High Probiotic Potential of Soil-Derived Bacillus safensis, Bacillus pumilus and Bacillus zhangzhouensis Strains: In...

The aim of this study was to isolate, molecularly identify, and comprehensively evaluate the probiotic potential of native Bacillus strains from the soil of Isfahan province, Iran. Using the thermal enrichment method, 23 Gram-positive spore-forming strains were isolated. Following preliminary safety screening, three superior strains (NF16, NF20, and NF24) were selected based on the absence of hemolytic activity (γ-hemolysis), lack of lecithinase production, and absence of enterotoxin genes (hbl and nhe). The strains were identified as Bacillus zhangzhouensis NF16, Bacillus pumilus NF20, and Bacillus safensis NF24 through 16 S rRNA genes sequence. Their 16 S rRNA gene sequences were deposited in the NCBI GenBank database under the accession numbers PX930946, PX930921, and PX930970, respectively. All three strains demonstrated excellent tolerance to simulated gastrointestinal conditions, the survival rates of the three tested strains ranged from 48.59% to 81.45% at pH 2 and from 81.09% to 88.60% in artificial gastric juice (pH 2.5), and high resistance to 0.3% bile salts (inhibition coefficient 0.30-0.38). They also exhibited strong adhesion to HT-29 cells (71.28-82.74%), high auto-aggregation (> 72% after 24 h), favorable cell surface hydrophobicity, and potent antioxidant activity (47.51-78.73% DPPH radical scavenging). Furthermore, the strains showed desirable antibiotic susceptibility profile and lacked enterotoxin genes, confirming their safety. In comparison with the commercial control strain Bacillus subtilis PXN^® 21^®, the native isolates were equal or superior in several key probiotic properties. These findings indicate that the soil-isolated B. safensis NF24, B. pumilus NF20, and B. zhangzhouensis NF16 are promising candidates for the development of robust and effective indigenous probiotics.