Genetic Structure and Phylogeographic analysis of six Anopheles (Diptera: Culicidae) mosquito species Using the Mitochondrial COI Gene Marker in the Northern districts of Tamil Nadu, India.

Species of Anopheles serve as the primary vectors of human malaria parasites worldwide. Accurate species identification is critical for the design and implementation of effective vector control strategies, particularly in regions where cryptic species complexes complicate surveillance and management. In this study, we examined the genetic diversity and phylogenetic relationships of six Anopheles species-An. annularis, An. pallidus, An. jamesi, An. subpictus, An. vagus, and An. tessellatus-collected in the northern districts of Tamil Nadu, India. Mitochondrial cytochrome oxidase I (COI) gene sequences were used to generate a phylogenetic tree, which revealed distinct conspecific species clusters. The mean of the lowest intraspecific and highest interspecific Kimura 2-parameter (K2P) distances was 0.003 and 0.158, respectively. Intraspecific polymorphism estimates using DnaSP v6.12.03 showed An. tessellatus complex exhibited numerous polymorphic and mutation sites, with a nucleotide diversity (π) of 0.046. The highest nucleotide diversity was observed in An. subpictus s.l. (0.078), whereas the lowest was in An. pallidus (0.007). Haplotype diversity (Hd) ranged from 0.735 in An. pallidus to 0.969 in An. vagus. Population genetic analyses confirmed considerable genetic diversity across populations. Evolutionary relationships among haplotypes were inferred using haplotype network analysis in PopART 1.7. Overall, this study highlights the genetic diversity and potential population expansion of six notable Anopheles species, offering insights into migration patterns, molecular biology, and vector competence that support species monitoring and region-specific vector control strategies.