Clostridium butyricum RH2 ameliorates diarrhea in juvenile mice under continuous antibiotic exposure by modulating gut microbiota and metabolome.

Antibiotic-associated diarrhea (AAD) is a self-limiting disorder triggered by antibiotic therapy in pediatric populations. Although multiple probiotics are clinically employed for AAD management, the therapeutic efficacy of Clostridium butyricum (C. butyricum) in pediatric AAD and its underlying mechanisms remain poorly characterized. This study aimed to establish a juvenile mice model of AAD and investigate the therapeutic potential of oral C. butyricum administration in juvenile mice subjected to continuous antibiotics exposure. We systematically assessed pathological changes in colonic tissue, colitis severity, intestinal epithelial barrier integrity, fecal metabolomic profiles, and gut microbiota diversity. Our analysis demonstrates that C. butyricum ameliorates intestinal inflammation, enhances barrier function by modulating the gut microbiota and its metabolites, and significantly alleviates diarrhea symptoms in juvenile AAD mice. Collectively, these findings indicate that the therapeutic benefits of C. butyricum are closely linked to its ability to tolerate continuous antibiotic exposure, providing a scientific rationale for its co-administration with antibiotics.IMPORTANCEClostridium butyricum demonstrates significant therapeutic potential for pediatric antibiotic-associated diarrhea (AAD) by dual modulation of gut microbiota and host physiology. This study reveals its capacity to alleviate intestinal inflammation, restore barrier integrity via upregulation of tight junction proteins and mucins, and rebalance gut microbiota linked to key anti-inflammatory metabolites-even under ongoing antibiotic exposure. These findings position C. butyricum as a targeted probiotic therapy for AAD, offering mechanistic insights to advance microbiome-driven interventions for antibiotic-induced diarrhea in children.