Characterization of Ovine herpesvirus 2 putative glycoprotein Ov8
Alhajri, Salim Mohamed
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Malignant catarrhal fever (MCF) is a lymphoproliferative usually fatal disease of ungulates caused by several gammaherpesviruses in the genus Macavirus including alcelaphine herpesvirus 1 (AlHV-1) and ovine herpesvirus 2 (OvHV-2). There is no treatment or vaccine for the disease. MCF is diagnosed by clinical history, histopathology, polymerase chain reaction and serology. However, all current serological assays utilize AlHV-1 antigens thus none are specific for detecting anti-OvHV2 antibodies. The lack of cell culture system to propagate OvHV-2 has hindered studies of OvHV-2 biology including its entry mechanism and in making of an OvHV-2 specific antibody detection assay. As an alternative method to study OvHV-2 glycoproteins responsible for membrane fusion as a part of the entry mechanism, we developed a virus-free cell-to-cell membrane fusion assay to identify the minimum required OvHV-2 glycoproteins to induce membrane fusion. OvHV-2 glycoproteins gB, gH and gL could induce membrane fusion together but not individually. Cell-cell membrane fusion was detected by both fluorescence microscopy and reporter gene expression. Additionally, open reading frame Ov8, unique to OvHV-2, was found to encode a transmembrane glycoprotein that can significantly enhance membrane fusion. These results are described in chapter 1. To overcome the lack of cell culture to grow OvHV-2, a recombinant fusion protein containing part of the OvHV-2-specific Ov8 glycoprotein was expressed and evaluated in an enzyme linked immunosorbent assay (ELISA). Using sheep serum samples, the assay showed relatively 100% sensitivity, 98.97% specificity, 99.07% positive predictive value, and 100% negative predictive value. The assay also showed very good agreement with a competitive inhibition (CI)-ELISA (kappa= 0.990 and 95% CI=0.971 - 1.000) that detects an epitope conserved among all MCF viruses. Using samples from bison with MCF, both the CI-ELISA and Ov8 ELISA showed 58.82% sensitivity and100% specificity. Importantly, 31.5% of samples not detected by CI-ELISA were detected in Ov8 ELISA. These results show that Ov8 ELISA has potential as an OvHV-2 specific diagnostic assay.