DNA probes for the rapid identification of medically important Candida species using a multianalyte profiling system.
Recently, a new flow cytometric technology to detect multiple DNA target sequences in a single microtiter well plate was developed [multianalyte profiling (MAP) System, Luminex Corp., Austin, TX]. DNA probes, directed to the internal transcribed spacer 2 region of ribosomal DNA, were therefore designed to detect and differentiate PCR amplicons from six medically important Candida species using this system. Each probe was covalently linked to one of 100 available microsphere (bead) sets. Biotinylated PCR amplicons were then hybridized to the complementary probe on each bead set. Bound amplicons were detected fluorometrically using a streptavidin-linked reporter dye, R-phycoerythrin. Specific hybridization was noted for all six Candida species probes (mean sample-to-background ratio+/-standard error: Candida albicans, 58.7+/-1.2; Candida tropicalis, 53.2+/-3.8; Candida glabrata, 46.9+/-2.1; Candida parapsilosis, 59.9+/-1.6; Candida krusei, 54.7+/-3.7 vs. 0.9+/-0.03 for all heterologous Candida species DNA targets and vs. 1.0+/-0.1 for samples containing water instead of DNA; P < 0.001). The limit of test sensitivity was 0.5 pg of DNA. A sample could be processed and analyzed within 1 h post-PCR amplification. Therefore, the multianalyte profiling system was rapid, sensitive and specific for the detection and differentiation of the most medically important species of Candida.
Mycotic Diseases Branch, Division of Bacterial and Mycotic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, GA 30333, USA.
Sanchita
S
Das
Teresa M
TM
Brown
Kathryn L
KL
Kellar
Brian P
BP
Holloway
Christine J
CJ
Morrison
D002175:Candida / Q000145:classification* / Q000235:genetics; D002177:Candidiasis / Q000382:microbiology; D015342:DNA Probes; D004271:DNA, Fungal / Q000032:analysis; D005434:Flow Cytometry; D006801:Humans; D008863:Microspheres; D016533:Mycological Typing Techniques / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011137:Polystyrenes; D012680:Sensitivity and Specificity; D013997:Time Factors
D015342:DNA Probes; D004271:DNA, Fungal; D011137:Polystyrenes