Intraspecies interactions of Streptococcus mutans impact biofilm architecture and virulence determinants in childhood dental caries.

Early childhood dental caries (ECC) is the most common chronic disease among children with a heavy disease burden among low socioeconomic populations. Streptococcus mutans is most frequently associated with initiation of ECC. Many studies report children with multiple S. mutans strains (i.e., genotypes) having greater odds of developing ECC, studies investigating intraspecies interactions in dental caries are lacking. In this study, the impact of intraspecies interactions on cariogenic and fitness traits of clinical S. mutans isolates are investigated using in-vitro and in-vivo approaches. Initially clinical S. mutans isolates of 10 children from a longitudinal epidemiological study were evaluated. S. mutans strains (G09 and G18, most prevalent) isolated from one child were used for subsequent analysis. Association analysis was used to determine if presence of multiple S. mutans genotypes within the first-year of colonization was associated with caries. Biofilm analysis was performed for single and mixed cultures to assess cariogenic traits, including biofilm biomass, intra-polysaccharide, pH, and glucan. Confocal Laser Scanning Microscopy (CLSM) and time-lapse imaging were used to evaluate spatial and temporal biofilm dynamics, respectively. A Drosophila model was used to assess colonization in-vivo. Mean biofilm pH was significantly lower in co-cultured biofilms as compared with monoculture biofilms. Doubling of S. mutans in-vitro biofilms was observed by CLSM and in-vivo colonization in Drosophila for co-cultured S. mutans. Individual strains occupied specific domains in co-culture and G09 contributed most to increased co-culture biofilm thickness and colonization in Drosophila. Biofilm formation and acid production displayed distinct signatures in time-lapsed experiments.