Abstract:Constipation is a widespread gastrointestinal disorder that significantly impacts individuals’ health and quality of life. Although various treatment options are available, many patients experience unsatisfactory results, creating a demand for alternative therapeutic strategies. This study explores the efficacy of Bifid Triple Viable Capsules, containing Bifidobacterium longum, Lactobacillus acidophilus, and Enterococcus faecium, in alleviating loperamide-induced constipation in a rat model. Rats were administered high or low doses of Bifid after induction of constipation with loperamide. Our findings demonstrate that high-dose Bifid treatment significantly improves body weight and intestinal propulsion rate in constipated rats. Histopathological analysis reveals that Bifid restores colon tissue integrity, reducing inflammation and maintaining the intestinal epithelial barrier. Using 16S rRNA sequencing, we identified an increase in the gut microbial diversity and composition, with higher levels of beneficial norank_f_norank_o_Coriobacteriales and Anaerofustis bacteria. Transcriptomic analysis of colon tissues showed that high-dose Bifid treatment modulates gene expression involved in immune system regulation and epithelial barrier integrity. Differentially expressed genes (DEGs) were enriched in pathways related to the immune response and integral component of membrane, particularly those associated with the intestinal immune network and bile acid metabolism. These results suggest that Bifid alleviates constipation by balancing the gut microbiota, regulating the gut innate immune response, and maintaining the intestinal epithelial barrier. Our study provides a foundational basis for further research and therapeutic applications of probiotics in treating gastrointestinal diseases.