Analysis of influencing factors and functional redundancy of bacterial community structure in the gut of folivorous larvae

Insect gut bacterial communities play an important role in host nutrient metabolism, detoxification and adaptive evolution. However, the driving mechanisms and functional correlations that shape and maintain the community structure still require systematic analysis. This study established a "folivorous larvae - plants - bacteria community" tripartite system to investigate the factors influencing gut bacterial community structure of folivorous larvae and structure-function relationships within these bacterial communities. Field collections spanned three distinct geographic regions (Beijing, Zhejiang, and Sichuan), yielding 304 larval specimens, 142 host plant samples, and 106 soil samples. DNA barcoding and morphological identification were utilized to determine species for both insects and plants. Through 16S rRNA amplicon sequencing (V3-V4 regions), we assessed how host taxonomy, host plants, and environmental microorganisms (phyllosphere/soil) shape insect gut microbiota. PICRUSt2 functional predictions were employed to test for functional redundancy in larval gut microbiota. The results showed that the effects of folivorous larvae species identity and host plants identity on gut microbiota structure intensified with taxonomic resolution (Order→Species), peaking at the species level. The taxonomic factors at the larval species level explained 30.27% of the structural variations of the gut bacterial community (R² = 0.3027, F = 3.2457, P < 0.001) and the taxonomic factors of host plants at the species level explained 28.73% of the variations (R² = 0.2873, F = 4.1882, P < 0.001), respectively. While phyllosphere microbiota, soil microbiota, and sampling sites significantly influenced gut community structure (P < 0.001), their effects were subordinate to larval species identity and dietary differences. At the species level, significant divergence in gut bacteria composition was detected across 65.37%~70.56% of folivorous larvae species pairs. But only 25.29%~26.71% of these pairs exhibited functionally distinct microbial profiles, demonstrating widespread functional redundancy within gut bacterial communities among these folivorous larvae.