Abstract
Visceral leishmaniasis (VL), one of the most important neglected tropical diseases, is caused by Leishmania donovani eukaryotic protozoan parasite of the genus Leishmania, the disease is prevalent mainly in the Indian sub-continent, East Africa and Brazil. VL can be diagnosed by PCR amplifying ITS1 and/or kDNA genes. The current study involved the optimization of Loop-mediated isothermal amplification (LAMP) for the detection of Leishmania DNA in human blood or tissue samples. Three LAMP systems were developed; in two of those the primers were designed based on shared regions of the ITS1 gene among different Leishmania species, while the primers for the third LAMP system were derived from a newly identified repeated region in the Leishmania genome. The LAMP tests were shown to be sufficiently sensitive to detect 0.1pg of DNA from most Leishmania species. The green nucleic acid stain SYTO16, was used here for the first time to allow real-time monitoring of LAMP amplification. The advantage of real time-LAMP using SYTO 16 over end-point LAMP product detection is discussed. The efficacy of the real time-LAMP tests for detecting Leishmania DNA in dried blood samples from volunteers living in endemic areas, was compared with that of qRT-kDNA PCR.
Highlights
The leishmaniasis are a group of diseases, caused by eukaryotic protozoan parasites of the genus Leishmania, transmitted by blood-sucking phlebotomine sand flies
ITS1 Sequences from different Leishmania species selected from GenBank, were aligned in order to identify the DNA sequences shared by all Leishmania species and, suitable for Loop-mediated isothermal amplification (LAMP) primers
In this study we evaluated the efficacy of several newly developed LAMP assays for detecting low concentrations of Leishmania DNA in biological samples
Summary
The leishmaniasis are a group of diseases, caused by eukaryotic protozoan parasites of the genus Leishmania, transmitted by blood-sucking phlebotomine sand flies. The most commonly targeted genes are the ribosomal ITS1 and the kinetoplast minicircle genes (kDNA) (el Tai et al, 2000; Nicolas et al, 2002; Rodgers et al, 1990; van Eys et al, 1992). These PCR assays have been developed for the detection of Leishmania DNA in a variety of clinical samples such as skin biopsies and smears, bone marrow and lymph node aspirates as well as peripheral blood. There is no generallyaccepted gold standard for Leishmania identification (Miller et al, 2014). Van der Auwera and Dujardin (2015), gave a comprehensive literature analysis for the different available and accepted methods used for Leishmania species detection in clinical and epidemiological studies
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
