Theodore J. Starr

Replication pattern of psittacosis virus in a tissue culture system

Abstract The growth curve of psittacosis virus in McCoy cells has been determined. There is a 17-hour latent period before virus production is demonstrable and maximum virus levels are achieved by 48 hours. Addition of either FU or FUDR had an inhibitory effect on the amount of virus subsequently produced, and this effect was reversible by uracil and thymidine (for FUDR only). These drugs probably act by immediately stopping the production of viable viral DNA, and the amount of virus ultimately synthesized by a culture may be a measure of the amount of viral DNA which had already been synthesized at the time of drug addition. By varying the time of drug addition it was found that there was a 10-hour lag between DNA synthesis and its incorporation into particles. This strongly suggests that psittacosis virus replicates by an assembly of subunits, not by binary fission, and hence is a true virus.

Electron and Fluorescence Microscopy of Mouse Hepatitis Virus.

Summary Livers of mice infected with mouse hepatitis virus (MHV 1) were examined by conventional staining procedures and by electron and fluorescence microscopy using acridine orange fluorochrome and fluorescein-tagged antibody. Preparations stained with acridine orange showed abnormal accumulations of RNA-staining material in cytoplasm of hepatic cells. Inclusion bodies of similar size, shape, and location were stained selectively with fluorescein-tagged antibody. Observations by electron microscopy showed virus particles in cytoplasm which measured 90 ± 20 mμ in diameter. The authors are indebted to Dr. R. H. Rigdon and his staff for invaluable technical assistance.

Propagation of duck hepatitis virus in tissue culture

Summary Duck hepatitis virus (DHV) survived storage for at least 21 days at 37°C. It survived treatment with di-ethyl ether and with fluorocarbon. DHV has been propagated through 25 consecutive passages in tissue cultures prepared with chick embryo explants. It was not propagated in cultures of trypsinized chick embryo cells or in cell lines of mammalian origin.

Susceptibility of Cortisone-treated Mice to Infection with Mouse Hepatitis Virus.

Summary A small proportion of mice inoculated with cortisone alone died or developed ascites and/or liver lesions. Bacteria were isolated from some of the livers showing cortisone-induced lesions. Homogenates of such livers were not infective for normal suckling Swiss mice. Cortisone acetate reduced susceptibility of normally resistant weanling Swiss mice to infection with the Gledhill strain of mouse hepatitis virus. The effectiveness of cortisone in decreasing host resistance was enhanced as dosage of the steroid was increased. There was a decrease in this cortisone effect as the time interval between virus and cortisone administration was increased from 1 to 5 days.

Micronuclei Formation in Tissue Cultured Cells Treated with Colchicine.

SummaryFormation of mitotic figures and micronuclei in a colchicine-treated human synovial cell line (McCoy) were quantitated at different times after treatment. After a lag period of 4 hours, mitotic figures increased in a roughly linear manner until the 12th hour. A rapid drop to control values by the 15th hour was correlated with the appearance of micronucleated cells. A second lag period of approximately 4 hours was followed by an apparently enhanced rate of formation of mitotic figures to the 30th hour after treatment. After this point their numbers gradually decreased so that at 72 hours virtually none was seen. Except for shifts induced by this second lag period, micronucleated cells continued to increase after the 12th hour at a more-or-less constant rate through the 30th hour. The micronucleated cells were terminal forms resulting from colchicine treatment except for a nearly constant fraction (10%) of polyploid giant cells which remained unchanged.Summary Formation of mitotic figures and micronuclei in a colchicine-treated human synovial cell line (McCoy) were quantitated at different times after treatment. After a lag period of 4 hours, mitotic figures increased in a roughly linear manner until the 12th hour. A rapid drop to control values by the 15th hour was correlated with the appearance of micronucleated cells. A second lag period of approximately 4 hours was followed by an apparently enhanced rate of formation of mitotic figures to the 30th hour after treatment. After this point their numbers gradually decreased so that at 72 hours virtually none was seen. Except for shifts induced by this second lag period, micronucleated cells continued to increase after the 12th hour at a more-or-less constant rate through the 30th hour. The micronucleated cells were terminal forms resulting from colchicine treatment except for a nearly constant fraction (10%) of polyploid giant cells which remained unchanged.

Gytochemical Effects of Substituted Pyrimidines on Psittacosis Virus.

Summary Cytochemical changes associated with psittacosis virus infection of tissue cultures were studied by acridine orange staining and fluorescence microscopy. Normal virus development was interrupted by the addition of 5-fluorouracil and 5-fluoroorotic acid which caused the accumulation of “fraudulent” RNA. It is suggested that the substituted pyrimidines interfered with development of intact virus through inhibition of protein synthesis.

Propagation of mouse hepatitis virus (Gledhill) in tissue culture.

Summary The Gledhill strain of mouse hepatitis virus (MHV 1) was passaged through 10 successive tissue cultures of newborn mouse kidney explants. As MHV 1 did not cause reproducible cytopathogenic effects, infectivity tests with newborn mice were used to detect propagation of this virus in serially passaged cultures. Observations suggest that virus was adsorbed by the proliferating explants and that cell-free culture fluids were infective. Total time of virus growth far exceeded decay rate of the virus incubated without tissue. After the tenth passage, the virus had been diluted 1012-fold which exceeded the extinction point of the original inoculum.