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Title
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Evaluation of a Pan-Leishmania spliced-leader RNA detection method in human blood and experimentally infected Syrian golden hamsters
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Author
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Abstract
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| Several methods have been developed for the detection of Leishmania, mostly targeting the minicircle kinetoplast DNA (kDNA). A new RNA real-time quantitative PCR (qPCR) assay was developed targeting the conserved and highly expressed spliced-leader (SL) mini-exon sequence. This study compared the limits of detection of various real-time PCR assays in hamsters infected with Leishmania infantum, in spiked human blood, and in clinical blood samples from visceral leishmaniasis patients. The SL-RNA assay showed an excellent analytical sensitivity in tissues (0.005 and 0.002 parasites per mg liver and spleen, respectively) and was not prone to false-positive reactions. Evaluation of the SL-RNA assay on clinical samples demonstrated lower cT values than the kDNA qPCR, an excellent interrun stability of 97%, a 93% agreement with the kDNA assay, and an estimated sensitivity, specificity, and accuracy of 93.2%, 94.3%, and 93.8%, respectively. The SL-RNA qPCR assay was equally efficient for detecting Leishmania major, Leishmania tropica, Leishmania mexicana, Leishmania guayensis, Leishmania panamensis, Leishmania braziliensis, L. infantum, and Leishmania donovani and revealed similar SL-RNA levels in the different species and the occurrence of polycistronic SL-containing transcripts in Viannia species. Collectively, this single SL-RNA qPCR assay enables universal Leishmania detection and represents a particularly useful addition to the widely used kDNA assay in clinical studies in which the detection of viable parasites is pivotal to assess parasitological cure. |
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Language
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English
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Source (journal)
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The journal of molecular diagnostics / American Society for Investigative Pathology; Association for Molecular Pathology [Bethesda, Md] - Bethesda, Md
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Publication
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Bethesda, Md
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2018
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ISSN
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1525-1578
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DOI
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10.1016/J.JMOLDX.2017.12.003
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Volume/pages
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20
:2
(2018)
, p. 253-263
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ISI
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000428608800014
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Pubmed ID
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29355825
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Full text (Publisher's DOI)
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