Character-based diagnostic keys, molecular identification and phylogenetic relationships of threadfin breams (family: Nemipteridae) based on mitochondrial genes from the Southern coromandel Coast, India.

DNA barcoding, primarily focusing on cytochrome c oxidase subunit I (COI) gene has been appraised as an effective tool for species identification. In this study, we focused on the marine fishes of Family Nemipteridae, one of the commercially important group distributed within the Coromandel Coast. The Partial sequences of COI and 16S rRNA of mitochondrial genes were analyzed for species identification and phylogenetic relationship of Nemipterus species (Nemipterus japonicus, Nemipterus peronii, Nemipterus bipunctatus, Nemipterus bathybius). Character-based identification approaches that categorize specimens to species using classification rules that compactly identify species in terms of key diagnostic nucleotides in selected gene sequences. Using the BLOG 2.0 software, species-specific diagnostic nucleotides were identified for the selected group of species. A data set of 198 mtCOI sequences was obtained from published resources and used to screen character-based molecular diagnostic keys for species in silico. Partial sequences of both the genes provided sufficient phylogenetic information to distinguish the four Nemipterus species indicating the usefulness of mtDNA-based approach in species identification. This study proves the use of mtDNA genes sequence-based approach is a support tool along with traditional taxonomy for identifying fish species at a faster pace.

Synthesis of Iron Nanoparticles Using Sargassum wightii Extract and Its Impact on Serum Biochemical Profile and Growth Response of Etroplus suratensis Juveniles.

The present study focuses on the green synthesis of iron nanoparticles using plant extracts as reducing, capping, and stabilizing agents. Aqueous seaweed extracts with the addition of iron solution were mixed using a magnetic stirrer which resulted in a color change indicating the formation of iron nanoparticles. The iron nanoparticles were successfully synthesized using Sargassum wightii extract. The synthesized iron nanoparticles were characterized by UV-Vis spectrophotometer, Fourier transform infrared spectroscopy (FTIR), and zeta potential techniques. The UV-Vis spectra showed a peak at 412 to 415 nm. Zeta potential revealed that the synthesized iron nanoparticles were negative and positive charges. FTIR spectroscopy analysis showed the presence of chemical bond and amide group likely to be responsible for the green synthesis of iron nanoparticles. The effect of nano-iron as a dietary iron source on the growth and serum biochemical profile of Etroplus suratensis fingerlings was evaluated. Iron nanoparticles were fed to E. suratensis fingerlings for 60 days with two levels 10 mg (T1) and 20 mg (T2) and a control group without iron nanoparticles. The highest WG% and SGR and lowest FCR were observed in the T2 group which is significantly different (p < 0.05) from other groups. The serum biochemical profile showed significantly increased activity on 20 mg/kg of nano-iron-supplemented diet. The findings of the present study concluded that supplementation of nano-iron at the 20 mg/kg level to the regular fish diet has a better impact not only on growth but also on the overall health of the fish.

Genetic stock identification in Perna viridis (Linnaeus1758) from the Indian Peninsula by using microsatellite markers.

BACKGROUND: Perna viridis (Linnaeus, 1758), the Asian green mussel, is native to the Asia-Pacific region. The species is extensively distributed in the Indian subcontinent and is a candidate species for aquaculture in the Southeast Asian region. Availability of genetic information on wild populations is essential for the effective conservation and management of Perna species. The present study assessed the genetic variation and population structure across the distribution range of this species from the Indian peninsula by using microsatellite markers to determine the genetic structuring among the species. METHODS: A total of 15 microsatellite loci with M13 labeling were used for the genetic characterization of P. viridis along Indian waters. Genotyped data were analyzed using analytical software to determine the genetic stocks and understand the genetic variability across the populations. RESULTS: We identified 15 polymorphic markers to understand the genetic stocks and variability across Perna populations. The mean value of the observed heterozygosity (Hobs: 0.741) for all populations was closer to the expected heterozygosity (Hexp: 0.75). The pairwise Fst values between the west and east coasts of India varied significantly, indicating the existence of significant genetic structure between the populations. CONCLUSIONS: Genetic stock identification using software analysis exhibited two distinct stocks, one along the west coast (Arabian Sea) and another along the east coast (Bay of Bengal). Bottleneck analysis indicated the genetic stability of species in the wild. P. viridis is a commercially vital species in Indian peninsular regions. The present study suggests the adoption of stock-specific relaying programs of the species from Indian waters in future studies.

Population connectivity and genetic structure of Asian green mussel, Perna viridis along Indian waters assessed using mitochondrial markers.

Perna viridis (Linnaeus, 1758), the Asian green mussel, belonging to the family Mytilidae is widely distributed along the Indian coast. The species is majorly found in southeastern countries and is considered an ideal candidate for aquaculture due to its high nutritional value and growth rate. Obtaining their genetic information is essential for their sustainable capture-based production. In the present study, genetic variation, population structure, and demographic processes of the populations across the distribution of this species were assessed using the mitochondrial DNA ATPase6 and cytb gene. In total, we selected 170 samples from five localities across the Indian subcontinent including Andaman Sea. Sequence analysis of partial cytb (885 bp) and ATPase6 (714 bp) genes revealed 45 and 58 haplotypes, respectively. The significant coefficient of genetic differentiation (FST: 0.255 for cytb and 0.252 for ATPase6) and analyses of molecular variance indicated three varieties of stocks, namely Arabian Sea, Bay of Bengal, and Andaman Sea. All the populations showed low nucleotide diversity, suggesting severe historical bottleneck events and high haplotype diversity, indicating population expansion. The genetic variation and demographic process reported in this study will form the baseline information for framing policies, which can be adopted while planning stock specific ranching and relaying programmes in the Indian subcontinent with view to enhance and manage the fishery.

Mitochondrial signatures for identification of grouper species from Indian waters.

Groupers are important commercial fish in many parts of the world. Accurate identification is critical for effective conservation assessment and fisheries management. Genetic barcodes provide a simple and reproducible method for the identification of species even in the absence of taxonomic expertise. The generation of reference barcodes from properly identified specimens is an important first step in this direction. Here, 36 species belonging to the subfamily Epinephelinae (Family: Serranidae) were collected from landings on the west coast of India and Port Blair, Andaman, and partial nucleotide sequence data of the mitochondrial cytochrome C oxidase subunit I (COI) gene was generated. Barcodes for 13 species were developed from Indian waters for the first time. Analysis using the COI gene produced phylogenetic trees in concurrence with other multi-gene studies. Epinephelus fasciatus and E. areolatus were found to be a species complex, as hypothesized in other studies. The DNA barcodes developed in the study can be used for identifying species within Epinehelinae, where taxonomic ambiguity still exists.

Hints for panmixia in Scomberomorus commerson in Indian waters revealed by mitochondrial ATPase 6 and 8 genes.

Scomberomorus commerson is an economically important migratory fish distributed worldwide. The genetic stock structure of S. commerson distributed along the Indian waters was identified using mitochondrial ATPase 6 and 8 genes. A total of 842 bp sequence of ATPase 6/8 genes obtained in this study revealed 23 haplotypes with mean low nucleotide diversity and high haplotype diversity. Co-efficient of genetic differentiation (FST) values obtained for pair wise populations were low and non-significant with an overall value of -0.02074. The high haplotype and low nucleotide diversity values together with mismatch distribution analysis suggested a history of genetic bottleneck events or founder effect, with subsequent population expansion in S. commerson. The findings of the present study indicated the panmixia nature of the species which can be managed as a unit stock in Indian waters.

Establishment and characterization of fin-derived cell line from ornamental carp, Cyprinus carpio koi, for virus isolation in India.

Cyprinus carpio koi fin (CCKF) cell line was established and characterized from the caudal fin tissue of ornamental common carp, C. carpio koi. This cell line has been maintained in L-15 medium supplemented with 15% foetal bovine serum (FBS) and subcultured more than 52 times over a period of 24 mo. The CCKF cell line consisted of epithelial cells and was able to grow at temperatures between 22 and 35°C with an optimum temperature of 28°C. The growth rate of these cells increased as the proportion of FBS increased from 2 to 20% with optimum growth at the concentrations of 15% FBS. Karyotype analysis revealed that the modal chromosome number of CCKF cells was 2n = 100. Partial amplification and sequencing of fragments of two mitochondrial genes 16S rRNA and COI confirmed that CCKF cell line originated from ornamental common carp. The CCKF cells showed strong reaction to the cytokeratin marker, indicating that it was epithelial in nature. The extracellular products of Vibrio cholerae MTCC 3904 and Aeromonas hydrophila were toxic to the CCKF cells and not susceptible to viral nervous necrosis virus (VNNV). These CCKF cells were confirmed for the absence of Mycoplasma sp. by polymerase chain reaction. Furthermore, 90% of viable cells could be effectively revived 4 mo after cryopreservation from CCKF cell population suggesting the possibility of long-term storage of the cells.

Mitochondrial ATPase 6/8 genes reveal genetic divergence in the Coilia dussumieri (Valenciennes, 1848) populations of north east and northwest coasts of India.

The golden anchovy, Coilia dussumieri, though possessing discontinuous distribution along northeast and northwest coasts of India, it is being managed as unit stock for fishery assessment purposes. By considering the need for stock specific management of the species, mitochondrial ATP synthase 6 and 8 (ATPase 6/8) genes were analyzed for delineating genetic stock structure of the species. Sequence analysis revealed a total of 34 haplotypes across four populations from both the east and west coasts of India. Haplotype diversity (h) was found in the range of 0.7421-0.9368. Similarly, nucleotide diversity (π) varied from 0.0012 to 0.0025. AMOVA results indicated a high total variance of 72.66% between east and west coast populations and less (1.34%) among populations within the respective coast. Phylogenetic tree constructed using pair wise FST also indicated the genetic divergence of populations of east and west coasts of India. The findings of the present study will be helpful in developing stock specific management measures for conservation and sustainable utilization of the species.

Microsatellite markers to determine population genetic structure in the golden anchovy, Coilia dussumieri.

Coilia dussumieri (Valenciennes, 1848) commonly called as golden anchovy, constitutes a considerable fishery in the northern part of both the west and east coasts of India. Despite its clear-cut geographic isolation, the species is treated as a unit stock for fishery management purposes. We evaluated 32 microsatellite primer pairs from three closely related species (resource species) belonging to the family Engraulidae through cross-species amplification in C. dussumieri. Successful cross-priming was obtained with 10 loci, which were sequenced for confirmation of repeats. Loci were tested for delineating the genetic stock structure of four populations of C. dussumieri from both the coasts of India. The number of alleles per locus ranged from 8 to 18, with a mean of 12.3. Results of pairwise F ST indicated genetic stock structuring between the east and west coast populations of India and also validated the utilization of identified microsatellite markers in population genetic structure analysis.