Development of dipstick enzyme linked immunosorbent assay for on-site sero-diagnosis of Japanese encephalitis in swine.

Japanese encephalitis (JE) is an important viral zoonotic disease in Asia, especially in rural and suburban areas where rice cultivation and pig farming coexist. Pigs serve as a suitable sentinel model, the surveillance of which could predict a potential JE outbreak in human population in the immediate vicinity. However, existing diagnostics like ELISA and VNT require sophisticated laboratory facilities which are more often not available in field conditions. In the present study, we aimed at developing recombinant non-structural (NS1) protein-based dipstick IgG ELISA as an on-site assay for sero-diagnosis of JE in swine. The assay was standardized by optimizing various parameters and the following conditions were found to be ideal including 1 µg of rNS1 protein in carbonate buffer per strip of nitrocellulose membrane comb; bovine serum albumin as blocking agent at 4 °C overnight; serum dilution of 1:10 and conjugate dilution of 1:5000 in skimmed milk powder. Relative diagnostic sensitivity and specificity of dipstick IgG ELISA was 100% and 92.9%, respectively. The dipstick assay was validated in three laboratories as per OIE guidelines. The storage life of dipstick was up to 7 months at 4 °C. The assay is easy to perform and the results can be interpreted with visual observation that precludes the need for absorbance reading equipment. The standardized dipstick assay was found promising for screening swine serum samples in field conditions. Timely detection of JE virus in swine will aid in predicting the outbreak in humans and thus in taking suitable preventive and control measures.