Rapid identification and differentiation of Candida albicans and Candida dubliniensis by capillary-based amplification and fluorescent probe hybridization.
We developed a rapid genotypic assay to differentiate the germ tube-positive yeasts Candida albicans and Candida dubliniensis. Fluorescently labeled nucleic acid probe binding and subsequent denaturation from the target site in the PCR amplicons produced characteristic peak melting temperatures (T(m)) that identified each species. Peak T(m)s of C. albicans (n = 69) and C. dubliniensis (n = 28) isolates produced in the presence of their respective probes were 61.04 +/- 0.64 degrees C and 60.52 +/- 1.01 degrees C (averages +/- standard deviations). No signal was generated when the C. albicans or C. dubliniensis probes were tested against DNA from their counterparts. Both probes reacted with Candida tropicalis DNA, but the T(m) was 51.85 +/- 0.05 degrees C with the C. albicans probe and 51.92 +/- 0.10 degrees C with the C. dubliniensis probe, differentiating C. tropicalis DNA from C. albicans and C. dubliniensis. A novel hybrid probe was designed to identify both species in a single reaction based on a 4 degrees C difference in peak T(m)s. Our assay is rapid (</=2 h) and allows reliable detection and differentiation of the two germ tube-positive Candida spp.
Selvarangan
Rangaraj
R
Departments of Laboratory Medicine. Medicine. Microbiology, University of Washington Medical Center, Seattle, Washington 98195-7110, USA.
Limaye
Ajit P
AP
Cookson
Brad T
BT
D002175:Candida / Q000145:classification* / Q000235:genetics; D002176:Candida albicans / Q000145:classification* / Q000235:genetics; D002177:Candidiasis / Q000382:microbiology; D015342:DNA Probes; D004271:DNA, Fungal / Q000032:analysis; D021903:DNA, Ribosomal Spacer / Q000032:analysis; D005456:Fluorescent Dyes; D006801:Humans; D016533:Mycological Typing Techniques; D016133:Polymerase Chain Reaction / Q000295:instrumentation* / Q000379:methods*; D012680:Sensitivity and Specificity; D013696:Temperature; D013997:Time Factors
D015342:DNA Probes; D004271:DNA, Fungal; D021903:DNA, Ribosomal Spacer; D005456:Fluorescent Dyes