Role of Plaque Microflora as a Dental Caries Predictor

Objective: Subtle differences in plaque microflora may have a role in evaluating caries risk assessment. This preliminary study used quantitative real-time PCR (QRTPCR) to examine differences in the plaque microflora from subjects who completed a 1yr longitudinal study on early childhood caries. Baseline plaque samples from subjects who were caries-free at baseline that either developed caries (n=6, Group A) or remained caries-free (n=6, Group B) after 1yr were assayed for the presence of specific bacterial species determined previously to be overabundant (Actinomyces naeslundii, Streptococcus mutans) or scant (S.mitis, S.gordonii) in carious lesions.

Methods: Plaque samples or control bacteria in cell lysis solution (1% Triton X-100, 20mM Tris-HCl, 2mM EDTA [pH 8.0]) were incubated with 20U of mutanolysin/ml and 0.2mg of lysozyme/ml at 37^oC for 2 hr, then boiled at 100^oC for 10 min. 20μl of a mixture containing 1μl of lysed cells, 1X TaqMan Universal PCR Master Mix (Applied Biosystems), 200nM (each) sense and antisense primer and 250nM TaqMan probe (5'FAM and 3'TAMRA labeled) were analyzed using the ABI PRISM 7000 sequence detection system (Applied Biosystems) with the following cycle profile: 50^oC for 2min, 95^oC for 10min, and 60 cycles of 95^oC for 15s and 58^oC for 1min. Serial dilutions (in triplicate) of each sample were assayed.

Results: Median % of total values for S.mutans and A.naeslundii (overabundant in caries) was 99.4% for Group A and 17.2% for Group B. The median % of total values for S.mitis and S.gordonii (scant in caries) was 0.59% for Group A and 82.8% for Group B. Mean %'s were not significantly different between the two groups in this small pilot study.

Conclusion: Preliminary results suggest examination of plaque microflora as a tool for caries risk assessment may be feasible but a larger cohort of species and multiple sampling timepoints may be necessary.