Characteristics of the Adenoviruses

Abstract

Investigation of the early period of viral multiplication permits observation of certain discernible stages in the reproductive cycle. Detailed analysis of these steps ((a) combination of virus with susceptible cells, (b) synthesis of new viral particles, and (c) dissociation or release of virus) yields data concerning basic characteristics of the agent. This information is pertinent for purposes of classification, and to stimulate hypotheses concerning the nature of the reactions of a virus with susceptible host cells under conditions of natural infections. To obtain evidence concerning the characteristics of multiplication of the adenoviruses (1-3), investigation of 4 prototype viruses of this group was undertaken. A preliminary report of these studies has been made (4); detailed presentation of the experimental results forms the basis of this paper. These data indicate that the types 1 to 4 adenoviruses studied adsorb at a relatively slow rate to the host cells, strain HeLa cells; that the latent or eclipse period of the multiplication cycle is relatively long and is longer for types 1 and 2 than for types 3 and 4; that the incremental period of viral propagation is relatively short; and that only a very small amount of the maximum virus detectable is dissociated from the host cells during and after the incremental phase of the multiplication cycle. Materials and Methods Tissue Cu]#ure.--A strain of HeLa cells was employed throughout this investigation (5)"For the majority of experiments, a growth fluid composed of 40 per cent human serum and Hanks's balanced salt solution (BSS) was used for propagation of cells in serial cultures by methods described in detail previously (6). In the latter part of this investigation cells were cultivated in Eagle's basal medium (7) to which was added human serum in a final concentration of I0 per cent. These cells, as compared with those previously employed, were equally susceptible to adenovirus infection. Cen cultures were initiated by inoculation of 30 ml of a *

Intracellular Localization of Type 4 Adenovirus

“…Tissue Culture.--Epithelial cells of the HeLa strain (6) were employed in all experiments and were propagated in either Eagle's basal medium (7) containing 10 per cent human serum, or as previously described (8), in a growth fluid composed of 40 per cent human serum and Hanks' balanced salt solution.…”

“…The amount of virus used for infection of cultures in bottles and the time which elapsed between infection and harvest varied among the experiments; these data are included in the description of experiments. Infectivity titrations were performed in HeLa cell culture tubes using 1:3.2 (10 -0.5 ) serial dilutions; the details of the methods have been given elsewhere (8). Employing this titration procedure it has been shown that a difference of 0.36 log units between 2 infectivity titers should occur by chance only once in 20 titrations (8).…”

“…Infectivity titrations were performed in HeLa cell culture tubes using 1:3.2 (10 -0.5 ) serial dilutions; the details of the methods have been given elsewhere (8). Employing this titration procedure it has been shown that a difference of 0.36 log units between 2 infectivity titers should occur by chance only once in 20 titrations (8).…”

“…These agents were propagated by serial passage in HeLa cells and stock viral pools were prepared and stored by methods reported earlier (8).…”

“…Therefore, it was of interest to determine whether the same series of events might not be true for adenovirus-infected cells. It has been shown that the period before the appearance of newly synthesized type 4 adenovirus in HeLa cells is inversely proportional to the amount of virus used to infect the cells (8). The "eclipse period" (elapsed time between viral inoculation and detection of newly formed virus) is 14 to 15 hours when maximal infecting doses of virus are used and the initial multiplication cycle is measured.…”