Background and Objective: Chabahar is in Southern Iran located near the Iran-Pakistan border. Since leishmaniasis is an emerging disease in this region, this study aimed to diagnose the disease and identify different species of Leishmania parasite in the patients referred to the central laboratory. Methods: This descriptive cross-sectional study was conducted in 2011-2012 on patients referred to the central laboratory in the city of Chabahar. The sampling of lesions, slide preparation, culture and PCR specific for kinetoplast DNA (kDNA), extracted from the media and slides, were performed. The data collected by a questionnaire were analyzed by the SPSS software. Results: The resulted bands from the 48 tested cutaneous leishmaniasis isolates were compared with the standard strains of Leishmania tropica, L. infantum and L. major. All 48 investigated bands were in the 620bp region, which is related to L. major. Conclusion: Since PCR has high sensitivity and specificity, it is recommended to use kDNA (present in a unique organelle called kinetoplast) for the routine diagnosis and treatment of the disease.