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Background: Cutaneous leishmaniasis (CL) is a vector-borne parasitic disease characterized by the presence of one or more lesions on the skin that usually heal spontaneously after a few months. Most cases of CL worldwide occur in Southwest Asia, Africa and South America, and a number of cases have been reported among troops deployed to Afghanistan. No vaccines are available against this disease, and its treatment relies on chemotherapy. The aim of this study was to characterize parasites isolated from Canadian soldiers at the molecular level and to determine their susceptibility profile against a panel of antileishmanials to identify appropriate therapies., Methodology/principal Findings: Parasites were isolated from skin lesions and characterized as Leishmania tropica based on their pulsed field gel electrophoresis profiles and pteridine reductase 1 (PTR1) sequences. Unusually high allelic polymorphisms were observed at several genetic loci for the L. tropica isolates that were characterized. The drug susceptibility profile of intracellular amastigote parasites was determined using an established macrophage assay. All isolates were sensitive to miltefosine, amphotericin B, sodium stibogluconate (Pentostam) and paromomycin, but were not susceptible to fluconazole. Variable levels of susceptibility were observed for the antimalarial agent atovaquone/proguanil (Malarone). Three Canadian soldiers from this study were successfully treated with miltefosine., Conclusions/significance: This study shows high heterogeneity between the two L. tropica allelic versions of a gene but despite this, L. tropica isolated from Afghanistan are susceptible to several of the antileishmanial drugs available.