Morphometric and Molecular Characterization of Mosquito Species from Different Larval Habitats in Gulshan Thana, Dhaka

Abstract

This comprehensive study, conducted in Gulshan Thana, Dhaka, investigated mosquito diversity, larval habitat characteristics, and cryptic species identification using an integrated approach combining wing morphometrics and molecular analysis. Larval habitats were systematically surveyed across natural and artificial breeding sites during pre-, peak-, and post-monsoon seasons over three consecutive years using a stratified random sampling design. GPS-referenced data analyzed in ArcGIS Pro revealed high-density breeding zones through kernel density and hotspot analyses. A total of 36 mosquito species across eight genera were identified, including 11 new records for Bangladesh: Aedes (Ae.) aegypti aegypti, Ae. aegypti var. luciensis, Ae. flavopictus, Culex (Cx.) annulirostris, Cx. pipiens, Cx. pipiens f. pipiens, Cx. pipiens f. molestus, Cx. pipiens pallens, Cx. pseudosinensis, Lutzia (Lt.) chiangmaiensis, and Ochlerotatus sp. The Breteau Index comparison between Ae. aegypti and Ae. albopictus was significant (U = 109.5, p = 0.032), peaking in September, with Korail slum identified as a year-round hotspot for Ae. aegypti. Habitat preferences varied, with Culex preferring drains (16.83%) and Aedes favoring water filter pots (27.07%). Morphometric analysis of wing landmarks (CVA and PCA) revealed significant interspecific variations in size and shape (p < 0.0001). Molecular characterization using COI and rRNA markers produced 33 COI and four ribosomal sequences, with an average length of 662.91 base pair. Genetic divergence among six mosquito genera was evaluated using the Kimura 2-parameter model, revealing an average pairwise distance of 21.62%. A Neighbor-Joining (NJ) tree was constructed with 1,000 bootstrap replicates using MEGA 11, to evaluate the robustness of the inferred clades. The Kruskal-Wallis test confirmed significant intergeneric variation (H = 33.14, p = 3.54 × 10⁻⁶). A median-joining network of 20 COI haplotypes revealed high intraspecific genetic diversity (HD = 1.000 ± 0.016; π = 0.11723 ± 0.00855). Notably, Ae. aegypti showed clear divergence from its cryptic subspecies: Ae. aegypti aegypti (distance: 0.21) and Ae. aegypti var. luciensis (distance: 2.15), with 2.30% divergence. The detection of geographically isolated taxa such as Cx. pipiens pallens and Ae. aegypti var. luciensis highlights ongoing ecological range expansion among invasive disease vectors.