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.

The research focuses on creating a specific protein, known as VP7, for African horse sickness virus and confirming the efficiency of an enzyme-linked immunosorbent assay (ELISA) in detecting antibodies against this protein in horse blood samples. The method presented is highlighted for its increased sensitivity in detecting the virus.

Producing the VP7 Antigen

• The researchers used Baculovirus, a type of virus that specifically infects insects, to express the VP7 protein from the African horse sickness virus (AHSV).
• The protein was then purified from the infected insect cells, to provide a pure sample of the VP7 antigen for use in their tests.

Testing the I-ELISA

• Indirect ELISA (I-ELISA) was then used to detect the presence of IgG-antibodies against the VP7 protein in horse blood samples.
• The test was applied to a range of samples, including blood from a horse infected with AHSV, foals born to vaccinated mares, guinea-pigs immunized with all nine serotypes of AHSV, and animals infected with other related orbiviruses.
• The I-ELISA demonstrated high accuracy in its results, showing no cross-reactivity with related orbiviruses and successfully identifying antibodies of all nine serotypes of AHSV.

Evaluating the Sensitivity and Specificity

• The true effectiveness of the I-ELISA was assessed by its ability to differentiate between vaccinated horses in areas affected by AHSV and unvaccinated horses in an AHS-free zone.
• Traditional virus neutralisation tests were used to categorize the blood samples as either positive or negative for antibodies against AHSV.
• The statistical measure, TG-ROC, was used to determine the most effective cut-off value for the test. The researchers chose a cut-off of 11.9% of the high positive control serum.
• At this cut-off value, the I-ELISA showed a sensitivity of 99.4% and specificity of 100%. The Jouden index was 0.99, indicating great accuracy in the test’s results.

Conclusion

• The I-ELISA provided a sensitive, accurate, and reliable method for detecting antibodies against AHSV in horses.
• This could prove beneficial in managing AHSV infections, particularly in providing early detection and monitoring of the antibody levels in infected and recently vaccinated horses.