Development of a recombinant glycoprotein E-based ELISA for detection of antibodies against Bovine alphaherpesvirus 1: a DIVA compliant.

Infectious bovine rhinotracheitis/infectious pustular vulvovaginitis (IBR/IPV) caused by Bovine alphaherpesvirus-1 belongs to the family Orthoherpesviridae subfamily Alphaherpesvirinae, and genus Varicellovirus. The use of IBR marker vaccines necessitates the development of diagnostic assays targeting glycoprotein E (gE) to differentiate infected animals from vaccinated ones. In the present study, the N-terminal one-third region of the gE gene was cloned and expressed in a prokaryotic expression system. The recombinant gE protein (~ 25 kDa) was confirmed to be immunogenic by western blot analysis using bovine hyperimmune serum. The purified recombinant protein was subsequently utilized for the development of an indirect ELISA. A total of 456 bovine serum samples, comprising 206 gE-positive and 250 gE-negative as determined by the IDEXX IBR gE ELISA, were employed for standardization of the in house rgE ELISA. Statistical analysis identified an optimal cut-off value of 45.92 percent positivity (PP), at which the assay demonstrated a diagnostic sensitivity of 84.9% and specificity of 91.2%. The area under the ROC curve was 0.914 at 95% confidence interval, indicating assay could be used for mass screening.