Introduction
Syngnathid fishes are a typical component of fish communities in the Black Sea shallow coastal waters, and are very important in terms of biodiversity. Seven species of pipefishes (Nerophis ophidion (Linnaeus, 1758), Syngnathus abaster Risso, 1827, S. acus Linnaeus, 1758, S. schmidti Popov, 1927, S. tenuirostris Rathke, 1837, S. typhle Linnaeus, 1758, S. variegatus Pallas, 1814) and one, Hippocampus hippocampus (Linnaeus, 1758) (Vasil’eva, 2007), or two (as believe Bilecenoglu et al. (2014) meaning the presence also of H. guttulatus Cuvier, 1829) species of seahorses, have been known from there.
These small fishes are also known to contribute to the helminths life cycle that extending to water birds and some predatory Black Sea fish (Gaevskaya & Korniychuk, 2003). Only a few specimens of the Black Sea Syngnathidae fish were studied by helminthologists in the middle of XX century; so, information on the species composition of Syngnathidae helminths in the sea is extremely scarce even in relation to the Black Sea coast of the Crimea – the Black Sea region, the parasite fauna there is being studied most thoroughly (Gaevskaya & Korniychuk, 2003). Some modern data on Syngnathidae helminth fauna in the Black Sea are available (Kornyychuk et al., 2016); nevertheless, corrections are needed now in taxonomic status of some parasite species had previously been listed as well as appropriate quantitative data.
Being small-size fishes, Syngnathidae are not active migrants (Svetovidov, 1964; Vasil’eva, 2007) and one could expect existence of different local population of Syngnathidae in the Black Sea (at least, there are evidence for special S. typhle population in the Sea of Marmara and the Black Sea (Wilson & Veraguth, 2010), with originality of their parasite fauna.
Syngnathid fishes are known as endangered part of some Black Sea coastal marine ecosystems: S. tenuirostris, S. typhle, H. hippocampus are included to the Red Book of Crimea (Ivanov, Fateryga, 2015) and Black Sea Red Data Book (Dumont, 1999), S. variegatus – to Red Book of Crimea (2015) and they can certainly be affected by physical degradation and destruction of their habitats, chemical pollutants, eutrophication, sedimentation of dispersed silts on seagrasses or other changes in water quality, intensive mining of sea sands, large-scale hydraulic construction, noise pollution, invasive species and even climate change (Boltachev & Karpova, 2012; Felipe et al., 2016) as well as by parasites and diseases and helminthosis are one of most important of them. Nevertheless, being important part of coastal marine ecosystems, they are not commercially important food fishes and that’s why Syngnathidae are still little studied fish hosts in the Black Sea. So, the purpose of the investigation was to study, based on original data, the modern state of helminth fauna of the Black Sea Syngnathidae off Crimean coasts, with special attention to helminths as possible fish disease agents, and to specify the role these fishes play in helminths parasite systems using quantitative indices.
Results
Fifteen helminth species were found from the studied pipefishes and seahorses inhabiting the biocenoses of the Black Sea shelf of the Crimea, they were six Trematoda, three Cestoda, four Nematoda and two Acanthocephala (Table 1). All the parasites found are heteroxenous (with multiple-host life-cycle) utilizing the fish hosts as definitive, intermediate or paratenic hosts.
Trematoda species recorded during the course of the study were two species of Cryptogonimidae Ward, 1917 family, namely Timoniella imbutiformis (Molin, 1859) Brooks, 1980 – maritae and metacercaria (in fish gills, muscles, serosa) and Aphallus tubarium (Rudolphi, 1819) Poche, 1926 maritae, three Heterophyidae Leiper, 1909 – Galactosomum lacteum (Jägerskiöld, 1896) Looss, 1899 metacercaria in fish gill muscles, Pygidiopsis genata Looss, 1907 metacercaria in fish hosts gut walls and Cryptocotyle concava (Creplin, 1825) Lühe, 1899 metacercaria in gills, serosa and fish gut wall as well as Opecoelidae gen. sp. metacercaria in gills, serosa and gut walls of fish.
The most diverse digenean fauna among all studied regions was registered in Karkinit Bay (five species).
The fauna of Syngnathidae cestodes near Crimean Black Sea coasts revealed to be very poor, it is represented by larvae of three species from three orders: Bothriocephalus gregarius Renaud Gabrion & Pasteur, 1983 nomen nudum (Bothriocephalidea Kuchta, Scholz, Brabec & Bray, 2008), Progrillotia dasyatidis Beveridge, Neifar & Euzet, 2004 (Trypanorhyncha Diesing, 1863) and Scolex pleuronectis Müller, 1788 (“Tetraphyllidea” Van Beneden, 1850 relics) species complex (Table 1). Of the three fish species examined, the larvae of all these cestodes were found only in S. abaster in the estuary of the Chernaya River. Infectiveness of S. abaster with B. gregarius plerocercoids is rather high (Table 1) but this parasite species was found only in 4 % of the examined fish. P. dasyatidis plerocercoids was found once and only in one specimen of S. abaster, this implies an accidental parasitization of this host.
Four nematodes species ending their development in waterfowl, marine and brackish water fish were found in Syngnathidae studied, they belong to three families: Anisakidae Skrjabin & Karokhin, 1945, Raphidascarididae Hartwich, 1954, and Acuariidae Railliet, Henry & Sisoff, 1912 (Table 1). Indexes of invasion of needlefish by nematode larvae were extremely low (Table 1) that means minor significance of Syngnathidae in the parasite systems of these helminths in the Black Sea.
All larvae of nematodes of the genus Contracaecum found in needlefish were at the third-stage larva of development; larvae of C. rudolphii (spiculigerum) Hartwich, 1964 were encapsulated in the intestinal walls and on the mesentery of host fish, and C. multipapillatum (Drasche, 1882) Lucker, 1941 larvae were localized in the bile ducts of fish.
Only one representative of the Nematoda Raphidascarididae family, H. aduncum, was found. In the studied Syngnathidae fish, most of the larvae were at third-stage of development and were located on the mesentery in the body cavity of the fish; once, an immature female was found in the intestinal lumen of S. abaster (Table 1).
Acanthocephalans in gut of Syngnathidae studied were represented by two species, Acanthocephaloides propinquus (Dujardin, 1845) Meyer, 1933 and Telosentis exiguus (von Linstow, 1901) (Table 1). All the found worms (A. propinquus, a male and T. exiguus, a mail and a female) were immature.
As some of the fishes studied were caught in marine protected areas, we for the first time list the parasites found there as a part of their fauna check-lists:
The Crimean Nature Reserve, “Swan Islands” branch:
Trematoda: Timoniella imbutiformis met., maritae; Aphallus tubarium maritae; Galactosomum lacteum met.; Cryptocotyle concava met.; Opecoelidae gen. sp. met.
Nematoda: Contracaecum rudolphii (spiculigerum) larvae; Contra caecum multipapillatum larvae; Hysterothylacium aduncum larvae III; Cosmocephalus obvelatus larvae III.
Acanthocephala: Telosentis exiguus.
The State Zoological Reserve “Kazachya Bay”
Trematoda: Timoniella imbutiformis met.; Pygidiopsis genata met.
Discussion
The pipefishes and a seahorse studied are Atlantic-Mediterranean species broadly distributed along Black Sea coasts and inhabiting seaweeds at depth up to about 20 m (Vasil’eva, 2007). In the Black sea they are known to feed with small-sized prey: fish fries and small-sized adult fishes and crustaceans (Svetovidov, 1964; Vasil’eva, 2007; Gürkan & Uncumusaoglu, 2012); on the other hand, they themselves are known in the Black Sea as food objects of rays Raja clavata Linnaeus, 1758 (Saglam & Bascinar, 2008), dolphins Delphinus delphis ponticus Barabasch-Nikiforov, 1935 (Svetovidov, 1964), turbots Scophthalmus maeoticus (Pallas, 1814) fry – under experimental conditions (Shishkina et al., 2007), sea gulls Larus argentatus Pontoppidan, 1763 (Sten’ko, 1983) that determine complex ways of helminth circulations in Black Sea coastal zone due to presence of pipefishes and seahorses.
Trematoda are the most constant and most numerous (meaning species number) part of the Syngnathidae helminth community (40 % of the total number of registered helminth species). They are represented mainly by larval stages (Table 1) of which three species finish development in birds and two are in fish. All species of trematodes, the metacercariae of which are found in needlefish, are broadly specific at this developmental stage to the Black Sea bottom pelagic fishes (generalists).
T. imbutiforme was found from Syngnathidae hosts in all localities, contributed substantially to the observed trematode community’s composition studied and is considered to be the most common (“core”) digenean species for the Black Sea S. typhle: maritae infect more than 50 % of the broadnosed pipefishes studied; at metacercarial stage this species play the same role for H. hippocampus as they were found in more than 15 % of long-snouted seahorses.
Other trematodes found in the studied Syngnathidae were recorded sporadically and at low rates of invasion. Most all of them are generalists, marine and brackish species (excepting marine species, A. tubarium, which is specific to Syngnathidae only at marita stage).
Black Sea seahorses, black-striped and broadnosed pipefishes revealed to be intermediate hosts of trematodes C. concava and P. genata (Heterophyidae) known as broad specific ones to bottom-dwelling and pelagic Black Sea fishes (generalists) (Gaevskaya & Korniychuk, 2003). The life cycle of these trematodes is complex and the adult parasites are located in the piscivorous bird’s intestine; of P. genata and C. concava are known from the area of Karkinit Bay in several bird species and are numerous: for example, in Larus argentatus abundance of P. genata was 47 and C. concava 331 ind. per host (Sten’ko, 1983) indicating that these helminths are core species in the local parasite fauna. The cercaria of these trematodes penetrate the skin of a fish, encyst there becoming metacercaria and marking parasitized individuals with visible to the naked eye black dots (produced by host melanocytes migrating through the cysts’ fibrous wall) and cause so called “black spot disease” (BSD) (Stunkard, 1930; Bush et al., 2001). Heavy infections by metacercaria can reduce fish host physical condition and survival (Sindermann, 1990).
The cestodes B. gregarius is a nomen nudum (Kuchta & Scholz, 2017) and before the publication of its description from the type host from the native range, we use this name accompanying it with this mark. B. gregarius is a typical marine species but it can be transferred into estuarine biocenoses by their hosts; earlier, larvae of this species were registered from S. abaster in the Lake Donuzlav (Miroshnichenko, 2008), the salinity of which is equal to that of the Black Sea (18 – 19 ‰) (Ivanyutin, 2019). We found B. gregarius plerocercoids without blastocyst both as at the Black Sea typical salinity (at the mouth of the Chernaya River) as in fresh waters (upstream of the Chernaya River).
According to (Solonchenko, 1982), the only second intermediate hosts of B. gregarius in the Black Sea are Gobiidae. Probably, the black-striped pipefish is also the second intermediate host for this cestode species as the cestode larvae we found in pipefish were at the plerocercoid stage without blastocyst, had a well-formed scolex with slit-like bothria, and the infection of S. abaster was high (Table 1). In addition, planktonic crustaceans, in particular, Acartia (Acartiura) clausi Giesbrecht, 1889 (the first intermediate host of B. gregarius) are included in the spectrum of food items for both Gobiidae and Syngnathidae (Svetovidov, 1964; Solonchenko, 1982; Vasil’eva, 2007). And, finally, the final host for B. gregarius, the kalkan flounder Scophthalmus maeoticus (Pallas, 1814), feeds on both Gobiidae (Svetovidov, 1964; Vasil’eva, 2007) and Syngnathidae (Shishkina et al., 2007). Consequently, the infection of the flounder can occur both through the gobies that dominate in its diet, and through the pipefishes. As S. abaster can feed on larvae and juvenile fish, including Gobiidae (Didenko et al., 2018), infection of black-striped pipefish with B. gregarius plerocercoids can occur not only through crustaceans, but also through infected small gobies.
The P. dasyatidis larvae we found in the Black Sea S. abaster were at the plerocercoid stage without blastocyst. The life cycle of this cestode species is not known; we recorded earlier it’s plerocercoids in 8 species of Black Sea teleost fishes from other families (second intermediate hosts), with much higher indexes of invasion (Polyakova et al., 2017; Polyakova, 2020). The very low infection rates of black-striped pipefish with P. dasyatidis larvae implies that S. abaster is an occasional host for these cestodes. There are no data on the feeding of Dasyatis pastinaca (the final host of this cestode species) by pipefishes and seahorses; however, Hippocampus sp. and Syngnathus sp. are known from the stomachs of other Black Sea rays, Raja clavata Linnaeus, 1758, (Saglam & Bascinar, 2008). We hope, further detailed studies of Cestodes in Syngnathidae fish will help to reveal the role of these fish in the life cycle of B. gregarius and P. dasyatidis in the Black Sea.
Larvae of Contracaecum Railliet & Henry, 1912 nematodes were previously recorded in the Black Sea from S. typhle and S. abaster and were identified as C. microcephalum (Chernyshenko, 1955, Gaevskaya et al., 1975, Mange, 1993). Previously (Korniychuk et al., 2016) we also mentioned C. microcephalum (Rudolphi, 1809) l. and C. rudolphii Hartwich, 1964 l. from both S. typhle and H. guttulatus on the basis of conventional morphological features (Gaevskaya et al., 1975; Moravec, 1994). Nevertheless, now we reinvestigated the collected material and believe the larvae of nematodes of the genus Contracaecum from the pipefish and the seahorse we previously mentioned as C. microcephalum and C. rudolphii (Korniychuk et al., 2016), morphologically corresponding to the larvae of two another species of the genus, namely, C. rudolphii (spiculigerum) and C. multipapillatum; their presence in the Black Sea is confirmed on the basis of analysis of nucleotide sequences of partial LSU rDNA and ITS rDNA (Pronkina & Spiridonov, 2018, 2020). So, it is impossible now to say exactly what species these larvae mentioned by (Chernyshenko, 1955, Gaevskaya et al., 1975, Mange, 1993) belong to, but we have no reasons to deny it wasn’t C. microcephalum.
The range of definitive hosts of Contracaecum spp. nematodes is quite wide; adults and LIV parasitize fish-eating birds and marine mammals. Piscivorous birds are the most significant (main) final hosts in the Azov-Black Sea basin; C. rudolphii is more common in cormorants Phalacrocorax Brisson, 1760 and C. multipapillatum – in pelicans Pelicanus Linnaeus, 1758 (Smogorzhevskaya, 1976; Kornyushin et al., 2004). The first intermediate hosts of C. rudolphii were identified only in experiments (crustaceans) (Mozgovoy et al., 1968; Moravec, 2009); the natural first intermediate hosts for both C. rudolphii and C. multipapillatum have not been found yet. Contracaecum spp. third-stage larvas are known from marine, brackish and freshwater fish. We assume that the main way of realizing the life cycle of these nematodes is trixenous and fish in the Black Sea (including Syngnathidae) are obligatory intermediate hosts and definitive hosts (fish-eating birds) become infected while eating them.
Syngnathidae infection with nematodes H. aduncum third-stage larva occurs when fish eat infected planktonic crustaceans (first intermediate hosts). In this case, the nematodes penetrate the intestinal walls of the fish and encyst on the serous membrane of their internal organs, i.e. Syngnathidae are the second intermediate hosts of these nematodes. The find of an immature H. aduncum female in the intestinal lumen of S. abaster speaks in favor of the infection of this pipefish via infected pelagic planktivorous fish. Nematodes C. obvelatus belongs to fam. Acuariidae which finish their development in birds and are known as epizootics agents of domestic and wild ducks (Smogorzhevskaya, 1976; Kornyushin et. al., 2004). The larvae of these nematodes can enter the final host, birds, in two ways: directly through amphipods and mysids (first intermediate hosts) or through fish (paratenic hosts) previously infected by nematodes larvae via amphipods and mysids (Moravec, 1994). Probably, both ways are possible in the case of pipefish infection.
Acanthocephala A. propinquus and T. exiguus are generalists, but A. propinquus mature and abundant mainly in Gobiidae (Kvach, 2006) and T. exiguus – in pelagic fish, mainly in Atherinidae (Belofastova & Grintsov, 2005). Judging by the extremely low rates of infection and as the worms we found were not sexually mature, pipefishes and seahorses are occasional or abortive hosts for the both species of acanthocephalans in the studied areas of the Black Sea. Nevertheless, immature A. propinquus could move via food nets: it is known that the final host of A. propinquus, the grass goby Zosterisessor ophiocephalus (Pallas, 1814), becomes infected not only when eating isopods Idotea balthica (Pallas, 1772) (the first intermediate host), but also due to cannibalism and predation on other fish, including Syngnathidae (Smirnov, 1986).
As mainly larval helminthes persist in Syngnathidae fishes, we believe no reason to expect significant seasonal fluctuations in the composition of syngnathids helminths fauna.
As pipefishes and seahorses are fishes with low mobility and small home ranges, marine reserves are thought to be effective for conservation purposes. Conservation statuses for H. hippocampus is “no available information”, “vulnerable” for S. abaster, “least concern” for S. typhle (Yankova et al., 2014). In the region studied, the pipefishes and the seahorse occur in at least two marine protected areas, namely Crimean Nature Reserve (“Swan Islands” branch in Karkinit Gulf) and State Zoological Reserve “Kazachya Bay” in aquatoria of Sevastopol city. Moreover, all the studied Syngnathidae species are abundant there and are the basis part of the faunistic diversity in the western part of Karkinit Gulf (Belogurova et al., 2020).
List of major threats protected marine fishes are affected in the Black Sea (Yankova et al., 2014) doesn’t include fish diseases and helminthoses. Nevertheless, it was shown (Rosenqvist & Johansson 1995; Mazzi 2004) that Cryptocotyle-BSD affect S. typhle female fecundity (unparasitized females provide fertilizing more eggs) and sexual attractivity (males preferentially mate with fish females with lower Cryptocotyle cysts loads). Some researchers associate BSD also with growth retardation, body weight loss and higher mortality of young infected fish, which could regulate the host population density (Lemly & Esch, 1985). Anisakidae larvae (for example, C. rudolphii) are also known to cause lesions of fish host pancreas due to mechanical compression and displacement of liver in the case of high infection (destruction of the parenchyma, disorders of blood microcirculation, fatty degeneration of hepatocytes) and inflammation of the bile ducts (Hauck & May, 1977; Popuk, 2017). So, some helminths we found from Syngnathids in the frames of this study are potentially important as threats in marine protected aquatoria being potential agents of pipefishes and seahorses’ diseases.
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