Analytical Performance of a Loop-Mediated Isothermal Amplification Assay for Leishmania DNA Detection in Sandflies and Direct Smears of Patients with Cutaneous Leishmaniasis
Loop-mediated isothermal amplification (LAMP) is ideal for the detection of DNA as it is a quick and easy-to-perform test that does not require complex or sophisticated equipment or infrastructure. However, the application of this technique in the detection of DNA has not been comprehensively analyz...
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| Published in: | The American journal of tropical medicine and hygiene Vol. 98; no. 5; pp. 1325 - 1331 |
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| Main Authors: | , , , , , , |
| Format: | Journal Article |
| Language: | English |
| Published: |
United States
Institute of Tropical Medicine
01-01-2018
The American Society of Tropical Medicine and Hygiene |
| Subjects: | |
| Online Access: | Get full text |
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| Summary: | Loop-mediated isothermal amplification (LAMP) is ideal for the detection of
DNA as it is a quick and easy-to-perform test that does not require complex or sophisticated equipment or infrastructure. However, the application of this technique in the detection of
DNA has not been comprehensively analyzed to date (analytical validation). Our objective was to evaluate the sensitivity and analytical specificity (anticipated reportable range [ARR], the limit of detection [LoD], and accuracy) of LAMP targeting the 18S rRNA gene in the diagnosis of six New World
species. We then applied the validated LAMP assay across 50 samples of sandflies and 50 direct smears from a recent outbreak of cutaneous leishmaniasis in Colombia to determine its diagnostic performance. The LAMP assay exclusively amplified the DNA of
spp., and an ARR of between 1 × 10
and 1 × 10
equivalent parasites/mL was determined. An LoD of 1 × 10
equivalent parasites/mL was established and there was no statistically significant variation in terms of accuracy. Finally, a sensitivity of 100% in direct smears and sandflies samples was calculated and a specificity of 90.9% for direct smears using microscopy as reference and 96.8% for sandflies using real-time polymerase chain reaction as reference were determined. To our knowledge, this is the first attempt to analytically validate a LAMP test to detect
DNA, which showed good diagnostic potential from sandflies and direct smear samples. |
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| Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Financial support: We thank the Departamento Administrativo de Ciencia, Tecnología e Innovación ‘COLCIENCIAS’ for funding the Project “Fortalecimiento de la capacidad diagnóstica, de investigación y de vigilancia de enfermedades transmisibles emergentes y reemergentes en Colombia” grant number 757-13. Authors’ addresses: Cielo M. León, Marina Muñoz, Carolina Hernandez, and Juan David Ramírez, Facultad de Ciencias Naturales y Matemáticas, Grupo de Investigaciones Microbiológicas, Universidad del Rosario, Bogotá, Colombia, E-mails: cmlr8527@gmail.com, claudiamarina23@gmail.com, dcahernandez@gmail.com or dcahernandezc@gmail.com, and juand.ramirez@urosario.edu.co. Juan H. Tabares, Departamento de Salud Publica, Universidad Nacional de Colombia, Bogotá, Colombia, E-mail: jhtabaresg@unal.edu.co. Carolina Florez and Martha S. Ayala, Grupo de Parasitología, Instituto Nacional de Salud, Bogotá, Colombia, E-mails: aflorez@ins.gov.co and mayalas@ins.gov.co. |
| ISSN: | 0002-9637 1476-1645 |
| DOI: | 10.4269/ajtmh.17-0808 |