Diagnostic accuracy of direct agglutination test, rK39 ELISA and six rapid diagnostic tests among visceral leishmaniasis patients with and without HIV coinfection in Ethiopia.

Mekibib Kassa,Bruno C Bremer Hinckel,Johan Van Griensven,Pascal Mertens,Arega Yeshanew,Wim Adriaensen,Saïd Abdellati,Wasihun Hailemichael,Florian Vogt,Lieselotte Cnops,Dorien Van Den Bossche,Ermias Diro,Zvi Bentwich

doi:10.1371/journal.pntd.0008963

Abstract

Diagnosis of a first-time visceral leishmaniasis (VL) infection in Ethiopia is established by use of a rapid diagnostic test (RDT) detecting antibodies against rK39, direct agglutination test (DAT) and microscopy according to the national algorithm. The performance of individual tests and algorithm is variable and depends on several factors, one being HIV status. Limited data are available on the performance of tests in VL-HIV coinfected patients. Assessment of the performance of DAT (ITM-A), rK39 ELISA (Serion) and six RDT (Onsite Leishmania Ab CTK, Antigen ICT Xinjier, IT Leish Biorad, Kalazar Detect Inbios, rK39 IgG1 Coris, rk28 IgG1 Coris) for the diagnosis of VL was done on a panel of 91 stored serum and plasma samples of 'first-episode' suspected VL patients, with HIV coinfection (n = 51) and without (n = 40). A combined reference standard was used: either positive microscopy on tissue aspirates, or in case of negative microscopy, positive PCR results on the aspirate slide. Additionally, endemic healthy controls (n = 20), non-endemic controls (n = 10) and patients with confirmed malaria infection (n = 10) were tested for specificity evaluation. Sensitivities ranged from 69.2% for DAT (applied cut-off ≥ 1/3200) to 92.2% for the Onsite RDT, whereas specificities ranged from 20.0% for Kalazar Antigen ICT to 100% for IT Leish and rK39 IgG1. Sensitivities from all assays decreased upon stratification according to HIV status but was only significantly different for rK39 Serion ELISA (p-value 0.0084) and the Onsite RDT (p-value 0.0159). In conclusion, performance of commercially available assays for VL on samples from Northern-Ethiopian patients varied widely with a substantial decrease in sensitivity in the VL-HIV coinfected group. Clear guidelines on minimal performance criteria of individual tests and algorithms are needed, as well as which reference standard should be used to determine the performance.

Highlights

Leishmaniasis presents as a spectrum of diseases and is caused by obligate intracellular protozoa of the genus Leishmania which is transmitted by sand flies of the genus Phlebotomus
In northwest Ethiopia the rates of visceral leishmaniasis (VL) patients coinfected with HIV ranges from 20 to 40%
Limited data are available on the sensitivity and specificity of antibody and antigen detection tests for VL diagnosis in HIV coinfected

Summary

Introduction

VL is caused by members of the L. donovani complex, the closely related species L. infantum and L. donovani, the latter being responsible for kala-azar in East Africa. Clinical diagnosis of VL is inaccurate and can be difficult in endemic settings as several causes of febrile splenomegaly exist, notably malaria [3]. A direct parasitological diagnosis of VL is obtained through microscopy, culture and PCR on tissue aspirates. Molecular techniques have been shown to have the highest diagnostic sensitivity in aspirate or tissue samples, and to be highly specific. Alternative diagnosis by urinary antigen detection or antibody detection through direct agglutination test (DAT), immunofluorescence assays (IFA) and enzyme-linked immunosorbent assays (ELISA) require experienced laboratory technicians, specific laboratory equipment and is time consuming. Rapid diagnostic tests (RDTs) for Leishmania antibody detection have been implemented in endemic settings for over a decade. Modest improvement in sensitivity of rK28 RDTs over rK39 RDTs was reported in East Africa [11]

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