| Summary: | The white-back planthopper (WBPH), <i>Sogatella furcifera</i>, mainly harms rice and occurs in most rice regions in China and Asia. With the use of chemical pesticides, <i>S. furcifera</i> has developed varying degrees of resistance to a variety of pesticides. In our study, a chlorpyrifos-resistant population (44.25-fold) was built through six generations of screening with a sublethal dose of chlorpyrifos (LD<sub>50</sub>) from a field population. The expression levels of ten selected resistance-related P450 genes were analyzed by RT-qPCR and found that <i>CYP408A3</i> and <i>CYP6CS3</i> were significantly more expressed in the third instar nymphs of the XY17-G5 and XY17-G6 populations, about 25-fold more than the Sus-Lab strain, respectively (<i>p</i> < 0.01). To elucidate their molecular function in the development of resistance towards chlorpyrifos, we cloned two P450 full lengths and predicted their tertiary protein structures. <i>CYP408A3</i> and <i>CYP6CS3</i> were also downregulated after injecting <i>dsCYP408A3</i>, <i>dsCYP6CS3</i>, or their mixture compared to the control group. Moreover, the mortality rates of the <i>dsCYP6CS3</i> (91.7%) and the mixture injection treatment (93.3%) treated by the LC<sub>50</sub> concentration of chlorpyrifos were significantly higher than the blank control group (51.7%) and <i>dsCYP408A3</i> injection treatment (69.3%) at 72 h (<i>p</i> < 0.01). Meanwhile, the P450 enzyme activities in the dsRNA treatments were lower than that in the control (XY17-G6) (<i>p</i> < 0.01). Therefore, the P450 gene <i>CYP6CS3</i> may be one of the main genes in the development of chlorpyrifos resistance in <i>S. furcifera</i>.
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