Maintenance of Gut Microbial Balance via the Kynurenine Pathway Improves Larval Performance and Resistance to Bacillus thuringiensis in Spodoptera exigua.

Abstract

The gut microbiota is a key determinant of insect physiology, influencing nutrition, immunity, and interactions with plants and pathogens. In Lepidoptera, larval gut communities are dynamic, but a core microbiota, often dominated by Enterococcus species, persists across instars. In Spodoptera littoralis, the enzyme kynurenine 3-monooxygenase (KMO) regulates gut bacterial composition via 8-hydroxyquinoline-2-carboxylic acid (8-HQA), a secreted iron-chelating compound. To investigate whether this mechanism is conserved in Noctuidae, we generated Spodoptera exigua kmo^-/- mutants using CRISPR/Cas9 and analyzed bacterial communities in foregut, midgut, hindgut, and oral secretions by 16S metabarcoding, using RNA-derived cDNA for gut samples and DNA for oral secretions due to lower microbial biomass. The kmo deletion abolished 8-HQA production, reduced bacterial diversity, and collapsed compartment-specific bacterial communities in the gut, while also being associated with Enterococcus dominance in oral secretions. Fitness assays revealed that kmo^-/- larvae exhibited reduced weight gain on artificial diet, and higher mortality and delayed growth when fed on pepper leaves. Moreover, kmo^-/- larvae were threefold more susceptible to Bacillus thuringiensis, consistent with an interaction between host physiological state, gut microbial homeostasis, and pathogen susceptibility. Dietary supplementation with 8-HQA partially mitigated, but did not fully rescue, growth deficits. Our results demonstrate that the kynurenine pathway and 8-HQA production are crucial for maintaining gut microbial homeostasis, particularly within Enterococcus, thereby supporting larval development, dietary adaptation, and pathogen resilience. These findings reveal a conserved mechanism in noctuid moths linking host metabolism, microbiota regulation, and ecological performance, emphasizing the interplay between host genetics, microbiota composition, and environmental stressors.