Comparative genomic and phenotypic profiling of commercial probiotic Enterococcus faecium strains: insights into safety and functional attributes.

Abstract

Enterococcus faecium is widely used in probiotics but can also act as an opportunistic pathogen, necessitating strain-specific safety and efficacy assessments. While previous studies have characterized individual E. faecium probiotic strains, few have provided an integrated genomic and phenotypic comparison of multiple commercial strains from the Asian market. This study conducted a comparative genomic and phenotypic analysis of five E. faecium strains isolated from commercial probiotic products alongside five reference strains. Whole-genome sequencing revealed that the probiotic-derived strains, which were identified as Enterococcus lactis, clustered into two distinct phylogenetic clades (Clade I: ST-812; Clade II: ST-76), corroborated by Average Nucleotide Identity and Multilocus Sequence Typing analyses. These clades exhibited coherent functional genomic profiles, with Clade I enriched in genes for carbohydrate metabolism and dietary fiber degradation, while Clade II showed a different emphasis, including carotenoid biosynthesis. Crucially, all probiotic strains lacked high-risk antibiotic resistance genes (vanA) and major virulence determinants (esp, hyl, gelE) found in the clinical control strain ATCC 51559, and were phenotypically susceptible to the tested β-lactams (penicillin, ampicillin), vancomycin, and tetracycline, and lacked acquired resistance genes for WHO-defined critically important antibiotics. In vitro probiotic property assays further showed significant inter-strain variability in gastric acid tolerance, bile salt resistance, and bile salt hydrolase (BSH) activity, underscoring the strain-specific nature of functional performance. Our integrated analysis indicates a strong link between genomic identity, functional potential, and safety profile in commercial E. faecium probiotics. This study provides a detailed genomic framework for strain-level safety assessment and supports the informed selection of E. faecium strains for probiotic applications.