Use of PCR-Restriction Fragment Length Polymorphism Analysis To Identify the Main New WorldLeishmaniaSpecies and Analyze Their Taxonomic Properties and Polymorphism by Application of the Assay to Clinical Samples

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Abstract
At least 13 characterizedLeishmaniaspecies are known to infect humans in South America. Five of these parasites are transmitted in the sylvatic ecotopes of the whole French Guianan territory and responsible for cutaneous leishmaniasis. For the diagnosis of cutaneous leishmaniasis, restriction fragment length polymorphism (RFLP) analyses have shown promising results. Thus, the end of the small subunit and internal transcribed spacer 1 of the rRNA genes were sequenced and targeted by PCR-RFLP analysis in the 10 main New World (NW)Leishmaniaspecies from the two subgenera. Then, the procedure was tested on 40 samples from patients with cutaneous leishmaniasis, and its results were compared with those of conventional methods. (i) The results of this simple genus-specific method were in agreement with those of previous isoenzyme analyses. (ii) This method distinguished the most medically relevantLeishmaniaspecies with only one enzyme (RsaI). (iii) This method could be performed directly on human biopsy specimens (sensitivity of 85.7%). Performing NWLeishmaniaspecies typing rapidly and easily in the field constitutes a very valuable improvement for detection ofLeishmaniaspp. Revealing great diversity with several enzymes, this method could also be useful for taxonomic, ecological, and epidemiological studies in space and time.