PxSpätzle3 Regulates the Toll Pathway To Affect <i>Bacillus thuringiensis</i> Susceptibility of <i>Plutella xylostella</i>

Spätzle is an important messenger in the Toll pathway of the insect innate immune system. However, the function of Spätzle in regulating the Toll pathway in <i>Bacillus thuringiensis</i> (Bt) susceptibility of <i>Plutella xylostella</i> is unclear. In this study, we cloned the <i>Spätzle3</i> gene of <i>P. xylostella</i>. Molecular docking and yeast two-hybrid experiments indicated that PxSpätzle3 combined with PxToll6 to regulate the Toll pathway. After knocking out <i>PxSpätzle3</i>, the expression of downstream Toll pathway genes and antimicrobial peptides (AMPs) decreased. Antibacterial assays showed that PxGloverin2 could inhibit Bt8010. Further bioassays revealed that the susceptibility of the mutant to Bt8010 was significantly higher than that of the wild type. Intriguingly, the gut bacteria <i>Enterobacter</i> sp. EbPXG5 enhanced the lethality of Bt8010 to the mutant <i>P. xylostella</i>. Our findings clarify that PxSpätzle3 activates the Toll pathway by binding with PxToll6, regulates AMP production, and affects the susceptibility of <i>P. xylostella</i> to Bt8010. Additionally, our study uncovers the role of gut bacteria in this process. These insights provide new ideas for the sustainable control of pests.