Isolation and characterization of Bacillus spp. M001 for potential application in turbot (Scophthalmus maximus L.) against Vibrio anguillarum

The aim of this study was to screen Bacillus strains from the guts of Scophthalmus maximus, Paralichthys olivacues, Epinephelus coioides and Clupanodon punctatus, for use as probiotics in aquaculture. Eight Bacillus strains were screened, and strain M001 was selected for probiotic study based on its antagonistic activity against multiple aquatic bacterial pathogens including Vibrio anguillarum, V.campbellii, V.vulnificus, V.parahamolyticus, Streptococcus sp. and Edwardsiella tarda. M001 was identified as B.amyloliquefaciens based on the biochemical tests and 16S rRNA gene analysis. In vitro experiments revealed that M001 was able to grow at a wider range of temperature, pH and salinity and was capable to use turbot mucus as nutrient for growth. Additionally, M001 isolate greatly inhibited the growth of V.anguillarum by producing antibacterial substances and was acid tolerance, non-antibiotic resistance and non-harmful. Thereafter, the potential probiotic effect of M001 was tested in turbot by dietary administration of M001 at a dose of 10(8)CFUg(-1) diet for 42days. No significant differences of weight gain, specific growth rate and feed ratio were found in the M001-diet group of fish compared with control fish, but which increased, respectively, by 5.5%, 4.7% and 7.0% after 42days of feeding. Several digestive enzyme activities were found to increase significantly in the M001-diet group, including protease and amylase activities in hepatopancreas, protease activity in intestine and lipase activity in stomach (P<0.05). Sera superoxide dismutase activity and total protein content (P<0.05) were also increased significantly (P<0.05) in the M001-diet group. The challenge experiment showed that the M001-diet group of fish showed a relative per cent of survival of 62.7% against V.anguillarim infection. The Bacillus M001 identified from this study has good potential to provide vibriosis control as probiotic feed additive for turbot aquaculture.