Immunomodulatory effects of dietary Bacillus coagulans in grouper (Epinephelus coioides) and zebrafish (Danio rerio) infected with Vibrio vulnificus

Groupers bred for mariculture are of high economic value in Taiwan. Unfortunately, these fish are highly susceptible to bacterial infection, and the use of antibiotics to control disease has had the unintended consequence of generating antibiotic-resistant bacteria. Herein, we examined whether it is possible to enhance grouper immunity through application of Bacillus coagulans, a probiotic bacterial species. In order to study its effects on immunity, B. coagulans was isolated from lactobacillus mixtures and administered live to grouper (Epinephelus coioides) and zebrafish (Danio rerio). Fish were fed for 30 days on a diet consisting of B. coagulans (10⁴, 10⁶, 10⁸, or 10¹⁰ colony-forming units (cfu) of B. coagulans in 50 ml of media) mixed with 50 g of eel powder. After 30 days, disease resistance against Vibrio vulnificus (204) infection and expression of immune-related genes were determined. We found that oral administration of B. coagulans significantly enhanced expression of several immune-related genes, including myeloid differentiation factor (MyD)88, interleukin (IL)-1β, tumor necrosis factor (TNF)-2, and IL-1β in grouper, and Toll-like receptor (TLR) 4, TNF-α, TRAM 1, and nuclear factor (NF)-κB in zebrafish. Treatment with B. coagulans significantly reduced mortality at 24, 48, 72, and 96 h after challenge with V. vulnificus (204). In vitro studies revealed that B. coagulans possesses bactericidal activity against Enterococcus faecalis (10066), Staphylococcus aureus subsp. (10780), Streptococcus agalactiae (S47, S48, 819), Escherichia coli (10675), Klebsiella oxytoca (13985), Pseudomonas aeruginosa (19660), and V. vulnificus (204, 12905, YJ016). On average, 10⁷ colony-forming units of B. coagulans were required for bactericidal activity. In conclusion, our results indicate that dietary intake of B. coagulans enhances expression of immune-related genes and protects grouper and zebrafish against V. vulnificus (204) infection.