Development and Validation of Point of Care Diagnostics for the Rapid Detection of Multiple Species of Leptospira at Resource-Limited Areas

Leptospirosis is a life threatening, emerging, infectious zoonotic disease of humans and livestock all over the world. The diagnosis of this disease is frequently ineffective. Conventional methods of diagnosis, gold standard bacterial culture and microscopic agglutination test (MAT) are being used. These tests are time-consuming and the test for the disease can be positive more than 2 weeks after the onset of the disease. Present study was taken up to circumvent the problems and disadvantages in the methods/protocols being used currently in leptospirosis diagnosis. This study aimed to develop a point of care lateral flow (LFA) and indirect ELISA assays to detect disease-specific antibodies in the whole blood/serum/plasma for the rapid diagnosis of acute infection. The Leptospira specific lipopolysaccharide (LPS) was used in combination with recombinant multi-epitope membrane protein as an antigen candidate for the detection of disease specific antibodies. The developed lateral flow (LFA) and indirect ELISA assays were compared with gold standard MAT and commercial ELISA kit. The MAT confirmed standard reference sera were used for validation of developed assays. A total of 223 positive and 115 non-positive samples for leptospirosis were used in the present validation. Indirect ELISA assay developed in the present study showed higher sensitivity and specificity of 98.65% and 100% respectively. In contrast, the sensitivity and specificity of the commercial ELISA and LFA were 96.41%, 97.39%, 87.44% and 98.26% respectively. The results obtained proved that LFA and indirect ELISA assays were more effective than the conventional methods for the diagnosis of acute leptospirosis, especially within the onset and useful for the point of care diagnosis at resourcelimited areas.