Screening of antagonistic bacteria against visceral white-spots disease of Larimichthys crocea and preliminary study on its biological characteristics

In order to achieve the biological prevention and control of visceral white-spots disease of Larimichthys crocea, and promote the reduction of drug use in aquaculture, we isolated and screened the probiotics with an antagonistic effect on Pseudomonas plecoglossicid, which is a pathogen of visceral white spot disease in L. crocea, from the intestine of healthy L. crocea. The strains were screened by agar diffusion method, identified by physiological and biochemical characteristics and molecular biology analysis, and evaluated for hemolysis, drug sensitivity, safety, enzyme production ability and broad-spectrum antibacterial activity. Thirty-seven strains of potential probiotics were isolated and the three most potent strains were further characterized for their probiotic potential, named as P1-17, P2-33 and P3-11. The three most promising isolates were identified by sequencing the 16S rRNA gene and physiological and biochemical characteristics. Strains P1-17 and P2-33 were identified as Bacillus velezensis and P3-11 was identified as Enterococcus faecalis. According to the hemolytic test and disk diffusion method, none of the three strains had sigificant hemolytic rings, containing few drug resistance factors, so they had no potential pathogenicity. The result from the artificial infection safety test confirmed that these three strains of antagonistic bacteria had no pathogenicity to healthy L. crocea. The results of the antimicrobial spectrum show that two strains of Bacillus had an antagonistic effect on common aquatic pathogens such as Vibrio alginolyticus, V. harveyi and Photobacterium damselae. Moreover, two strains of Bacillus could produce amylase and protease, while the strain of E. faecalis only had an antagonistic effect on P. plecoglossicida. The study provides a scientific basis for the subsequent screening and application of intestinal probiotics in L. crocea.