Identification of a novel cytochrome P450 gene,CYP321E1from the diamondback moth,Plutella xylostella(L.) and RNA interference to evaluate its role in chlorantraniliprole resistance

Insect cytochrome P450 monooxygenases (P450s) play an important role in catalysis of many reactions leading to insecticides resistance. Our previous studies on transcriptome analysis of chlorantraniliprole-resistant development in the diamondback moth,Plutella xylostellarevealed that up-regulation of cytochrome P450s are one of the main factors leading to the development of chlorantraniliprole resistance. Here, we report for the first time a novel cytochrome P450 geneCYP321E1, which belongs to the cytochrome P450 gene family CYP321. Real-time quantitative PCR (RT–qPCR) analyses indicated thatCYP321E1was expressed at all developmental stages ofP. xylostellabut was highest in the fourth-instar larvae; furthermore, the relatively high expression was observed in the midgut of the fourth-instar larvae, followed by fat bodies and epidermis. The expression ofCYP321E1inP. xylostellawas differentially affected by three representative insecticides, including alphamethrin, abamectin and chlorantraniliprole. Among them, the exposure to chlorantraniliprole resulted in the largest transcript level of this cytochrome P450 gene. The findings suggested potential involvement ofCYP321E1in chlorantraniliprole resistance ofP. xylostella. To assess the functional link ofCYP321E1to chlorantraniliprole resistance, RNA interference (RNAi)-mediated gene silencing by double stranded RNA (dsRNA) injecting was used. Results revealed that injection delivery of dsRNA can greatly reduce gene expression after 24 h. As a consequence of RNAi, a significant increment in mortality of larvae injectedCYP321E1dsRNA was observed after 24 h of exposure to chlorantraniliprole. These results strongly support our notion that this novel cytochrome P450 gene plays an important role in chlorantraniliprole detoxification in the diamondback moth and is partly responsible for its resistance.