Minute virus of mice. I. Procedures for quantitation and detection.
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Biomedical subjects
Publications and source records attributed to W P Rowe.
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Tissue culture cells of a Rous virus-induced rat tumor undergo syncytium formation when placed in contact with mouse embryo cell cultures infected with murine leukemia viruses. This phenomenon can be used as a cytopathic end point for isolating and titrating these viruses in tissue culture. The principle should be applicable to detection of leukemia viruses of other species.
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A tissue culture cell system for isolation and identification of members of the murine leukemia virus group (the complement fixation for murine leukemia test) was modified to permit the isolation of naturally occurring virus from leukemic and normal mice. The important factors for increasing the sensitivity of the test were the use of National Institutes of Health (NIH) strain Webster Swiss embryo cell cultures and the selection of rat-immune sera having complement-fixing antibodies to tissue culture antigens of both the Gross and FMR subgroups. In all, 163 strains of mouse leukemia virus, from 11 inbred mouse strains, have been isolated. Representative virus isolates were shown to possess the properties of the murine leukemia virus group; i.e., they were chloroform-sensitive, noncytopathic agents which replicated in mouse embryo tissue culture and produced group-reactive, complement-fixing antigen and budding C-type particles visible by electron microscopy. These viruses could serve as helpers in the rescue of Moloney sarcoma virus genome from non-producer hamster sarcoma cells, yielding pseudotypes. All of the 19 field isolates tested were neutralized by Gross passage A antiserum but not by potent antisera to the Moloney, Rauscher, and Friend strains. Virus was recovered regularly from embryos and from the plasma and spleen of adult mice of high leukemic strains. In low leukemic mouse strains, different patterns of virus detection were observed. In C3H/He mice, virus was occasionally present in embryos and was found in 40% of adult spleens. BALB/c mice were virus-negative as fetuses or weanlings, but spleens of more than half of the mice over 6 months of age yielded virus. NIH mice have never yielded virus. In reciprocal matings between AKR and BALB/c mice, virus recovery from embryos was maternally determined. The development of tissue culture isolation procedures made possible for the first time the application of classical infectious disease methods to the study of the natural history of murine leukemia virus infection.
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Lymphocytic choriomeningitis (LCM) virus was observed by electron microscopy in thin sections of infected tissue cultures. The particles were pleomorphic and varied greatly in size. The smaller particles (50 to 200 nm) appeared to be spherical, whereas the largest (over 200 nm) were often cup-shaped. All particles contained one to eight or more electron-dense granules which were removed by ribonuclease. The particles were formed by budding from the plasma membrane and appeared to have spikes. The morphological evidence suggests that LCM should be considered as belonging to the presently unclassified group of lipoprotein-enveloped ribonucleic acid viruses.
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A quantitative immunofluorescent procedure for detection of viral antigen was used to study the potentiation of AAV-1 by Ad.7. AAV viral antigen formed only when the cells were also infected with adenovirus, and only in cell culture systems in which the adenovirus infection proceeded to completion. Ad. 7 infection of AGMK. cell cultures did not potentiate AAV unless the Ad. 7 infection was itself potentiated by SV40. Dose-response studies indicated that a single AAV particle and a single infectious Ad. 7 particle sufficed to initiate AAV antigen synthesis. Sequential inoculation studies showed that AAV antigen formed simultaneously with Ad. 7 viral antigen when the AAV was inoculated any time between 15 hr before to 10 hr after the Ad. 7, both antigens appearing about 15 hr after inoculation of Ad. 7. The AAV-1 antigen formation had a minimum latent period of 5 hr, as seen with Ad. 7 preinfection of 10 hr or more. When UV-irradiated Ad. 7 was used as helper, the AAV antigen still appeared simultaneously with the Ad. 7 viral antigen, even though the latter was delayed by 23 hr compared to nonirradiated virus. When the early replicative events of both viruses were allowed to proceed in FUDR-inhibited cells, and then the FUDR inhibition was reversed, AAV antigen formed within 2 hr, which was 3 hr before the Ad. 7 viral antigen appeared. It was inferred that the event in the adenovirus cycle that renders a cell competent to synthesize AAV occurs after the 10th hr and may be temporally associated with replication of the adenovirus DNA.
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