IntroductionThe gut microbiota is a critical modulator of host energy metabolism and immune regulation. Although exercise modulates microbial composition, the temporal dynamics and strain-specific impact of acute high-intensity exercise remains unclear.MethodsThis study investigated dynamic alterations in gut microbiota following acute high-intensity exercise in BALB/c and C57BL/6 mice, focusing on energy and gut health-related genera. Age-matched male mice underwent a 30-min high-intensity treadmill run. To capture temporal dynamics, colonic content samples were collected at 0, 30, and 60 min post-exercise for 16S rRNA gene sequencing.ResultsExercise induced significant remodeling of gut microbiota composition in both strains, with a notable post-exercise elevation in the Bacteroidetes/Firmicutes ratio, particularly in C57BL/6 mice (C57Cr vs. C57T0, p < 0.01; C57Cr vs. C57T60, p < 0.05; BCCr vs. BCT30, p < 0.05). Both alpha and beta diversity metrics revealed significant exercise-induced microbial changes, with C57BL/6 mice showing a more pronounced increase in diversity at 30 and 60 min post-exercise (OTUs and observed species, all p < 0.001) compared to BALB/c mice. Functional analysis revealed strain-specific responses: BALB/c mice exhibited upregulated enzymes involved in energy metabolism but impaired immune regulation, indicating compromised intestinal barrier integrity and delayed energy recovery. In contrast, C57BL/6 mice displayed enhanced anti-inflammatory capacity, accelerated energy recovery through enrichment of energy metabolism-related genera, and maintained gut integrity through the proliferation of beneficial bacteria. The potential important genus Muribaculum identified as a key exercise-responsive taxon across time points, potentially associated with gut motility. The role of Muribaculum warrants further using multi-omics investigation.DiscussionThese findings provide a foundation for identifying genetic loci influencing exercise microbiota interactions, thereby providing a theoretical basis for microbiota-based personalized exercise interventions.