Bifidobacterium longum Subspecies infantis M63 Enhances the Colitis-Alleviating Effect of 2′-Fucosyllactose by Restoring Intestinal Barrier Function and Microbial Metabolism

Both 2′-fucosyllactose (2′-FL) and Bifidobacterium can modulate gut health; however, whether their combination provides strain-dependent protection against colitis remains unclear. This study aimed to compare the protective effects of 2′-FL alone and in combination with Bifidobacterium longum subsp. infantis M63 or Bifidobacterium breve M16V in dextran sulfate sodium (DSS)-induced colitis. Forty-eight male C57BL/6J mice were randomly assigned to six groups: negative control, DSS model control, 5-aminosalicylic acid positive control, 2′-FL, 2′-FL + M63, and 2′-FL + M16V. The interventions were administered for two weeks, with 2.5% DSS supplied in the drinking water during the second week. Colitis-related indices, colonic histopathology, inflammatory cytokines, oxidative-stress markers, intestinal barrier-associated gene expression, gut microbiota composition, lactate, and short-chain fatty acids (SCFAs) were evaluated. Compared with the model control, 2′-FL and both synbiotic combinations ameliorated colonic tissue injury and goblet-cell loss. Both combinations significantly reduced IL-1β, whereas 2′-FL and both combinations reduced IL-6 and TNF-α. The combinations also increased catalase and glutathione peroxidase activities and decreased malondialdehyde levels. 2′-FL alone increased Occludin and RELMβ expression, whereas 2′-FL + M63 showed the strongest effects on tight-junction and mucus-barrier restoration, including increased Claudin-1, Claudin-2, ZO-1, TFF3, and MUC2 expression. All 2′-FL-containing interventions increased microbial diversity, while 2′-FL + M63 increased Bacteroidota, reduced Proteobacteria, and enriched Muribaculaceae and Ruminococcaceae. Both synbiotic combinations increased lactate and all measured SCFAs; 2′-FL + M63 produced the highest acetate level, whereas 2′-FL + M16V produced the highest propionate level, and both combinations markedly increased butyrate. These findings demonstrate strain-dependent protective effects, with 2′-FL + M63 showing the greatest overall efficacy in restoring intestinal barrier function and microbial metabolism, supporting its development as a candidate functional synbiotic food formulation.