Study on the Mechanism of Action of Baicalein in Inhibiting the Invasion of Streptococcus agalactiae

Streptococcus agalactiae, also known as Group B Streptococcus (GBS), is a major pathogen responsible for mastitis in dairy cows. It causes persistent and difficult-to-treat mammary infections, leading to reduced milk production. Baicalein, a flavonoid compound, exhibits anticancer, anti-inflammatory, and antibacterial activities; however, its specific mechanism of action against GBS remains unclear. This study aimed to investigate the mechanism by which baicalein inhibits GBS invasion of bovine mammary epithelial cells (bMECs). The results showed that baicalein at concentrations of 4 μg/mL or higher effectively inhibited 50% of the invasion of bMECs by GBS strain HB31 and exerted a concentration-dependent inhibitory effect on bacterial adhesion. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of baicalein against HB31 were both greater than 1024 μg/mL. Therefore, the antibacterial effect of baicalein alone may not fully account for its mechanism; other pathways likely contribute to the reduced invasiveness of GBS. To elucidate the mechanism by which baicalein inhibits GBS invasiveness, this study investigated both bacterial metabolism and gene expression. Metabolomic analysis revealed that baicalein treatment led to the downregulation of amino acid metabolites, including alanine and aspartic acid, as well as nucleotide metabolites such as adenine and UMP in GBS HB31. Additionally, the NADH/NAD+ ratio increased while ATP levels decreased, indicating that the overall metabolic activity of GBS was suppressed. Transcriptomic analysis focused on changes in invasion-associated virulence genes. The results showed that the expression of pbsP, an invasion-associated virulence gene, was significantly reduced, while the expression of hylB and cfb showed downward trends that did not reach statistical significance. In contrast, the expression of cylE and the two-component system vicKR was upregulated. The upregulation of cylE may be related to baicalein-induced oxidative stress in HB31. Furthermore, HB31 suppressed Nrf2-HO-1 mRNA expression, whereas baicalein activated the Nrf2 signaling pathway and reduced HB31-induced IL-6 and NF-κBmRNA expression. These findings provide new insights for the development of anti-virulence therapeutic strategies targeting GBS.