Biochemical characterisation of Australian orbiviruses

There are over 30 antigenically distinct orbiviruses found in Australia, including members in the bluetongue virus (BTV) and epizootic hemorrhagic disease of deer virus (EHDV) serogroups. Genomic RNA profiles were analysed by polyacrylamide gel electrophoresis (PAGE) on both 10% Laemmli and tris-borate-EDTA-(TBE)-urea gels. There was considerably more variation in the RNA profiles in Laemmli gels than was apparent in the TBE-urea gels. Since the latter system separates on molecular size, then presumably migration in the Laemmli gels may depend upon molecular weight (MW) and conformation. Analyses of 35S-methionine labeled proteins in virus-infected cells was carried out by PAGE in 10 to 20% gradient Laemmli gels. Twelve to 15 virus-specific labeled protein bands were observed in cells infected with orbiviruses. A detailed analysis of Australian BTV 1 isolates was made to identify these proteins. In addition to previously reported proteins (P1 to P8A) an additional low MW protein, P9, was observed (approx. MW 12,000). In the EHDV serogroup, 3 viruses (Ibaraki, CSIRo402 and CSIRo439), which were very closely related by virus neutralization tests and in protein-PAGE, were distinct in their migration by RNA-PAGE. Analyses of the individual RNA segments by 1-dimensional T1-ribonuclease oligonucleotide mapping showed minor differences between Ibaraki and the Australian EHDV isolates, suggesting that the 3 viruses have similar 3'-terminal RNA sequences. These studies suggest that Ibaraki (IBA) virus is closely related to but distinct from the Australian isolates.