NDLI: Comparison between Quantitative Nucleic Acid Sequence-Based Amplification, Real-Time Reverse Transcriptase PCR, and Real-Time PCR for Quantification of Leishmania Parasites▿

Content Provider	PubMed Central
Author	Meide, Wendy Van Der Guerra, Jorge Schoone, Gerard Farenhorst, Marit Coelho, Leíla Faber, William Inge, Peekel Schallig, Henk
Abstract	DNA or RNA amplification methods for detection of Leishmania parasites have advantages regarding sensitivity and potential quantitative characteristics in comparison with conventional diagnostic methods but are often still not routinely applied. However, the use and application of molecular assays are increasing, but comparative studies on the performance of these different assays are lacking. The aim of this study was to compare three molecular assays for detection and quantification of Leishmania parasites in serial dilutions of parasites and in skin biopsies collected from cutaneous leishmaniasis (CL) patients in Manaus, Brazil. A serial dilution of promastigotes spiked in blood was tested in triplicate in three different runs by quantitative nucleic acid sequence-based amplification (QT-NASBA), quantitative real-time reverse transcriptase PCR (qRT-PCR), and quantitative real-time PCR (qPCR). In addition, the costs, durations, and numbers of handling steps were compared, and 84 skin biopsies from patients with suspected CL were tested. Both QT-NASBA and qRT-PCR had a detection limit of 100 parasites/ml of blood, while qPCR detected 1,000 parasites/ml. QT-NASBA had the lowest range of intra-assay variation (coefficients of variation [CV], 0.5% to 3.3%), while qPCR had the lowest range of interassay variation (CV, 0.4% to 5.3%). Furthermore, qRT-PCR had higher r 2 values and amplification efficiencies than qPCR, and qPCR and qRT-PCR had faster procedures than QT-NASBA. All assays performed equally well with patient samples, with significant correlations between parasite counts. Overall, qRT-PCR is preferred over QT-NASBA and qPCR as the most optimal diagnostic assay for quantification of Leishmania parasites, since it was highly sensitive and reproducible and the procedure was relatively fast.
Related Links	http://dx.doi.org/10.1128/jcm.01416-07
Ending Page	78
Page Count	6
Starting Page	73
File Format	PDF
ISSN	00951137
e-ISSN	1098660X
Journal	Journal of Clinical Microbiology
Issue Number	1
Volume Number	46
Language	English
Publisher	American Society for Microbiology (ASM)
Publisher Date	2008-01-01
Access Restriction	Open
Rights Holder	American Society for Microbiology (ASM)
Subject Keyword	Microbiology (medical) Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Microbiology (medical)

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