Identification and Functional Characterization of CYP4D2 Putatively Associated with β-Cypermethrin Detoxification in Phortica okadai

Abstract

Phortica okadai, a polyphagous pest, serves as a vector for Thelazia callipaeda in China. Currently, there are no effective control strategies for this vector. Agricultural pest control may cause P. okadai to become a threat due to the development of pesticide resistance. Cytochrome P450s (CYP450) plays a significant role in detoxifying xenobiotics in insects. In this study, we performed RNA sequencing of P. okadai exposed to β-cypermethrin for 0 and 1 h and then gene cloning of the five up-regulated CYP450 genes. Three CYP450 genes were successfully cloned, and their expression patterns in different developmental stages and in different tissues were analyzed by RT-qPCR. Pocyp4d2 was observed to have the highest expression in the midgut (fold change 2.82 for Pocyp4d2, 2.62 for Pocyp49a1, and 1.77 for Pocyp28d2). Functional analysis was carried out according to overexpression in S2 cells from the pfastbac1 vector and RNAi with siRNA. The results of the CCK8 assay indicated that the overexpression of the recombinant protein PoCYP4D2 suppressed the decrease in S2 cell viability due to β-cypermethrin. The expression levels of PoCYP4D2 decreased significantly, and the mortality rates increased from 6.25% to 15.0% at 3 h and from 15.0% to 27.5% at 6 h after Pocyp4d2-siRNA injection. These results suggest that Pocyp4d2 may be an essential key gene in the metabolism of β-cypermethrin in P. okadai. This study constitutes a foundation to explore further the functions of P. okadai CYP450 genes in insecticide metabolism.

Genome-Wide Comparative Analysis of the Cytochrome P450 Monooxygenase Family in 19 Aphid Species and Their Expression Analysis in 4 Cereal Crop Aphids

“…According to tissue expression profiles of Tribolium castaneum , TcCYP6BQ8 is mostly expressed in the head and integument of both larvae and adults [ 53 ]. In another research, Pocyp4d2 had the highest expression in the midgut of Phortica okadai [ 54 ]. It was reported that SsubCYP341A and SsubCYP341B_ortholog1 were more highly expressed in the pheromone gland than in the female body of Scopula subpunctaria [ 49 ].…”