title: Preparation of recombinant African horse sickness virus VP7 antigen via a simple method and validation of a VP7-based indirect ELISA for the detection of group-specific IgG antibodies in horse sera.
abstract: This paper describes the production and purification of a group-specific recombinant protein VP7 of African horse sickness virus serotype 3 (AHSV-3) and validation of an I-ELISA for the detection of IgG-antibodies to VP7 in horse sera. Baculovirus-expressed VP7 crystals were purified from infected insect cells. Analytical accuracy of the I-ELISA was examined using sera (n = 38) from an experimentally infected horse, from foals born to vaccinated mares, from guinea-pigs immunized with nine serotypes of AHSV, and from sera of animals infected with other orbiviruses. Compared to traditional serological assays, the I-ELISA was more sensitive in detection of the earliest immunological response in an infected horse and declining levels of maternal immunity in foals. Antibodies to all nine serotypes of AHSV could be detected. Cross-reactivity to related orbiviruses was not observed. Diagnostic accuracy of the I-ELISA was assessed by testing sera from vaccinated horses (n = 358) residing in AHS-enzootic areas and from unvaccinated horses (n = 481) residing in an AHS-free area. Sera were categorised as positive or negative for antibodies to AHSV using virus neutralisation tests. The TG-ROC analysis was used for the selection of the cut-off value. At a cut-off of 11.9 of the high positive control serum (percentage positivity), the I-ELISA specificity was 100%, sensitivity 99.4%, and the Jouden index was 0.99.
authors.affiliation: Department of Biochemistry, Onderstepoort Veterinary Institute, P/Bag X 5, Onderstepoort, Onderstepoort 0110, South Africa. marees@arc.agric.za
authors.forename: Sonja
authors.initials: S
authors.lastname: Maree
authors.forename: Janusz T
authors.initials: JT
authors.lastname: Paweska
mesh_terms: D000355:African Horse Sickness / Q000175:diagnosis* / Q000276:immunology*; D000356:African Horse Sickness Virus / Q000276:immunology*; D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000276:immunology* / Q000302:isolation & purification; D016367:Baculoviridae / Q000235:genetics; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005822:Genetic Vectors; D006736:Horses; D007112:Immunity, Maternally-Acquired; D007074:Immunoglobulin G / Q000097:blood; D009500:Neutralization Tests; D011786:Quality Control; D011994:Recombinant Proteins; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014758:Viral Core Proteins / Q000276:immunology* / Q000302:isolation & purification; D014765:Viral Vaccines / Q000276:immunology
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D014758:Viral Core Proteins; D014765:Viral Vaccines; C069264:VP7 major core antigen, African horsesickness virus