Exploring synergistic effect of bacteriophages with probiotics against multidrug resistant Salmonella Typhimurium in a simulated chicken gastrointestinal system using metagenomic- and culturomic approaches.

Salmonella is a critical foodborne pathogen that significantly affects global food safety and the poultry industry. The reliance on antibiotics to control Salmonella infections has led to the emergence of antibiotic-resistant bacteria, necessitating the development of alternative strategies. The present study explored the synergistic potential of combining bacteriophages and probiotics as a dual approach for Salmonella suppression and gut microbiota modulation. Using a multi-omics approach integrating culture and metagenomics, we isolated Lactobacillus reuteri J2M1, a robust probiotic strain with strong acid and bile tolerance, intestinal adhesion, and a safety profile. Concurrently, two bacteriophages, SLAM_phiST45 and SLAM_phiST56, targeting Salmonella Typhimurium were selected based on their broad host range, enhanced inhibitory effects as a cocktail, and genomic safety validated through whole-genome sequencing. In a model that simulated the gut environment of poultry, the combined application of bacteriophages and L. reuteri J2M1 resulted in significant changes in the microbial community at the genus level. Although the application of bacteriophages alone effectively reduced Salmonella populations, it also led to the proliferation of genera that contain potential pathogens, such as Clostridium perfringens. In contrast, the co-application of L. reuteri J2M1 mitigated these negative effects by promoting the growth of beneficial genera, including Oscillibacter and Clostridium butyricum, which are associated with anti-inflammatory properties and gut health. These findings demonstrate that combining bacteriophages with probiotics suppresses pathogens as well as contributes to the establishment of a balanced gut microbial community. This study highlights the utility of a multi-omics approach for probiotic discovery, the efficacy of bacteriophage cocktails in pathogen suppression, and the benefits of their synergistic application with probiotics. Although in vivo validation is required, these in vitro results provide a robust foundation for the development of sustainable and effective alternatives to antibiotics for poultry production.