Phenotypic and 16 S rRNA-based characterization of cultured oral Bacillus isolates from adults with periodontitis

Culture-based information on oral Bacillus recovered from adults with periodontitis remains limited, particularly for endospore-forming isolates related to the Bacillus cereus sensu lato complex. We characterized 11 Bacillus isolates (PCP01-PCP11) recovered from purposively selected oral swab specimens from adults with periodontitis. The isolates were evaluated for morphology, motility, catalase and oxidase activities, hemolysis, thermotolerance, nitrate reduction, lecithinase activity, and 16 S rRNA gene sequence similarities. All isolates were gram-positive, endospore-forming rods; 10/11 were motile, 11/11 were catalase-positive, 8/11 exhibited β-hemolysis, 8/11 grew at 50 °C, 9/11 reduced nitrate, and 6/11 were lecithinase-positive. All isolates grew under both aerobic and anaerobic conditions, supporting a facultatively anaerobic growth phenotype. Primers 27 F/1492R generated approximately 1.5-kb amplicons, from which an approximately 909-bp common high-quality region was retained after bidirectional sequencing and trimming. BLASTn and neighbor-joining analyses supported affiliation with Bacillus, with several isolates showing the closest sequence affinity to the reference taxa within the B. cereus sensu lato. Because partial 16 S rRNA sequences provide insufficient resolution within the B. cereus sensu lato complex, reference matches were interpreted only as genus- or group-level affinities for the species. Sex-stratified phylogenetic patterns were considered descriptive because female- and male-derived isolates were analyzed against different reference sets, precluding population-level inferences from the analysis. Overall, this study provides preliminary phenotypic and phylogenetic characterization of cultured oral Bacillus and highlights the need for systematic sampling and genome-resolved taxonomic classification of oral Bacillus species.