Characterization of RKZ isolate of ovine herpesvirus 1

Cytopathic effect (CPE) characterized mainly by foci of rounded cells was o bserved in cultures of primary plexus choroideus cells from healthy lamb fo llowing cryopreservation. it was possible to transmit the infectious agent to other primary cells of ovine origin by co-cultivation with infected cell s. By indirect immunofluorescence microscopy it was found that high percent age of sheep (65-80% in 3 different herds from Slovakia) are infected with this infectious agent. Electron microscopy of cells with CPE revealed the p resence of herpesvirus particles. Viral DNA was isolated from infected cell s using pulse-field gel electrophoresis and further used as probe in Southe rn blot analysis. The probe reacted specifically only with DNA from cells i nfected with Ovine herpesvirus 1 (OvHV-1) but not with DNA of other ruminan t herpesviruses. Some of the HindIII restriction fragments of DNA of the ob tained OvHV-1 isolate denominated RKZ were cloned. Part of the H9 clone was sequenced identifying a gene that encoded a polypeptide homologous to cons erved herpesvirus VP23 structural protein. From comparison of the sequence of this clone with VP23 sequences of other herpesviruses it was deduced tha t OvHV-1 might be classified within the Rhadinovirus genus of the Gammaherp esvirinae subfamily. The sequencing of the H9 clone of DNA of RKZ isolate e nabled establishment of sensitive and highly specific polymerase chain reac tion (PCR) assay for detection of OvHV-1.