Building a DNA barcode library of commercially exploited fish genetic resources (FGR) of the Gomti River, Ganga...

The Gomti River is a vital tributary of the largest Gangetic Riverine system, flowing in the northern part of Uttar Pradesh, India. This habitat serves as one of India's most precious repository for fish genetic resources (FGR), contributing significantly to regional biodiversity, and is a source of livelihood. It has diverse climatic conditions, which have led to a distinctive fish community structure and the presence of native species. The application of DNA barcoding for assessing FGR in the north Himalayan region, particularly in UP, presents a notable research gap. This study established the first DNA barcodes of voucher specimens identified through classical taxonomy, creating a molecular catalogue of commercially important fish species. DNA barcoding was accomplished using the cytochrome oxidase subunit I gene (COI) for species identification and phylogenetic analysis. We identified 37 commercially important fish species (34 native & 3 exotics) using classical taxonomy and molecular biology, which belonged to 8 orders, 18 families, and 29 genera. The IUCN conservation status revealed that 30 species (81.08%) were categorized as Least Concern (LC), followed by 4 species (10.81%) as Near Threatened (NT), 2 species (5.41%) as Vulnerable (VU), and 1 species (2.70%) as Endangered (EN). Cypriniformes order and Cyprinidae family emerged as the most dominant groups, contributing 40.54% and 29.73%, respectively. Average read lengths of all sequences were 630.82 bp (range 578-655 bp). Overall nucleotide composition of Adenine (A) = 25.81%, Thymine (T) = 28.88%, Cytosine (C) = 27.53%, and Guanine (G) = 17.78%. Average GC content (54.69%) was higher than AT content (45.31%). As expected, the uncorrected p-genetic distances showed a progressive increase in genetic divergence from lower to higher taxonomic levels. p-distance values ranged from 0-0.48% (mean 0.06%) within species, 0-11.37% (1.56%) within genera, 0-12.27% (5.00%) within families, and 0-15.31% (8.88%) within orders. Among families, Cyprinidae (12.27%) and Danionidae (11.57%) exhibited the highest genetic divergence. Whereas at the order level, the greatest divergence was observed in Perciformes (15.31%), followed by Siluriformes (15.22%). The constructed phylogenetic tree shows that all species are clearly separated based on similar groups and clustered into independent clades. Assemble Species by Automatic Partitioning (ASAP) analysis identified ten partitions with the best ASAP score (4.00). It reveals a barcode gap of 1-9% (0.01-0.09) and delimits approximately 31-37 putative fish species. The GTR + G + I model revealed a strong transition bias in the COI gene of the barcoded fishes, with higher pyrimidine substitutions and transition/transversion ratios (k₁ = 10.80; k₂ = 26.98; R = 3.09). Our dataset comprised 110 COI nucleotide sequences with 592 positions in the final alignment, including 236 polymorphic sites and 39 distinct haplotypes. We also estimated haplotype diversity (Hd = 0.975) and nucleotide diversity (π = 0.194). Our findings highlighted that efficacy of traditional taxonomy and DNA barcoding method (also known as integrative taxonomy) is complementary to each other for identifying and authenticating fish taxa. Additionally, the developed COI library will be valuable for future environmental DNA (eDNA) studies for detecting indigenous and exotic fish species and useful for nature conservation. We conclude that it serves as an effective tool for sustainable conservation, fishery management, and future monitoring of freshwater fish genetic resources.