Genetic diversity and population structure of the threatened chocolate mahseer (Neolissochilus hexagonolepis McClelland 1839) based on SSR markers: implications for conservation management in Northeast India.

Neolissochilus hexagonolepis (MacClend 1839), commonly known as chocolate mahseer, is an ecologically threatened fish species. The species is reported from Southeast Asia and in India, it is distributed across the Northeast region. The present study was carried out to develop species-specific novel microsatellite markers using next-generation sequencing and to assess the genetic diversity of wild chocolate mahseer populations distributed through Northeastern India. A total of 25 polymorphic loci (mean PIC = 0.933) were amplified in 194 individuals belonging to seven different populations. We observed high genetic diversity across the loci with mean observed (Ho) and expected heterozygosity (He) of 0.557 and 0.939 respectively. The studied loci didn't show significant deviation from Hardy-Weinberg equilibrium. Genetic analyses indicate substantial pairwise Nei's genetic distance and moderate to high levels of genetic differentiation among populations (mean FST = 0.23). The structure, factor and cluster analysis identified five major clusters that can be considered as different conservation units while formulating any management measures. Furthermore, the migration analysis inferred that there is no active migration among the studied populations. Results suggested that two populations i.e. Dikrong river (Arunanchal Pradesh) and Umiam river have high genetic diversity. These populations can be utilized for the breeding programme to achieve substantial genetic variations in the descendant populations. The Nongbareh populations showed very less genetic vigor and need an immediate attention for conservation. The SSR markers developed in the present study will provide a valuable resource for future population genetic assessment and implementation of effective conservation strategies for wild chocolate mahseer.

Molecular identification and genetic diversity analysis of Chocolate mahseer (Neolissochilus hexagonolepis) populations of Northeast India, using mitochondrial DNA markers.

The population genetic structure and genetic diversity of Neolissochilus hexagonolepis were studied using three mitochondrial genes (CoxI, Cytb, ATPase 6/8). A total of 120 individuals representing nine populations from different drainages of Northeast India were used for the study. Thirty-three distinct haplotypes were identified from concatenated gene analysis. The total haplotype and nucleotide diversities are 0.8880 and 0.0280, respectively. The analysis of molecular variance (AMOVA) reveals that the main variation (89.33%) was among populations. Most of the populations showed high polymorphisms, parsimony and haplotype diversity which indicate genetically healthy stocks in the wild. The genetic differentiation patterns were consistent with geographical distributions. Pairwise FST comparison of populations showed significant genetic differentiation (0.9088, p < .05). The pattern of haplotype network and phylogenetic tree revealed six major groups. Results suggested that chocolate mahseer populations in Northeast India having high haplotype diversity and genetic differentiation can be utilized in breeding programs to maintain genetic diversity in the descendant populations. The present study would be beneficial for sustainable management, stock-specific strategies for breeding and conservation of the wild population of N. hexagonolepis in future.

Transcriptome profiling and expression analysis of immune responsive genes in the liver of Golden mahseer (Tor putitora) challenged with Aeromonas hydrophila.

Transcriptome profiling has been used to decipher the novel mechanisms behind immune responses of the fishes. However, the molecular mechanism underlining immune response in mahseer is not studied so far. Fishes are greatly affected by bacterial pathogens such as Aeromonas hydrophila. In this study, transcriptome response of golden mahseer (Tor putitora) infected with A. hydrophila was examined using paired end Illumina sequencing of liver tissue to understand the immune response of the fish. The de novo assembly generated 61,042 unigenes ranging from 200 to 9322 bp in length and an average length of 463 bp. The gene ontology annotations resulted a total of 131,826 term assignments to the annotated transcriptome including 60,846 (46.16%) allocations from the biological process; 21,603 (16.39%) from molecular function and 49,377 (37.46%) from cellular components. Differential gene expression analysis of the transcriptome data from challenged and control group revealed 1104 upregulated and 1304 down-regulated unigenes. The differentially expressed genes were mainly involved in the pathways including cell surface receptor signaling, TH1 and TH2 cell differentiation, pathogen recognition, and immune system process/defense response especially complement cascade. Twelve unigenes including ankyrin, serum amyloid, hsp4b, STAT3, complement factor c3 and c7 were validated using qPCR and found differentially expressed in accordance with in silico expression analysis. The results obtained in this study will provide the first and crucial information on the molecular mechanism of mahseer fishes against bacterial infection.

The complete mitogenome of brown trout (Salmo trutta fario) and its phylogeny.

The complete mitochondrial genome of Salmo trutta fario, commonly known as brown trout, was sequenced using NGS technology. The mitochondrial genome size was determined to be 16 677 bp and composed of 13 protein-coding gene (PCG), 22 tRNAs, 2 rRNA genes, and 1 putative control region. The overall mitogenome composition of S. trutta fario is A: 28.13%, G: 16.44%, C: 29.47%, and T: 25.96% with A + T content of 54.09% and G + C content of 45.91%. The gene arrangement and the order are similar to other vertebrates. The phylogenetic tree constructed using 42 complete mitogenomes of Salmonidae fishes confirmed the position of the present species under the genus Salmo of subfamily Salmoninae. NGS platform was proved to be a rapid and time-saving technology to reveal complete mitogenomes.

Molecular cloning and expression profile of snow trout GPDH gene in response to abiotic stress.

Glycerol-3-phosphate dehydrogenase (GPDH) gene possibly plays a key role for cold acclimation process in snow trout during winter months when water temperature goes down to 4-5 °C. In this study, 1,012 bp nucleotide fragment of GPDH gene was obtained from two snow trout species (Schizothorax richardsonii and S. niger; family: Cyprinidae), distributed in several Himalayan rivers. The gene encoded a protein of 334 amino acids. The encoded protein sequence was very similar to GPDH of Danio rerio (94.36 %) using BLASTx searches. In S. richardsonii the qRT-PCR showed highest expression in muscle tissue followed by liver and also revealed 19 fold gene expression in liver tissue under cold (5 °C) in comparison with warm (15 °C) condition. The elevated expression levels of GPDH cDNA on cold treatment furthermore suggest that GPDH plays a role in stress related responses in S. richardsonii. The phylogenetic analysis showed that the two snow trout species GPDH share the same clade with characterized GPDHs from other teleost fishes suggesting a common evolutionary origin and a similar catalytic function. In addition, the Ka/Ks ratios of these sequences suggested that they are under purifying selection. Moreover, the expression profile of GPDH gene among co generic species of genus Schizothorax showed that GPDH cDNA expression was highest in S. richardsonii and lowest in S. esocinus which gives an indication of species specific adaptation in relation to different geographical areas.