Rio Bravo (RB) virus has been assigned to the family on the basis of its antigenic relatedness to other members of this family. RB virus, unlike most members of the , is believed not to have an arthropod vector. We examined biochemical and biophysical characteristics of RB virus to determine whether it should be assigned to the and to compare it with arthropod-borne flaviviruses. Purified RB virus banded at a density of 1.18 g/ml in sucrose and had a sedimentation coefficient of about 200 S. Virions, negatively stained with ammonium molybdate, were spherical, had diameters of 42 nm, and appeared to be surrounded by envelopes bearing surface projections. The loss of infectivity after infectious virus was incubated with diethyl ether or sodium deoxycholate confirmed the presence of envelopes. Partially purified RB virions contained single-stranded RNA, lacking 3′ poly(A) tracts, that sedimented in a 15% to 30% sucrose gradient as one discrete band with a sedimentation coefficient of about 40 S. Most of the viral proteins in preparations of purified virus and in immunoprecipitates had similar electrophoretic mobilities and glycosylation patterns to known flavivirus proteins. Therefore, they were assigned the following tentative designations using the nomenclature for flavivirus proteins: gp52 and gp47, envelope proteins; gp46, non-structural protein 1;p25, gp20(prM), precursor to membrane protein; gp < 18K. Putative core and membrane proteins were not identified. These physical and biochemical characteristics of RB virus are remarkably similar to those of the arthropod-borne members of the and they confirm the classification of RB virus in this family. This is the first report of biochemical and physical properties of a non-arthropod-borne member of the .


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