DefinePK

Abstract

Euconocephalus pallidus (Redtenbacher, 1891), a katydid species belonging to the family Tettigoniidae, plays a significant ecological role across diverse habitats in Pakistan. Despite its ecological importance, molecular data on its genetic diversity and evolutionary relationships remain scarce. To address this knowledge gap, the present study investigates the genetic variation of E. pallidus using DNA barcoding of the mitochondrial Cytochrome c Oxidase I (COI) gene. PCR amplicons were purified and sequenced unidirectionally using the Sanger sequencing method. Raw sequences were processed, aligned using ClustalW, and compared with reference sequences from the NCBI database via BLAST analysis. Phylogenetic relationships were inferred using the Maximum Likelihood method based on the Tamura-Nei model in MEGA11 software. BLAST results showed high sequence identity between the obtained COI sequences and those of closely related species, supporting the morphological identification of E. pallidus . The resulting phylogenetic tree placed E. pallidus in a clade with congeners such as E. nasutus and E. varius , supported by moderate bootstrap values. Furthermore, the observed intra-population genetic variation suggests potential phylogeographic structuring within the sampled population. This study provides novel insights into the genetic diversity and evolutionary positioning of E. pallidus in Pakistan. It represents the first molecular assessment of this species using DNA barcoding in the ecological context of the Bahawalpur division, addressing a critical gap in molecular data for Orthopteran insects in the region.