H ₂ O ₂ -producing commensal streptococci disrupt Streptococcus mutans–Candida albicans synergism

Microbial homeostasis is maintained by the antagonistic capacity of commensal bacteria against cariogenic pathogens. In the oral cavity, commensal Streptococcus , dominant colonizers of the tooth surface, can produce hydrogen peroxide (H 2 O 2 ), modulating virulent cross-kingdom biofilm formation. To investigate their ecological role, clinical isolates from dental plaque were compared with reference strains, including Streptococcus oralis ATCC 35037 and S. oralis subsp. tigurinus J22 to determine their H 2 O 2 -producing capabilities. The antagonistic potential of S. oralis against Streptococcus mutans and Candida albicans was evaluated using microbial and biochemical assessments. In a saliva-coated hydroxyapatite disc model, S. oralis strains were co-cultured with S. mutans and C. albicans . A high H 2 O 2 -producing S. oralis J22 inhibited EPS formation in S. mutans and yeast-to-hypha transition in C. albicans , thereby reducing EPS-mediated bacterial–fungal cell colocalization. Time-lapse confocal imaging revealed that S. oralis J22 dominated the biofilm through H 2 O 2 -mediated antagonistic interactions. In contrast, the inhibitory effect of S. oralis strains lacking the spxB gene on cross-kingdom biofilms was significantly reduced. These data provide ecological insights into how physicochemical properties of early colonizing commensals shape the structure and virulence of cross-kingdom oral biofilm through antimicrobial-mediated antagonistic activity. IMPORTANCE The co-existence of S. mutans and C. albicans accelerates the development of severe early childhood caries, particularly under frequent sucrose exposure. This study demonstrates that early colonizing and antimicrobial-producing oral commensal bacteria can disrupt these pathogenic interactions by modulating their physicochemical associations. These findings highlight the potential of enhancing commensal bacteria as part of novel caries prevention strategies. Further characterization of the functional oral microbiota, especially clinically relevant oral commensals, could advance the development of diagnostic biomarkers and microbiome-targeted therapeutics to prevent painful and costly oral diseases.