Structured Exercise Modulates Gut Microbiota Composition and Protects Against Diet-Induced Dysbiosis in a Rat Model

<b>Background/Objectives:</b> Dietary composition and physical activity are major determinants of gut microbiome structure, and dysbiosis is strongly associated with metabolic disorders. While both diet and exercise independently influence the gut microbiome, their interactive effects-particularly across different exercise modalities-remain incompletely understood. This study investigated the combined effects of diet type (normal chow [NC] vs. high-fat diet [HFD]) and exercise modality (control [C], voluntary [V], and forced [F]) on gut microbiota composition in rats. <b>Methods:</b> Sixty-three Wistar rats were randomized into six groups according to diet and exercise status. Fecal samples were collected and analyzed using full-length 16S rRNA gene sequencing (Oxford Nanopore Technologies). Alpha and beta diversity metrics were calculated, and taxonomic composition was assessed at phylum and genus levels. <b>Results:</b> HFD groups exhibited significantly higher alpha diversity than NC groups (Shannon index: 3.47-3.63 vs. 2.76-2.94, <i>p</i> < 0.001), with forced exercise associated with a greater diversity than voluntary exercise. Beta-diversity analysis confirmed diet as the dominant factor influencing microbial structure (PERMANOVA <i>p</i> = 0.001), with exercise providing an additional modulatory effect. Firmicutes, Bacteroidota, Deferribacterota, and Proteobacteria predominated, with Firmicutes decreasing under HFD. Forced exercise significantly enriched beneficial genera, including <i>Akkermansia</i> (detected exclusively in exercised HFD groups; <i>p</i> = 0.03), <i>Blautia</i>, <i>Coprococcus</i>, and <i>Roseburia</i>. <i>Akkermansia</i> abundance correlated positively with exercise distance (<i>p</i> < 0.001) and negatively with body weight (<i>p</i> < 0.01). <b>Conclusions:</b> Structured exercise, particularly forced treadmill training, attenuates HFD-associated dysbiosis and promotes the beneficial gut bacteria that is associated with metabolic health. These findings highlight exercise modality as a critical factor in dietary strategies targeting gut microbiome modulations.