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Molecular diagnosis of Old World leishmaniasis: Real-time PCR based on tryparedoxin peroxidase gene for the detection and identification of Leishmania spp

Author(s): Sharifeh Khosravi , Saied Hossein Hejazi , Mortaza Hashemzadeh , Gilda Eslami & Hossein Yousofi Darani

Journal: Journal of Vector Borne Diseases
ISSN 0972-9062

Volume: 49;
Issue: 1;
Start page: 15;
Date: 2012;
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Keywords: Cutaneous leishmaniasis | diagnosis | Iran | peroxidase | RT-PCR | tryparedoxin

Background & objectives: Rapid and accurate diagnosis and identification of Leishmania sp causing cutaneousleishmaniasis is crucial in control and therapeutic programs. The problem of diagnosis with traditional methodsis that they have a low sensitivity or time consuming but molecular techniques would be an alternative methodfor rapid and accurate diagnosis. In this work, tryparedoxine peroxidase gene-based real-time PCR was used foraccurate identification of Leishmania spp causing Old-World cutaneous leishmaniasis.Methods: In this study, biopsies of specimens were taken from the ulcerative sites in 100 patients and used fordirect microscopy, culture in NNN or fixed in alcohol for identification of Leishmania spp using tryparedoxinperoxidase gene-based realtime PCR (qPCR).Results: Using direct microscopy and culture method, Leishmania parasites were isolated from 68 out of 100patient samples. However, 13 patients with negative finding on traditional tests, had positive results on RT-PCRtest. After melting curve analysis of PCR product, Leishmania major in 75 and L. tropica in 4 cases wereidentified. The sensitivity and specificity of RT-PCR for diagnosis of cutaneous leishmaniasis was 98.7 and59.8%, respectively.Conclusion: Results of this study showed that RT-PCR was the most sensitive diagnostic test for cutaneousleishmaniasis and represents a tool for rapid species identification.

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