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

Abstract

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.

Keywords: Culex quinquefasciatus; carboxylesterase; gene expression; in vitro protein expression; metabolism; permethrin resistance.