Integration of microbiome and metabolome reveals the anti-fatigue mechanism of <em>Lactiplantibacillus plantarum</em>

Exercise fatigue is a phenomenon where the body loses its ability to sustain a specific exercise level or intensity. It arises from multiple factors that negatively impact the body. Although probiotics have been shown to alleviate exercise-induced fatigue, the underlying mechanisms for this effect are not yet fully understood. This study aimed to investigate the anti-fatigue effects of <em>Lactiplantibacillus plantarum</em> SHY21-2 (SHY21-2) in a mouse model and to elucidate the underlying mechanisms. The anti-fatigue effects of SHY21-2 were evaluated by measuring muscle glycogen and serum fatigue markers, with the underlying mechanisms subsequently elucidated through an analysis of mitochondrial function, gut microbiota, and metabolites. The results showed that SHY21-2 significantly enhanced running and swimming endurance, reduced serum fatigue markers including blood levels of lactate (LA), blood urea nitrogen (BUN), and creatine kinase (CK), and increased muscle glycogen levels. It also upregulated mitochondrial biogenesis (peroxisome proliferator-activated receptor-γ co-activator-1α), fusion (mitofusin 2), fission (fission 1), and mitophagy proteins (PTEN induced putative kinase 1, Parkin). SHY21-2 reshaped the gut microbiota, enriching <em>norank_f_Muribaculaceae </em>and <em>Alloprevotella</em> while reducing fatigue-associated genera. Metabolomic analysis revealed 10 upregulated metabolites, including hesperetin and tectorigenin, which were positively correlated with beneficial microbiota. These findings suggest that the anti-fatigue effects of SHY21-2 are associated with enhanced mitochondrial function, as well as with changes in the gut microbiota and host metabolites. This supports its potential as a functional probiotic for endurance improvement.