Combination of bacteriophage–probiotics alleviates intestinal barrier dysfunction by regulating gut microbiome in a chick model of multidrug-resistant Salmonella infection

Background: The rapid emergence of multidrug-resistant Salmonella in poultry demands alternative control strategies beyond conventional antibiotics. In this study, we evaluated a combination of lytic Salmonella-infecting bacteriophages (SLAM_phiST45 and SLAM_phiST56) and a probiotic bacterium Limosilactobacillus reuteri (SLAM_LAR11) in a chick model challenged with Salmonella enterica serovar Typhimurium infection. Results: Co-administration with two-phage cocktail and a probiotic showed markedly reduced Salmonella colonization in the gut and systemic organs of chicks, comparable to the effect of phage-only treatment. In contrast with phage-only treatment, the combined therapy significantly improved the rate of body-weight change from the day of infection to necropsy (P < 0.0001) and alleviated infection-associated splenomegaly (P = 0.028) and hepatomegaly (P = 0.011). In the ileum, the villus height-to-crypt depth ratio (VH/CD) increased significantly (P = 0.044). In the colon, expression of tight-junction genes OCLN (P = 0.014), TJP1 (P < 0.0001), and MUC2 (P = 0.011) was elevated, whereas the pro-inflammatory cytokine IL6 was reduced (P = 0.018). These improvements were accompanied, in the cecum, by trends toward decreases in Escherichia–Shigella (P = 0.09) and Clostridium (P = 0.16) and a trend toward an increase in Blautia (P = 0.11); additionally, in the ileum, Lactobacillus (P = 0.037) and Blautia (P = 0.016) increased significantly, yielding a more balanced microbiota than with phage-only treatment. Consistently, levels of functional metabolites, including acetic acid (LDA = 3.32) and lactic acid (LDA = 5.29), were increased. Conclusion: Taken together, these findings demonstrate that phage–probiotic co-administration not only enhances the clearance of multidrug-resistant Salmonella more effectively than phage treatment alone but also promotes intestinal health, highlighting its potential as an antibiotic-alternatives strategy to improve intestinal health and ensure food safety in poultry production systems.