Abstract
Visceral leishmaniasis (VL) is considered a major public health concern in Brazil and several regions of the world. A recent advance in the diagnosis of infectious diseases was the development of loop-mediated isothermal amplification (LAMP). The aim of this study was to develop and evaluate a new LAMP assay for detection of K26 antigen-coding gene of L. donovani complex. A total of 219 blood samples of immunocompetent patients, including 114 VL cases and 105 non-VL cases, were analyzed for the diagnosis of VL in the present study. Diagnostic accuracy was calculated against a combination of parasitological and/or serological tests as a reference standard. The results were compared to those of kDNA Leishmania-PCR. The detection limit for the K26-Lamp assay was 1fg L. infantum purified DNA and 100 parasites/mL within 60 min of amplification time with visual detection for turbidity. The assay was specific for L. donovani complex. Sensitivity, specificity, and accuracy were 98.2%, 98.1%, and 98.2%, respectively, for K26-LAMP and 100%, 100%, and 100%, respectively, for kDNA Leishmania-PCR. Excellent agreement was observed between K26-LAMP and kDNA Leishmania-PCR assays (K = 0.96). A highly sensitive and specific LAMP assay targeting K26 antigen-coding gene of L. donovani complex was developed for diagnosis in peripheral blood samples of VL patients.
Highlights
Visceral leishmaniasis (VL) is a Neglected Tropical Disease (NTD) considered a significant public health problem in Brazil and several regions of the world [1]
Over 90% of VL human cases reported in the Americas occur in Brazil, where more than 41,000 cases were recorded between the years 2000 and 2011, with 3,322 deaths [2]
This study reports the development and evaluation of a loop-mediated isothermal amplification (LAMP) assay for the detection of K26 antigen-coding gene of L. donovani complex in the blood samples of VL patients and compares the results to those of kDNA Leishmania-Polymerase chain reaction (PCR), a test used in reference centers for VL diagnosis in Brazil, exhibiting high sensitivity and specificity
Summary
Visceral leishmaniasis (VL) is a Neglected Tropical Disease (NTD) considered a significant public health problem in Brazil and several regions of the world [1]. Direct agglutination test (DAT), indirect fluorescent antibody test (IFAT), enzyme-linked immunosorbent assay (ELISA), and immunochromatographic tests (ICT) are currently used for VL diagnosis [3, 4]. In general, these methods present a number of limiting factors such as (i) the possibility of cross-reaction with other diseases, including Chagas disease and tegumentary leishmaniasis; (ii) persistence of positive antibody titers over long periods, even after treatment; (iii) low sensitivity in immunocompromised patients; (iv) expensive infrastructure and advanced technical skills required to conduct some these tests [5]
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