Comparative efficacy of recombinant VP6 protein based in-house Latex Agglutination test with other diagnostic assays for detection of Rotavirus A from calves, piglets and children

Abstract

In this study, VP6 protein of rotavirus A (RVA) was expressed in the prokaryotic system for the development of indigenous Latex Agglutination Test for diagnosis of RVA gastroenteritis in animals. Polyclonal anti-rVP6 IgG were raised in rabbits; purified and conjugated to carboxylated beads via covalent coupling for development of in-house LAT. Clinical utility of in-house developed LAT was evaluated on 313 stool samples collected from calves, children and piglets of Bareilly, Uttar Pradesh, India. In-house LAT yielded consistent results at weekly intervals for 2 months. Best visual perception of agglutination was observed when 5 μL beads coupled with 200 μg anti-rVP6 IgG and 10 μL of antigen reacted within reaction time of 2 minutes. Relative sensitivity and specificity of in-house LAT w.r.t RT-PCR was 65.45 % and 95.73 %, respectively; w.r.t commercial LFA was 75.55 % and 95.14 %, respectively and w.r.t RNA-PAGE was 70.27 % and 92.39 %, respectively. The kappa agreement between LAT and RT-PCR was 0.65 (substantial); between LAT and LFA was 0.7 (substantial) and between LAT and RNA-PAGE was 0.56 (moderate). Overall RVA stool positivity from children, calves and piglets with 4 assays was found to be 40 % (40/100), 9 % (9/100) and 16.81 % (19/113), respectively. Higher positivity was recorded in male (45.90 %, 28/61) than in female (30.76 %, 12/39) children. Developed LAT has fulfilled the WHO criteria for point-of-care testing with satisfactory efficacy for detection of RVA. This may serve as a preliminary assay for epidemiological surveillance of RVA antigen in determining rotavirus outbreaks in animal herds under resource-poor settings.