Longitudinal dynamics of ethanol-resistant microbes and sporulation in the human gut

Abstract Background A considerable proportion of human gut bacteria use some form of dormancy to survive unfavorable conditions such as oxygen exposure, antibiotic treatment, and inflammation. Dormancy strategies enable persistence, allow recolonization following disruptions, and facilitate transmission between individuals. In this study, we explored the composition, characteristics, and longitudinal dynamics of the dormant community in the gut as determined by ethanol resistance. We conducted a six-month longitudinal study on nine healthy male adults, who were sampled every two weeks. Gut microbiota composition was determined with 16S rRNA amplicon sequencing in untreated (bulk microbiota) and ethanol treated samples. A rigorous algorithm was implemented to determine the ethanol-resistant and non-resistant community within the gut microbiota. Results We detected ethanol-resistant gut bacteria in eight distinct phyla, with the highest number of OTUs observed in the phylum Bacillota , and substantial representation in Actinomycetota and Pseudomonadota . In general, ethanol-resistant taxa exhibited higher relative abundance and longer within-individual persistence compared to non-resistant taxa. Ethanol resistance indicated a higher rate of sharing between individuals, independent of phylum. Ethanol-resistant Bacillota also exhibited longer persistence within the gut over the six-months period in comparison to ethanol non-resistant Bacillota and other ethanol-resistant or ethanol non-resistant phyla. Finally, the analysis of sporulation frequency of highly prevalent ethanol-resistant Bacillota members revealed that sporulation occurred in each individual synchronously among different community members but varied significantly between individuals. Major shifts in sporulation frequency were associated with well-known disturbances such as antibiotic therapy and travel. Conclusion Our findings highlight that sporulation of Bacillota members and alternative dormancy strategies employed by non- Bacillota members contribute to bacterial persistence in the human gut and facilitate transmission between unrelated individuals. Moreover, we show that sporulation frequency (specific to Bacillota ) is a dynamic, community-level process shaped by host- and time-specific environmental factors rather than intrinsic, species-specific traits.