Characterisation of orbiviruses using molecular and electron microscopic techniques

Abstract

Orbiviruses are frequently isolated in northern Australia. They are arthropod-borne viruses, with a double stranded, segmented RNA genome. The Orbivirus genus is classified into serogroups on the basis of immunological tests. Their insect hosts can include mosquitoes,Culicoides (biting midges) and ticks, and the vertebrate hosts include a range of native and domesticated animals, depending on the virus serogroup. Most of the Orbiviruses identified in Australia are non-pathogenic in animals, but members of one serogroup, bluetongue virus (BTV), can cause severe disease in sheep. At present the ranges of the insect hosts of BTV and the sheep populations in Australia don't coincide, but with climatic changes this may alter. Consequently, Orbiviruses in general and BTV in particular, are of major importance to the agricultural industries in Australia. Better tests are needed to identify virus serotypes within each serogroup, especially for BTV, and a method is described for E.coli expression of a region of the outer coat protein of BTV1 considered to contain virus neutralising epitopes, and the use of the expressed peptide to raise antiserum in a rabbit. The antiserum showed some specificity for BTV1 in immunofluorescence tests and by immunoelectronmicroscopy, but it was concluded this was not a practical method for deriving serotype-specific antisera.

Identification of Orbiviruses by molecular techniques may be quicker, more efficient, and more useful than current serological tests. A commonly isolated Orbivirus serogroup in northern Australia is the Wongorr serogroup. No information on the molecular characteristics of the group was available, so RNA profiles of the prototype Wongorr virus (WV) were made, one of the serogroup-specific virus proteins was partially sequenced, and oligonucleotide primers were designed for PCR. With this information the Wongorr serogroup was compared with nine other Orbivirus serogroups by PCR, RNA profiles and multiple sequence alignments. Several WV isolates were also compared with each other, and two distinct clusters of isolates were identified. These were separated geographically, with the Northern Territory WV isolates showing consistent differences from the southern isolates. A previously unidentified Orbivirus, Paroo River virus has been shown to be a WV by PCR and sequencing, and the position of Picola virus in the Wongorr serogroup has been confirmed. Two other NT Orbivirus isolates were identified as Corriparta viruses by PCR and sequencing, and a third unidentified Orbivirus has been shown provisionally to be Mitchell River virus.

Date of Award	Mar 1995
Original language	English
Supervisor	Karen Gibb (Supervisor)