Azadirachtin Inhibits Nuclear Receptor in the Prothoracic Gland to Block Larval Ecdysis in the Fall Armyworm, .

Azadirachtin has been used to control agricultural pests for a long time; however, the molecular mechanism of azadirachtin on lepidopterans is still not clear. In this study, the fourth instar larvae of fall armyworm were fed with azadirachtin, and then the ecdysis was blocked in the fourth instar larval stage (L4). The prothoracic glands (PGs) of the treated larvae were dissected for RNA sequencing to determine the effect of azadirachtin on ecdysis inhibition. Interestingly, one of the PG-enriched genes, the nuclear hormone receptor 3 (), was decreased after azadirachtin treatment, which plays a critical role in the 20-hydroxyecdysone action during ecdysis. To deepen the understanding of azadirachtin on ecdysis, the was knocked out by using the CRISPR/Cas9 system, while the mutants displayed embryonic lethal phenotype; thus, the stage-specific function of during larval molting was not enabled to unfold. Hence, the siRNA was injected into the 24 h L4 larvae to knock down . After 96 h, the injected larvae were blocked in the old cuticle during ecdysis which is consistent with the azadirachtin-treated larvae. Taken together, we envisioned that the inhibition of ecdysis in the fall armyworm after the azadirachtin treatment is due to an interference with the expression of in PG, resulting in larval mortality. The results in this study specified the understanding of azadirachtin on insect ecdysis and the function of in lepidopteran .