Animal gut microbiota affect host physiology and behaviour. In eusocial Hymenoptera, where colony-level integrity is preserved via a nestmate discrimination system based on cuticular hydrocarbon mixtures, microorganismal effects may influence social dynamics. Although nestmate recognition has undergone a thorough exploration during the last four decades, few studies have investigated the putative role of gut microbes. Here we integrate metagenomic, chemical and behavioural approaches to test

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... ther gut microbes affect nestmate recognition in Acromyrmex echinatior leaf-cutting ants. Treating workers with a sterile diet or with antibiotics resulted in a substantial alteration of their gut microbial communities. In pairwise social interactions, untreated vs. antibiotic-treated nestmates behaved more aggressively than other nestmate and non-nestmate pairs, suggesting that the suppression of microbes indirectly alters chemical social cues and triggers aggressive behaviour. Chemical analyses on treated individuals revealed a decrease in the abundance of two metapleural gland antifungal compounds, and we confirmed the correspondence between aggression levels and chemical profile differences. Feeding microbiota-remodelled ants with conspecific faecal droplets partially restored the original bacterial communities. Furthermore, non-nestmates fed with faecal droplets from different colonies were unusually aggressive compared to pairs fed with faecal droplets from the same colony. This suggests that chemicals derived from microbial strains may shape nestmate recognition, opening novel questions about the role of microorganisms in the evolution of social behaviour.