Switching from a low-fat and high-fiber diet to a Western-style high-fat and high-sugar diet causes microbiota imbalances that underlay many pathological conditions (i.e., dysbiosis). Although the effects of dietary changes on microbiota composition and functions are well documented, their impact in gut bacterial evolution remains unexplored. We followed the emergence of mutations in Bacteroides thetaiotaomicron, a prevalent fiber-degrading microbiota member, upon colonization of the murine gut under different dietary regimens. B. thetaiotaomicron evolved rapidly in the gut and Western-style diet selected for mutations that promote degradation of mucin-derived glycans. Periodic dietary changes caused fluctuations in the frequency of such mutations and were associated with metabolic shifts, resulting in the maintenance of higher intraspecies genetic diversity compared to constant dietary regimens. These results show that dietary changes leave a genetic signature in microbiome members and suggest that B. thetaiotaomicron genetic diversity could be a biomarker for dietary differences among individuals.