Gene expression profiles of the Southern house mosquito Culex quinquefasciatus during exposure to permethrin

Insecticide resistance is a major obstacle to the management of diseasevectoring mosquitoes worldwide. The genetic changes and detoxification genes involved in insecticide resistance have been extensively studied in populations of insecticide-resistant mosquitoes, however few studies have focused on the resistance genes upregulated upon insecticide exposure and the possible regulation pathways involved in insecticide resis- tance. To characterize the changes in gene expression during insecticide exposure, and to investigate the possible connection of known regulation pathways with insecticide resistance, we conducted RNA-Seq analysis of a highly permethrin-resistant strain of Culex quinquefasciatus following pcrmethrin exposure. Gene expression profiles revealed a total of 224 upregulated and 146 downregulated genes when compared to a blank acetone carrier treated control, respectively, suggesting that there were multiple, but specific genes involved in permethrin resistance. Functional enrichment analysis showed that the upregulated genes contained multiple detoxification genes including a glutathione S-transferase and multiple eytochrome P450 genes, as well as several immune-related genes, while the downregulated genes consisted primarily of proteases and carbohydrate metabolism and transport. Further analysis showed that permethrin exposure resulted in a decrease in the expression of serum storage proteins and likely represented a delay in the development of the fourth instar possibly due to a decrease in feeding. This effect was more pronounced in an insecticide-resistant strain than in an insecticide-susceptible strain and may represent a behavioral mechanism of insecticide resistance in Culex mosquitoes.

Molecular and functional characterization of three novel carboxylesterases in the detoxification of permethrin in the mosquito, Culex quinquefasciatus

Carboxylesterases (CarEs) belong to a super family of multifunctional enzymes associated with the degradation of endogenous and exogenous compounds. Many insect CarEs are known to play important roles in catalyzing the hydrolysis of organophosphates (OPs), carbamates, and synthetic pyrethroids (SPs). The elevation of esterase activity through gene amplification and overexpression of estα2 and estβ2 genes contributes to the development of resistance to OP insecticides in the mosquito Culex quinquefasciatus. Three additional CarE genes are upregulated in permethrin-resistant Cx. quinquefasciatus according to an RNA-seq analysis, but their function remains unknown. In this study, we, for the first time, characterized the function of these three novel genes using in vitro protein expression, an insecticide metabolism study and molecular docking analysis. All three CarE genes were significantly overexpressed in resistant mosquito larvae, but not adults, compared to susceptible strain. No gene copy differences in these three genes were found in the mosquitoes tested. In vitro high-performance liquid chromatography (HPLC) revealed that CPIJ018231, CPIJ018232, and CPIJ018233 metabolized 30.4% ± 2.9%, 34.7% ± 6.8%, and 23.2% ± 2.2% of the permethrin, respectively. No mutations in resistant strains might significantly affect their CarE hydrolysis ability. A docking analysis further confirmed that these three CarEs from resistant strain all potentially metabolize permethrin. Taken together, these three carboxylesterase genes could play important roles in the development of permethrin resistance in Cx. quinquefasciatus larvae through transcriptional overexpression, metabolism, and detoxification.