Date of Degree
PhD (Doctor of Philosophy)
Tooth decay is a serious health risk and a significant contributor to health care costs in both industrialized and developing nations. Tooth decay is the end result of a change in the balance of plaque ecology towards more acidogenic and aciduric bacterial species. The primary force facilitating this change is an increase in the amount and frequency of simple carbohydrate, in particular, sucrose ingestion by the host. The acidity of plaque increases after host ingestion of fermentable carbohydrates and this promotes demineralization of the tooth enamel which is restored by salivary buffering and mineral deposition. Frequent and prolonged periods of low plaque pH drive cycles of enamel homeostasis towards demineralization, which ultimately leads to the formation of dental caries. Streptococcus mutans is the main etiologic agent in the development of dental caries. The cariogenic potential of S. mutans is based on their ability to produce and tolerate large amounts of acid and to adhere to and accumulate large numbers on the surface of a tooth. They are capable of efficiently fermenting a variety of simple carbohydrates and can produce high concentrations of acid, even in a low pH environment. However, it is the ability of S. mutans to rapidly synthesize copious amounts of water-insoluble and water-soluble glucan from dietary sucrose, which allow the bacteria to accumulate large enough numbers to dominate the dental plaque and significantly lower the plaque pH. Synthesis of glucan is mediated by glucosyltransferase enzymes and is crucial to sucrose-dependent adherence and to the cariogenicity of S. mutans. S. mutans also makes four non-GTF glucan-binding proteins: GbpA and GbpD are secreted and released proteins that contain a region that is homologous to the glucan-binding domains of the Gtf enzymes, and GbpC is a cell wall bound protein that confers the property of dextran-dependent aggregation during stressful conditions, and GbpB whose glucan-binding properties appear secondary to its role in cell-wall metabolism. It was hypothesized that Gbps A, C, and D primarily function to shape the architecture of S. mutans biofilms which in turn affects the cariogenicity of S. mutans. To test this hypothesis, a panel of Gbp mutants was constructed from S. mutans strain UA130 that encompasses all deletions of Gbps (GbpA, GbpC and GbpD) individually and in combination. Specific pathogen-free rats were infected with the WT S. mutans UA130 strain along with each of the Gbp mutants, were fed a high sucrose diet for seventy days, and were then scored for caries. Significant attenuation of caries was observed in some but not all gbp mutants. Biofilms were also grown and analyzed via confocal microscopy and COMSTAT image analysis software. Architectural differences were found with all of the gbp mutants when compared to the wild-type, most notably the mutant strains lost significant biofilm depth. Several of the architectural parameters correlated with caries attenuation. It was concluded that deletion of one or more Gbps resulted in a partial loss of the cohesive properties of S. mutans biofilms and changes in biofilm architecture. In several cases this resulted in significant attenuation of cariogenicity but not a complete loss. The architectural changes that resulted from this loss of biofilm cohesiveness and the specific combinations of Gbp deletions that lead to significant attenuation suggested specific roles for each Gbp in biofilm formation. Furthermore, the attenuation of Gbp mutant strains could not be explained by differences in acidogenicity or aciduricity among the mutants. Therefore it was concluded that Gbps A, C and D make profound contributions to biofilm architecture and changes in biofilm architecture, as a result of loss of Gbp-mediated cohesion, affects S. mutans cariogenicity.
Copyright 2010 David John Lynch
Lynch, David John. "An analysis of the role of glucan-binding proteins in Streptococcus mutans biofilm architecture and caries development." dissertation, University of Iowa, 2010.