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|Title:||Studies on the REOvirus type 2 - human amnion cell system|
|Authors:||Oie, Herbert Kazuto|
|Abstract:||REOvirus type 2 (REO-2), strain D-5, and its interaction with « continuous line of human amnion cells (RA) were investigated. The investigations were carried out to obtain information on the effects of several physicochemical agents on the virion, the early interaction between virus and cell, and some of the consequences stemming from this virus-cell interaction. The virion was found to be resistant to ethyl ether (30% and 50%). It was quite stable at temperatures below 250°C, but at 37°C and 60°C, its infectivity titers were reduced by one-half in 5 days and 30 seconds, respectively. For the first 2-3 minutes, sonication of virus preparations at 20 Kc resulted in increased infectivity titers, but a rapid decrease in titers resulted with prolonged treatment. The virus was stable for at least 7 days over a pH range of 2-9 but was rapidly destroyed at pH 11. UV-irradiation inactivated the virus very rapidly, reducing its infectivity titer by one-half in less than 15 seconds. The rate of adsorption of virus to cell was found to be inversely related to the volume of inoculum and directly related to the temperature. The elution of adsorbed virus from cells was found to depend on the exposure multiplicity and the temperature of incubation. At multiplicities of 10 or more, significant elution was observed; at 1 or less, no significant elution occurred. At 370 C, elution occurred but a stable cell-virus complex was formed within 5 minutes. At lower temperatures, elution occurred but at slower rates. No significant desorption occurred at 40 C. Single cycle growth studies of REO-2 in RA cells revealed an eclipse period of 9 hours and maximal yields of virus obtained in about 36 hours. In comparison, REO-3 in RA cells was found to have an eclipse period of 6 hours and maximal yields obtained in 24-26' hours. Immunofluorescent and cytochemical (acridine orange) examinations of REO-2 infected RA cells revealed the following: (1) the appearance of viral antigen as early as 4 hours post-infection (p.i.), (2) a maximal number of antigen-containing cells at 9 hours p.i., (3) the appearance of orthochromatically green-staining inclusion bodies at 6 hours p.i., and (4) maximal number of cells containing inclusion bodies in 12 hours. Infection of RA cells with either active or UV-inactivated REO-2 at low multiplicities (1-5) resulted in the production of an anti-viral substance which was found to possess many characteristics in common with interferon. In addition, REO-2 was found to be sensitive to the inhibitory effect of this anti-viral agent. When HeLa, RA, or BSC-1 cells were exposed to high multiplicities of UV-inactivated REO-2 preparations, rapid death of cells was observed. This cytotoxic phenomenon was found to have the following characteristics: (1) morphological changes were indistinguishable from that due to viral cytopathic effect, (2) first signs of the effect could be seen at about 3-4 hours p.i. and culminated in 9-10 hours, and (3) the intensity and extent of the effect was dependent on exposure multiplicity. Only UV-inactivated virus preparations produced the effect; neither heat-inactivated nor active virus was observed to produce cytotoxicity. Data obtained strongly implicate the UV-inactivated viral particle as the cytotoxic agent. The significance of the new information obtained and some ramifications in REOvirus research are discussed. In addition, suggestions for further investigations are presented.|
Thesis (Ph. D.)--University of Hawaii, 1968.
Bibliography: leaves 93-98.
xii, 98 l graphs
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|Appears in Collections:||Ph.D. - Microbiology|
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