Antimicrobial effects and metabolomic analysis of metabolites of Lacticaseibacillus paracasei SHY-X4 isolated from Koumiss in Inner Mongolia and its potential application in milk preservation

Milk and its derivatives are important foodstuffs and are recognized as high-risk categories prone to microbial contamination. Here, we investigated the antibacterial activity and antimicrobial properties of metabolites of Lacticaseibacillus paracasei SHY-X4 isolated from Koumiss in Inner Mongolia. The results showed that the SHY-X4 metabolites had good tolerance to acid (pH = 3), alkali (pH = 9) and heat (105 ℃) treatment and exhibited broad-spectrum antimicrobial activity against a wide range of Gram-positive and Gram-negative pathogens. Further analysis of key antimicrobial components by non-targeted metabolomics revealed 52 potential antimicrobial metabolites (variable importance in prediction (VIP) >1, adjusted P < 0.05, and absolute fold change (FC) >1.3) in SHY-X4, mainly comprising organic acids and peptides. Notably, both physiological and phenotypic perspectives revealed that the inhibitory effect of SHY-X4 metabolites on the growth and biofilm formation of Staphylococcus aureus ATCC25923 and Escherichia coli O157:H7 NCTC12900 positively contributed to the impact on cell viability and metabolic activity, disrupting the integrity of cell membranes and leading to cellular deformation. When applied to the milk models, SHY-X4 metabolites significantly inhibited the growth of pathogens and spoilage bacteria, such as S. aureus, E. coli O157:H7, and Enterobacteriaceae. They also maintained the pH, nutritional composition, and antioxidant activity of pasteurized milk for at least 15 d during refrigerated storage at 4 ℃. Overall, these findings provide insights into the potential application of SHY-X4 metabolites in the dairy industry for combating microbial contamination.