Generation and Characterization of Microcosm Biofilms Modelling Pathogen-Specific Plaques

Dental plaque is a heterogeneous site-specific biofilm comprised of hundreds of species, many uncultivated. Pathogen content is also site-specific. Objectives: To generate dental plaque microcosm biofilms reflecting of differing pH, caries and periodontal species content, including uncultured species. Methods: Plaque microcosms were cultured supplied with artificial saliva (DMM) supplemented to vary the pHmax (with 10mM urea, or 5 or 10mM glucose), and with sucrose supplied for 6 min every 4h, 8h, or not supplied. The microbiota were analyzed by checkerboard analysis, culture and uncultured species-specific PCR. Results: Plaques supplied with DMM supplemented with 10 mM urea had a pHmax above 7.0 irrespective of sucrose frequency. Plaques in the neutral to slightly acid range showed a highly complex flora similar to natural plaque by Checkerboard analysis, and were different in each growth condition. High levels of Gram negative and Peptostreptococci spp. were present, and in some plaques, Tannerella forsythia was present. In the urea-supplemented high pHmax plaques, uncultured taxa that have been associated with periodontitis (Megasphaera, Dialaster, Desulfobulus, Lachnospiracae ) were detected by PCR. In one plaque Entamoeba gingivalis was also detected by TEM. DMM supplementation with 5mM glucose and 4-hourly sucrose pulses gave plaques with an eco-catastrophic pH collapse to a pHmax of 4.5. With DMM-10 mM glucose, both 8- and 4-hourly sucrose yielded plaques with a pHmax of 3.8, a pH found in the depth of carious lesions. Almost all CKB and PCR-probed species had disappeared leaving mainly L. plantarum, S. vestibularis, S. mitis II , and low levels of S. oralis, S. sanguis, S. gordonii , and the OP11 phylotype. Conclusion: Environmental conditions that model different locations in the oral cavity generate highly realistic self-organized plaque microcosm biofilms of distinctive complex microbial populations, including uncultured species, and species relevant to periodontitis and to dental caries.