[Quantitative real-time polymerase chain reaction for the diagnosis of invasive fungal disease].
OBJECTIVE
To establish the diagnostic assays of quantitative real-time polymerase chain reaction (PCR) for the diagnosis of invasive fungal disease and evaluate their accuracy in clinical samples.


METHODS
Three assays have been developed for the diagnosis of clinically prevalent fungi, Aspergillus spp. and Candida spp.. Their analytical sensitivity and specificity were evaluated. Twenty-one serum samples from invasive fungal disease patients and non-invasive fungal disease patients were analyzed by the diagnostic assays and the accuracy was evaluated.


RESULTS
Three assays were managed to run at the same condition. The universal fungi assay was able to detect, but not differentiate between most of the pathogenic fungi, including A. fumigatus, A. flavus, A. niger, C. albicans, C. parapsilosis, C. tropicalis, C.kruseii, C.glabrata, Cryptococcus neoformans, Rhizomucor variabilis, Rhizopus arrhizus and Penicillosis marneffei. The assays of Aspergillus spp. and Candida spp. were able to detect, but not differentiate between A. fumigatus, A. flavus, A. niger and C. albicans, C. parapsilosis, C. tropicalis. The detection limits of assays were 4 pg of A. fumigatus gDNA, 2 copies of A. fumigatus and 2 copies of C. albicans respectively. The results of quantitative real-time PCR matched well with each other and were identical to the classical procedures in all patients. Among the 11 patients with invasive fungal disease, the pathogens were identified down to a genus level in 2 invasive aspergillosis patients and 2 invasive candidiasis patients. And the other 7 patients could be diagnosed with invasive fungal disease by the universal fungi assay. All the 10 serum samples from non-invasive fungal disease patients were revealed as negative.


CONCLUSIONS
The universal fungi assay is helpful in screening while the assays of Aspergillus spp. and the Candida spp. may identify the pathogens down to a genus level.
Department of Dermatology, Peking University First Hospital, Research Center for Medical Mycology, Peking University, Beijing 100034, China.
Qiao-yun
QY
Lu
Jin
J
Yu
Lu-juan
LJ
Gao
Gang
G
Zheng
Ruo-yu
RY
Li
D017931:DNA Primers; D004271:DNA, Fungal / Q000032:analysis* / Q000235:genetics; D005658:Fungi / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D009181:Mycoses / Q000175:diagnosis* / Q000382:microbiology; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D017931:DNA Primers; D004271:DNA, Fungal