| Summary: | This study aimed to isolate, identify, and evaluate the probiotic properties of <i>Bacillus</i> species from honey of the stingless bee <i>Heterotrigona itama</i>. <i>Bacillus</i> spp. were isolated from five different <i>H. itama</i> meliponicultures, and the isolates were characterized through Gram-staining and a catalase test. Tolerance to acidic conditions and bile salt (0.3%), hydrophobicity, and autoaggregation tests were performed to assess the probiotic properties of the selected isolates, <i>B. amyloliquefaciens</i> HTI-19 and <i>B. subtilis</i> HTI-23. Both <i>Bacillus</i> isolates exhibited excellent antimicrobial activity against both Gram-positive and Gram-negative bacteria and possessed significantly high survival rates in 0.3% bile solution for 3 h. Their survival rates in acidic conditions were also comparable to a commercial probiotic strain, <i>Lactobacillus rhamnosus</i> GG. Interestingly, the hydrophobicity and autoaggregation percentage showed no significant difference from <i>L. rhamnosus</i> GG, a commercial probiotic strain. The results from this study suggest that <i>B. amyloliquefaciens</i> HTI-19 and <i>B. subtilis</i> HTI-23 isolated from stingless bee honey have considerably good probiotic properties. Therefore, more studies should be done to investigate the effects of these bacteria cultures on gastrointestinal health.
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