Quantitative Isolation of Fungi from Root Canals: A Methodological Study

Fungi have been implicated in cases of failed endodontic treatment. The prevalence of yeasts in root canals has been reported to be between 7% and 55%. Since the oral flora includes yeasts such as Candida species, this wide range may be the result of contamination from the oral cavity. We describe a method for isolation and quantification of yeasts that controls for contamination from oral flora. Objective: To quantitatively determine the presence and to identify the yeasts species in the root canals of endodontic patients with pulp necrosis and apical periodontitis. Methods: Oral flora was evaluated for the presence of Candida species with quantitative culturing of an oral rinse. HIV+/AIDS subjects with cfu>400 were included. HIV- subjects served as controls. After isolation with a rubber dam, the tooth surface was sampled for presence of adherent yeasts. Tooth surface was disinfected and sampled again. The pulp chamber was accessed with sterile new burr without water spray, and a sample was collected from the root canal. The yeasts were quantified using colony forming assay. The organisms were speciated using an API 20 C AUX Kit (BioMérieux) and by germination. Results: Candida was present in the oral cavity of 37% of HIV- subjects, with counts ranging 8-110 cfus per rinse, and none were found in root canals. Candida was present on the tooth surface of HIV+/AIDS subjects, and was completely eliminated by disinfection. Importantly, Candida species were recovered from root canals of HIV+/AIDS subjects as pure cultures, and were the same as those found in the PBS oral rinse. Conclusion: This methodology assures us that Candida species recovered from root canals were not contaminants. Our results indicate that Candida species may play a role in endodontic disease of individuals with HIV+/AIDS .

Supported by the Campbell Foundation and the UCLA Start up Grant