An Innovative Field-Applicable Molecular Test to Diagnose Cutaneous Leishmania Viannia spp. Infections

Cutaneous and mucosal leishmaniasis is widely distributed in Central and South America. Leishmania of the Viannia subgenus are the most frequent species infecting humans. L. (V.) braziliensis, L. (V.) panamensis are also responsible for metastatic mucosal leishmaniasis. Conventional or real time PCR...

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Bibliographic Details
Published in:	PLoS neglected tropical diseases Vol. 10; no. 4; p. e0004638
Main Authors:	Saldarriaga, Omar A, Castellanos-Gonzalez, Alejandro, Porrozzi, Renato, Baldeviano, Gerald C, Lescano, Andrés G, de Los Santos, Maxy B, Fernandez, Olga L, Saravia, Nancy G, Costa, Erika, Melby, Peter C, Travi, Bruno L
Format:	Journal Article
Language:	English
Published:	United States Public Library of Science 26-04-2016 Public Library of Science (PLoS)
Subjects:	Biology and Life Sciences Chromatography, Affinity Control Copyright Cutaneous leishmaniasis Deoxyribonucleic acid Diagnosis Diagnostic tests DNA DNA Primers - genetics DNA, Kinetoplast - genetics DNA, Protozoan - genetics Engineering and Technology Humans Infections Laboratories Leishmania Leishmania - genetics Leishmania - isolation & purification Leishmaniasis Leishmaniasis, Cutaneous - diagnosis Medicine and Health Sciences Metastasis Microscopy Molecular Diagnostic Techniques - methods Nucleic Acid Amplification Techniques Oligonucleotide Probes - genetics Parasites Parasitic diseases People and places Point-of-Care Systems Polymerase chain reaction Research and Analysis Methods Sensitivity and Specificity Surveillance Tropical diseases Viannia ASW, South America Latin America
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Summary:	Cutaneous and mucosal leishmaniasis is widely distributed in Central and South America. Leishmania of the Viannia subgenus are the most frequent species infecting humans. L. (V.) braziliensis, L. (V.) panamensis are also responsible for metastatic mucosal leishmaniasis. Conventional or real time PCR is a more sensitive diagnostic test than microscopy, but the cost and requirement for infrastructure and trained personnel makes it impractical in most endemic regions. Primary health systems need a sensitive and specific point of care (POC) diagnostic tool. We developed a novel POC molecular diagnostic test for cutaneous leishmaniasis caused by Leishmania (Viannia) spp. Parasite DNA was amplified using isothermal Recombinase Polymerase Amplification (RPA) with primers and probes that targeted the kinetoplast DNA. The amplification product was detected by naked eye with a lateral flow (LF) immunochromatographic strip. The RPA-LF had an analytical sensitivity equivalent to 0.1 parasites per reaction. The test amplified the principal L. Viannia species from multiple countries: L. (V.) braziliensis (n = 33), L. (V.) guyanensis (n = 17), L. (V.) panamensis (n = 9). The less common L. (V.) lainsoni, L. (V.) shawi, and L. (V.) naiffi were also amplified. No amplification was observed in parasites of the L. (Leishmania) subgenus. In a small number of clinical samples (n = 13) we found 100% agreement between PCR and RPA-LF. The high analytical sensitivity and clinical validation indicate the test could improve the efficiency of diagnosis, especially in chronic lesions with submicroscopic parasite burdens. Field implementation of the RPA-LF test could contribute to management and control of cutaneous and mucosal leishmaniasis.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-3 content type line 23 ObjectType-Undefined-2 ObjectType-Feature-2 Conceived and designed the experiments: OAS ACG PCM BLT. Performed the experiments: OAS ACG. Analyzed the data: OAS ACG PCM BLT. Contributed reagents/materials/analysis tools: RP GCB AGL MBdLS OLF NGS EC. Wrote the paper: OAS ACG PCM BLT. Reviewed the manuscript, made comments and recommended additions or modifications: RP GCB AGL NGS. The authors have declared that no competing interests exist.
ISSN:	1935-2735 1935-2727 1935-2735
DOI:	10.1371/journal.pntd.0004638