Open AccessDNA probes for the rapid identification of medically important Candida species using a multianalyte profiling system
Author(s) -
Das Sanchita,
Brown Teresa M.,
Kellar Kathryn L.,
Holloway Brian P.,
Morrison Christine J.
Publication year - 2006
Publication title -
fems immunology & medical microbiology
Language(s) - English
Resource type - Journals
eISSN - 1574-695X
pISSN - 0928-8244
DOI - 10.1111/j.1574-695x.2005.00031.x
Subject(s) - candida krusei , amplicon , biology , candida parapsilosis , candida tropicalis , candida glabrata , candida albicans , polymerase chain reaction , microbiology and biotechnology , dna , microtiter plate , hybridization probe , internal transcribed spacer , ribosomal rna , genetics , gene
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 .