Epigenetic phase variation in the gut microbiome enhances bacterial adaptation

Abstract The human gut microbiome within the gastrointestinal tract continuously adapts to variations in diet, medications, and host physiology. A strategy for bacterial genetic adaptation is epigenetic phase variation (ePV) mediated by bacterial DNA methylation, which can regulate gene expression, enhance clonal heterogeneity, and enable a single bacterial strain to exhibit variable phenotypic states. Genome-wide and site-specific ePVs have been characterized in human pathogens’ antigenic variation and virulence factor production. However, the role of ePV in facilitating adaptation within the human microbiome remains poorly understood. Here, we comprehensively cataloged genome-wide and site-specific ePV in human infant and adult gut microbiomes. First, using long-read metagenomic sequencing, we detected genome-wide ePV mediated by complex structural variations of DNA methyltransferases, highlighting those associated with antibiotics or fecal microbiota transplantation. Second, we analyzed a collection of public short-read metagenomic sequencing datasets, uncovering a great prevalence of genome-wide ePV in the human gut microbiome. Third, we quantitatively detected site-specific ePVs using single-molecule methylation analysis to identify dynamic variation associated with antibiotic treatment or probiotic engraftment. Finally, we performed an in-depth assessment of an Akkermansia muciniphila isolate from an infant, highlighting that ePVs can regulate gene expression and enhance the bacterial adaptive capacity by employing a bet-hedging strategy to increase tolerance to differing antibiotics. Our findings indicate that epigenetic modifications are a common strategy used by gut bacteria to adapt to the fluctuating environment. Competing Interest Statement R.H.V. is a co-founder of Bloom Science (San Diego, CA) Footnotes 1. We have added additional Supplementary Data to confirm that the ePV events originate from the same strain. 2. We have incorporated new analyses of genetic variations across ciprofloxacin-treated passages of Akkermansia muciniphila, providing evidence for an alternative pathway by which the strain may tolerate ciprofloxacin. 3. We have made minor revisions to several figures to improve readability. Data availability All sequencing data generated in this study have been submitted to NCBI SRA with the accession number PRJNA1207694, and will be made publicly available upon publication.