Brown_2026_Pestic.Biochem.Physiol_217_106832

Insecticide resistance in Aedes aegypti is driven by multiple mechanisms, yet the dynamics of their development under defined selection regimes remain unclear. Here, we established laboratory selection lines from a field-collected parental strain (AeStA) using permethrin, malathion, or alternating exposures to these two insecticides across successive generations. Continuous selection with permethrin rapidly produced extremely high resistance (resistance ratio, RR = 2700) within five generations, whereas malathion selection yielded only low-level tolerance (RR = 3). Alternating permethrin and malathion exposures delayed the escalation of permethrin resistance and produced only modest malathion tolerance. Molecular analyses revealed that permethrin-selected and alternately selected strains carried nearly fixed vgsc kdr mutations (V1016I and F1534C) after the first generation. Despite their early fixation, resistance continued to rise, implicating additional mechanisms. Synergist assays and transcriptomic analysis confirmed that cytochrome P450s (notably CYP6BB2 and CYP325v1 proteins) and carboxylesterases contributed to permethrin detoxification. The combination of target-site substitutions and metabolic detoxification likely acted synergistically to generate the exceptionally high permethrin resistance observed. By contrast, no carboxylesterase gene (ace-1) mutations were detected in malathion-selected strains; modest overexpression of CYP6BB2 and CYP6M11 was insufficient to drive strong resistance, underscoring the absence of synergism between target-site and metabolic mechanisms. These laboratory selection experiments demonstrate how resistance trajectories in Ae. aegypti are shaped by the interaction of target-site and metabolic mechanisms. They provide insights into resistance processes under controlled laboratory conditions, while not serving as direct predictions for operational mosquito control programs.