Study of bacterial viability within human supragingival dental calculus

BACKGROUND: There is evidence that supragingival calculus contains unmineralized channels and lacunae. The purpose of this study was to investigate the viability of bacteria within these areas. METHODS: Supragingival calculus harvested from patients with moderate to severe chronic periodontitis was immediately frozen to -70 degrees C. Six samples were cryosectioned, stained with a bacterial viability kit, and examined with fluorescence microscopy. Controls comprised heat treatment of cryosections prior to staining. Four additional samples were stained and examined whole in a confocal laser scanning microscope (CLSM). Nine additional samples were prepared for bacterial culture, after initial irradiation with ultraviolet light to kill viable organisms on the covering plaque layer. Test samples were crushed to expose internal bacteria, while two controls were used without crushing. RESULTS: Viable bacteria, as identified using the bacterial viability stain, were found within cavities/lacunae in supragingival calculus cryosections. Similar results were obtained from whole calculus samples using CLSM. Of the nine experimental samples where bacterial culture was attempted, five provided positive bacterial culture under both aerobic and anaerobic conditions; one showed positive growth under aerobic conditions only; while one showed no bacterial growth. The controls showed no bacterial growth. CONCLUSIONS: From this study, it appears that viable aerobic and anaerobic bacteria may be present within supragingival calculus, specifically within the internal channels and lacunae. Clinically, this may be important, since incomplete removal of supragingival calculus may expose these reservoirs of possible pathogenic bacteria and be a factor in the recurrence of periodontal diseases after treatment.