The Susceptibility Status of Aedes aegypti (Diptera: Culicidae) Mosquitoes in Malaysia on Pyrethroid and Organophosphate Insecticides with First Detection of T1520I Mutation

Background Aedes aegypti , the primary vector of dengue, has developed widespread resistance to insecticides, posing a threat to the efficacy of vector control programs. This study assessed the susceptibility status of Ae. aegypti across Malaysia from seven dengue hotspot localities and characterized knockdown resistance ( kdr ) mutations, including the first detection of T1520I. Methods Susceptibility was assessed using World Health Organization (WHO, 2022) bioassays with four insecticides: deltamethrin (0.03%), permethrin (0.4%), pirimiphos-methyl (60 mg/m²), and malathion (5%). Knockdown times (KT₅₀ and KT₉₅) were determined using probit regression, and resistance ratios were calculated relative to a laboratory-susceptible strain. Genomic DNA was extracted from survivors, and sequencing of the voltage-gated sodium channel (VGSC) gene was conducted to detect knockdown resistance (kdr) mutations. Results All field populations remained susceptible to malathion, with mortality ≥ 95%, though one Johor strain (ABS) showed suspected resistance (95%). Pirimiphos-methyl resistance was widespread, with mortality as low as 6% (ABS). Whereas resistance to pyrethroids was pronounced, with deltamethrin mortality ranging from 22% (ABS) to 95% (AA), and permethrin from 0% (TMB, ABS) to 95% (AA). Knockdown assays revealed high resistance ratios, particularly for permethrin (TMB: RR₅₀=9.98, RR₉₅=14.98; ABS: RR₉₅=41.46). Sequencing identified multiple kdr mutations, including S989P, V1016G, F1534C, A1007G, and the novel detection of T1520I in Malaysian populations. F1534C was strongly associated with permethrin resistance, while V1016G and S989P predominated under deltamethrin exposure. Conclusions Aedes aegypti populations in Malaysia exhibit widespread pyrethroid resistance and emerging organophosphate resistance, underpinned by multiple kdr mutations. The first detection of T1520I underscores the evolving genetic basis of resistance. These findings highlight the urgent need for integrated resistance management, including molecular surveillance and insecticide rotation, to sustain effective dengue control.