Fusobacteria are able to co-aggregate with numerous bacterial genera. In particular, Fusobacterium nucleatum (F.n) is thought to act as a “microbial bridge” as it co-aggregates with early and late colonizers of dental plaque. Objectives: Studies in our laboratory showed that under certain environmental conditions, F.n forms co-aggregates that may lead to the subsequent establishment of homogenous biofilms. The aims of this study were to identify environmental conditions that promote co-aggregation and investigate the nature of the interactions between cells. Methods: F. nucleatum ATCC 10953 was grown in continuous culture under controlled conditions. Environmental pH was varied and growth at 7.4 was compared to that at 8.1 or 6.4. Cellular biomass, intracellular polymer synthesis (IPS), co-aggregation and hydrophobicity were measured at each growth condition. SEM was used to assess biofilms grown on enamel slabs in the culture vessel. Results: Growth at pH 7.4 in the presence of glucose (20mM) produced cells containing significant IPS levels (250umol glucose/g protein). Growth at pH 8.1 resulted in, co-aggregation but no detectable IPS. Growth in the absence of glucose at pH 8.1 showed that carbohydrate is not required for biofilm formation. SEM analysis of enamel slabs revealed dense biofilms of elongated cells but those from growth at pH 7.4 displayed minimal cell attachment. Cellular hydrophobicity increased significantly in co-aggregated cells (34%±7) compared to cells grown at pH 7.4 (15%±5). Hemin inhibited co-aggregation whereas glucosamine, trypsin and lactose were ineffective. Conclusion: Conditions in the diseased periodontal pocket exhibit increased alkalinity with pocket depth and severity of the host inflammatory response. This study reports a relationship between environmental pH and biofilm formation. Growth at elevated pH promotes co-aggregation and biofilm formation. Co-aggregation appears linked to increased cell-surface hydrophobicity.
We wish to thank the Australian Dental Research Foundation for their support.