关键词: 美国白蛾, 细胞色素P450, 氯虫苯甲酰胺, 耐受性, RNAi

Abstract: 【Aim】 To reveal the functions of the cytochrome P450 (CYP450）genes, HcCYP4S47 and HcCYP332A29, in Hyphantria cunea in response to chlorantraniliprole stress.【Methods】The diets containing LC₃₀ (0.06 mg/L) of chlorantraniliprole were fed to the 3rd instar larvae of the laboratory population, northern population (Tieling population from Liaoning), and southern population (Dawu population from Xiaogan, Hubei) of H. cunea via mixing the pesticide into the diets, and the surviving larvae were collected at 6, 12, 24 and 48 h after feeding treatment, respectively. The CYP450 activity in the 3rd instar larvae of H. cunea was determined. The expression levels of HcCYP4S47 and HcCYP332A29 in the 3rd instar larvae of the three geographical populations after exposure to 0.06 mg/L of chlorantraniliprole were analyzed using RT-qPCR. Additionally, HcCYP4S47 and HcCYP332A29 in the 3rd instar larvae of the three geographical populations were silenced by RNAi technology, and the expression levels of target genes were detected by RT-qPCR after dsRNA injection. The diets containing 0.06 mg/L of chlorantraniliprole were fed to the gene-silenced 3rd instar larvae of the three geographical populations, and the survival rates of H. cunea larvae were recorded at 12, 24, 36, 48, 60 and 72 h after feeding.【Results】 After exposure to 0.06 mg/L of chlorantraniliprole, the CYP450 activities in the 3rd instar larvae of the northern and southern populations of H. cunea were higher than those in the laboratory population at four treatment time points (6, 12, 24 and 48 h). Specifically, the CYP450 activity in the 3rd instar larvae of the northern population in the chlorantraniliprole treatment group exhibited the highest levels at 24 and 48 h after treatment, while that in the 3rd instar larvae of the southern population in the chlorantraniliprole treatment group peaked at 12 and 24 h after treatment. Phylogenetic analysis indicated that HcCYP4S47 and HcCYP332A29 belong to the CYP4 and CYP3 subfamilies, respectively. The expression levels of HcCYP4S47 and HcCYP332A29 were significantly higher in the 3rd instar larvae of the northern and southern populations than those in the 3rd instar larvae of the laboratory population. After exposure to 0.06 mg/L of chlorantraniliprole, the expression levels of HcCYP4S47 and HcCYP332A29 were significantly up-regulated in the 3rd instar larvae of the laboratory population as compared to those in the control group, peaking at 12 h post treatment. The expression levels of HcCYP4S47 in the 3rd instar larvae of the southern and northern populations were significantly down-regulated at 6, 12 and 48 h after chlorantraniliprole treatment, that in the northern population was significantly up-regulated and that in the southern population showed no significant change at 24 h after chlorantraniliprole treatment, as compared to that in the control group. The expression level of HcCYP332A29 in the 3rd instar larvae of the northern population in the chlorantraniliprole treatment group was significantly decreased at 6 and 12 h, but significantly up-regulated at 48 h, as compared to that in the control group. The expression level of HcCYP332A29 in the 3rd instar larvae was significantly decreased at 6 and 12 h after chlorantraniliprole treatment but significantly up-regulated at 48 h after chlorantraniliprole treatment as compared to that in the control group. In the 3rd instar larvae of the southern population, the expression level of HcCYP332A29 was significantly down-regulated at 6 h after chlorantraniliprole treatment, followed by a significant increase at 12 and 24 h. Silencing HcCYP332A29 and HcCYP4S47 increased the susceptibility of the 3rd instar larvae of H. cunea to chlorantraniliprole. 【Conclusion】 HcCYP332A29 and HcCYP4S47 play crucial roles in the response of H. cunea to chlorantraniliprole stress.

Key words: Hyphantria cunea, cytochrome P450; chlorantraniliprole; tolerance, RNAi