A method for mapping the distribution pattern of cariogenic streptococci within dental plaque in vivo

This study was carried out to develop a method for mapping the distribution of cariogenic oral streptococci, Streptococcus mutans and Streptococcus sobrinus, from the outermost to the innermost plaque. Ten consenting subjects were asked to form plaque by abstaining from tooth brushing over 3 days within in situ plaque-generating devices, which were placed on the upper molars. The plaque formed in the devices was separated into 8-10 layered fractions (100 microm thick). Genomic DNA was extracted from each plaque fraction by a commercial DNA purification kit and used for the amplification of the 16S ribosomal RNA gene sequences by polymerase chain reaction (PCR) with universal primers. The products were then amplified by PCR with S. mutans- or S. sobrinus-specific nested primers. The final products were separated on agarose gels, stained and photographed to confirm the existence of S. mutans and S. sobrinus. The results showed that S. mutans was detected in the plaque obtained from all of the 10 subjects and S. sobrinus in the plaque of 7 subjects. However, the distribution patterns of fractions positive for S. mutans and S. sobrinus varied among the subjects, with a tendency for frequent detection of both species in the outer to middle layers of dental plaque. There were no plaque fractions in which only S. sobrinus was found. This method could be useful to map the distribution of cariogenic microorganisms and to estimate the bacterial ecology for oral biofilm.