You are here:
DETECTION AND IDENTIFICATION OF PATHOGENIC CANDIDA SPECIES IN WATER USING FLOW CYTOMETRY COUPLED WITH TAQMAN PCR
Citation:
Brinkman, N, R A. Haugland, J Santo Domingo, AND S J. Vesper. DETECTION AND IDENTIFICATION OF PATHOGENIC CANDIDA SPECIES IN WATER USING FLOW CYTOMETRY COUPLED WITH TAQMAN PCR. Presented at American Society for Microbiology, Orlando, FL, May 20-24, 2000.
Impact/Purpose:
1. Evaluate the efficacy of QPCR technology for detecting low level microbiological contaminants in water supplies
2. Provide additional data on the range of pathogenic or potentially pathogenic species of fungi in community distribution system and hospital water samples.
Description:
As the incidence of human fungal infection increases, the ability to detect and identify pathogenic fungi in potential environmental reservoirs becomes increasingly important for disease control. PCR based assays are widely used for diagnostic purposes, but may be inadequate for analyses of environmental samples due to the presence of inhibitors. Another method, flow cytometry, allows samples to be rapidly screened for organisms that exhibit target antibodies. These organisms can be recovered using the single cell sorting capability of the flow cytometer. The limitation of the flow cytometry method is that different related and even unrelated species may share the target characteristics used for sorting and thus cannot be distinguished. In this study we tested the use of flow cytometry for isolation of pathogenic Candida species from water samples coupled with TaqMan PCR for identification of the recovered organisms. Using flow cytometry, it was demonstrated that a fluorescently labeled Candida species, including C. albicans, C. tropicalis, C. parasilosis, C. viswanathii, C. sojae, C. dubliniensis, C. maltosa, C. haemulonii, and C. lusitaniae exhibit sufficiently higher fluorescence levels than unrelated yeasts to allow their selective recovery. Different TaqMan PCR assays for each of these target species were shown to be specific and were indicated to be sensitive enough to detect as few as 1 - 20 cells per reaction, depending on the target species. The combination of the two methods described here provides a useful approach for identifying pathogenic Candida species in environmental samples.