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Aeromonas veronii  is an emerging aquatic pathogen causing hemorrhagic septicemia in humans and animals. Probiotic is an effective strategy for controlling enteric infections through reducing intestinal colonization by pathogens. Here we report that the consumption of  Bacillus velezensis  regulated the intestinal innate immune response and decreased the degree of intestinal inflammation damage caused by the  A. veronii  in Crucian carp. In this study, we isolated four strains of  B. velezensis , named C-11, S-22, L-17 and S-14 from apparently healthy Crucian carp, which exerted a broad-spectrum antimicrobial activity inhibiting both Gram-positive and Gram-negative bacteria especially the fish pathogens.  B. velezensis  isolates showed typical  Bacillus  characteristics by endospore staining, physiological and biochemical test, enzyme activity analysis (amylase, protease, and lipase), and molecular identification. Here,  Bacillus -containing dietary was orally administrated to Crucian carp for 8 weeks before  A. veronii  challenge. Immunological parameters and the expression of immune-related genes were measured at 2, 4, 6, 8, and 10 weeks post-administration. The results showed that  B. velezensis  was found to promote the increase in the phagocytic activities of peripheral blood leukocytes (PBLs) and head kidney leukocytes (HKLs), as well as the increase in interleukin 1β (IL-1β), IL-10 and tumor necrosis factor α (TNF-α) concentration of serum. Lysozyme levels (113.76 U/mL), ACP activity (25.32 U/mL), AKP activity (130.08 U/mL), and SOD activity (240.63 U/mL) were maximum ( P  < 0.05) in the  B. velezensis  C-11 treated group at 8 week. Our results showed that Crucian carp fed with the diet containing  B. velezensis  C-11 and S-22 developed a strong immune response with significantly higher ( P  < 0.05) levels of IgM in samples of serum, mucus of skin and intestine compared to  B. velezensis  L-17 and S-14 groups. Moreover,  B. velezensis  spores appeared to show no toxicity and damage in fish, which could inhabit the gut of Crucian carp.  B. velezensis  restrained up-regulation of pro-inflammation cytokines (IL-1β, IFN-γ, and TNF-α) mRNA levels in the intestine and head kidney at final stage of administration, and the expression of IL-10 was increased throughout the 10-week trial.  A. veronii  infection increased the population of inflammatory cells in the intestinal villi in the controls. In contrast, numerous goblet cells and few inflammatory cells infiltrated the mucosa in the  B. velezensis  groups after challenge with  A. veronii . Compared with  A. veronii  group,  B. velezensis  could safeguard the integrity of intestinal villi. The highest post-challenge survival rate (75.0%) was recorded in  B. velezensis  C-11 group. The present data suggest that probiotic  B. velezensis  act as a potential gut-targeted therapy regimens to protecting fish from pathogenic bacteria infection.   IMPORTANCE  In this work, four  Bacillus velezensis  strains isolated from apparently healthy Crucian carp, which exhibited a broad-spectrum antibacterial activity especially the fish pathogens. Administration of  B. velezensis  induced the enhancement of the intestinal innate immune response through reducing intestinal colonization by pathogens. The isolation and characterization would help better understand probiotic can be recognized as an alternative of antimicrobial drugs protecting human and animal health.

frontiers in microbiology

Effect of Bacillus velezensis on Aeromonas veronii-Induced Intestinal Mucosal Barrier Function Damage and Inflammation in Crucian Carp (Carassius auratus)