Investigation into the formation and susceptibility of Candida albicans biofilms and denture plaque in vitro
Microbial biofilms are wide-spread in nature and the organisms within them are often able to cause acute and chronic human diseases. The nature of biofilms is complex and the organisms behave differently from their planktonic counterparts. Many studies have been carried out on bacterial biofilms related to oral diseases, but little work has focused upon the potential clinical role of fungal and mixed fungal and bacterial biofilms. The aims of this project were to develop an in vitro model to investigate the development and structure of fungal biofilms and mixed fungal and bacterial biofilms associated with Denture related Stomatitis (DRS) and the effectiveness of antimicrobial agents against them. Single species Candida albicans and denture plaque derived from patients with DRS was grown in biofilms on dental materials in a Constant Depth Film Fermentor. Initial colonisation (2 - 6 h) of C. albicans was dependent upon the material and the roughness of the surface on which the biofilm formed. Higher numbers of yeast cells were present on enamel compared to dentine or acrylic whilst the highest numbers of cells were found on the roughest surfaces during early biofilm development. Confocal microscopy revealed that the hyphal form was associated with biofilm development and structure. Candida albicans grown in the biofilm phase of growth was more resistant to fluconazole, miconazole and chlorhexidine than when grown planktonically, although drug resistance varied between agents and biofilm growth phases. Microcosm biofilms containing similar proportions of genera to those of DRS lesions were developed. Further, the delivery of DRS-associated treatments was mimicked and it was demonstrated that a combination of miconazole and chlorhexidine was able to reduce the total bacterial count by 3 logio and the number of Candida species by over 2 logio. It can be concluded from this work that the initial phase of biofilm formation has a significant role to play in the development of biofilms on various oral surfaces and their subsequent susceptibility. Further, the development of an in vitro denture plaque model allowed the determination of the likely clinical effectiveness of treatment regimes.