A sensitive and a rapid multiplex polymerase chain reaction for the identification of Candida species in concentrated oral rinse specimens in patients with diabetes

Abstract

Attached

Objectives: Oral candidiasis is being frequently recognized in patients with diabetes, and is associated with multiple pathogens induding C andid a albicans, C andid a parapsilosis, C an d id a g la b ra ta and C andida tropicalis. The aim of this study was to evaluate a usefulness of a Multiplex Polymerase Chain Reaction as a rapid diagnostic tool for identification of four oral Candida pathogens in patients with diabetes. Materials and methods: A multiplex PCR was optimized to identify four C an d id a species in concentrated oral rinse samples. Common reverse primer, ITS4 and four species-specific forward primers targeting ITS1 and TTS2 regions of yeast genome were used. Species-spedfic single amplicon were detected by agarose gel electrophoresis. Performance efficacy of multiplex PCR was compared with phenotypic identification. Results: Out of 100 oral rinse samples, 72 were culture positive and of these 43 were at risk of oral Candida infection (>600cfu/ml). Multiple C an d id a species including C. albicans, C. parapsilosis and C. tropicalis were identified in 22 samples which had risk of oral Candida infection. In total, 85 patients were positive for Candida by multiplex PCR and of them 49 had multiple C andid a species. All 43 colonized specimens were also positive by multiplex PCR. C. albicans was the most predominant organism (75/85) followed by C. parapsilosis (47/85), C. tropicalis (17/85) and C. g la b ra ta (6/85). In specimens with multiple species, the two most common organisms were C albicans and C. parapsilosis. Multiplex PCR yielded a sensitivity of 10 Candida cells/ml of oral rinse sample. Conclusions: Multiplex PCR is found to be rapid, sensitive and specific than phenotypic identification methods in discriminating multiple C andid a species in oral rinse specimens.