Spodoptera frugiperda, commonly known as the fall armyworm (FAW), is a major pest across the globe due to its broad host range and distribution worldwide. We investigated the function of microRNAs (miRNAs) in the detoxification of insecticides, with a specific focus on its susceptibility to chlorantraniliprole which is widely utilized insecticide for its management. miRNAs are small non-coding RNA molecules, crucial for post-transcriptional regulation of gene expression. This study aims to elucidate the impact of these miRNAs on the expression of cytochrome P450 genes, which play a significant role in conferring insecticide resistance. We identified notable changes in the abundance of two specific miRNAs, sfr-miR-10465-5p and sfr-miR- 10476-5p through RNA sequencing, after chlorantraniliprole exposure. These miRNAs exhibited significantly high expression in the fat body tissue, while showing relatively lower expression in the head, midgut, and malpighian tubules. Further analysis suggested that these miRNAs might target specific cytochrome P450 genes, like CYP4C1 and CYP4C21, which are known to play a role in insecticide resistance development. Experimentation with miRNA mimics through microinjection revealed a notable increase in the survival rates of S. frugiperda larvae when subjected to chlorantraniliprole exposure, with a significant reduction in CYP4C1 and CYP4C21 gene expression levels. This suggests a direct connection between the miRNAs and the increased tolerance of Spodoptera larvae to the insecticide. Our research presents the complex function of miRNAs in gene expression regulation related to insecticide resistance, offering valuable insights into the molecular mechanisms of chlorantraniliprole resistance in S. frugiperda. These findings pave the way for further investigations into miRNA roles and their potential in managing pesticide resistance in agricultural pests.

The fall armyworm, Spodoptera frugiperda, has developed extremely high levels of resistance to chlorantraniliprole and other classes of insecticides in the field. As microRNAs (miRNAs) play important roles in various biological processes through gene regulation. we examined the miRNA profile of S. frugiperda in response to Chlorantraniliprole, Indoxacarb and Thiamethoxam. Transcriptome analysis showed significant changes in the abundance of some miRNAs after treatment of S. frugiperda larvae with LC20 concentrations of three insecticides. A total of 197 miRNAs were systematically identified from S. frugiperda, and 16, 9, 2 miRNAs were differentially expressed after treatments of three insecticides. Importantly, three miRNAs were significantly downregulated and three were upregulated by RT-qPCR after treatment the LC50 of three insecticides with S. frugiperda larvae. Microinjection of agomirs of these six miRNAs into S. frugiperda larvae resulted in significant changes in mortality rates when exposed to three insecticides. Additionally, we also screened potential target genes for some of differentially expressed miRNAs, which may play important roles in insecticide resistance development. These findings provide valuable insights into the molecular mechanisms of insecticide resistance and underscore the potential of miRNAs as targets for the development of novel pest control strategies in S. frugiperda.