Methods incorporating a polymerase chain reaction and restriction fragment length polymorphism and their use as a ‘gold standard’ in diagnosing Old World cutaneous leishmaniasis

Abstract

Three polymerase chain reaction (PCR) assays — kinetoplast DNA (kDNA) PCR, 7SL PCR, ITS1 PCR — were compared for their ability to detect leishmanial parasites in the skin tissue aspirates of 212 Palestinian suspect cases of cutaneous leishmaniasis (CL). The kDNA PCR was the most sensitive, detecting 156/170 (91.8%) of positive samples confirmed by a set ‘gold standard’ but of poor specificity in identifying the species of Leishmania. The 7SL PCR detected 154/170 (90.5%) and the ITS1 PCR only 108/170 (63.5%) of the true positives, and both were 100% specific. The highest Kappa coefficient agreement (95% CI) was obtained with the 7SL PCR (0.792 ± 0.098). Restriction analysis of the products from the ITS1 PCR and 7SL PCR enabled identification of species of Leishmania. L. tropica was the predominant cause of CL, with L. major being less frequent.