Bacillus spp. Isolated from Miang as Potential Probiotics in Nile Tilapia Culture—In Vitro Research

Among 79 <i>Bacillus</i> spp. isolated from Miang, a fermented tea in north Thailand, 17 <i>Bacillus</i> strains were selected with probiotic potential in Nile tilapia culture based on the capabilities of bacteriocin production and associated antimicrobial activities against fish pathogens, <i>Aeromonas hydrophila</i> and <i>Streptococcus agalactiae</i>. However, only six isolates were selected for further extensive studies based on the strength of their antimicrobial activities and their tolerance against simulated gastrointestinal conditions. The molecular identification by 16S rRNA gene sequence analysis revealed that five isolates, K2.1, K6.1, K7.1, K15.4, and K22.6, were <i>Bacillus tequilensis,</i> and the isolate K29.2 was <i>Bacillus siamensis</i>. <i>B. siamensis</i> K29.2 showed complete susceptibility to antibiotics tested in this study, while <i>B. tequilensis</i> K 15.4 showed moderate resistance to some antibiotics; therefore, both strains were selected as potential probiotic bacteria. <i>B. tequilensis</i> K15.4 and <i>B. siamensis</i> K29.2 were capable of the production and secretion of extracellular protease and polysaccharide degrading enzymes, including cellulase, xylanase, and β-mannanase. The tannin tolerant test also demonstrated their ability to grow on selective agar plates and secrete cellulase and β-mannanase in the presence of hydrolyzable tannin. In addition, in vitro digestion of commercial fish substrate revealed that the extracellular enzymes produced by both strains efficiently reacted with feed protein and polysaccharides. Based on the results from this study, <i>B. siamensis</i> K29.2 was deemed to have the highest potential multifunctional probiotic qualities for application in Nile tilapia culture, while the antibiotic-resistant gene in <i>B. tequilensis</i> K15.4 must be clarified before field application.