Biological Properties of <i>S. warneri</i> KYS-164 Isolated from Kefir Grains

<i>Staphylococcus worderi</i> KYS-164, isolated from homemade Tibetan kefir grains, produces bacteriocin-like inhibitory substances (BLIS), which are peptides with antimicrobial properties, but have not been fully characterized. The research on BLIS will lay the foundation for mining new bacteriocins. In this study, the optimal culture conditions for the production of highly active BLIS were found to be incubation at 30 °C and 120 rpm, and the most effective extraction method was ammonium sulfate precipitation (ASP) using ammonium sulfate at 80% saturation. The postantibiotic effect (PAE) of BLIS on <i>Staphylococcus aureus</i> CICC 10384 is significant, with a 4 × MIC BLIS concentration able to prolong the PAE to 2.39 h. BLIS has excellent biosafety, with no deleterious effects observed at 8 × MIC concentration. Gas chromatography-ion mobility spectrometry (GC-IMS) was used to analyze the volatile compounds synthesized by <i>Staphylococcus warneri</i> KYS-164 during its growth. Hydroxycitronellal, ethyl pyruvate, and α-pinene were found to be unique substances produced by this strain, which can provide fresh, refreshing floral and fruity aromas as well as strong pine and resinous aromas in the process of kefir grain fermentation of milk. Analysis of the <i>S. warneri</i> KYS-164 genome provided insights into the major metabolic pathways in which genes expressed in this strain are involved. This study represents the first isolation of <i>S. warneri</i> KYS-164 from kefir grains prepared by Tibetan families, and provides a comprehensive analysis of its physicochemical properties. This research provides a solid foundation for better understanding and utilization of <i>S. warneri</i> KYS-164.