Involvement of cytochrome P450 gene CYP333B119v2 in the resistance of Cydia pomonella to lambda-cyhalothrin

Extensive studies have demonstrated that cytochrome P450 enzymes play a critical role in the metabolic detoxification and resistance development of pests against insecticides; however, relative to P450 genes in Clan 3, knowledge on mitochondrial Clan members remains limited. Here, we report the involvement of a CYP333 family member, CYP333B119v2 , in conferring lambda -cyhalothrin resistance in Cydia pomonella . Phylogenetic analysis shows that CYP333B119v2 clusters with CYP333 family members of Lepidoptera, it containing three highly conserved P450 domains: the Heme-binding region, C-Helix, and K-helix. Spatiotemporal expression profiling indicates that CYP333B119v2 transcription levels increase progressively during larval development, peaking at the fourth instar, with significant expression in the midgut, the primary detoxification tissue. Relative to susceptible population (SS), CYP333B119v2 expression is 2.73-fold higher in lambda -cyhalothrin moderate resistant population (LCR) and is significantly induced by sublethal doses of lambda -cyhalothrin . Knockdown of CYP333B119v2 enhances larval susceptibility to lambda -cyhalothrin. Molecular docking based on AlphaFold models revealed strong binding interactions between CYP333B119v2 and lambda -cyhalothrin, with diverse non-covalent interactions involved in the binding process. These findings suggest that CYP333B119v2 may contribute to lambda -cyhalothrin resistance in C. pomonella . This study is the first report implicating a mitochondrial CYP333B subfamily member in the resistance mechanisms of Lepidoptera, advancing our understanding of P450-mediated metabolic resistance and providing new insights into pest management strategies.