Graduate Thesis Or Dissertation


The Oregon sockeye salmon virus (IHN) : A. Replication and autointerference. B. The effect of temperature on infection in Kokanee salmon (Oncorhynchus nerka) and on virus stability Public Deposited

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  • Since the isolation of the Oregon sockeye salmon virus (OSV) in 1958, extensive investigations have been undertaken to characterize the properties of this virus. The results of these investigations have indicated that OSV is a single-stranded RNA virus which contains essential lipids and has a density of 1.16 gm/cm³ in sucrose. In negative stained, glutaraldehyde-fixed preparations, OSV has been identified as a bullet shaped particle 166 to 181 nm in length and 91 to 98 nm in diameter. OSV has been tentatively placed in the Rhabdovirus group. OSV has been found to be antigenically indistinguishable from infectious hematopoietic necrosis (IHN) virus, a bullet shaped virus of sockeye salmon and rainbow trout. Events occurring during a single cycle of OSV replication were investigated. OSV was found to adsorb rapidly to the cell monolayer in the first 15 minutes of exposure. By 60 to 75 minutes after virus exposure under the conditions used, adsorption appeared to be essentially completed. A single-cycle growth curve of OSV indicated that free and total virus increase in an exponential manner. Total virus reached maximum titer in 24 hours while free virus did not reach maximum titer until 48 to 72 hours after infection. Progeny virus appear to be retained in the cell and released gradually into the surrounding medium. The titer attained by free and total virus was nearly the same. Total viral RNA synthesis during a single cycle of OSV replication was monitored by incorporation of uridine-5-³ into acid-precipitable RNA. Total viral RNA in actinomycin D treated monolayers increased exponentially from 8 to 17 hours after infection then stabilized. Accumulation of total viral RNA corresponded well to the total virus growth curve. RNA incorporation into infectious virus continued after total viral RNA reached a maximum. Cellular RNA synthesis was reduced by greater than 99% in the presence of actinomycin D. Indirect fluorescent antibody staining indicated that OSV developed in the cytoplasm. Fluorescent granules appeared at 2 hours after virus infection and increased in size and number on longer incubation. The fluorescence observed was confined to the cytoplasm with no fluorescence observable in the nucleus. Direct evidence for the occurrence of the autointerference phenomenon with OSV was demonstrated with serial undiluted virus passage. Serial undiluted virus passage titers were consistently 10 to 100 times lower than parallel titers achieved in serial diluted virus passage indicating that autointerference occurs with OSV. Thin section preparations of cells infected with OSV indicated a morphological difference in virus particles produced using diluted virus inoculum and undiluted virus inoculum (autointerfering conditions). Three types of particles were observed. OSV-I particles were 188 nm in length by 70 nm in diameter; OSV-II particles were 118 by 69 nm; and OSV-III particles were 81 by 66 nm. In cells exposed to diluted virus inoculum, essentially only OSV-I particles were observed. In cells exposed to undiluted virus inoculum, all three types of particles were observed. OSV-III particles were found in the greatest number while OSV-I particles were found in the least number. Negative stained virus preparations gave some insight into the nature of the OSV nucleocapsid. The striated component of the virus appears to be helical in configuration. The striations are 2.4 nm in width with periodicity of 5.2 nm. Situated within and observed extruding from the striated component, there appears to be filamentous material which in these preparations appeared to have a random configuration. It is not known if the striated and filamentous components are continuous. Thin section preparations indicated the presence of 12 nm surface projections extending radially from the surface of the virus particle. Virus particles were observed in cytoplasmic vesicles and were observed budding through surface membranes into the surrounding medium. No inclusion bodies were observed in either the cytoplasm or the nucleus. No virus particles were observed in the nucleus. The thermal stability of OSV was investigated in three aqueous environments. The virus became more unstable with increasing temperature. All virus titers decreased exponentially. The effect of temperature on OSV infection in kokanee salmon was investigated. The optimum temperature range for the progress of the infection was 12.2°C to 15.0°C. At temperatures above 17.8°C, the percent mortality decreased significantly. At temperatures below 9.4°C, the percent mortality was very high, but the mean time for death was greatly increased. The antigenic relationship between OSV and Egtved virus, a bullet shaped virus of rainbow trout, was investigated by the cross plaque neutralization test. No neutralization of the heterologous virus was observed with either antiserum. By this method OSV and Egtved virus appear to be antigenically unrelated.
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