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Fish are highly diverse aquatic organisms that are integral to global food systems and have been extensively scientific investigation. The region of Dir and Malakand Khyber Pakhtunkhwa was examined for the first time, expanding the cytochrome c oxidase I (COI) gene as a DNA barcoding tool for accurate species documentation in conjunction with traditional morphological diagnostic techniques. Fish specimens were collected from three distinct local streams including, River Panjkora, the Gumbat Khwar River and the Shogram Khwar River in Malakand Division, Khyber Pakhtunkhwa (KP), Pakistan. A total of fourteen species (Barilius vagra, Garra gotyla, Channa punctata, Channa marulius, Crossocheilus diplocheilus, Pethia conchonius, Carassius auratus, Schizothorax plagiostomus, Glyptothorax cavia, Glyptosternon reticulatum, Paraschistura cristata, Paraschistura abdolii, Mastacembelus armatus and Acanthocobitis botia) belonging to 13 genera, five families and four orders were identified by using morphological and COI based analysis. The diversity indices indicated rich ichthyofauna, with a Simpson index of 0.92 and a Shannon–Wiener index of 0.80. S. plagiostomus showed the highest relative density, while A. botia was the least abundant. COI sequencing produced a 594 bp fragment with an overall GC content of 46.37%, containing 41% variable sites and a transition/transversion ratio of 1.5. The mean genetic divergence was 0.24%, supporting clear species-level differentiation. Furthermore, phylogenetic analysis of each family was performed to validate the usefulness of the COI in species identification and their evolutionary relationship. This study demonstrates the effectiveness of the integration of molecular analysis with a traditional morphological taxonomic method to identify fish robustly and reliably as well as the need to have advanced techniques such as mitochondrial genome methods and the use of a broader geographical region.
Jan et al. (Thu,) studied this question.