Molecular Tools for Diagnosis of Visceral Leishmaniasis: Systematic Review and Meta-Analysis of Diagnostic Test Accuracy

Abstract

e Molecular methods have been proposed as highly sensitive tools for the detection of Leishmania parasites in visceral leishmaniasis (VL) patients. Here, we evaluate the diagnostic accuracy of these tools in a meta-analysis of the published literature. The selection criteria were original studies that evaluate the sensitivities and specificities of molecular tests for diagnosis of VL, adequate classification of study participants, and the absolute numbers of true positives and negatives derivable from the data presented. Forty studies met the selection criteria, including PCR, real-time PCR, nucleic acid sequence-based amplification (NASBA), and loop-mediated isothermal amplification (LAMP). The sensitivities of the individual studies ranged from 29 to 100%, and the specificities ranged from 25 to 100%. The pooled sensitivity of PCR in whole blood was 93.1% (95% confidence interval [CI], 90.0 to 95.2), and the specificity was 95.6% (95% CI, 87.0 to 98.6). The specificity was significantly lower in consecutive studies, at 63.3% (95% CI, 53.9 to 71.8), due either to true-positive patients not being identified by parasitological methods or to the number of asymptomatic carriers in areas of endemicity. PCR for patients with HIV-VL coinfection showed high diagnostic accuracy in buffy coat and bone marrow, ranging from 93.1 to 96.9%. Molecular tools are highly sensitive assays for Leishmania detection and may contribute as an additional test in the algorithm, together with a clear clinical case definition. We observed wide variety in reference standards and study designs and now recommend consecutively designed studies. Visceral leishmaniasis (VL), or kala-azar, is a vector-borne disease that is caused by the protozoan parasites Leishmania donovani and L. infantum. The disease is transmitted to humans by the bite of infected phlebotomine sandflies. VL is a neglected tropical disease with an estimated 200,000 to 400,000 new cases and 20,000 to 40,000 deaths annually (1). More than 90% of all VL cases occur in Bangladesh, Brazil, India, Ethiopia, Sudan, and South Sudan. The clinical picture consists of fever, weight loss, fatigue, and general weakness; patients may present with enlarged lymph nodes, hepatomegaly, and splenomegaly. As VL is a fatal condition when left untreated and treatments have high toxicity, a diagnostic test that is both highly sensitive and specific is required. The gold standard for diagnosing VL is mainly the demonstration of parasites by microscopic examination of cultures of splenic aspirates. Splenic aspirates can be associated with hemorrhage, and the process should be carried out only with access to surgical facilities. For this reason bone marrow and lymph node aspirates are commonly taken for parasitological diagnosis. The specificity of these methods is high, but the sensitivity varies depending on the type of specimen, i.e., approximately 93 to 99% for the spleen, 53 to 86% for bone marrow, and 53 to 65% for lymph (2). In addition, the sensitivity and specificity of parasitological testi...

Health-seeking behaviour, diagnostics and transmission dynamics in the control of visceral leishmaniasis in the Indian subcontinent

“…These tests, along with standard polymerase chain reaction, are able to detect circulating DNA in the blood of individuals who are actively infected. Studies show that molecular tests are very sensitive, and although they have low specificity, it may be that these are infections that are below the limit of detection of the reference standard, and therefore in fact true positives 4 . Diagnostics that are able to detect asymptomatic infection and PKDL may also be important, depending on the relative role of transmission in these groups.…”

“…This definition is used partly because of the low specificity of the rK39 test to identify infection rather than exposure, and to differentiate kala-azar from other fever-causing aetiologies. For the control of kala-azar, it has been proposed that a diagnostic test that detects active infection rather than the immune response to infection could be used 4 . Testing earlier could identify more patients and interrupt transmission by early treatment.…”

Budgetary impact of diagnostic tests for visceral leishmaniasis in Brazil

“…Furthermore, the procedure is invasive, and its sensitivity is 77% 3 . Molecular diagnosis using standard polymerase chain reaction (PCR) may be less invasive than parasitological examination of bone marrow, as it presents high performance in whole blood (sensitivity of 93.1% and specificity of 95.6%), according to a meta-analysis study by Ruiter et al 7 . However, it also requires a complex infrastructure and specialized professionals and, thus, the technique is restricted to reference centres.…”

Diagnosing Neglected Tropical Diseases in HIV Coinfection

“…Causes of false-negative diagnosis may be low parasite load, lack of a well-trained expert to identify the parasites, and delayed diagnosis. [7][8][9] Another limitation of microscopic methods is the identification of species of the causal agent, which is not possible for leishmaniasis and Chagas disease but possible for malaria, despite high error rates. 10 Other diagnostic methods have improved aggregate sensitivity and specificity relative to conventional techniques.…”