Enrichment by Serum of Black-pigmented Anaerobes in a Microbial Community

Periodontal diseases are associated with increased levels of proteolytic and obligately anaerobic, Gram negative bacteria. Some of the putative pathogens can be found at healthy sites, albeit in very low levels. Changes in key environmental factors may be responsible for perturbing the natural balance that exists among the sub-gingival microflora, thereby favouring the selection and enrichment of these pathogens. Objective: to investigate the ability of serum (as a surrogate of gingival crevicular fluid, GCF) to enrich for putative periodontal pathogens within a microbial community. Methods: Ten bacteria, representative of those found in health and disease, were grown as a mixed culture in an anaerobic chemostat, maintained automatically at pH 7.0 „b 0.1, on a mucin-based growth medium. Viable counts were determined on selective and non-selective agar plates. Protease activity of the whole consortium was measured using synthetic substrates. Once steady-state growth was established, pH-control was terminated, and serum was introduced into the culture medium (10% final concentration), and changes in culture parameters were monitored over time. Results: During steady-state conditions before the serum addition, the culture was dominated by streptococci, Veillonella dispar and Fusobacterium nucleatum; levels of black-pigmented anaerobes and total protease activity were relatively low. Following the addition of serum, the culture became more anaerobic (the Eh fell from -330mV to -390mV), and bacterial metabolism resulted in the pH being raised to ca. pH7.5. These environmental changes were associated with an increase in the total biomass of the community, especially in black-pigmented anaerobes, and the culture became dominated by Porphyromonas gingivalis; total protease activity also rose. Conclusion: The increased flow of GCF during the host inflammatory response can favour the growth of some of the putative periodontal pathogens, and result in their ability to out-compete organisms associated with oral health.