A model for clinical evaluation of anti-microbial effects of agents on plaque colonization

Purpose: To evaluate a new improved method of microbial analysis in a cross -over clinical study by investigating the efficacy of a single application of an essential oil-containing (EOC) dentifrice as compared to its vehicle control (VC) over a 6-hr period. Materials and Methods: Acrylic stents with retention areas for 7, 3mm x 3mm hydroxyapatite (HA) squares were fabricat ed for 12 subjects. 30 min following stent placement, one HA square was rem oved and sampled for viable microflora. After stent replacement, subjects w ere assigned either the EOC dentifrice or its VC and brushed under supervis ion for 1 min. Stents remained in place for the next 6 hrs. A HA square was removed at hourly intervals for 6 hrs following brushing. The microflora w as analyzed for total anaerobes on Schaedler's media, for total Gram-negati ve anaerobes on Schaedler-NV selective media, and for total Fusobacterium s pecies on CVE selective media. Plates were incubated anaerobically at 37 de grees C for 2-5 days. Colony forming units were calculated. For each time p oint, pairwise t-tests were performed using the adjusted means and the pool ed error term from the analysis of variance (ANOVA). Results: Treatment wit h the EOC dentifrice resulted in a statistically reduced plaque growth. Dif ferences were seen as reductions in: (1) total Gram-negative anaerobes seen from 1-6 hrs ( P less than or equal to 0.011), (2) total anaerobic bacteri a which achieved significance at 3 hrs and continued through 6 hrs (P less than or equal to 0.005), and (3) Fusobacterium species as seen from 4-6 hrs (P less than or equal to 0.002).