPAEVA1 and DMITRY ATOPKIN2,3*

1A. N. Severtsov Institute of Ecology and Evolution, Leninskij prosp. 33, 119071 Moscow, Russia 2Federal Scientific Center of the East Asia Terrestrial Biodiversity, Far Eastern Branch of the RAS, 100-letija prosp. 159, 690022 Vladivostok, Russia 3Department of Cell Biology and Genetics, Far Eastern Federal University, Sukhanova Street 8, 690051 Vladivostok, Russia

Received 22 November 2019; revised 15 January 2020; accepted for publication 8 February 2020

A new species, Skrjabinopsolus nudidorsalis sp. nov. is described from the sterlet Acipenser ruthenus, caught in the River Volga basin (Russia). This species differs from previously described congeners by the absence of vitelline follicles on the dorsal side of the body. The complete 18S rRNA and partial 28S rRNA gene sequences obtained for S. nudidorsalis are the first molecular data for the family Deropristidae. The results of phylogenetic analysis indicate that Deropristidae is sister to the Monorchiidae + Lissorchiidae group. The results of the phylogenetic study contradict the current taxonomic hypothesis that Deropristidae belongs to the superfamily Lepocreadioidea and allow inclusion of this family in Monorchioidea. The morphological similarity of deropristids to other monorchioids is recognizable from the presence of a bipartite internal seminal vesicle, spinous cirrus and a voluminous, armed metraterm.

ADDITIONAL KEYWORDS: 18S – 28S –Monorchioidea – new species – River Volga – Skrjabinopsolus – sturgeons.

INTRODUCTION Ivanov, 1936, which was collected from the intestines of the Russian sturgeon Acipenser gueldenstaedtii Acipenseriformes – sturgeons and paddlefishes – are von Brandt & Ratzeburg, 1833, the starry sturgeon one of most ancient groups of ray-finned fish and are Acipenser stellatus Pallas, 1771, the sterlet Acipenser of great scientific and economic importance (Birstein ruthenus Linnaeus, 1758 and the beluga Acipenser et al., 1997; Betancur et al., 2017). Currently, these fish huso Linnaeus, 1758, caught in the River Volga delta species are present in Europe and large parts of Asia (Ivanov & Murygin, 1936). The year of description and North America. The parasite fauna of sturgeons of this trematode genus and species is not indicated and paddlefishes is unique and conditioned by the correctly in previous publications, except that of presence of host-specific high-level taxa (Skrjabina, Shulman (1954). The first time A. S. Ivanov presented 1974; Choudchury & Dick, 2001; Evans et al., 2008). data on Skrjabinopsolus was at the 230th session of In the present study, we focus on trematodes of the the Standing Committee for the Study of Helminth genus Skrjabinopsolus Ivanov, 1936. This genus was Fauna of the USSR Academy of Sciences on 19 May described based on the type species, S. acipenseris 1934. However, these materials were not published. The original description of this taxon was published in the paper of Ivanov & Murygin (1936). *Corresponding author. E-mail: [email protected] [Version of record, published online 21 March 2020; http:// At various times, nine species of trematodes zoobank.org/ urn:lsid:zoobank.org:pub: 093E8C8F-A240-49AC- have been assigned to the genus Skrjabinopsolus, 9384-4BF0B3C27AEB] namely S. acipenseris, S. semiarmatus (Molin, 1858),

S. skrjabini Osmanov, 1940, S. manteri (Cable, 1952), MATERIAL AND METHODS S. minor Bykhovskaya-Pavlovskaya & Mikailov, Sample collection 1969, S. elongatus (Madhavi, 1974) (with synonyms of S. indicus Gupta & Ahmad, 1976 and S. kurotchkini Worm specimens were collected in the course of a Parukhin, 1976 as proposed by Hafeezullah, 1984) parasitological investigation of 12 specimens [total and S. sanyaensis Shen, 1990 (e.g. Bykhovskaya- length (TL) = 38–54 cm] from the sterlet, caught in Pavlovskaya & Mikailov, 1969; Hafeezullah, 1984; Shen, the River Oka (River Volga basin) near the village 1990). Skrjabinopsolus elongatus is now considered a of Kletino, Ryazan Oblast, Russia, in June 2019. All member of the monorchiid genus Opisthodiplomonorchis applicable international guidelines for the care and Madhavi, 1974 (Choudhury & Dick, 1998; Madhavi & use of animals were followed by the authors (Directive

Bray, 2018). Skrjabinopsolus sanyaensis does not show 2010/63/EU of the European Parliament and of Downloaded from https://academic.oup.com/zoolinnean/article/190/2/448/5810955 by guest on 02 October 2020 the diagnostic characters of either Skrjabinopsolus or the Council of 22 September 2010 on the protection the Deropristidae. Therefore, its systematic position is of animals used for scientific purposes). Catching still unclear (Choudhury & Dick, 1998). of sterlet was carried out by the employees of the The basic features of S. acipenseris, S. skrjabini, Russian Federal Research Institute of Fisheries and S. minor and S. manteri (body and organ size, Oceanography (FSBSI ‘VNIRO’, Moscow) in accordance gonad arrangement, extension of the vitelline fields, with the monitoring programme of the ichthyofauna of distribution of the uterus, egg size, etc.) are highly the middle course of the River Oka. variable (Bykhovsky & Dubinina, 1954; Shulman, The worms collected for morphological study were 1954; Bykhovskaya-Pavlovskaya & Mikailov, fixed in hot 70% ethanol, stained with acetocarmine, 1969, Skrjabina, 1974; Bunyatova & Mikailov, dehydrated in dimethyl phthalate, mounted in Canada 1991; Choudhury & Dick, 1998). In this respect, balsam and studied with the aid of a light microscope Skrjabina (1974) and Choudhury & Dick (1998) Axio Imager A1 (Zeiss AG, Oberkochen, Germany). All considered S. acipenseris, S. skrjabini and S. minor measurements were made in micrometres based on 12 as junior synonyms of S. semiarmatus. Bunyatova adults and four juvenile specimens, unless otherwise & Mikailov (1991) considered that S. manteri and indicated. The measurements of the adults are provided S. semiarmatus are also conspecific. Choudhury as the range of followed by the mean values in round & Dick (1998) recognized S. manteri as a Nearctic brackets and the measurements for the holotype in subspecies of S. semiarmatus – S. semiarmatus square brackets. For the juvenile specimens, only the manteri. At the same time, Choudhury & Dick (2001) range is provided. Specimens were deposited in the considered S. manteri as an independent species Museum of Helminthological Collections at the Centre but without additional argument. Thus, the genus of Parasitology of the A. N. Severtsov Institute of Skrjabinopsolus is most widely considered to consist Ecology and Evolution (IPEE RAS) in Moscow, Russia. of two valid species – S. semiarmatus, identified Specimens destined for molecular analysis were fixed in Europe, Western Asia and the Russian Far East in 96% ethanol and stored at –18 °C. (Skrjabina, 1974; Yukhimenko & Belyaev, 2002; Noie, 2011), and S. manteri, found in North America (Choudhury & Dick, 1998, 2001). Examination of further specimens from museum According to the present view on systematics of collections the class Trematoda, this genus belongs to the family We studied the morphology of the following museum Deropristidae of the superfamily Lepocreadioidea specimens of Skrjabinopsolus to clarify the differential (Bray, 2005; Bray et al., 2009; Bray & Cribb, 2012). diagnosis of a new species: However, this viewpoint has not been verified with molecular data. • Photos of holotype, paratype and two voucher In 2019, trematode specimens belonging to the specimens of S. manteri from North American genus Skrjabinopsolus were found by E. Voropaeva sturgeons, the National Museum of Natural History in sterlet from the River Volga basin (Russia). (USNM 1337876, USNM 1337877, USNM 1380936, These parasites differ from their congeners by USNM 1384527), Washington DC, United States. a number of morphological characteristics that Photos were kindly provided by Dr Anna Philips. allow us to consider these worms as a new species, • Eighteen voucher specimens of S. semiarmatus Skrjabinopsolus nudidorsalis. In the present paper, (Molin, 1858). Whole-mounted adult specimens we provide a description, drawings and results of the from the beluga, the starry sturgeon and the molecular phylogenetic analysis of this species with bastard sturgeon Acipenser nudiventris Lovetsky, clarifying phylogenetic relationships of the family 1828 caught in the Caspian Sea, April 1964 (IPEE Deropristidae. RAS 1082–1088, 1092).

Unfortunately, the procedure of descriptions of 3.1 (Guindon & Gascuel, 2003) and MrBayes v.3.1.2 S. semiarmatus s.s. and two other nominal species, software (Huelsenbeck et al., 2001), respectively. The namely S. acipenseris and S. skrjabini, were performed best nucleotide substitution model, GTR+G+I, was without designation of the place of storage of type estimated with jModeltest v.2.1.5 software (Darriba specimens (see: Molin, 1858; Ivanov & Murygin, 1936; et al., 2012) for both ML and BI algorithms. Bayesian Osmanov, 1940). These specimens are absent in the most analysis was performed using 10 000 000 generations, presumable depositaries: the central helminthological with two independent runs. Summary parameters and collections of Moscow (A. N. Severtsov Institute of the phylogenetic tree were calculated with a burn-in Ecology and Evolution; K. I. Skrjabin All-Russian of 250 000 generations. Support of the phylogenetic Institute of Helminthology), St. Petersburg (Zoological relationships was estimated using posterior probabilities

Institute RAS), Vena (Naturhistorisches Museum) and (Huelsenbeck et al., 2001) for both ML and BI algorithms. Downloaded from https://academic.oup.com/zoolinnean/article/190/2/448/5810955 by guest on 02 October 2020 Berlin (Museum für Naturkunde). The type specimens The phylogenetic relationships were inferred from of S. minor were placed to the Helminthological our data, along with the nucleotide sequences of the Laboratory of Zoological Institute of Azerbaijan SSR, combined complete 18S rRNA gene and partial 28S Baku (see: Bykhovskaya-Pavlovskaya & Mikailov, rRNA gene of other trematode specimens obtained from 1969), but these specimens have apparently been lost the NCBI GenBank database (Table 1). The tree was (A. Manafov, personal communication). rooted with species from the Opisthorchioidea.

Molecular and phylogenetic study Total DNA was extracted from three adult specimens RESULTS of the potential new species, S. nudidorsalis, using the Phylogeny ‘hot shot’ technique (Truett, 2006). Our sample represents the first species of the Polymerase chain reaction (PCR) was used to amplify family Deropristidae to be represented by molecular the 18S rRNA gene with the primers 18S-8 (5’-GCA data. In both the ML and BI analyses, based on GCC GCG GTA ACT CCA GC-3’) and 18S-A27 (5’-CCA concatenated complete 18S rRNA and partial 28S TAC AAA TGC CCC CGT CTG-3’) (Littlewood & Olson, rRNA genes, Deropristidae appears as a member 2001). The initial PCR reaction was performed in a of the well-supported Monorchioidea clade, within total volume of 20 µL and contained 0.25 mM of each which this family forms a sister relationship with primer, approximately 10 ng of total DNA in water, the Monorchiidae + Lissorchiidae group (Fig. 1). 10X Dream Taq buffer, 1.25 mM dNTPs and one unit The Monorchioidea clade is closely related to the of Dream Taq polymerase (Thermo Scientific, USA). Brachycladioidea + Opecoeloidea + Gorgoderoidea + Amplification of a 2000-base pair (bp) fragment of 18S Haploporoidea group, with high Bayesian support for rDNA was performed in a GeneAmp 9700 (Applied both ML and Bayesian trees. Biosystems, USA) with a 5-min denaturation at 96 °C, Representatives of the Lepocreadioidea formed 35 cycles of 1 min at 96 °C, 20 s at 58 °C and 5 min at a distinct clade, sister to the (Brachycladioidea + 72 °C and a 10-min extension at 72 °C. Negative and Opecoeloidea + Gorgoderoidea + Haploporoidea) + positive controls using both primers were included. Monorchioidea clade. Trematodes of the Apocreadioidea The 28S rRNA gene was amplified with the primers are basal to all the above-mentioned superfamilies, DIG12 (5’-AAG CAT ATC ACT AAG CGG -3’) and with high statistical support for both ML and Bayesian 1500R (5’-GCT ATC CTG AGG GAA ACT TCG-3’) analyses (Fig. 1). The new species is supported and is (Tkach et al., 2003). The master mix for the PCR described below. reaction was identical to that described above for 18S rRNA gene. Amplification of a 1200-bp fragment of 28S rRNA gene was performed in a T-100 thermocycler (Bio-Rad, USA) with a 3-min denaturation at 94 °C, 40 TAXONOMY cycles of 30 s at 94 °C, 30 s at 55 °C and 2 min at 72 °C Family Deropristidae Cable & Hunninen, 1942 and a 7-min extension at 72 °C. Negative and positive controls using both primers were included. Genus Skrjabinopsolus Ivanov, 1936 Sequences were assembled with SeqScape v.2.6 Skrjabinopsolus nudidorsalis sp. nov. software. Alignments and estimation of the number of variable sites and sequence differences were (Figs 2A–D, 3A, 4A) performed using MEGA 7.0 (Kumar et al., 2016). Phylogenetic analyses of the nucleotide sequences ZooBank registration LSID: http://www.zoobank. were performed using the maximum likelihood (ML) org/urn:lsid:zoobank.org:act:0CB540AC-F471- and Bayesian inference (BI) algorithms with PhyML 4C92-AEC4-80F5AD9F5A53

Table 1. List of previously published sequences used in the phylogenetic analysis

Species 18S rRNA gene 28S rRNA gene Reference

Acanthocolpidae Pleorchis polyorchis (Stossich, 1889) DQ248202 DQ248215 Bray et al. (2005) Pleorchis uku Yamaguti, 1970 DQ248203 DQ248216 Bray et al. (2005) Stephanostomum baccatum (Nicoll, 1907) DQ248205 DQ248218 Bray et al. (2005) Stephanostomum bicoronatum (Stossich, 1883) DQ248212 DQ248225 Bray et al. (2005) Stephanostomum cesticillus (Molin, 1858) DQ248213 DQ248226 Bray et al. (2005) Stephanostomum interruptum Sparks & Thatcher, 1958 DQ248210 DQ248223 Bray et al. (2005) Downloaded from https://academic.oup.com/zoolinnean/article/190/2/448/5810955 by guest on 02 October 2020 Stephanostomum gaidropsari Bartoli & Bray, 2001 DQ248208 DQ248221 Bray et al. (2005) Stephanostomum minutum (Looss, 1901) DQ248211 DQ248224 Bray et al. (2005) Stephanostomum pristis (Deslongchamps, 1824) DQ248209 DQ248222 Bray et al. (2005) Stephanostomum tantabiddii Bray & Cribb, 2004 DQ248207 DQ248220 Bray et al. (2005) Tormopsolus orientalis Yamaguti, 1934 DQ248204 DQ248217 Bray et al. (2005) Aephnidiogenidae Tetracerasta blepta Watson, 1984 L06670 – Blair & Barker (1993) – FJ788494 Bray et al. (2009) Allocreadiidae Allocreadium neotenicum Peters, 1957 JX983204 JX977132 Bray et al. (2012) Apocreadiidae Homalometron armatum (MacCallum, 1895) AY222130 AY222241 Olson et al. (2003) Homalometron synagris (Yamaguti, 1953) AJ287523 – Cribb et al. (2001) – AY222243 Olson et al. (2003) Neoapocreadium splendens Cribb & Bray, 1999 AJ287543 – Cribb et al. (2001) – AY222242 Olson et al. (2003) Schistorchis zancli Hanson, 1953 AY222129 AY222240 Olson et al. (2003) Atractotrematidae Atractotrema sigani Durio & Manter, 1969 AJ287479 – Cribb et al. (2001) – AY222267 Olson et al. (2003) Pseudomegasolena ishigakiense Machida & Kamiya, 1976 AJ287569 – Cribb et al. (2001) – AY222266 Olson et al. (2003) Brachycladiidae Brachycladium goliath (van Beneden, 1858) KR703279 KR703279 Briscoe et al. (2016) Zalophotrema hepaticum Stunkard & Alvey, 1929 AJ224884 – Cribb et al. (2001) – AY222255 Olson et al. (2003) Dicrocoeliidae Brachylecithum lobatum (Railliet, 1900) AY222144 AY222260 Olson et al. (2003) Dicrocoelium dendriticum (Rudolphi 1819) Y11236 – Sandoval H.H. (un- published) – AF151939 Tkach et al. (2000) Lyperosomum collurionis (Skrjabin & Isaichikov, 1927) AY222143 AY222259 Olson et al. (2003) Gorgocephalidae Gorgocephalus kyphosi Manter, 1966 AY222126 AY222234 Olson et al. (2003) Gorgoderidae Degeneria halosauri (Bell, 1887) AJ287497 – Cribb et al. (2001) – AY222257 Olson et al. (2003) Gorgodera sp. AJ287518 – Cribb et al. (2001) – AY222264 Olson et al. (2003) Nagmia floridensis Markell, 1953 AY222145 AY222262 Olson et al. (2003) Gyliauchenidae Paragyliauchen arusettae Machida, 1984 AY222127 – Olson et al. (2003) – FJ788503 Bray et al. (2009) Enenteridae Enenterum aureum Linton, 1910 AY222124 AY222232 Olson et al. (2003)

Table 1. Continued

Species 18S rRNA gene 28S rRNA gene Reference

Koseiria xishaensis Gu & Shen, 1983 AY222125 AY222233 Olson et al. (2003) Haploporidae Elonginurus mugilis Lü, 1995 MH763777 MH763761 Atopkin et al. (2019) Hapladena nasonis Yamaguti, 1970 AY222146 AY222265 Olson et al. (2003) Haploporus benedeni (Stossich, 1887) FJ211228 FJ211237 Blasco-Costa et al. (2009) Lepocreadiidae Preptetos caballeroi Pritchard, 1960 AJ287563 – Cribb et al. (2001) Downloaded from https://academic.oup.com/zoolinnean/article/190/2/448/5810955 by guest on 02 October 2020 – AY222236 Olson et al. (2003) Preptetos trulla (Linton, 1907) AY222128 AY222237 Olson et al. (2003) Lissorchiidae Lissorchis kritskyi Barnhart & Powell, 1979 AY222136 AY222250 Olson et al. (2003) Monorchiidae Cableia pudica Bray, Cribb & Barker, 1996 AJ287486 – Cribb et al. (2001) – AY222251 Olson et al. (2003) Diplomonorchis leiostomi Hopkins, 1941 AY222137 AY222252 Olson et al. (2003) Lasiotocus typicum (Nicoll, 1912) AJ287474 – Cribb et al. (2001) – AY222254 Olson et al. (2003) Provitellus turrum Dove & Cribb, 1998 AJ287566 – Cribb et al. (2001) – AY222253 Olson et al. (2003) Opecoelidae Halosaurotrema halosauropsi (Bray & Campbell, 1996) [access AJ287514 – Cribb et al. (2001) as Gaevskajatrema halosauropsi] – AY222207 Olson et al. (2003) Macvicaria macassarensis (Yamaguti, 1952) AJ287533 – Cribb et al. (2001) – AY222208 Olson et al. (2003) Peracreadium idoneum (Nicoll, 1909) AJ287558 – Cribb et al. (2001) – AY222209 Olson et al. (2003) Outgroup Caecincola parvulus Marshall & Gilbert, 1905 AY222123 AY222231 Olson et al. (2003) Haplorchis pumilio (Looss, 1896) KX815125 KX815125 Le et al. (2017) Metagonimus miyatai Saito, Chai, Kim, Lee & Rim, 1997 HQ832626 HQ832635 Pornruseetairatn et al. (2016) Metagonimus yokogawai (Katsurada, 1912) HQ832630 HQ832639 Pornruseetairatn et al. (2016) Opisthorchis felineus (Rivolta, 1884) MF077357 MF099790 Dao et al. (2017) Opisthorchis viverrini (Poirier, 1886) JF823987 JF823990 Thaenkham et al. (2011)

Type-host: Acipenser ruthenus Linnaeus, 1758 Site of infection: Intestines of adult specimens and (Actinopterygii: Acipenseridae), the sterlet. stomach of juvenile specimens.

Type locality: Oka River near Kletino village, Ryazan Prevalence: 66.7% (N = 12). Oblast, Russia (54°59′55″ N, 41°10′56″E). Intensity of infection: 1–297 worms/infected host Type and voucher materials: Holotype IPEE RAS specimen. 1319 (whole-mounted adult specimen), 11 paratypes, IPEE RAS 1319–1324 (whole-mounted adult Representative DNA sequences: Sequences of three specimens) and five voucher specimens, IPEE RAS representatives were deposited in the NCBI database: 14279 (four juvenile trematodes and 14 280 one complete 18S rRNA gene (MN700959–MN700961) adult specimen). and partial 28S rRNA gene (MN700996–MN700998).

© 2020 The Linnean Society of London, Zoological Journal of the Linnean Society, 2020, 190, 448–459 A NEW SPECIES OF DEROPRISTID TREMATODE 453 Downloaded from https://academic.oup.com/zoolinnean/article/190/2/448/5810955 by guest on 02 October 2020

Figure 1. Phylogenetic position of Skrjabinopsolus nudidorsalis, constructed by Bayesian inference and maximum likelihood (ML/BI) analyses of 18S+28S rRNA genes sequences alignment. Nodal support shower for ML/BI algorithms, respectively. References for data retrieved from GenBank, are listed in Table 1.

One 18S rRNA gene sequence (MN700959) possesses 1.39) [1: 1.39]. Oesophagus 142–212 (166) [212]. a single transition G/A. Sequences of 28S rRNA gene Intestinal bifurcation at about midlevel of forebody. were identical to each other. Caeca terminate blindly near posterior end of body. Testes two, entire, ellipsoidal, tandem, separated, Etymology: Latin compound adjective nudidorsalis, from in posterior half of hindbody; anterior testis 283–524 nudus, naked, and dorsalis, dorsal, indicating a lack of (319) × 191–262 (216) [326 × 255], posterior testis 319– the fields of vitelline follicles on the dorsal side of the body. 630 (398) × 163–276 (215) [425 × 226]. Post-testicular region 4.9–15.5 (10.3) [8.4]% of hindbody length. Description of adult specimens: Body elongate, Cirrus-sac elongate, 587–814 (698) × 120–205 (147) flattened dorsoventrally, length 3.48–5.28 (4.08) [616 × 156], reaches into anterior hindbody, contains [3.61] mm, maximum width 0.47–0.70 (0.57) [0.59] bipartite internal seminal vesicle, tubular pars mm at level of ventral sucker. Forebody 24.2–37.7 prostatica surrounded by extensive field of prostatic (32.1) [37.7]% of body length. Tegument spinous. cells, and long invaginated cirrus. Ejaculatory duct Eye-spot pigment scattered at oesophageal level. absent and pars prostatica directly joining proximal Oral sucker subglobular, 212–283 (239) × 219–290 end of invaginated cirrus. Cirrus covered by thin (247) [212 × 226]. Ventral sucker rounded, 170– cytoplasmic filaments and numerous large spines; 262 (205) × 163–276 (209) [198 × 205], sessile, pre- spines length 31–47. Vas deferens absent; vasa equatorial, slightly smaller than oral sucker. Oral efferentia directly joining seminal vesicle. Cirrus-sac sucker to ventral sucker width ratio 1: 0.68–0.95 to ventral sucker length ratio 1: 0.25–0.33 (1: 0.29) [1: (1: 0.84) [1: 0.91]. Mouth opening subterminally. 0.32]. Genital atrium distinct. Genital pore median, Prepharynx distinct, 106–184 (154) [142]. Pharynx immediately anterior to ventral sucker. large, 149–212 (172) × 149–212 (178) [177 × 163]. Ovary entire, rounded to pyriform, 142–262 Pharynx to oral sucker width ratio 1: 1.27–1.62 (1: (187) × 127–226 (174) [180 × 198], pretesticular,

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Figure 2. Skrjabinopsolus nudidorsalis. A, holotype, whole view. B, juvenile voucher specimen, whole view. C, terminal genitalia of voucher specimen, dextro-lateral view. D, ovarian complex of paratype, dorsal view. Abbreviations: c, cirrus (partially evaginated); fc, fertilization chamber; lc, Laurer’s canal, mpo, middle part of oviduct; mt, metraterm, om, oötype with Mehlis’ gland, ov, ovary; pc, prostatic cells, pp, pars prostatica, s, sphincter; sc, spines of cirrus, sm, spines of metraterm; sr, canalicular seminal receptacle, sv, bipartite internal seminal vesicle; vr, common vitelline reservoir. Scale bars: A 1 mm, B 0.4 mm, C 0.15 mm, D 0.2 mm.

© 2020 The Linnean Society of London, Zoological Journal of the Linnean Society, 2020, 190, 448–459 A NEW SPECIES OF DEROPRISTID TREMATODE 455 separated from anterior testes. Ovicapt weakly Excretory pore dorsosubterminal; excretory vesicle developed. Proximal part of oviduct forms tubular elongate oval, occupies 56–73% of hindbody length. fertilization chamber, terminates with sphincter. Middle part of oviduct tubular, receives Laurer’s Remarks: The presence of a short genital atrium, canal and common vitelline duct. Distal part of which does not expand behind the midline of the oviduct forms ootype. Laurer’s canal opens sinistroventral sucker, absence of a crown of large hooks dorso-sublaterally at the level of the ovary or Mehlis’ near the mouth, and absence of a lateral bulge armed gland, occasionally at the level of just the anterior with large spines on the anterior end of the body in to anterior margin of Mehlis’ gland. Canalicular combination with common features for all deropristids seminal receptacle saccular, large, usually between (external seminal vesicular absent, canalicular ovary and anterior testis and partly overlaps them seminal receptacle present, cirrus-sac present, cirrus Downloaded from https://academic.oup.com/zoolinnean/article/190/2/448/5810955 by guest on 02 October 2020 dorsally. Mehlis’ gland compact, anterodorsal to ovary, and metraterm strongly spined, testes in hindbody, contiguous or partly overlapping, median. Uterus suckers unspecialized, ventral sucker pre-equatorial, extends posteriorly to or beyond posterior testis, uterus reaches close to posterior end of body, terminates with massive metraterm. Metraterm 283– vitellarium not reaching posteriorly beyond posterior 517 (402) × 92–205 (153) [403 × 142], runs spiral from testis) are evidence of the new species belonging to the sinistroventral to sinistrodorsal side of cirrus-sac, genus Skrjabinopsolus (compare with: Bray, 2005). sheathed in gland cells. Internal surface of metraterm Skrjabinopsolus nudidorsalis differs from the two with thin cytoplasmic filaments and numerous large congeners by the arrangement of the fields of vitelline spines; spine length 19–38. Metraterm to cirrus- follicles. The arrangement of these fields is limited to sac length ratio 1: 1.08–2.08 (1: 1.75) [1: 1.53]. Eggs the ventral side of the body in published descriptions operculate, 46–51 (48.4) × 25–28 (26.9); shell provided of S. semiarmatus and S. manteri (Ivanov & Murygin, with numerous granular tubercles. Vitellarium 1936; Chulkova, 1939; Osmanov, 1940; Cable, 1952; follicular; follicles in two fields distributed along Bykhovsky & Dubinina, 1954; Edelényi, 1963, 1967, ventral and lateral sides of the hindbody, overlapping 1974; Agapova, 1966; Žitňan, 1966; Čanković et al., caeca and extend from the level of the middle third of 1968; Bykhovskaya-Pavlovskaya & Mikailov, 1969; the cirrus-sac to, or beyond, the midlevel of the anterior Schell, 1970; Mikailov, 1975; Bunyatova & Mikailov, testis, but do not reach the midlevel of the posterior 1991; Bray, 2005). According to our data obtained testis; occasionally single follicles may be present on during study of museum specimens of S. semiarmatus the dorsal side of the hindbody. and S. manteri, the fields of vitelline follicles in these Excretory pore dorsosubterminal; excretory vesicle species are located ventrally, laterally and dorsally. In elongate, oval or claviform, extends to, or beyond, the S. manteri, the left and right fields on the dorsal side posterior margin of the posterior testis, but does not of the body are separated, but in S. semiarmatus they reach beyond the midlevel of the posterior testis. can be separated or confluent (Fig. 3B, C). In the new species, the vitelline fields are absent on the dorsal Description of juvenile specimens: Body elongate, side of the body (Fig. 3A). flattened dorsoventrally, blunt at the anterior end In addition, S. nudidorsalis differs from and tapering at the posterior end; length 550– S. semiarmatus in the surface structure of egg shells. 663, maximum width 150–213 at the level of the The egg shell in the new species has numerous anterior quarter of the hindbody. Forebody 54.7– granular tubercles, but in S. semiarmatus it has 58.3% of body length. Tegument spinous. Eye-spot distinct irregular ridges (Fig. 4A, B). pigment scattered at oesophageal level. Oral sucker subglobular, 85–91 × 82–101. Ventral sucker rounded, sessile, postequatorial, smaller than oral sucker, DISCUSSION 69–76 × 69–76. Oral sucker to ventral sucker width ratio 1: 0.75–0.85. Mouth opening subterminal. Prepharynx The juvenile specimens of S. nudidorsalis described distinct, 25–44. Pharynx large, 57–63 × 57–69. Pharynx in this study are interpreted as being excysted to oral sucker width ratio 1: 1.09–1.89. Oesophagus metacercariae migrated from digested bodies of short, 16–31. Intestinal bifurcation in third-quarter of the second intermediate hosts in the stomach of forebody. Caeca terminate blindly near posterior end the sterlet. Body size, postequatorial position of the of body. ventral sucker and the developmental condition of the Primordiums of testes entire, oblique or almost reproductive system indicate that these specimens tandem, separated, in middle third of hindbody. were in the metacercarial stage. Metacercariae of Primordium of ovary entire, pretesticular, separated the Skrjabinopsolus have been described in only two from anterior testis, median. Primordium of terminal publications. Peters (1961) provided a description genitalia dorsally to ventral sucker. of the metacercaria of S. manteri from unidentified

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Figure 3. Microphotograph of dorsal side of the body. A, paratype of Skrjabinopsolus nudidorsalis; the microphotographs demonstrate the absence of dorsal vitelline follicles in the new species. B, paratype of S. manteri (©2019 F. Goetz, D. Weimer). C, voucher specimen of Skrjabinopsolus semiarmatus. Scale bars 0.5 mm. freshwater oligochaetes and Komarova (1968) reported respectively. However, Bunyatova & Mikailov (1991) on the metacercaria of S. skrjabini (= S. semiarmatus) and Choudhury & Dick (1998) suggested that the life from eurigaline mysid shrimp Limnomysis benedeni cycle of S. semiarmatus can be realized in a freshwater Czerniavsky, 1882. However, parasite species environment too. Undoubtedly, the new species is identification in the work of Komarova (1968) is a freshwater parasite, because the place where the not convincing, because the metacercaria described sterlet was caught is separated from the Caspian Sea by this author does not correspond with adults of by a large section of the River Volga (downstream S. semiarmatus s.l. in the position of the primordia of migrations of sterlet are limited; see: Sokolov & Vasil’ev, the gonads and vitellarium. 1989) and cascades of the Volga’s hydroelectric dams. Skrjabina (1974) attributed S. semiarmatus and A record of juvenile specimens of S. nudidorsalis also S. manteri to marine and freshwater ecological groups, suggests that infection with this parasite occurred at

© 2020 The Linnean Society of London, Zoological Journal of the Linnean Society, 2020, 190, 448–459 A NEW SPECIES OF DEROPRISTID TREMATODE 457 Downloaded from https://academic.oup.com/zoolinnean/article/190/2/448/5810955 by guest on 02 October 2020 Figure 4. Microphotographs of the egg shell. A, paratype of Skrjabinopsolus nudidorsalis. B, voucher specimen of Skrjabinopsolus semiarmatus. Scale bars: 10 μm. the place where the sterlet was caught. The conclusions into the Monorchioidea, because long-tailed, biocellate of Bunyatova & Mikailov (1991) and Choudhury & cercariae with thick-walled excretory vesicle and Dick (1998) about the possibility of completion of the without a stylet are typical of some monorchiids S. semiarmatus life-cycle in freshwater environments (e.g. Cremonte et al., 2001; Gilardoni et al., 2011). are based on records of this species in a freshwater The basal position of Deropristidae relative to other riverine sterlet. It is possible that the data in the Monorchioidea members is consistent with the literature on Skrjabinopsolus in the freshwater ancient origin of the definitive hosts of deropristid riverine sterlet actually refer to S. nudidorsalis. trematodes – acipenseriform and anguilliform fish. The genus Skrjabinopsolus was originally placed These fish groups appeared earlier than the lineage in the family Acanthocolpidae (Ivanov & Myrygin, of clupeocephalan fishes (Hughes et al., 2018), 1936). Skrjabin (1958) moved this genus to the which includes most of the definitive hosts of the family Deropristidae and placed deropristids into the Monorchiidae and Lissorchiidae. superfamily Lepocreadioidea. A few years earlier, Cable & Hunninen (1942) had originally allocated deropristids as a subfamily of Lepocreadiidae. Most authors agreed with K. I. Skrjabin’s view on the family affiliation of ACKNOWLEDGEMENTS Skrjabinopsolus spp. and the superfamily affiliation The authors are deeply thankful to Drs Ilya of deropristids (Peters, 1961; Skrjabina, 1974; Brooks I. Gordeev, Andrey D. Bykov and Dmitry M. Palatov et al., 1985; Gibson, 1996; Choudhury & Dick, 1998; (Russian Federal Research Institute of Fisheries and Bray, 2005; Bray et al., 2009; Bray & Cribb, 2012). Oceanography, Moscow) for help in collecting samples We obtained molecular evidence of the of S. nudidorsalis from the River Oka, and Research phylogenetic position of the genus Skrjabinopsolus Zoologist Dr Anna J. Phillips for essential help with and extrapolated this to the family Deropristidae, examination of the S. manteri collection stored in the based on morphologically substantiated monophyly National Museum of Natural History, Washington DC. of deropristids (see: Choudhury & Dick, 1998). The The present study was financialy supported by the results of phylogenetic analysis allow the consideration Russian Science Foundation, project No: 17-74-20074. of Deropristidae as a family in the superfamily Conflict of interest: The authors declare that they Monorchioidea, which is sister to the Monorchiidae have no conflict of interest to declare. + Lissorchiidae group (Fig. 1). The morphological similarity of deropristids to most other monorchioid trematodes (monorchiids and lissorchiids) is in REFERENCES the anatomy of the terminal genitalia, namely the Agapova AI. 1966. The parasites of fishes from ponds in presence of a cirrus-sac with bipartite seminal vesicle Kazakhstan. Alma-Ata: Izdatelstvo Nauka Kazakhskoi SSR. and spinous cirrus, and also in the large spinous Atopkin DM, Besprozvannykh VV, Ha ND, Nguyen HV, metraterm (see: Bray, 2008; Madhavi, 2008). Among Khamatova AY, Vainutis KS. 2019. Morphometric and Deropristidae, cercariae have been described only for molecular analyses of Carassotrema koreanum Park 1938 Deropristis inflata (Molin, 1859) and S. manteri (see: and Elonginurus mugilus Lu 1995 (Digenea: Haploporidae) Cable & Hunninen, 1942; Seitner, 1951; Peters, 1961). Srivastava, 1937 from the Russian Far East and Vietnam. Cercariae of these species possess a long tail, two Parasitology Research 118: 2129–2137. eyespots, an excretory vesicle with well or moderately Betancur RR, Wiley EO, Arratia G, Acero A, Bailly N, Miya M, thick wall and have no stylet. These features of Lecointre G, Ortí G. 2017. Phylogenetic classification of bony cercariae do not impede the inclusion of deroprestids fishes. BMC Evolutionary Biology 17: 162.

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