NADPH-cytochrome P450 reductase involved in the multi-resistance of Aphis spiraecola to imidacloprid and lambda-cyhalothrin

NADPH-cytochrome P450 reductase (CPR) serves as the obligate electron donor for cytochrome P450 enzymes and is essential for xenobiotic metabolism and detoxification. However, its involvement in insecticide resistance of Aphis spiraecola remains unclear. In this study, the open reading frame (ORF) of the A. spiraecola CPR gene was cloned and characterized. qRT-PCR analysis revealed that AsCPR was most abundantly expressed in apterous adults of the susceptible (SS), imidacloprid-resistant (IM-R) and lambda-cyhalothrin-resistant (LC-R) strains, with higher transcript levels in the hemolymph and midgut. In different strains, AsCPR expression was stably induced by both imidacloprid and lambda-cyhalothrin, with induction levels ranging from 1.21 to 4.9-fold. Furthermore, SPc-mediated RNAi of AsCPR achieved silencing efficiencies exceeding 40% at 12 h post-injection in both resistant strains. This knockdown significantly reduced P450 enzyme activity by 18.4% in the IM-R strain and 16.6% in the LC-R strain. Accordingly, the sensitivities of the IM-R strain to imidacloprid and the LC-R strain to lambda-cyhalothrin were significantly increased, with LC 50 values decreasing by 57.6% and 49.8%, respectively, compared with the ds GFP control group. Finally, the recombinant AsCPR protein was expressed in vitro , and its activity was assessed. The results indicate that AsCPR plays a key role in the development of multi-resistance of A. spiraecola to pyrethroid and neonicotinoid insecticides.