Polymorphism in the HASPB Repeat Region of East African Leishmania donovani Strains

Arie Zackay,Zewdu Hurissa,Charles L Jaffe,Ermias Diro,Asrat Hailu,Teklu Weldegebreal,Dagimawie Tadesse,Abdelmajeed Nasereddin,Sisay Yifru,Yegnasew Takele,Aysheshm Kassahun,Workagegnehu Hailu,Paul Andrew Bates

doi:10.1371/journal.pntd.0002031

Abstract

Background/ObjectivesVisceral leishmaniasis (VL) caused by Leishmania donovani is a major health problem in Ethiopia. Parasites in disparate regions are transmitted by different vectors, and cluster in distinctive genotypes. Recently isolated strains from VL and HIV-VL co-infected patients in north and south Ethiopia were characterized as part of a longitudinal study on VL transmission.Methodology/Principal FindingsSixty-three L. donovani strains were examined by polymerase chain reaction (PCR) targeting three regions: internal transcribed spacer 1 (ITS1), cysteine protease B (cpb), and HASPB (k26). ITS1- and cpb - PCR identified these strains as L. donovani. Interestingly, the k26 - PCR amplicon size varied depending on the patient's geographic origin. Most strains from northwestern Ethiopia (36/40) produced a 290 bp product with a minority (4/40) giving a 410 bp amplicon. All of the latter strains were isolated from patients with HIV-VL co-infections, while the former group contained both VL and HIV-VL co-infected patients. Almost all the strains (20/23) from southwestern Ethiopia produced a 450 bp amplicon with smaller products (290 or 360 bp) only observed for three strains. Sudanese strains produced amplicons identical (290 bp) to those found in northwestern Ethiopia; while Kenyan strains gave larger PCR products (500 and 650 bp). High-resolution melt (HRM) analysis distinguished the different PCR products. Sequence analysis showed that the k26 repeat region in L. donovani is comprised of polymorphic 13 and 14 amino acid motifs. The 13 amino acid peptide motifs, prevalent in L. donovani, are rare in L. infantum. The number and order of the repeats in L. donovani varies between geographic regions.Conclusions/SignificanceHASPB repeat region (k26) shows considerable polymorphism among L. donovani strains from different regions in East Africa. This should be taken into account when designing diagnostic assays and vaccines based on this antigen.

Highlights

Parasites belonging to the Leishmania donovani complex, L. donovani and L. infantum, are the main causative agents of visceral leishmaniasis (VL), known as kalaazar
In East Africa and India, VL is primarily caused by L. donovani, and believed to be an anthroponosis, while in other regions, where VL is caused by L. infantum, this disease is a zoonosis with dogs and wild canids acting as reservoir hosts [4]
The L. infantum/L. donovani protein has been used for diagnosis of visceral leishmaniasis, and is a putative vaccine candidate for this disease

Summary

Introduction

Parasites belonging to the Leishmania donovani complex, L. donovani and L. infantum (synonym = L. chagasi), are the main causative agents of visceral leishmaniasis (VL), known as kalaazar This disease is invariably fatal if not properly diagnosed and treated. Northwestern Ethiopia (NW) accounts for ,60% of the VL cases [3], and a majority of the HIV - VL co-infections, with the disease focused in the Metema - Humera region near the Sudanese border. This is a semi-arid region, with extensive commercial monoculture, and scattered Acacia - Balanite forests.

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