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Intra- and Inter-Serogroup Genetic Relatedness of Orbiviruses. II. Blot Hybridization and Reassortment in vitro of Epizootic Haemorrhagic Disease Serogroup, Bluetongue Type 10 and Pata Viruses

D. K. Bodkin

Yale Arbovirus Research Unit, Department of Epidemiology and Public Health, Yale University School of Medicine, New Haven, Connecticut 06510, U.S.A.

Viruses which belong to the epizootic haemorrhagic disease (EHD), bluetongue and Eubenangee serogroups of orbiviruses exhibit low level cross-reactions in some serological tests. Although Pata virus cross-reacts at low levels with members of the EHD and bluetongue serogroups, it was assigned originally to the Eubenangee serogroup. RNA-RNA blot hybridization data, however, suggest that Pata virus is not a member of the Eubenangee serogroup. In this study, the genetic relatedness of the EHD serogroup viruses, bluetongue virus type 10 (BTV-10) and Pata virus was assessed by RNA-RNA blot hybridization and by gene reassortment experiments in vitro. The five members of the EHD serogroup examined were highly related by reciprocal RNA-RNA blot hybridization. Genes 1, 3, 4 and 6 to 9 were highly conserved with three unique types of gene 2, four variant types of gene 5 and two variant types of gene 10. Geographical boundaries could not be correlated with sequence relatedness because viruses isolated from the same locality were distant relatives when compared with another virus isolated on a different continent. The significance of the unique and variant genes is discussed. The EHD isolates, BTV-10 and Pata virus exhibited distinct profiles on agarose and polyacrylamide gel electrophoresis, and they are related distantly as shown by weak hybridization signals in blot hybridization. Gene 2 was a unique gene among the EHD isolates, BTV-10 and Pata virus. One BTV-10 gene hybridized more strongly to gene 9 of the EHD viruses than the other BTV-10 genes, and its role in encoding the cross-reactive antigen is discussed. Intra-serogroup gene reassortment in vitro was demonstrated in the six crosses among EHD serogroup members. In contrast, gene reassortment was not observed in inter-serogroup crosses between EHD 1 and BTV-10, BTV-10 and Pata virus, and between EHD 1 and Pata virus. Correlation of blot hybridization and gene reassortment indicated that viruses must share high sequence conservation in the majority of their genes before genetic interaction is likely. The usefulness of blot hybridization as an indicator of the likelihood of gene reassortment is discussed. These hybridization and gene reassortment data indicated that Pata virus is not a member of the bluetongue or EHD serogroups and that it should be assigned to the ungrouped set of orbiviruses. The hybridization and gene reassortment data suggest that members of the EHD serogroup, BTV-10 and Pata virus represent three distinct gene pools, and the role of reassortment in the generation of genetic diversity is discussed. The minor antigens shared among these gene pools probably reflect their common ancestry.

Keywords: Orbivirus, RNA-RNA blot hybridization, Reoviridae

Present address: Department of Microbiology and Molecular Genetics, Harvard University Medical School, 25 Shattuck Street, Boston, Massachusetts 02115, U.S.A.

Received 18 June 1987; accepted 7 September 1987.