Immuno-molecular study of cutaneous leishmaniasis: an alarming and emerging disease of Nepal

Abstract

Cutaneous Leishmaniasis is a vector borne, parasitic disease caused by the bite of an infected sand fly. The disease is rare in Nepal with only few cases reported till date. The diagnosis of CL might be difficult, in particular in areas where the disease is uncommon and where other skin diseases with similar clinical symptoms occur. The main objective of the study is to improve the diagnosis of CL and the identification of causal Leishmania species and studying the immune response of the patients. A total of 17 patients presenting with cutaneous lesions suggestive of CL were sampled for parasitological diagnosis by direct examination (DE), kinetoplast DNA (kDNA) nested PCR (CSB1X/CSB2X and 13Z/LiR primers). Immunological status was analyzed by flow cytometer and compared with healthy controls. The data were statistically analyzed using graph pad prism ver.7. A total of 17 patients with age ranging from 8 years to 85 years were included in the study. Most patients were seen from January to September and had 1-3 lesions (duration = 3 to > 12 months) that involved mainly the face (64.70%). Mean age was 36 ± 22.05 years. Most patients were in the age group 21-40 years (35.29%) followed by ≤20 years (29.41%). Male: Female ratio was 1.4:1. LD bodies were observed in (11.76%) of the cases. Out of 17 cases only 5.88% had travel history outside Nepal in Leishmaniasis endemic country. PCR positive cases was 58.82%. The diagnostic criteria of CL in KDNAPCR werebased on observation of 720bp, 680bp and 560bp for Leishmania donovani complex, Leishmania infantum and Leishmania major respectively. Immunological study showed significant difference in CD8+ T cells in CL patients with active lesions but not in CD4+ T cells and in B cells between the CL patients and the healthy controls. Cutaneous leishmaniasis is not common in Nepal. So, it is often neglected and misdiagnosed. It is in an increasing trend. The PCR-based assays used increased the speed and sensitivity of the diagnosis of CL as well as in species identification compared to the conventional techniques.

Key words: Cutaneous leishmaniasis, kinetoplast DNA, flow cytometer, Leishmania, nested PCR.