Midgut microorganisms transformed into hemolymph pathogens: Mediating DUOX-ROS immune response to regulate the insecticidal activity of Bacillus thuringiensis against Spodoptera exigua

Bacillus thuringiensis (Bt) is a microbial insecticide that disrupts intestinal microflora homeostasis and is widely used in agricultural pest control. Dual oxidase-reactive oxygen species (DUOX-ROS) can help insects resist pathogenic infections and maintain intestinal microbiota balance. However, it remains uncertain whether the intestinal microbiota regulates the DUOX-ROS immune pathway in the hemolymph and thereby influences Bt insecticidal activity. In this study, Bt oral infection inhibited the formation of vesicles (Ves), caused the loss of microvilli (Mi), and resulted in pore formation in the midgut, all of which aim to destroy the integrity of the midgut of Spodoptera exigua larvae. It also led to a significant increase in the bacterial load in the midgut and hemolymph, including loads of two isolated cultivable gut bacterial strains, Bacillus cereus HB1 and Enterococcus mundtii HB1. Bt oral infection suppressed the expression of DUOX-ROS pathway genes and ROS levels in the hemolymph of S. exigua. The introduction of Bt GS57, B. cereus HB1, and E. mundtii HB1 into the hemocoel of S. exigua caused significant larval mortality, downregulated the expression of DUOX-ROS pathway genes in hemolymph without enhancing ROS levels, and activated midgut-related DUOX-ROS immune response. Taken together, these results demonstrate that Bt oral infection disrupts midgut integrity and represses the DUOX-ROS immune response in the hemolymph, potentially allowing midgut bacteria to transform into hemolymph opportunistic pathogens and regulating the insecticidal activity of Bt, which enriches the insecticidal mechanism of Bt and provides a new perspective for controlling insects.