Bile Salt Tolerance Determines Intestinal Colonization Efficacy of Heyndrickxia coagulans: A Phenotypic and Genomic Study

The probiotic efficacy of H. coagulans relies on the bile salt tolerance of its vegetative cells, yet direct evidence linking this trait to intestinal colonization remains limited. This study integrated phenotypic screening, in vitro gastrointestinal simulation, in vivo colonization assays, and comparative genomics to address this gap. Among 50 strains, two highly bile salt-tolerant isolates (ATCC 7050 and Idrc019) were identified. In vitro assays using a simulated gastrointestinal model demonstrated that the spores of tolerant strains exhibited a significantly higher germination rate in the intestinal phase. Subsequently, in vivo time-course experiments demonstrated that tolerant strains exhibited superior intestinal proliferation and modulated the gut microbiota by enriching beneficial genera such as Blautia. Comparative genomic analysis revealed five variable genes associated with bile salt tolerance. Notably, BF29_941 (encoding a pilus assembly protein) was significantly upregulated under bile salt stress, suggesting a potential role in cell aggregation as a tolerance mechanism. These findings establish bile salt tolerance as a critical determinant of intestinal colonization in H. coagulans.