Rapid identification of Candida dubliniensis using a species-specific molecular beacon.
Candida dubliniensis is an opportunistic fungal pathogen that has been linked to oral candidiasis in AIDS patients, although it has recently been isolated from other body sites. DNA sequence analysis of the internal transcribed spacer 2 (ITS2) region of rRNA genes from reference Candida strains was used to develop molecular beacon probes for rapid, high-fidelity identification of C. dubliniensis as well as C. albicans. Molecular beacons are small nucleic acid hairpin probes that brightly fluoresce when they are bound to their targets and have a significant advantage over conventional nucleic acid probes because they exhibit a higher degree of specificity with better signal-to-noise ratios. When applied to an unknown collection of 23 strains that largely contained C. albicans and a smaller amount of C. dubliniensis, the species-specific probes were 100% accurate in identifying both species following PCR amplification of the ITS2 region. The results obtained with the molecular beacons were independently verified by random amplified polymorphic DNA analysis-based genotyping and by restriction enzyme analysis with enzymes BsmAI and NspBII, which cleave recognition sequences within the ITS2 regions of C. dubliniensis and C. albicans, respectively. Molecular beacons are promising new probes for the rapid detection of Candida species.
Park
S
S
Public Health Research Institute, New York, NY 10016, USA.
Wong
M
M
Marras
S A
SA
Cross
E W
EW
Kiehn
T E
TE
Chaturvedi
V
V
Tyagi
S
S
Perlin
D S
DS
D002175:Candida / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D002176:Candida albicans / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D002177:Candidiasis / Q000382:microbiology*; D004262:DNA Restriction Enzymes / Q000378:metabolism; D005456:Fluorescent Dyes; D006801:Humans; D008967:Molecular Biology / Q000379:methods; D015335:Molecular Probes; D016133:Polymerase Chain Reaction / Q000379:methods; D019105:Random Amplified Polymorphic DNA Technique; D012680:Sensitivity and Specificity; D013045:Species Specificity
D005456:Fluorescent Dyes; D015335:Molecular Probes; D004262:DNA Restriction Enzymes