Use of the recombinant k39 immunochromatographic strip-test for serological diagnosis of visceral leishmaniasis: experience from Apulia, southern Italy

Abstract number: P944

Monno R., Bruno R., Di Molfetta E., Bottalico P., Rizzo C., De Vito D., Fumarola L.

Objective: Visceral leishmaniasis (VL) is endemic in our area, Apulia, Southern Italy. The aim of this study was to evaluate a commercially available membrane-based immunoassay (MBI) using a recombinant protein rk39 as antigen in comparison to the immunofluorescent antibody assay (IFA) for the detection of specific anti-Leishmania antibodies.

Methods: A total of 264 individuals including 19 patients with VL (3 of whom HIV-positive), 67 with suspected VL and 40 healthy controls were tested. VL diagnosis was made by microscopic demonstration of Leishmania amastigotes in Giemsa stained smears from bone marrow aspirates and/or on the presence of specific antibodies. In a HIV-positive patient diagnosis was performed by the PCR on peripheral blood. IFA and MBI were performed in all 264 sera according to manufacturer's instructions.

Results: The MBI was positive in all the 19 patients with VL (diagnosed by the presence of antibodies by IFA and/or the presence of amastigotes in bone marrow aspirate) and negative in sera from the remaining individuals. The sensitivity and specificity of MBI was 100%, respectively. The IFA failed to detect anti-Leishmania antibodies in the first serum sample of a HIV-positive patient but antibodies were detected in a second serum sample obtained 7 days later. On this patient the MBI was positive on the first serum sample tested. The MBI works well also on HIV–Leishmania co-infected patients in whom low yield of serological studies may be a most characteristic finding. Follow up performed on 7 patients with VL showed that both antibodies to Leishmania detected by IFA and MBI remain at detectable levels up to 12–24 months. A dilution trial performed on sera of 2 patients showed that a positive reaction by MBI was detectable at serum dilution up to 20,480, indicating that a strong immuno-response is mounted against the rk39 antigen.

Conclusions: The MBI is a reliable test for diagnosis of leishmaniasis and is not only a good alternative in a setting where a reliable enzyme immunoassay or IFA tests are not available, but also in well equipped laboratories because positive results can be obtained in 10 minutes. The cost of MBI in our experience was approximately 8.43 euro per test, compared with approximately 12 euro for IFA. MBI appears highly sensitive, specific, rapid and cost-effective for the serological diagnosis of VL in our location where VL has a low endemicity.