Endemic Cyprinid Research Articles

Development and characterization of an embryonic cell line from endangered endemic cyprinid Honmoroko Gnathopogon caerulescens (Sauvage, 1883).

Establishing a cell line from endemic species facilitates the cell biological research of these species in the laboratory. In this study, an epithelium-like cell line RME1 was established from the blastula-stage embryos of the critically endangered cyprinid Honmoroko Gnathopogon caerulescens, which is endemic to ancient Lake Biwa in Japan. To the best of our knowledge, this is the first embryonic cell line from an endangered fish species. This cell line is well adapted to grow at 28°C in the culture medium, which was successfully used for establishing testicular and ovarian cell lines of G. caerulescens, and has displayed stable growth over 60 passages since its initiation in June 2011. Although RME1 did not express the genes detected in blastula-stage embryos, such as oct4, sox2, nanog, and klf4, it showed a high euploidy rate (2n = 50; 67.2%) with normal diploid karyotype morphology, suggesting that RME1 retains the genomic organization of G. caerulescens and can prove to be a useful tool to investigate the unique properties of endangered endemic fishes at cellular level.

Molecular cloning and antibacterial activity of hepcidin from Chinese rare minnow (Gobiocypris rarus)

Hepcidins, a kind of cysteine-rich antimicrobial peptides, play important roles in host immunological processes and iron regulation, which have been identified from several fish species. The rare minnow (Gobiocypris rarus), an endemic cyprinid fish in China, has been used extensively as model animal in laboratory. However, little is known about its hepcidin. Here, we report the cloning and characterization of a hepcidin gene from the liver of Chinese rare minnow. The full-length cDNA of rare minnow hepcidin is 662 bp, which contains an ORF of 273 bp encoding a prepropeptide of 90 amino acid residues. The predicted prepropeptide contains three domains: a signal peptide of 24 amino acids, a prodomain of 41 amino acids, and a mature peptide of 25 amino acids. Sequence alignment showed eight conserved cysteine residues in the mature peptide, which formed four disulfide bonds in spatial structure. The deduced structure of mature peptide showed a high degree of homology to the human hepcidin. Phylogenetic analysis showed that it had a close relationship with zebrafish hepcidin, and clustered in a clade with these from Cyprinidae. Synthetic peptide of rare minnow hepcidin could inhibit the growth of Gram positive bacterium Staphylococcus aureus and Gram negative bacteria Escherichia coli and Aeromonas hydrophila. These results suggested that rare minnow hepcidin had typical structure of hepcidins and antibacterial activity. It could participate in innate immune response as an antibacterial agent and be used as antibiotic substance.

Length–weight and length–length relationships for three endemic cyprinidspecies of the Aegean region (Turkey) with proposed standard weight equations

Empirical standard weight equations and length-weight and length-length relationships were estimated for Barbus pergamonensis, Capoeta bergamae, and Ladigesocypris irideus, three endemic cyprinid species of Turkey that are still not widely studied. To the best knowledge of the authors, empirical standard weight equations for the three species and L-W parameters for L. Irideus are given here for the first time. In this study, length and weight data for B. Pergamonensis, C. Bergamae, and L. Irideus were collected throughout the watercourses of Mugla Province and were used to develop species-specific Ws equations. The results were for B. Pergamonensis (total length [TL] range: 6-22 cm) log10Ws= -5.713 + 3.718 log10TL - 0.166 (log10TL)2, for C. Bergamae (TL range: 8-29 cm) log10Ws= -5.224 + 3.311 log10TL - 0.083 (log10TL)2, and for L. Irideus (TL range: 6-10 cm) log10Ws= -13.298 + 12.116 log10TL - 2.471 (log10TL)2. This study also reported new maximum TLs for the three species, together with species-specific length-weight and length-length equations.

Vitellogenin in the male Lake Van fish (Chalcalburnus tarichi Pallas, 1811).

Chalcalburnus tarichi is an endemic cyprinid species living in Lake Van, Turkey. In the present study, we investigated vitellogenin (Vtg) in the plasma of adult male C. tarichi, sampled monthly from Edremit, Campus and Çarpanak sites of Van Edremit Gulf (VEG) of lake, between October 2004 and May 2005. The fish living around VEG winters in deep water around Edremit site. University and Çarpanak sites are also on fish migration way from VEG to fresh water (Karasu River). The levels of measured plasma Vtg changed from trace levels to 26.35µg/mL. The highest level of plasma Vtg was detected in April, with a concentration of 21.60±1.29µg/mL. We also performed immunohistochemical Vtg staining in the liver sections of male fish and determined Vtg positive hepatocytes in the liver of 1/10 sampled fish. Our results indicated that C. tarichi living in Lake Van might be exposed to endocrine-disrupting chemicals.

The complete mitochondrial genome of Platysmacheilus nudiventris (Teleostei, Cyprinidae, Gobioninae)

The complete mitochondrial genome of Platysmacheilus nudiventris (Teleostei, Cyprinidae, Gobioninae), an endemic Chinese cyprinid fish, was first determined by DNA sequencing based on the PCR fragments. The total length of the mitochondrial genome is 16,603 bp, including 22 transfer RNA genes, 2 ribosomal RNA genes, 13 protein-coding genes and a non-coding control region (D-loop). The overall-based composition is 30.2% A, 26.9% T, 16.7% G and 26.1% C, with high A+T content (57.1%). The results will provide useful data for further studies on species identification, evolutionary and population genetic studies of P. nudiventris and its related species.

Development of sixteen novel polymorphic microsatellite markers in Rhinogobio ventralis (Teleostei, Cyprinidae, Gobioninae)

In the present study, we developed 16 polymorphic microsatellite loci in Rhinogobio ventralis, an endemic cyprinid fish in the upper reaches of the Yangtze River. The polymorphism of each locus was assessed in 32 individuals collected from Jiangjin section of the Yangtze River. A total of 126 alleles were detected and the number of alleles per locus ranged from 2 to 16. The observed and expected heterozygosities ranged from 0.161 to 0.968 and from 0.154 to 0.928, respectively. Two loci significantly deviated from Hardy–Weinberg equilibrium. No significant linkage disequilibrium was found between pairs of loci. These polymorphic microsatellites will be useful tools in the study of genetic diversity and population structure of R. ventralis.

Distribution, threats and conservation status of the Wayanad Mahseer, Neolissochilus wynaadensis (Day, 1873) (Teleostei: Cyprinidae): an endemic large barb of the Western Ghats, India

The Wayanad Mahseer Neolissochilus wynaadensis (Day, 1873) is an endemic cyprinid fish that occurs in the upland streams and rivers of the southern region of the Western Ghats. This species has been listed as Critically Endangered on the IUCN Red List of Threatened Species due to its restricted distribution and heavy declines in populations. Like many large cyprinids of the Western Ghats, N. wynaadensis is poorly known and documented, with very few verified records and voucher specimens. Based on specimens recently collected from Wayanad, Kerala, the type locality, as well as two additional locations in the Kodagu District of Karnataka; we provide information on the current distribution, phylogenetic position, threats and conservation. An updated conservation assessment of this species following the IUCN Red List categories and criteria is also provided.

EFFECTS OF FLOW REGULATION AND NON‐NATIVE SPECIES ON FEEDING HABITS OF EURASIAN OTTER Lutra lutra IN MEDITERRANEAN TEMPORARY RIVERS

ABSTRACTIn Mediterranean temporary rivers, ecological resources greatly fluctuate because of the high hydrological variability throughout the year. However, flow regulation prevents this natural regime and commonly entails associated non‐native species, which change the structure of aquatic communities. Nonetheless, few studies have tested the interaction of these two disruptive factors (flow regulation and non‐native species) and their synergistic effects on the Eurasian otter (Lutra lutra) diet at the river scale. The aim of this study was to compare the seasonal feeding habits of the otter between a temporary non‐regulated stretch and two regulated stretches invaded by non‐native species in a Mediterranean water course. The Bullaque River (Guadiana River Basin, central Spain) was seasonally sampled for otter spraints and prey abundance assessed from December 2009 to November 2010. Three stretches were considered: high (source, non‐regulated), medium (transition, regulated) and low (confluence, regulated). Diet varied from native prey in the high stretch (amphibians, insects and endemic cyprinids) to non‐native species in the low stretch (red‐swamp crayfish Procambarus clarkii and pumpkinseed sunfish Lepomis gibbosus). Seasonally, ingested biomass of native prey increased in spring. Diet was more diverse in the high stretch. Otters neutrally selected native cyprinids in the high stretch throughout the year, whereas crayfish was selected in the other two stretches. Overall results showed that flow regulation and non‐native species have increased prey availability for the otter; however, this paper highlights the importance of maintaining natural regimes in Mediterranean temporary rivers to conserve native communities and thus minimally impact food webs in Iberian freshwaters. Copyright © 2014 John Wiley & Sons, Ltd.

THREATENED FISHES OF THE WORLD: Squalius malacitanus Doadrio and Carmona 2006 (Cyprinidae)

Squalius malacitanus is an endemic cyprinid fish from the south of the Iberian Peninsula. This species is endangered due to habitat destruction, damming and low water availability.

Two new Dactylogyrus species (Monogenea: Dactylogyridae) D. angorae and D. turcicus from Capoeta angorae and C. mauricii endemic fishes in Turkey.

Two new Dactylogyrus spp., D. angorae and D. turcicus have been described from the gills of the endemic cyprinid fishes, Capoeta angorae and C. mauricii from the Anatolian Region of Turkey. The shape of the anchors, hooks and the copulation organ of both species differ from those of the Dactylogyrus spp. found on other Capoeta species known from the Ponto-Caspian and Tigris-Euphrates regions.

Development of empirical standard weight equation for Pursak chub Squalius pursakensis, an endemic cyprinid species of Northwest Anatolia

Indices of condition enable the evaluation of the well-being of fish, with the assumption that heavier fish of a given length are in a better condition. Relative weight (Wr) is one of these indices; it is calculated by comparing the actual weight of a specimen with the ideal weight of a specimen in good physiological condition of the same length from the same species, i.e. standard weight (Ws). In this research, length and weight data over the distribution range for Pursak chub Squalius pursakensis, an endemic species distributed in the Sakarya and Porsuk drainages in Northwest Anatolia (Turkey), were used to compute a Ws equation by means of the empirical percentile (EmP) method. The Ws equation obtained was log10 Ws = -4.657 + 2.614 log10 TL + 0.127 (log10 TL)2, and the total length range of application was 80-340 mm. Since the EmP Ws equation was not influenced by length variation, the use of this equation to compute the relative weight (Wr) for S. pursakensis throughout its area of distribution is suggested.

THREATENED FISHES OF THE WORLD: Achondrostoma salmantinum Doadrio and Elvira, 2007 (Cyprinidae)

Achondrostoma salmantinum is a small, endemic cyprinid fish from the Iberian Peninsula. It occurs in some tributaries of the Douro river basin in the Salamanca province (western Spain) and is currently endangered due to receding water levels resulting from extensive agriculture, construction of dams and weirs and water transfers.

Development of empirical standard weight equation for Pursak chub Squalius pursakensis, an endemic cyprinid species of Northwest Anatolia

Indices of condition enable the evaluation of the well-being of fish, with the assumption that heavier fish of a given length are in a better condition. Relative weight (W_{r}) is one of these indices; it is calculated by comparing the actual weight of a specimen with the ideal weight of a specimen in good physiological condition of the same length from the same species, i.e. standard weight (W_{s}). In this research, length and weight data over the distribution range for Pursak chub Squalius pursakensis, an endemic species distributed in the Sakarya and Porsuk drainages in Northwest Anatolia (Turkey), were used to compute a W_s equation by means of the empirical percentile (EmP) method. The W_s equation obtained was log_{10} W_s = -4.657 + 2.614 log_{10} TL + 0.127 (log_{10} TL)^{2}, and the total length range of application was 80-340 mm. Since the EmP W_s equation was not influenced by length variation, the use of this equation to compute the relative weight (W_r) for S. pursakensis throughout its area of distribution is suggested.

THREATENED FISHES OF THE WORLD: Squalius castellanus Doadrio, Perea and Alonso 2007 (Cyprinidae)

Squalius castellanus is an endemic cyprinid fish from the Iberian Peninsula, with a distribution area restricted to the River Tagus (Gallo sub-basin). This species is critically endangered mainly due to pollution and fluctuations of the water regime.

Distribution, threats and conservation status of Hypselobarbus thomassi (Day, 1874), a poorly known cyprinid fish of the Western Ghats freshwater ecoregion

The Red Canarese Barb, Hypselobarbus thomassi (Day, 1874) is an endemic cyprinid fish of the rivers of the Western Ghats of India, which has been listed as ‘Critically Endangered’ in the IUCN Red List of Threatened Species. Like many of its congeners, H. thomassi is poorly known with very few verified records and voucher specimens in the past decades. Based on fresh materials collected from three west flowing rivers of Kerala, we provide information on its identity, distribution, phylogenetic position, threats and conservation. An updated conservation assessment of this species following the IUCN Red List criteria is also provided.

The status of Lake Lanao endemic cyprinids (Puntius species) and their conservation

The endemic cyprinid species flock of Lake Lanao, Philippines is a classic example of explosive evolution. Based on historical surveys, most of the 18 endemic cyprinid species in Lake Lanao are now believed to be extinct. We compared results of landing and market surveys conducted in 2008 to information from historical surveys. Field sampling and fishermen interviews provided additional information on the status of the endemic and other fish species in Lake Lanao. Puntius lindog and Puntius tumba were the only two endemic species recorded in 2008, accounting for 0.01 % and 0.04 % respectively of the total weight harvested. Our discovery that at least two endemic cyprinids remain in Lake Lanao is surprising and significant as the lake is increasingly imperiled by human disturbances, exploitation and introduced species. Protecting and conserving the remaining endemic species are important management priorities.

Response to fish specific reproductive hormones and endocrine disrupting chemicals of a Sertoli cell line expressing endogenous receptors from an endemic cyprinid Gnathopogon caerulescens

Fish Sertoli cells play a critical role in spermatogenesis by mediating androgen and progestogen signaling. Their hormonal response, however, considerably differ among species. Therefore it would be ideal to use Sertoli cells originated from the fish of interest to investigate the effects of hormones as well as endocrine disrupting chemicals (EDCs). The aim of this study was to investigate the responses to reproductive hormones and EDCs of a Sertoli cell line that we established from an endemic cyprinid Gnathopogon caerulescens. As the Sertoli cell line expressed endogenous androgen and progestogen receptors, we were able to detect hormone responses by transfecting only a reporter vector (pGL4.36) expressing luciferase under the control of the mouse mammary tumor virus-long terminal repeat (MMTV-LTR) promoter into the cell line. Unlike previous reporter gene assays using fish steroid hormone receptors expressed in mammalian cell lines, luciferase activities were induced by the fish specific androgen (11-ketotestosterone) and progestogen (17α,20β-dihydroxy-4-pregnen-3-one), but not by testosterone and progesterone, at physiologically relevant concentrations. Furthermore, we found 4-nonylphenol (NP) but not bisphenol A showed strong anti-androgenic effects, implying that NP may have direct anti-androgenic effects on fish Sertoli cells in vivo. This is the first evidence, to the best of our knowledge, of anti-androgenic effects of NP in a fish Sertoli cell line. In addition, neither NP nor BPA showed anti-progestogenic effects. These results suggest that the Sertoli cell line established from the fish of interest can be a useful in vitro tool for investigating the mechanisms of reproductive hormones and EDCs in the specific fish.

Morphological Variation Among Wild Populations of Chinese Rare Minnow (Gobiocypris rarus): Deciphering the Role of Evolutionary Processes

Gobiocypris rarus Ye et Fu ( 1983 ) is an endemic cyprinid fish in China, and is considered to be an endangered species. From a conservation perspective, its population structure is interesting. In the present study, morphological variation, including morphometric and meristic traits, was assessed among wild samples collected in the upper Yangtze River basin. There were no significant meristic differences between sexes or among populations, except for scales in lateral line (LS). However, there were significant morphometric differences not only between sexes but also among populations. In discriminant function analysis, the first four discriminant functions explained 75.5% and 78% of the among-population morphometric variation for males and females, respectively. Almost all the truss network morphometric traits significantly contributed to population discrimination. By using all of the truss network morphometric traits, overall random assignments of male and female individuals into their original population were both 73.5%. In addition, the degree of differentiation in phenotypic traits (PST) significantly exceeds that in neutral molecular markers (FST). However, no significant correlation between PST and FST was found in males or females. Overall, these results suggest that two evolutionary processes, including phenotypic plasticity and natural selection, may contribute to the morphological patterns observed in G. rarus.

THREATENED FISHES OF THE WORLD: Iberochondrostoma almacai COELHO, MESQUITA and COLLARES-PEREIRA, 2005 (Cyprinidae)

Iberochondrostoma almacai is an endemic cyprinid fish from the southwest of the Iberian Peninsula, with a distribution area restricted to three independent Portuguese river basins. This species is critically endangered due to water abstraction, pollution, habitat destruction and introduction of exotic species. Captive breeding for restocking purposes is already being conducted.

Molecular cytogenetic analysis of the Appenine endemic cyprinid fish Squalius lucumonis and three other Italian leuciscines using chromosome banding and FISH with rDNA probes

Karyotype and other chromosomal characteristics of the Appenine endemic cyprinid fish, Toscana stream chub Squalius lucumonis, were analysed using conventional banding and FISH with 45S and 5S rDNA probes. The diploid chromosome number (2n = 50) and karyotype characteristics including pericentromeric heterochromatic blocks and GC-rich CMA(3)-positive sites corresponding to both positive Ag-NORs and 45S rDNA loci on the short arms of a single medium-sized submetacentric chromosome pair were consistent with those found in most European leuciscine cyprinids. On other hand, 5S rDNA FISH in the Toscana stream chub and three other Italian leuciscines, S. squalus, Rutilus rubilio and Telestes muticellus, revealed a species-specific hybridization pattern, i.e. signals on four (S. lucumonis), three (S. squalus and R. rubilio) and two (T. muticellus) chromosome pairs. Whereas all the species shared the 5S rDNA loci on the largest subtelocentric chromosome pair, a "leuciscine" cytotaxonomic marker, S. lucumonis showed both classes of rDNA loci tandem aligned on the short arms of chromosome pair No. 12. The present findings suggest that the observed high variability of 5S rDNA loci provides a powerful tool for investigation of karyotype differentiation in karyologically conservative leuciscine fishes.