A Toll7‐1‐Spätzle7‐1 pathway in Spodoptera litura against Bacillus thuringiensis

BACKGROUND Insect immunity plays a key role in defense against Bacillus thuringiensis (Bt). While the Toll/Toll‐like receptor (TLR) pathway is conserved for innate immunity, its specific role in immune defense against Bt remains poorly characterized. Furthermore, the functional distinctions between the prototypical Toll/TLR subtypes – single cysteine cluster (scc) and multiple cysteine cluster (mcc) – have not been thoroughly investigated. RESULTS Here, we identified 12 Toll and 12 Spätzle (Spz, the Toll ligand) genes in Spodoptera litura and classified them via genome‐wide phylogenetic analysis. All SlTolls are under negative selection, with mccTolls experiencing stronger purifying selection. S. litura mccTolls are intronless genes densely clustered on one chromosome, while a subset of Spz genes (group 2 Spzs) are clustered on another chromosome. S. litura mccTolls possess more structural motifs with regular arrangements, analyses of protein–protein interaction and molecular docking revealed that mccTolls exhibit broader ligand‐binding specificity. SlToll7‐1 (an mccToll) and SlSpz7‐1 (a group 2 Spz) were significantly upregulated in the larval midgut following exposure to Bt toxins, and SlToll7‐1 interacted with SlSpz7‐1. RNAi‐mediated knockdown of either SlToll7‐1 or SlSpz7‐1 significantly suppressed the expression of antimicrobial peptide (AMP) genes and increased larval susceptibility to Bt. CONCLUSION We demonstrate that the Toll7‐1‐Spz7‐1 pathway in S. litura is critical for defense against B. thuringiensis . Our findings also provide evolutionary insights into the diversification of mcc‐ and scc‐Tolls/TLRs.