Molecular and Biochemical Characterization of Insecticide Resistance in Field-Caught Aedes aegypti from Al-Madinah Al-Munawarah, Saudi Arabia

The global resurgence of mosquito-borne diseases such as dengue, chikungunya, and Zika represents a growing threat to public health, largely due to the rapid evolution of insecticide resistance in Aedes aegypti populations. This study investigates the molecular and biochemical mechanisms driving pyrethroid and DDT resistance in field-collected Aedes aegypti females from Al-Madinah Al-Munawarah, Saudi Arabia. Fifty adult female mosquitoes were analyzed to detect mutations in the voltage-gated sodium channel (Vgsc) gene and to evaluate detoxifying enzyme activities. Molecular genotyping revealed the coexistence of three major knockdown resistance (kdr) mutations─S989P, V1016G, and F1534C─occurring in both homozygous and heterozygous combinations. Approximately 36% of the samples exhibited triple mutant genotypes, indicating intense selection pressure and widespread fixation of resistance alleles within the local population. Complementary biochemical assays demonstrated elevated levels of catalase, glutathione S-transferase (GST), and esterases, alongside decreased acetylcholinesterase (AChE) activity, compared with the susceptible laboratory strain. These findings confirm the synergistic contribution of target-site insensitivity and metabolic detoxification to the development of resistance phenotypes. Collectively, this work provides the first integrated molecular–biochemical evidence of multiple resistance mechanisms in Ae. aegypti from Al-Madinah. The results highlight an urgent need for adaptive vector management strategies that incorporate molecular surveillance, insecticide rotation, and nonchemical interventions to mitigate resistance evolution. This study contributes valuable insights into the regional dynamics of resistance in the Arabian Peninsula and emphasizes the necessity for sustainable, evidence-based mosquito control programs.