Abstract

Accurate species identification is fundamental for understanding insect pest diversity and developing effective management strategies. Wheat aphids are among the most destructive insect pests worldwide, yet comprehensive molecular confirmation of species diversity in Bangladesh has been lacking. This study employed an integrative taxonomic approach combining DNA barcoding and morphological characterization to identify wheat aphid species collected from major wheat-growing regions of Bangladesh. Aphid specimens were sampled from 11 locations across four districts during the 2024-2025 wheat growing season. A partial fragment of the mitochondrial cytochrome C oxidase subunit I (COI) gene was amplified and sequenced, followed by BLAST analysis, phylogenetic reconstruction using the Maximum Likelihood method, genetic distance estimation, and detailed morphometric analyses. COI sequence analysis identified five wheat aphid species: Sitobion avenae, Rhopalosiphum padi, R. maidis, R. rufiabdominalis, and Hysteroneura setariae. Sitobion avenae was the predominant species, occurring across all surveyed regions. BLAST similarity ranged from 99.68% to 100%, confirming reliable species-level identification. Phylogenetic analysis grouped all specimens into five well-supported species-specific clades with strong bootstrap support (97-100%), corroborating molecular identification. Pairwise genetic distance analysis revealed very low intraspecific divergence (0.0-0.18%) and clear interspecific divergence (4.9-11.9%), indicating strong genetic discrimination among species. Morphological and morphometric analyses further distinguished the five species, with significant interspecific differences (P < 0.0001) observed for body length, antennal length, tubercle width, cauda length, and cornicle length. The concordance between molecular and morphological evidence demonstrates the robustness of the integrative taxonomic framework. This study provides the first comprehensive molecular and morphological characterization of wheat aphid species in Bangladesh, establishing a reliable baseline for biodiversity assessment, pest surveillance, insecticide resistance monitoring, and the development of evidence-based integrated pest management strategies to support sustainable wheat production amid changing agroecological conditions.