Cytochrome b and Molecular Typing of Leishmania spp. in a Passive Sampling of Suspected Patients with Cutaneous Leishmaniasis in Sistan and Baluchestan Province, Eastern Iran

Abstract

Background: Despite the high prevalence and drug resistance of disease in Sistan and Baluchestan Province of Iran, the species of cutaneous leishmaniasis (CL) has not been identified. In the present study, cytochrome b (Cyt b) was used in Sistan and Baluchestan to find species of Leishmania in suspected patients of CL using PCR-RFLP and DNA sequencing. Methods: This study was conducted from Oct 2015 to Oct 2016. The samples were collected from the individuals clinically suspected to CL and referred to Iran Shahr, Chabahar, Khash, Zabol, Zahedan, Mirjaveh, and Nikshahr health centers. Overall, 700 Giemsa-Stained slides from the wound of patients suspected of CL were passive collected and examined under a light microscope at×1000. After DNA extraction, positive samples were used for Cyt b detection by PCR-RFLP to determine the parasite species. One hundred positive slides were selected for molecular studies. Among positive samples, 20% were sequenced. To compare the results of sequences, molecular evolutionary genetic analysis (MEGA6) was used. Results: Overall, 53 samples were identified as L.‎ major and 47 samples (47%) L. tropica. Cyt b in L. major and L. tropica is converted to 400 and 480 bp and 130, 215 and 535 bp pieces respectively. In the isolated L. tropica and L. major, nucleotide changes were 3-5 (mainly in wobble site). Conclusion: Infection was more related to L. major. PCR-RFLP method has a high sensitivity for diagnosis of Leishmania species.

Keywords

• Leishmania major
• Leishmania tropica
• Cytochrome b
• PCR
• DNA sequencing
• Iran