Barbatula Toni Research Articles

Species diversity and structure of the fish community in different types water bodies of the Ergaki natural park (Western Sayan)

The Yenisei is one of the largest rivers on our planet. Flowing from south to north, it crosses all the natural zones of Siberia. The Yenisei basin covers the mountains and intermountain hollows of the Altai-Sayan mountainous country, the plateaus of Central and the lowlands of Western Siberia. The ichthyofauna of the flat part of the Yenisei basin is relatively well studied, while the data on the fish of the water bodies of the Western Sayan which are part of the Yenisei basin is limited in the available literature, and includes information only about the fish of some lakes. In this work, we studied of the ichthyofauna and the fish community of different types of water bodies in a specially protected natural area in the central part of the Western Sayan, in the Ergaki Natural Park — the Nizhnyaya Buiba and Bolshaya Oya rivers, and information about the ichthyofauna of Lake Oyskoye was clarified. In the studied water bodies, three species of fish (typical for rivers and lakes of the mountains of southern Siberia) were identified: Arctic grayling Thymallus arcticus (Pallas, 1776), Siberian loach Barbatula toni (Dybowski, 1869) and Eurasian minnow Phoxinus phoxinus (Linnaeus, 1758). Previously there was no information about the presence of Siberian loach in water bodies on the territory of the Ergaki Natural Park in the available scientific literature. The abundance and biomass of fish species varies significantly in different water bodies. Arctic grayling is common or dominant in rivers, Eurasian minnow is super dominant in the Nizhnyaya Buiba river and in the Oyskoe lake. In the Bolshaya Oya river, the superdominant species is the Siberian loach. Eurasian minnow in the Oyskoe lake is large of size.

Environmental DNA unveils deep phylogeographic structure of a freshwater fish.

Phylogeography bears an important part in ecology and evolution. However, current phylogeographic studies are largely constrained by limited numbers of individual samples. Using an environmental DNA (eDNA) assay for phylogeographic analyses, this study provides detailed information regarding the history of Siberian stone loach Barbatula toni, a primary freshwater fish across the whole range of Hokkaido, Japan. Based on an eDNA metabarcoding on 293 river water samples, we detected eDNA from B. toni in 189 rivers. A total of 51 samples, representing the entire island, were then selected from the B. toni eDNA-positive sample set for the subsequent analyses. To elucidate the phylogeographic structure of B. toni, newly developed eDNA metabarcoding primers (Barba-cytb-F/R) were applied to these samples, specifically targeting their haplotypic variation in cytochrome b. After a bioinformatic processing to mitigate haplotypic false positives, a total of 50 eDNA haplotypes were identified. Two regionally restricted, genetically distinct lineages of the species were revealed as a result of phylogeographic analyses on the haplotypes and tissue-derived DNA from B. toni. According to a molecular clock analysis, they have been genetically isolated for at least 1.5 million years, suggesting their ancient origin and colonisation of Hokkaido, presumably in the glacial periods. These results demonstrate how freshwater fishes can alter their distributions over evolutionary timescales and how eDNA assay can deepen our understanding of phylogeography.

New Data on Biology of Siberian Stone Loach Barbatula toni (Nemacheilidae) in Langeri River (Sakhalin)

New information on size composition, feeding, and spawning of Siberian stone loach Barbatula toni in a large watercourse in the northeast of Sakhalin Island is reported. The recorded maximal absolute body length and weight comprised 200 mm and 53.3 g, respectively. Predation and cannibalism in large fish specimens have been revealed.

РАЗНООБРАЗИЕ И ДИНАМИКА РЫБНЫХ СООБЩЕСТВ В БАССЕЙНАХ ГОРНЫХ ПРИТОКОВ АМУРА

Between 1975 and 2021 the diversity of fish communities of mountain rivers was studied in seven areas of the river Amur drainage: “Onon”, “Upper Zeya”, “Upper Bureya”, “Upper Amgun”, “Lower Amgun”, “Khingan”, “Lower Amur and Ussuri”. In these areas, various reaches of typical mountain watercourses were surveyed: 1) upper reaches (24), 2) “middle” (17) and 3) “lower” (10); 9 small mountain tributaries of lowland rivers – Amur and Ussuri, 5 mountain lakes and 6 floodplain reservoirs. According to the results of the research, 6 main types of communities were identified, including river ones: 1) upper reaches, 2) “middle” parts, 3) “lower” parts, 4) small mountain tributaries of large lowland rivers. Lake communities are represented by two main types of communities: 5) mountain lakes and 6) floodplain water bodies. In river communities, 30 fish species and one lamprey species were found; river communities are dominated by Barbatula toni, Cottus szanaga, Brachymystax tumensis, species of the genus Thymallus, as well as Rhynchocypris lagowskii and Phoxinus phoxinus. Upper reaches communities include from 4 to 12–13 species. Barbatula toni, Cottus szanaga, Brachymystax tumensis, and species of the Thymallus dominate here. In the communities of the middle parts of the rivers, there are from 7 to 17 species. Up to 23 species and one lamprey species are found in the lower parts. In the upper reaches and middle sections of the rivers, the composition of the community changes during the year as a result of seasonal migrations. In communities of mountain lakes 6 species of fish were found; three species are most common: Barbatula toni, Cottus szanaga, Brachymystax tumensis. 8 species inhabit floodplain water bodies, dominated by Rhynchocypris percnurus, Perccottus glenii, Misgurnus mohoity. The specificity of the region's coenotic diversity is determined by its position in the Amur basin, the presence or absence of mountain and floodplain lakes, as well as geomorphological and paleogeographic features.

Length–weight relations of ten freshwater fish species (Actinopterygii) from Abashiri River basin, eastern Hokkaido, Japan

Length–weight relations (LWRs) were estimated for ten freshwater fish species such as gin-buna, Carassius langsdorfii Temminck et Schlegel, 1846; lake minnow, Rhynchocypris percnura (Pallas, 1814); Siberian loach, Barbatula toni (Dybowski, 1869); Japanese smelt, Hypomesus nipponensis McAllister, 1963; masu salmon, Oncorhynchus masou (Brevoort, 1856); rainbow trout, Oncorhynchus mykiss (Walbaum, 1792); whitespotted char, Salvelinus leucomaenis (Pallas, 1814); ninespine stickleback, Pungitius pungitius (Linnaeus, 1758); hana-kajika, Cottus nozawae Synder, 1911; and a species of goby Rhinogobius sp. Specimens were collected once a month except in the snow season from the Abashiri River basin, eastern Hokkaido, between June 2007 and November 2011. Fishes were captured by an electrofishing device (Smith–Root Inc., Model 12-b). The estimated allometric coefficient b values ranged from 2.790 (ninespine stickleback) to 3.294 (hana-kajika), and r2 values ranged from 0.772 (lake minnow) to 0.994 (goby). All the LWRs were highly significant, with P < 0.001. Besides, the study provides the first estimates of LWRs for the Siberian loach, Japanese smelt, masu salmon, whitespotted char, hana-kajika, and the goby.

Биология рыб среднего течения реки Киренги

Kirenga River is the large (746 km long) right tributary of upper part of Lena River. River headwaters situated relatively close to west coast of Lake Baikal among the high peaks of the Baikal Ridge at 1470 m a.s.l. Fishes were collected from the Kirenga’s middle reaches during the investigations of biota of Tukolon’ nature protected territory along the 25 km section of riverbed (starting from approx. 375 km upstream from the mouth) in June 2013, June and September 2014, Oktober 2015 and April 2016. According to available data the ichthyofauna at this river section consists of 20 species in 11 families: Far Eastern brook lamprey Lethenteron reissneri, dace Leuciscus leuciscus baicalensis, roach Rutilus rutilus, Czekanowski's minnow Rhynchocypris czekanowskii, Amur minnow Rhynchocypris lagowskii, Eurasian minnow Phoxinus phoxinus, loach Cobitis melanoleuca, Siberian stone loach Barbatula toni, Northern pike Esox luceus, whitefish Coregonus pidschian, tugun Coregonus tugun, roundfish Prosopium cylindraceum, Baikal-Lena grayling Thymallus baicalolenensis, taimen Hucho taimen, lenok Brachymystax lenok, burbot Lota lota, sculpin (spotted sculpin, alpine bullhead) Cottus cf. poecilopus, Siberian sculpin Cottus sibiricus, ruff Gymnocephalus cernuus, perch Perca fluviatilis. Lamprey, burbot and sculpin in our findings were reported from guts of other piscivorous fich species but tugun and Czekanowski's minnow were not registered. Lenok and Baikal-Lena grayling are still most numerous in the river. Low abundance is characteristic to lamprey, loach, Siberian stone loach, burbot, sculpin and Siberian sculpin. Recent growth features in most species are generally similar with the ones for fishes from Kirenga’s low reaches and main course of Upper Lena. Their ages of maturation mostly coincides with other large streams in East Siberia.

Cobitidae (Вьюновые) и Balitoridae (Балиторовые) Среднего Амура в водоёмах Еврейской автономной области и сопредельных приграничных территорий Китая

The article reports on the results of a study on the ichthyological diversity observed in the middle part of the Amur basin on the territory of the Jewish Autonomous Region (JAO) and the adjacent border territory of the People's Republic of China. The research covers the issues of diversity, distribution and ecological specificity of fish for the families Cobitidae and Balatoridae in the water bodies and watercourses of the region. The research summarizes and amends information on the distribution of the Cobitidae and Balatoridae families in the Bastak Nature Reserve (Russian Federation), and in the Honghe and the Bachadao reserves (China), as well as in a number of rivers of the JAO (Amur, Dobraya, Bijan, Bira, Ikura, Tunguska, In, Zabelovka, Gari, etc.), and rivers of the adjacent Chinese territory (Amur, Yalu, Sungari). The paper confirms the families’ habitat within the location under study, and provides information on the distribution of eight members of the family Cobitidae: Cobitis lutheri (Rendahl, 1935), Cobitis melanoleuca (Nichols, 1925), Cobitis choii (Kim et Son, 1984), Misgurnus anguillicaudatus (Cantor, 1842), Misgurnus nikolskiy (Vasilieva, 2001), Misgurnus mohoity (Cantor, 1842), Paramisgurnus dabrianus (Dabry de Thiersant, 1872), Parabotia mantschurica (Berg, 1907), and three members of the family Balatoridae: Barbatula toni (Dybowski, 1869), Lefua costata (Kessler, 1876), Lefua pleskei (Herzenstein, 1887). The article contains data for the observation period from 2001 to 2019. The information is new for the area under investigation.

Рыбы семейств Cobitidae (вьюновые) и Balitoridae (балиторовые) в экотопах заповедника «Бастак»

The article provides the results of studying the fish fauna of the Bastak reserve. Based on the analysis of field ichthyologic collections in 2001-2019, the author described the species composition, distribution and habitat specificity in the nature reserve reservoirs - of the fish from the Cobitidae and Balitoridae families. Within the reserve and its conservation zone, it was found the habitat of four representatives of the family Cobitidae and two fish from the family Balitoridae. The research, made in 2019, first discovered the Paramisgurnus dabrianus (Dabry de Thiersant, 1872) and the round-tailed form of Barbatula toni (Dybowski, 1869) in the territory and protection zone of the Bastak nature reserve. There, it was also found some new habitats of species belonging to the families.

Биология рыб озера Фролиха (Северный Байкал, Восточная Сибирь)

The fish fauna of Lake Frolikha comprises 12 species confirmed by our recent studies: roach Rutilus rutilus (L.), minnow Phoxinus phonixus (L.), stone loach Barbatula toni (Dyb.), spine loach Cobitis melanoleuca Nichols, pike Esox lucius L. lenok Brachymystax lenok (Pallas), Arctic charr Salvelinus alpinus (L.), Baikal black grayling Thymallus baicalensis (Dyb.), burbot Lota lota (L.), sand sculpin Leocottus kesslerii (Dyb.), stone sculpin Paracottus knerii (Dyb.) and perch Perca fluviatilis L. Biological peculiarities of these fishes are determined by unique geographical position and limnic characteristics of Lake Frolikha. Being a typical glacial lake it is located at rather low altitude a.s.l. and is adjacent to the unparalleled ecosystem of Lake Baikal. An underdeveloped shallow coastal zone with sparse aquatic vegetation, which is characteristic of glacial lakes, determines some biological traits of Lake Frolikha cyprinids. As compared with other mountain lakes of Baikal Rift Zone (BRZ), roach and minnow have lower rates of linear and weight growth as well as unusual population sex ratios. Male to female ratio in roach is 2:1 and in minnow it is 1:1 whereas in other lakes it reaches 1:4 and 1:10 respectively. The presence of native Baikal species in Lake Frolikha provided abundant food for piscivores: sand sculpins are the dominant food item of lenok, Arctic charr and perch. Lenok and perch demonstrate prominent linear and weight growth rate in the lake. In other lakes of BRZ with less abundant prey fish species Arctic charr typically diverge into several trophic forms, but in Lake Frolikha they established a monomorphic mostly piscivorous population. Charr from Lake Frolikha are relatively small and short-lived representatives of the large form of Transbaikalian charr. Their unique feature is river spawning, which is not observed in any other charr population within BRZ. Due to special protection regime of the territory, which minimizes catch losses, several fish species maintain natural unaffected structure of their populations evidenced by the abundance of old fish. The population of lenok is represented by 19 age groups, the population of roach – by 15 groups and that of burbot – by 12 groups. Alternatively, populations of small-sized prey species such as minnow and sand sculpin, which are heavily influenced by numerous predators include fewer age groups than in other mountain lakes of BRZ.

Разнообразие рыб в техногенных водных объектах горных территорий бассейна Амура

Human impact on the population and diversity of fish in the mountainous regions of the Amur River basin has been increasing due to mining, deforestation, construction, and the development of hydroelectric power stations and reservoirs. Hundreds of man-made water bodies are located here, with hardly any research conducted on fish diversity in them. Meanwhile, the ichthyofauna of mountainous areas, in comparison with the flatland fauna, is less resistant to adverse influences. The taxonomic and coenotic diversity of fish was studied at 15 locations in 37 technogenic water bodies (18 settling ponds, three ponds, eight flooded quarries, five diverted or otherwise severely altered major watercourses, as well as in a drainage ditch of a minor tributary, a reservoir, and tailings) in the valleys of mountain and foothill tributaries of the Amur. In addition, natural streams and one lake were examined at each location; over 270 fish specimens of 21 species were collected. The taxonomic diversity of fish in technogenic water bodies includes 21 species from 17 genera, nine families and six orders. The order of Cypriniformes is represented by 12 species, Salmoniformes – by 5; the rest are represented by one species each. All species have been found in quarries and ponds; Lagovskiy minnow Rhynchocypris lagowskii (found at 11 sites) and lake minnow Phoxinus (Eupallasella) percnurus (9), siberian stone loach Barbatula toni (11) and Amur spleeper Perccottus glenii (6) are most common. Loach mainly lives here temporarily, other species inhabit these water bodies permanently; they have been breeding here, their populations increasing in numbers. Some reservoirs with a high level of water exchange provide a temporary habitat for typical river species, i.e. grayling, lenok, taimen, and Amur sculpin. The coenotic diversity of man-made reservoirs in terms of its species composition is close to the ichthyocenoses of the lakes in the lower parts of the valleys. As for artificial watercourses, 11 species were found there; most are present there temporarily. The composition of fish communities in watercourses is represented by depleted patterns of river ichthyocenoses. The number of species and their composition in technogenic water bodies depends on the geographic location of the site, the diversity of natural water bodies at the location–primarily, the presence or absence of lakes, the extent and pattern of watercourse alterations, and specific characteristics of the man-made water ponds and watercourses. Most species spread to the technogenic water bodies from local rivers, some–from lakes located nearby, while others were likely to be carried over by human hand and birds.

The complete mitochondrial genome of Barbatula nuda and B. toni (Teleostei: Nemacheilidae)

In this study, we cloned and sequenced the complete mitochondrial genome of Barbatula nuda and Barbatula toni with 16,620 and 16,619 nucleotides, respectively. The complete mitogenomes of the two species both include 13 protein-coding genes (PCGs), 2 ribosomal RNA genes, 22 transfer RNA genes, and a non-coding control region. All PCGs of the two species are initiated with ATG or GTG (only for COI) and ended with complete (TAA and TAG) or incomplete (TA and T) stop codons. The complete mitochondrial genomes would provide important molecular data for further phylogenetic analyses.

On the diversity of fishes in mountain lakes of the Amur basin

Results of the study of 11 lakes situated in the mountainous part of the Amur River basin are reported. Six fish species, namely, blunt-snouted lenok Brachymystax tumensis, Baikal-Lena grayling Thymallus baicalolenensis, Siberian stone loach Barbatula toni, Lagowski’s minnow Rhynchocypris lagowskii, Czekanowski’s minnow Rh. czekanowskii, and Amur sculpin Cottus szanaga, were found in five lakes. Four species were found in one lake (Lake Pereval’noe, basin of the Amgun’ River), and two species were found in each of the remaining lakes. Lenok and Amur sculpin were the most common species. The fauna of Lake Okonon (basin of the river Zeya) was the most specific and did not have analogs among the lake faunas; Czekanowski’s minnow and Baikal black grayling were found in this lake. The lenok ecotype that formed in the mountain lakes differed from the river lenok in some regards. The potential pathways of formation and protection of mountain lake ichthyocenoses of the Amur basin are discussed.

Mass immigration of juvenile fishes into a small, once-dried tributary demonstrates the importance of remnant tributaries as wintering habitats

The winter ecology of stream fishes is still poorly understood, especially in non-salmonid species. A small tributary (<4 m in width, <1 km in length) of the Otofuke River (central Hokkaido, northern Japan) dewatered in the summer of 2013, which allowed us to evaluate immigration after the summer draught. More than 10000 juveniles of Siberian stone loach Barbatula toni, Japanese dace Tribolodon spp., and non-native rainbow trout Oncorhynchus mykiss immigrated to the tributary in the four months following the summer dewatering. This suggests that even remnant tributaries may be critical wintering habitats for some juvenile fishes.

Morphological comparison and DNA barcoding of four closely related species in the genera Misgurnus and Paramisgurnus (Cypriniformes: Cobitidae)

To evaluate the feasibility of morphological and genetic identification of the closely related species in the genera Misgurnus and Paramisgurnus, the morphological characters of four species in these genera and DNA barcoding of five loaches (P. dabryanus, M. anguillicaudatus, M. bipartitus, M. mohoity, and Barbatula toni) were investigated. Twelve morphological characters were measured in 542 individuals to perform the comparative analysis. Among these characters, only the caudal peduncle length (LCP) revealed significant difference (P < 0.05) among these four species. The clustering based on morphological characters formed two clusters (P. dabryanus and M. anguillicaudatus; M. bipartitus and M. mohoity). A total of 186 COI fragments for the five loaches investigated were sequenced and analyzed. The results showed that interspecific K2P distance was much higher than intraspecific distance within the five species. Bayesian inference of phylogeny showed that individuals of these species were divided into five specific clades. Meanwhile, the COI fragments exhibited 22 character attributes for the differentiation of the five loach species based on character-based method. Our results suggested that DNA barcoding based on COI can be used as an efficient identifier of these five loach species; the combination of distance-based method, Bayesian inference and character-based approach provides higher resolution of identification at species level.

Proteocephalid tapeworms (Cestoda: Onchoproteocephalidea) of loaches (Cobitoidea): Evidence for monophyly and high endemism of parasites in the Far East

The parasite fauna of loaches (Cypriniformes: Cobitoidea), a group of small bottom-dwelling freshwater fishes with a mostly Eurasian distribution, remains a largely unknown quantity. Here we revise the taxonomy of tapeworms of the genus Proteocephalus Weinland, 1858 (Cestoda: Proteocephalidae) that had been found in loaches from the Palaearctic Region (Central Europe, Japan and Russia [Primorsky Region]). Molecular phylogenetic analysis based on two nuclear (ssr- and lsrDNA) and two mitochondrial genes (cox1 and rrnL) revealed a monophyletic group consisting of four valid species nesting within the Proteocephalus-aggregate: (i) Proteocephalus sagittus (Grimm, 1872) from Barbatula barbatula (Europe, Russia and Tajikistan), (ii) Proteocephalus demshini n. sp. from Barbatula toni (Russian Far East — Primorsky Region), (iii) Proteocephalus midoriensis Shimazu, 1990 from Lefua echigonia (Japan) and L. costata (Russia) (new host and geographical record), and (iv) Proteocephalus misgurni n. sp. from Misgurnus anguillicaudatus (Russia; Primorsky Region). Proteocephalus sagittus and P. demshini, and P. midoriensis and P. misgurni were recovered as sister taxa, respectively. Proteocephalus sagittus and P. demshini are characterized by having proglottids that are wider than long, an elongate to pyriform cirrus-sac and the vitelline follicles that form wide lateral bands. Proteocephalus midoriensis and P. misgurni are characterized by having proglottids that are more elongate and an ovoid to almost spherical cirrus-sac and the vitelline follicles forming narrow lateral bands. Proteocephalus demshini differs from P. sagittus in the posterolateral extent of the vitelline follicles, whereas P. misgurni can be distinguished from P. midoriensis mainly by the relative size of the ovary, posterior extent of the vitelline follicles and width of the scolex. Unlike most species of the Proteocephalus-aggregate that possess an apical sucker, all species from loaches are devoid of any apical organ. The existence of two new species in loaches from the Primorsky Region of Russia indicates high endemism of fish parasites in this region. A key to the identification of recognized species from loaches is provided.

Sequence and organization of the complete mitogenome of a Siberian stone loach, Barbatula toni (Dybowsky, 1869) (Cypriniformes: Balitoridae)

In this study, we determined the complete mitogenome sequence of Siberian stone loach, Barbatula toni (Dybowsky, 1869). The total length of mitogenome is 16 623 bp, which consists of 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and 1 control region (D-loop). The genome organization and gene order were identical to that of the typical vertebrates. The control region harbors conserved sequence blocks (CSB-D, E, F, CSB-1, CBS-2 and CBS-3) and TA-nucleotide microsatellite repeats in its 3′ end. The complete mitogenome may provide important molecular data for further phylogenetic analyses for higher taxa of teleost fishes, especially for the fishes in order Cypriniformes.

Feeding of blunt-snouted lenok Brachymystax tumensis Mori, 1931 (Salmoniformes, Salmonidae), in Korbokhon Lake (Left Bureya River, Khabarovsk Krai) in summer

Characteristics of feeding of blunt-snouted lenok Brachymystax tumensis in the mountain moraine Korbokhon Lake (Left Bureya River, Khabarovsk krai) in summer is presented. It was established that individuals at an age of 3+ to 13+ use all accessible seasonal types of preys both near bottom and in surface water layers as food. In total the contents of all studied stomachs, 88 components of animal origin (bottom invertebrates, lenok roe, larvae, pupae, and imagoes of aquatic and terrestrial insects), as well as three plant components, were found. With age, the spectrum of feeding of lenok widens. In stomachs of individuals at an age of 9+ to 13+, besides invertebrates, bearded stone loach Barbatula toni and fur small murine rodents were recorded. Selectivity of feeding with respect to weight and type of preys was revealed: lenok older than 9+ prefers larger food items; individuals at an age of 11+ to 12+ more frequently include beetles and other terrestrial insects in the diet, and the ratio of aquatic and terrestrial food items changes in favor of the latter.

Illustrated checklist of fishes from the Shubuto River System, southwestern Hokkaido, Japan

A checklist of fish fauna, comprising 40 species representing 15 families and 9 orders, was compiled from field, museum, and literature surveys of the Shubuto River System, southwestern Hokkaido, Japan. This area approximates the boundary of southern/northern affinities of Japanese freshwater fishes. All primary freshwater species, are listed except for Gymnogobius castaneus and all anadromous and marine amphidromous species that are scientifically presumed to occur naturally in the area. The list contains two invasive exotic species, Cyprinus carpio and Oncorhynchus mykiss, and two Siberian primary freshwater species, Lefua nikkonis and Barbatula toni, which are thought to mark the boundary of southern and northern affinities of Japanese freshwater fishes. Rhynchocypris perenurus, another Siberian primary freshwater species with a similar presumed range, was not recorded. The voucher specimen species included Platycephalus sp. 2, which is suggested to have the northernmost extended range.

White linear markings on the abdomen of river loaches (Nemacheilidae) — potential usage for the identification of individuals

Former studies demonstrated that the river loach species Lefua sp. and Lefua echigonia (Nemacheilidae) have thin white linear markings from the pectoral fin base to the ventral fin base on both right and left sides of the abdomen that can be used to identify individuals in laboratory and in the field. In the present study we demonstrate the existence of such markings and the uniqueness of their shape in reared individuals of a further six species of Nemacheilidae, but found them absent in three species of Balitoridae. Furthermore, the long-term stability of the shape of the markings was examined in reared individuals of the nemacheilid Barbatula toni over two years. The white line markings offer a non-invasive tool for the individual recognition of nemacheilid species, some of which are threatened with extinction.

A sperm cryopreservation protocol for the loach Misgurnus anguillicaudatus and its applicability for other related species

The aim of the present study was to establish a protocol of sperm cryopreservation in Misgurnus anguillicaudatus and verify the applicability of the obtained protocol in other loach species. We evaluated the following parameters: inseminating dose, thawing temperatures (20, 25 and 30 °C for 10 s), extenders (loach or cyprinid extenders), internal cryoprotectants (dimethyl sulfoxide (DMSO), dimethylacetamide (DMA), glycerol (Gly), ethylene glycol (EG), and methanol (MeOH) at 0, 5, 10 and 15%), external cryoprotectants (bovine serum albumin 1 and 2%; sucrose 0.5 and 1%; glucose 0.5 and 1%; glycine 0.5 and 1%), activating solutions (distilled water, dechlorinated tap water, 25 mM NaCl and 50 mM NaCl), and hatchability of the eggs when fertilized with fresh or cryopreserved sperm. After the evaluation of these parameters, we optimized the cryopreservation using the following procedure: thawing temperature at 25 °C for 10 s; loach or cyprinid extenders; methanol at 10 or 15% as internal cryoprotectants; glycine 0.5% or bovine serum albumin 1% as external cryoprotectants and 50 mM NaCl for sperm activation. Using this procedure, the fertilizability of the post-thawed sperm was 47% in comparison to the fresh sperm, at the minimum inseminating dose (687.65 spermatozoa egg −1 mL −1). Based on this protocol, sperm from other loach species Lefua nikkonis, Misgurnus mizolepis and Barbatula toni were cryopreserved successfully.