Tracking gene flow in the gut microbiome reveals phage activation and host-range expansion in response to diet

Abstract Horizontal gene transfer (HGT) drives microbial evolution, but what determines when genes move between species within complex communities remains unclear. We developed a junction-resolved metagenomic framework using short-read sequencing to distinguish mobile genetic element activation from de novo transfer and track both longitudinally. In a defined human gut community colonizing mice, mobile element activation was widespread but successful transfer was rare. A prophage carried by invading Phocaeicola vulgatus transferred reproducibly to resident species when the community resisted invasion, but not when dietary perturbation allowed the invader to expand. Transfer required prophage activation and persistence of a permissive recipient and coincided with diversification of a phage receptor-binding protein. Dietary polysaccharides reproduced prophage-associated fitness costs in vitro , and rhamnogalacturonan I induced cell lysis consistent with productive prophage activation. The framework also detected interspecies transfer retrospectively in longitudinal human metagenomes. Thus, diet and community ecology can redirect gene flow through the gut microbiome.