taxonID	type	description	language	source
03F3E74EFFDAFFE7CEA85785FE2BFC2F.taxon	description	We found infection with L. intestinalis in freshwater bream from Lakes Onego, Ladoga, Svyatozero and Konchezero. This is the first record of this cestode in bream from these water bodies. Infection indices could not be calculated because the material from these lakes was limited and collected in different years. In Lake Syamozero infection rates of bream with L. intestinalis were low: prevalence 8.9 % with ratio 1.52, intensity of infection from 1 to 3 tapeworms per fish, abundance 0.12 (Fig. 2). These values are similar to those recorded in 1975, when prevalence was 6.7 % and mean abundance was 0.07 (Novokhatskaya et al., 2008). In June 2024, we also examined several individuals of roach R. rutilus and white bream Blicca bjoerkna but did not find any plerocercoids of L. intestinalis in them. Phylogenetic analysis We obtained partial sequences of two mtDNA regions, COI (396 bp) and Cyt b 405 bp), from 28 individuals of L. intestinalis from Karelia. They were used for phylogenetic reconstruction together with the previously published sequences of Ligula spp. from different hosts and localities (Table 1, Fig. 3). Ecologica Montenegrina, 80, 2024, 21 - 37 Table 1 Ab 738 PQ 356679 PQ 329020 Abramis brama Russia: Karelia, Ab 739 PQ 356680 PQ 329021 Abramis brama Russia: Karelia, Ab 740 PQ 356681 PQ 329022 Abramis brama Russia: Karelia, 126 a OP 933968 OP 908173 Abramis brama Russia: 126 b OP 933988 OP 908193 Abramis brama Russia 126 c OP 933969 OP 908174 Abramis brama Russia 126 g OP 933970 OP 908175 Abramis brama Russia: 126 h OP 933971 OP 908176 Abramis brama Russia: brama _ Lip 20 OP 408033 OP 390380 Abramis brama Czech Republic: brama _ Lip 1 OP 408034 OP 390381 Abramis brama Czech Republic: brama _ Lip 2 OP 408035 OP 390382 Abramis brama Czech Republic: brama _ Lip 14 OP 408036 OP 390383 Abramis brama Czech Republic: brama _ Rimov 1 OP 408037 OP 390384 Abramis brama Czech Republic: EE 1 Ab JQ 279121 JQ 279085 Abramis brama Estonia EE 2 Ab EU 241192 EU 241275 Abramis brama Estonia: EE 3 ab EU 241160 EU 241276 Abramis brama Estonia: EE 4 Ab EU 241195 EU 241294 Abramis brama Estonia: EE 5 Ab JQ 279122 JQ 279086 Abramis brama Estonia: FR 30 Ab EU 241201 EU 241259 Abramis brama France: RU 3 Ab EU 241212 EU 241252 Abramis brama Russia: RU 4 Ab EU 241158 EU 241253 Abramis brama Russia: RU 5 Ab EU 241210 EU 241309 Abramis brama Russia: RU 8 Ab EU 241211 EU 241310 Abramis brama Russia: CZ 14 Ab EU 241182 EU 241263 Abramis brama Czech Republic: Nove CZ 16 Ab EU 241179 EU 241283 Abramis brama Czech Republic: Nove CZ 17 Ab EU 241183 EU 241284 Abramis brama Czech Republic: Nove CZ 18 Ab EU 241184 EU 241285 Abramis brama Czech Republic: Nove Table 1 CZ 24 Ab EU 241180 EU 241267 Abramis brama Czech Republic: Nove OK OP 933980 OP 908185 Rutilus rutilus Czech Republic UA 3 OP 933986 OP 908191 Rutilus rutilus Ukraine: CZ 90 Rr EU 241178 EU 241282 Rutilus rutilus Czech Republic: S 4 OP 933972 OP 908177 Rutilus rutilus Iran: Alborz CZ 7 Rr EU 241159 EU 241278 Rutilus rutilus Czech Republic: CR 22 OP 933981 OP 908186 Rutilus rutilus France: FR 67 Aa JQ 279124 JQ 279088 Alburnus alburnus CZ 106 Pc EU 241167 EU 241244 Podiceps cristatus eryth _ Most 1 OP 408040 OP 390387 Scardinius erythrophthalmus France: Czech Republic: Czech Republic: blicc _ Lip OP 408038 OP 390385 Blicca bjoerkna Czech Republic: MCf-FB 04919371 MW 602520 Pusa hispida saimensis Finland: Lake IE 2 Rr EU 241206 EU 241250 Rutilus rutilus Ireland: Lough GB 2 Pp EU 241175 EU 241304 Phoxinus phoxinus United Kingdom: Ru OP 933995 OP 908201 Hemiculter lucidus Russia: Lake CN 1 Hb EU 241153 EU 241229 Hemiculter bleekeri China: lac AU 1 Gt EU 241146 EU 241222 Galaxias truttaceus Australia Au 2 OP 933951 OP 908156 Galaxias maculatus Australia: Goodga Au 3 OP 933952 OP 908157 Galaxias maculatus Australia: TN 63 Ps JQ 279139 JQ 279102 Pseudophoxinus callensis Tunisia: Remel, IE 3 Gg EU 241188 EU 241305 Gobio gobio Ireland: Lough ALG 1 Bc JQ 279109 JQ 279074 Barbus sp. Algeria IE 4 Gg EU 241208 EU 241290 Gobio Gobio Ireland: Lough H 11 OP 933994 OP 908199 Neogobious Hungury: ALG 2 Bc EU 241143 EU 241219 Barbus sp. Algeria: Hamiz TN 62 Ps JQ 279138 JQ 279101 Pseudophoxinus callensis Tunisia: Joumine CN 4 Nt EU 241157 EU 241237 Neosalanx taihuensis China: Zhanghe Ecologica Montenegrina, 80, 2024, 21 - 37 Table 1 CN 8 OP 933997 OP 908203 Neosalanx taihuensis China CA 19 Sa EU 241150 EU 241224 Semotilus atromaculatus CA 5 Sa EU 241149 EU 241226 Semotilus atromaculatus Oregon _ C 4 OP 934005 OP 908211 Rhinichthys osculus CA 1 Cp EU 241152 EU 241228 Couesius plumbeus Canada Canada Canada: Mckenzie Canada ET 2 OP 934000 OP 908206 Barbus sp. Ethiopia ET 5 OP 934001 OP 908207 Barbus sp. Ethiopia K 1 CO OP 934009 OP 908215 Barbus sp Kenya 1 c OP 934010 OP 908216 Rastrineobola argentea Kenya C 2 OP 934011 OP 908217 Rastrineobola argentea Kenya C 3 OP 934012 OP 908218 Rastrineobola argentea Kenya 4 c OP 934013 OP 908219 Rastrineobola argentea Kenya 5 c OP 934014 OP 908220 Rastrineobola argentea Kenya 6 c OP 934015 OP 908221 Rastrineobola argentea Kenya 7 c OP 934016 OP 908222 Rastrineobola argentea Kenya 8 c OP 934017 OP 908223 Rastrineobola argentea Kenya 9 c OP 934018 OP 908224 Rastrineobola argentea Kenya 10 c OP 934019 OP 908225 Rastrineobola argentea Kenya K 3 CO OP 934020 OP 908226 Rastrineobola argentea Kenya K 4 CO OP 934021 OP 908227 Rastrineobola argentea Kenya Tanz 2 OP 934022 OP 908228 Engraulicypris sardella Tanzania Tanz 3 OP 934023 OP 908229 Engraulicypris sardella Tanzania Tanz 8 a OP 934024 OP 908230 Engraulicypris sardella Tanzania Bn 151 OP 934025 OP 908231 Barbus anoplus South SA _ Mpum OP 934026 OP 908232 Barbus anoplus South DRC OP 934027 OP 908233 Barbus sp Democratic Republic Table 1 Namibia 2 OP 934028 OP 908234 Barbus paludinosus Namibia SA _ Limpop OP 934029 OP 908235 Barbus anoplus South Namibia 1 OP 934030 OP 908236 Barbus paludinosus Namibia SA _ Buff OP 934031 OP 908237 Barbus anoplus South Bn 159 OP 934032 OP 908238 Barbus anoplus South NZ 2 OP 934033 OP 908239 Gobiomorphus breviceps New NZS OP 934034 OP 908240 Gobiomorphus breviceps New Ecologica Montenegrina, 80, 2024, 21 - 37 The phylogenetic analysis (BI) and maximum Likelihood (ML) analysis of concatenated partial genes showed that there were six geographically distinct lineages within the monophyletic L. intestinalis complex (Fig. 3). This result is consistent with the previous studies (Bouzid et al. 2008; Štefka et al. 2009; Nazarizadeh et al., 2022, 2023). On the phylogenetic tree, all samples from Karelia were placed in the clade “ L. intestinalis Lineage A ”, which included parasites of various cyprinid fish from around the world. The clade position was supported by high posterior probability (1.0) and bootstrap support (100) (Fig. 3). Haplotype analysis Haplotype analysis was performed for the sequences of L. intestinalis from freshwater bream obtained in this study and those available in GenBank. The indices of genetic diversity in L. intestinalis datasets of concatenated sequences of cox 1 and cytb and separate datasets of the same genes are presented in Table 2. Haplotype analysis of the 53 concatenated sequences revealed 40 haplotypes. Most of the individual plerocercoids had their own unique haplotype (Fig. 4). Only six haplotypes were shared by two or more tapeworms. The most numerous haplotype was noted in four individuals of the parasite in Lake Syamozero as well as in a cestode from Lake Řimov in Czech Republic. One haplotype was identified in three tapeworms from different geographical localities (Estonia, Czech Republic, Rybinsk). Another haplotype was detected in three cestodes from Rybinsk. Two other haplotypes were found in two plerocercoids from Lake Syamozero each (Ab 643 / Ab 649 and Ab 665 / Ab 666). One haplotype was noted in two tapeworms: one from Lake Syamozero and the other from Lake Konchezero (Ab 657 and Ab 660). Out of the 40 haplotypes revealed in our study, 22 haplotypes were found in specimens from Karelian lakes, and 21 of them were unique. Each plerocercoid sampled from bream in Lake Ladoga and Lake Onego had its own unique haplotype (Fig. 4). Fifteen haplotypes, each corresponding to an individual tapeworm, were found in cestodes from Lake Syamozero (Fig. 4). Within Lake Syamozero, three haplotypes represented by the greatest number of tapeworms were sampled in the Kurmoila Bay (Fig. 1), which might be associated with the larger amount of material sampled in this locality. P-distances of concatenated cytb + cox 1 of samples from Karelian lakes varied from 0.1 to 2.3 %. P-distances of the same markers between the Karelian samples and the tapeworms from Rybinsk, Estonia, Czech Republic and France were 0.45 %, 0.75 %, 0.7 % and 0.5 %, respectively. Examination of partial sequences separately for cox 1 and Cyt b revealed different patterns (Table 2). All Cyt b sequences were divided into 26 haplotypes. Karelian tapeworms were represented by 13 haplotypes, including 10 unique ones. At the same time, one of the most common haplotypes was noted only in Karelian specimens from lakes Syamozero, Konchezero, Onego, Ladoga (Ab 650, Ab 656, Ab 657, Ab 660, Ab 661, Ab 663, Ab 667, Ab 740). One haplotype was shared by five cestodes from Syamozero (Ab 641 Ab 646 Ab 736 Ab 738 Ab 739) and worms from Czech lakes Rimov and Lipno. One haplotype of tapeworms from Syamozero and Onego (Ab 640, Ab 643, Ab 649) was shared with those from Rybinsk reservoir (RU 3 Ab) and Lake Pepsi (EE 2 Ab) in Estonia. Another haplotype was common for tapeworms from Lake Syamozero and Lake Pepsi (EE 4 Ab) in Estonia. The least variable site was cox 1: 17 haplotypes in total and 12 haplotypes in Karelia (Table 2). Five haplotypes were unique for L. intestinalis from Karelian bream, while all the others were shared with tapeworms from other geographical locations. The best-represented haplotype was found in 11 tapeworms from Karelia (Syamozero, Konchezero) and 11 cestodes from Rybinsk, Estonia, Czech Republic and France. Two less common haplotypes, each in a different group of worms, were identified in two cestodes from Syamozero and four tapeworms from Rybinsk. Similarly, two haplotypes were found in two different groups including tapeworms from Syamozero and Estonia. Haplotypes of cestodes from Ladoga and Svyatozero coincided with those from Czech reservoirs.	en	Lebedeva, Daria I., Kochneva, Albina A., Lysenko, Lydmila A., Kantserova, Nadezda P., Zaitsev, Dmitry O., Milyanchuk, Nikolay P., Sukhovskaya, Irina V. (2024): Mapping of Ligula plerocercoids in the freshwater bream Abramis brama in Lake Syamozero and some other lakes of Northwestern Russia. Ecologica Montenegrina 80: 21-37, DOI: 10.37828/em.2024.80.3, URL: https://doi.org/10.37828/em.2024.80.3
03F3E74EFFDAFFE7CEA85785FE2BFC2F.taxon	discussion	Discussion Our results indicate an expanding dispersal of the cestodes Ligula intestinalis parasitizing freshwater bream in Karelia. One of the reasons is the dispersal of the bream itself, which has been noted in ichthyological studies (Sterligova et al. 2016). In Lakes Onego, Ladoga, and Svyatozero, where we recorded L. intestinalis in bream for the first time, these parasites had been previously recorded in other fish: roach Rutilus rutilus and crucian carp Carassius carassius L., 1758 (Rumyantsev 2007) in Lakes Onego and Svyatozero and roach R. rutilus, vimba bream Vimba vimba L., 1758, blue bream Ballerus ballerus L., 1758 and bleak Alburnus alburnus L., 1758 (Rumyantsev & Mamontova 2008) in Lake Ladoga. The infection rates in all these water bodies were low (prevalence less than 7 %, mean abundance 0.1). In Lake Konchezero, freshwater bream was introduced in the 1960 s (Sterligova et al. 2016) and has acclimatised. Our results show that its parasitic fauna now includes L. intestinalis. Lake Syamozero was the only Karelian lake where plerocercoids of L. intestinalis have been recorded in bream before the present study. The only other host in which L. intestinalis has been noted in Lake Syamozero is bleak, A. alburnus, and infection indices are low (prevalence 6 %, mean abundance 0.06) (Novokhatskaya 2008). No infection of freshwater bream with L. intestinalis had been noted in Syamozero in the 1950 s (Shulman 1962) (Fig. 2), the first record dating back to 1973 (Malakhova & Ieshko 1977). Since that time, the abundance of this parasite has varied, the fluctuations being possibly associated with the state of the bream population. In 1970 s- 1990 s, it was mainly represented by immature individuals (about 70 %); the maturation rates were slow, and the fecundity was low. This long-term depression was probably due to fishing restrictions and eutrophication of the lake. The numbers of bream increased as a result of long-term ban on bream fishing, and there was not enough benthos, which is an energy-rich resource, for all the bream in the lake. Moreover, eutrophication, caused by the use of fertilizers, resulted in a depression of the benthic communities, while the abundance and biomass of plankton increased. Under these conditions, bream mostly fed on zooplankton, which is a low-energy resource. The role of copepods in bream diet became more significant, and the infection rates of bream with L. intestinalis plerocercoids increased correspondingly. The parasite probably depressed the growth rate of the host even further (Ieshko & Malakhova 1982; Novokhatskaya et al. 2008; Sterligova et al. 2016). Current infection rates of bream with L. intestinalis are similar to those from 1975, which indicates that the share of plankton in the bream diet is fairly high. We provided new gene sequences of L. intestinalis from A. brama and identified new haplotypes. Haplotype diversity was high both for the parasites from different countries and for Karelia (0.98 – 0.99), but the nucleotide diversity was low (0.000.006 – 0.007). Tajimaʼs D values were negative both in Europe and in Karelia, with statistically insignificant values, suggesting that L. intestinalis population in freshwater bream in the European part of Palearctic is genetically diverse and rapidly expanding. Our data support the hypothesis, based on historical demography modeling, that isolation with continuous gene flow is the most likely scenario of the divergence of L. intestinalis (Nazarizadeh et al. 2024). All the samples of plerocercoids involved in our study were placed into Lineage A of L. intestinalis (Nazarizadeh et al. 2023). The authors have suggested that L. intestinalis from freshwater bream have certain haplotypes that are almost never found in other cyprinids (Nazarizadeh et al. 2022). Having examined water bodies situated at a distance of 50 - 150 km from each other, they concluded that the differences in prevalence between fish host species in different lakes might be influenced not only by the parasite’s ecology but also by its genetic diversity (Nazarizadeh et al. 2022). We arrived at the same conclusion in this study. Different haplotypes of L. intestinalis from bream could be found in the same location in lake (e. g. Kurmoila Bay of Lake Syamozero), while the same haplotypes could be found in locations separated by a distance of 5 - 20 km within the lake. Nazarizadeh et al. (2022) note that the heterogeneity of the helminth population in the sample of bream is due to the fact that the material is collected in different seasons. We caught bream individuals in Kurmoila Bay for 12 days in June 2024 but our sample was also rather heterogeneous (22 haplotypes). Large numbers of bream spawn and migrate in the lake in the study period, and their populations mix, which affects the diversity of the parasites (Sterligova et al. 2016). In summer, the mixing and dispersal of the plerocercoids is facilitated by the feeding of young bream from different populations in shallow and well-warmed littoral areas and numerous bays of Lake Syamozero. Conditions are favourable there for copepods, which are the first intermediate hosts of L. intestinalis. Accumulations of young fish in such places attracts the final hosts, fish-eating birds. Large numbers of gulls (Black-headed Gull, Chroicocephalus ridibundus and Herring Gull, Larus argentatus), which are probably the main hosts of L. intestinalis, have been observed at Lake Syamozero (Sazonov 2004). In this way, the transmission of the parasite in the ecosystem is promoted. Similar results have been obtained in a study of Ligula circulation in some aquatic ecosystem in south-western Spain (Capasso et al. 2024). Variation in the occurrence of haplotypes of L. intestinalis in different locations may be due to the different species of fish-eating birds, their definitive hosts, as well as their migration pathways. Among the definitive hosts of Ligula intestinalis indicated by Dubinina (1980), different species of gulls, goosanders, grebes, cormorants, making both short- and long-distance migrations for wintering or nesting, are found in Northwestern Russia (Noskov et al. 2016). Nazarizadeh et al. (2022) suggest that fish-eating birds such as Great Cormorant P. carbo, grebes Podiceps auritus (Linnaeus), P. cristatus and P. nigricollis Brehm, Goosander Mergus merganser, and Common Pochard Aythya ferin a (Linnaeus) may be potential final hosts of L. intestinalis in the Czech Republic. So far, L. intestinalis has been noted in Karelia only in Great Crested Grebe Podiceps cristatus and Great Cormorant Phalacrocorax carbo (Dubinina 1980; Yakovleva et al. 2020) but this may be due to the limited scope of parasitological research. Though P. carbo has been shown to expand into the water area of Lake Ladoga, this bird has not yet been noted at Syamozero, Onego, Svyatozero and Konchezero (Lapshin & Mikhaleva 2021). Our molecular data on L. intestinalis indirectly support the connections between water bodies on the migratory routes of fish-eating birds, particularly, gulls, discovered by Noskov et al. (2016). The numbers of Herring Gull and Black-headed Gull in the Karelia has been increasing in recent decades (Zimin et al. 1993; Noskov et al. 2016). Numerous colonies of these birds are observed along the shores of many Karelian water bodies, and local residents report that gulls often feed on discarded fish. These species of fish-eating birds may transmit L. intestinalis both between lakes within Karelia and between Karelia and Europe. Black-headed Gull and Herring Gull, wintering on the southern coast of the North Sea and the Baltic Sea (Noskov et al. 2016), are likely to maintain a more homogeneous population of the parasite within the northern part of their range. This hypothesis is supported by the occurrence of the same haplotypes in bream in the Karelian water bodies examined in our study and in water bodies in other regions (Fig. 4). The study of the first intermediate hosts of L. intestinalis, i. e. crustaceans, would be interesting for identification of factors influencing the dispersal rate, the survival and the host specificity of this parasite. The life products of bird colonies strongly influence the zooplankton in the littoral zone of freshwater bodies (Krylov et al. 2012). The birds do not only release infective agents into water but also change the structure and abundance of zooplankton. These changes are likely to affect the implementation of the life cycle of L. intestinalis and the survival of its lineages / subspecies. Conclusion In this study, we obtained data on the occurrence of the cestode L. intestinalis in freshwater bream inhabiting several lakes in Northwestern Russia and examined the genetic structure of its plerocercoids using two mitochondrial genes (Cyt b and COI). Our results highlight the need to study this parasite in other fish of the region in order to understand its specificity to the second intermediate host. It is also important to obtain the data on the bird species that serve as the main infection vectors of L. intestinalis. These data would contribute to epidemiology, control and treatment options of Ligula infection. Acknowledgements We are grateful to our colleagues, particularly to Drs. Olga Sterligova, Eugeny Ieshko, Sergey Bugmyrin, and Fedor Fariseev (IB KarRC RAS), for their help with material collection. We extend our sincere thanks to the two anonymous reviewers for their valuable comments on the first version of the manuscript. The study was funded by the Russian Science Foundation, project no. 24 - 26 - 00251. Ethics Approval The paper does not contain any studies involving animal experiments. The wild animal study protocol was approved by the Institute of Biology of Karelian Research Centre, the Russian Academy of Sciences (protocol no. 7 of 8 July 2023). Research Fishing was under Permit of North-West Territorial Administration of the Federal Agency for Fishery (7820240317689) of 14 May 2024.	en	Lebedeva, Daria I., Kochneva, Albina A., Lysenko, Lydmila A., Kantserova, Nadezda P., Zaitsev, Dmitry O., Milyanchuk, Nikolay P., Sukhovskaya, Irina V. (2024): Mapping of Ligula plerocercoids in the freshwater bream Abramis brama in Lake Syamozero and some other lakes of Northwestern Russia. Ecologica Montenegrina 80: 21-37, DOI: 10.37828/em.2024.80.3, URL: https://doi.org/10.37828/em.2024.80.3
