Proactive Resistance Management Studies Highlight the Role of Cytochrome P450 Genes in the Resistance of Tuta absoluta Against Tetraniliprole

The diamide insecticide tetraniliprole is a valuable tool for managing major insect pests like the invasive tomato pinworm, <i>Tuta absoluta</i> (Meyrick). However, the mechanisms underlying tetraniliprole resistance, as well as its associated fitness costs, remain unclear. In this study, we assessed the fitness of tetraniliprole-resistant (TetraRS) and susceptible (SS) strains of <i>T. absoluta</i> and conducted Illumina RNA-seq to compare their transcriptomes. We also used nanocarrier-mediated RNA interference (RNAi) to knockdown P450 genes and evaluate their role in tetraniliprole resistance. After eight generations of selection, <i>T. absoluta</i> developed a 20.80-fold resistance to tetraniliprole, accompanied by fitness costs. RNA-seq analysis revealed 3332 differentially expressed genes (DEGs), with 1707 upregulated and 1625 downregulated in the TetraRS compared to the SS strain. Gene Ontology (GO) annotations showed significant enrichment in categories related to metabolic processes, cellular processes, catalytic activity, cellular anatomical entity, and binding. These genes were also identified in key KEGG pathways such as cytochrome P450, drug metabolism, carbon metabolism, oxidative phosphorylation, fatty acid metabolism, and protein processing. RT-qPCR analysis confirmed that P450 genes (<i>CYP405D1</i>, <i>CYP6AB269</i>, and <i>CYP4AU1</i>) were upregulated in TetraRS insects, in line with the RNA-seq results. Cytochrome P450 activity was significantly higher in the TetraRS strain than in the SS strain. Notably, nano-encapsulated dsRNA targeting these overexpressed P450 genes increased the susceptibility of <i>T. absoluta</i> to tetraniliprole. Further, cytochrome P450 activity was significantly reduced following silencing of P450 genes. These findings suggest that multiple genes and pathways, particularly P450 genes, contribute to tetraniliprole resistance in <i>T. absoluta</i>. This study provides valuable insights into the molecular mechanisms underlying insecticide resistance in this key pest species.