BackgroundDomestic dogs are the principal reservoir hosts of Leishmania infantum in regions where visceral leishmaniasis (VL) is endemic. Although serologic methods are frequently used for the screening of infected dogs, antibody-based tests require further assessment, due to lack of sensitivity and specificity. In this study, we employed a multi-antigen printing immunoassay (MAPIA) to compare the antibody responses to novel recombinant proteins of L. infantum with the potential for the detection of canine VL.FindingsMAPIA strips were prepared employing 12 recombinant proteins. Antibody reactivity to these antigens was compared using a panel of sera collected from clinically asymptomatic (n = 16) and symptomatic (n = 41) culture-positive animals. Our findings showed that the canine immune response to antigen differs between dogs and depends on infection status. Using this screening assay, when five out of the 12 antigens were combined, an overall 81% detection rate of L. infantum-infected dogs was achieved.ConclusionsWe conclude that MAPIA is an effective screening tool to rapidly select multiple antigens of diagnostic utility to be used in a more sensitive point of care diagnostic test such as the Dual-Path Platform (DPP) multiplex test for the rapid detection of infected dogs.
Background
Visceral leishmaniasis (VL) is a zoonotic disease caused by Leishmania infantum, for which dogs constitute the main urban parasite reservoir. Control measures and the treatment of canine visceral leishmaniasis (CVL) are essential to reduce VL cases. Early and accurate detection of L. infantum-infected dogs is crucial to the success of VL control. To improve the serological detection of L. infantum-exposed dogs, we evaluated the early diagnosis capacity of a recombinant protein (rLci5) in an immunosorbent assay (ELISA) to detect naturally infected dogs. Additionally, we evaluated the persistence of the positive results obtained by rLci5 ELISA in comparison to other conventional diagnostic test methods.
Methods
Serum samples obtained from 48 L. infantum-infected dogs involved in a cohort study were evaluated using different diagnostic methods (qPCR, EIE-LVC, DPP-LVC and splenic culture). The results were compared to rLci5 ELISA to determine its capacity to diagnose L. infantum infection at earlier infection time points. The persistence of positive diagnostic test results was also compared for each dog evaluated.
Results
rLci5 ELISA presented higher rates of positive results at early time points compared to the other diagnostic tests employed in the cohort study, as early as 24 months prior to detection by other tests. rLci5 ELISA positivity was 52.1% (25/48) at baseline, while qPCR was 35.4% (17/48), DPP-LVC 27.1% (13/48), EIE-LVC 22.9% (11/48) and culture only 4.2% (2/48). In at least one of the time points of the 24-month cohort study, rLci5 ELISA was positive in 100% (48/48) of the dogs, versus 83% (40/48) for qPCR, 75% (36/48) for DPP-LVC, 65% (31/48) for EIE-LVC and 31% (15/48) for culture. Investigating clinical signs in association with diagnostic test positivity, rLci5 ELISA successfully detected CVL in 62.9% (95/151) of the clinical evaluations with a score of 0–3, 64.3% (45/70) with scores between 4 and 7, and 73.7% (14/19) with scores > 7, providing higher rates of positivity than all other methods evaluated. Moreover, rLci5 ELISA presented the greatest persistence with respect to test positivity: 45.8% of the dogs evaluated.
Conclusion
Four diagnostic tests were compared to rLci5 ELISA, which presented earlier infection diagnosis and a greater persistence of positive test results. Accordingly, the use of the rLci5 ELISA can improve CVL diagnostic performance by detecting infected dogs sooner than other testing methods, with enhanced persistence of positive results over the course of the infection.
Graphic abstract
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