Dietary niche shapes bacterial community in Indo-Pacific ants

Ants are among the most ecologically diverse insects, especially in tropical forest ecosystems, yet what shapes their microbial associates remains poorly understood. Most research has focused on Neotropical ants, where strong microbial associations have been linked to shifts in diet-such as herbivory-and nesting ecology. In contrast, Indo-Pacific ants, which have independently evolved similar specialized lifestyles, remain largely unstudied for their microbial associations. Here, we integrate deep-coverage 16S rRNA sequencing with stable isotope (δ¹⁵N) analyses to characterize the microbiomes of 239 ants from 36 species across 24 genera from the Indo-Pacific region. These ants span a broad ecological range, from ground-dwelling predators to strict arboreal herbivores, allowing us to ask: which ecological traits shape bacterial diversity in tropical ants? Our results reveal that dietary niche (δ¹⁵N)-rather than nesting habitat-is the primary explanatory variable of microbial diversity and composition. Ants consuming protein-rich diets (predators and omnivores) exhibit significantly higher bacterial alpha diversity and distinct community composition profiles compared to their plant-based (herbivorous) counterparts, with Rickettsiales, Rhizobiales, and Enterobacterales as major contributors to these differences. Notably, herbivorous ants tend to harbor simpler, Enterobacterales-dominated microbiomes, whereas predators support more complex communities, frequently containing Rhizobiales bacteria. Phylogenetic comparative analyses suggest that the acquisition of specific bacteria, particularly Enterobacterales, is strongly correlated with evolutionary transitions toward herbivory. Our findings expand on previously documented trends in ant-microbe interactions while also offering new insights into how symbioses drive the evolution of dietary strategies in tropical ecosystems.IMPORTANCEHost-microbe interactions have played an integral role in the evolution of specialized lifestyles in insects. Ants, with their ecological diversity and broad microbial associations, offer a powerful model for studying these dynamics. However, most research has focused on Neotropical ant lineages, limiting our broader understanding of how microbes influence ant evolution. Our study addresses this gap by examining Indo-Pacific ants-an underexplored but ecologically rich group-and reveals that diet, rather than nesting habitat, is the primary driver of microbial diversity. Notably, our findings challenge established patterns: Rhizobiales are more frequently associated with predatory ants than herbivores, contrasting with trends in Neotropical taxa. Furthermore, phylogenetic analyses suggest Enterobacterales may have played a key role in the evolution of herbivory. These results underscore the value of expanding research beyond taxa in well-studied regions and show how microbial partnerships can both reinforce and reshape our understanding of lifestyle evolution in ants.