Ветеринария сегодня (Apr 2018)
DEVELOPMENT AND USE OF THE COMPETITIVE ELISA-BASED TEST SYSTEM FOR DETECTION OF ANTIBODIES TO GROUP A ROTAVIRUS IN LIVESTOCK SERA
Abstract
Test-system based on competitive enzyme-linked immunosorbent assay (ELISA) for detection of antibodies to Group A rotavirus in sera collected from livestock of various species (cattle, pigs) was developed. Bovine rotavirus TE87 strain of serogroup A adapted to continuous porcine embryo kidney cell culture was selected as an antigen for ELISA. The said strain has the infectivity titre of 7.0-7.5 lg TCID50/cm3. It is group-specific and the most technologically suitable for production antigen scaling-up. Specificity of prepared components, rotavirus antigen and hyperimmune rabbit serum, was confirmed by real-time PCR and immunoblotting. The development of competitive ELISA-based test-system included selection of optimal blocking buffer composition as well as optimal antigen dilution, specific hyperimmune serum and anti-species conjugate. Admissible optical density values of control sera and cut-off values were determined. The test-system was examined for its specificity and sensitivity as compared to those ones of INGENASA test-kit (Spain) by parallel testing of 203 cattle sera. Performed studies resulted in the development of sensitive (92%) and specific (94%) competitive ELISA enabling detection of antibodies against Group A rotavirus in cattle and porcine sera with 92.6% accuracy. A total of 2,259 cattle and porcine sera submitted from 48 farms located in various Russian Federation regions in 2016-2017 were tested with the developed test-system. Average proportion of Group A rotavirus-positive cattle and pigs was 42% and 84%, respectively.
