Comparative genomic and phenotypic analysis of human- and dairy cattle -derived Streptococcus agalactiae reveals potential cross-species infection capability

Streptococcus agalactiae is an opportunistic Gram-positive pathogen that can infect humans, dairy cattle, and other animals, posing a potential zoonotic risk. In this study, whole-genome sequencing was performed on 40 GBS strains isolated from dairy cows. The results showed high homology among strains collected from the same farm. Based on these findings, two representative dairy cattle-derived strains and one human-derived strain were selected for comparative whole-genome analysis to identify differences in antibiotic resistance and virulence genes. Additionally, we characterized the antibiotic resistance phenotypes, hemolytic phenotypes, invasion capacity, and the levels of oxidative stress markers and inflammatory factors in bovine mammary epithelial cells (BMECs) during infection for all three strains. The three strains HB27, HB31, and RW exhibited close phylogenetic relationships. All showed strong resistance to sulfonamide and aminoglycoside antibiotics but remained susceptible to cephalosporins, quinolones, and macrolides. The mefA gene was present in RW but absent in HB27 and HB31. RW exhibited higher hemolytic activity and cylE mRNA expression levels than HB27 and HB31, and the pilA and pilB genes were also present only in RW. HB31 demonstrated significantly faster invasion rates than HB27 and RW before the 2‑h invasion time point. The effects of the three strains on antioxidant markers and the protein and mRNA expression of inflammatory factors in BMECs were consistent, indicating that the human-derived S. agalactiae possesses the capacity to infect BMECs. Although human- and dairy cattle-derived GBS differ in virulence-associated genes and hemolytic activity, the human strain invades BMECs in vitro and induces host responses similar to those of dairy cattle strains. These findings suggest that human-derived GBS may have the potential for cross-species infection. To date, most studies on human- and dairy cattle-derived GBS have focused primarily on epidemiology. Even when addressing whether GBS is a zoonotic pathogen, direct evidence of cross-species transmission mechanisms remains limited. This study provides in vitro evidence that human-derived GBS can directly infect BMECs. However, whether cross-species transmission between human- and dairy cattle-derived GBS requires repeated exposure or can occur following a single exposure requires further investigation.