- Volume 8, Issue 2, 1970

In RHF-1 cells infected with either adenovirus 2 or 12, the formation of infectious virus and antigens decreased with each successive passage of cells until the virus was ultimately eliminated from the cultures. These cultures then emerged into a new phase in which some virus-induced proteins were present in at least a small proportion of cells. Adenovirus 2 fibre antigen persisted throughout the 15th subculture, whereas adenovirus 12 early (T) and late (fibre) antigens were carried throughout the 30th subculture over a period of 600 days. Virus-free but antigen-containing cells may therefore have possessed at least a portion of the virus genome. Shortly after the disappearance of virus, distinct multilayered foci of cells emerged in both lines. This phenomenon became a characteristic feature of the cultures only in the adenovirus 12 line.

In KB cells doubly infected with vaccinia virus and Frog virus 3 (FV 3) at 37°, a non-permissive temperature for FV 3, the replication of vaccinia virus DNA was inhibited. The inhibition, which was as effective with gamma-irradiated or heat-inactivated virus as with live FV 3, was dependent upon the multiplicity of infection with FV 3. A 50% inhibition of vaccinia DNA synthesis was observed when the cells were pretreated with 0.2 p.f.u./cell of purified FV 3. Inhibition was not related to a toxic effect of FV 3 on the cells, as there was no auto-inhibition of the replicating FV 3 DNA in KB cells incubated 3 hr at 37° before incubation at 26°.

Intracellular replication of murine sarcoma virus was followed by Luria-Latarjet experiments. In experiments with unsynchronized cultures ultraviolet sensitivity of the cells infected with murine sarcoma virus as focus centres increased with the lapse of time after infection, and 16 hr after infection u.v.-resistant cells appeared which were probably producing mature virus. The u.v.-resistant population continued to increase thereafter.

Ultraviolet sensitivities as focus centres of synchronized cells infected with murine sarcoma virus at different periods of the cell cycle were compared 23 hr after infection. The cells infected with the virus in the G1 phase remained more sensitive to u.v. light than those infected with the virus in the S phase, i.e. the latent period was longer after G1 phase infection than after S phase infection. An antimitotic agent colchicine, prevented the increase of u.v. resistance of the cells as focus centres. These facts suggest that murine sarcoma virus infection requires the division of the host cells and that virus replication is synchronized at this stage of the cell cycle.

Two types of experiment have been made during the early part of the latent period on a culture of cells singly infected with phage T4.

First, samples taken from the infected cells at different times were irradiated with ultraviolet light in order to obtain a set of Luria-Latarjet curves. Secondly, and with the same culture of infected cells, the kinetics of phage DNA synthesis was followed using a density gradient technique.

A comparison of these results suggests that the radiation stability shown by the Luria–Latarjet curves is first detectable at about the time that phage DNA synthesis commences. The radiation stability, however, increases at a rate which is much faster than the rate of replication of the parental phage DNA.

Disulphide-linked immunosorbents prepared from rabbit antisera against extracts of uninfected HeLa (ERK) cells specifically removed host-cell antigens from extracts of cells which had been infected with vaccinia virus. Similarly, disulphide-linked immunosorbents prepared from rabbit anti-vaccinia sera specifically removed vaccinia-specific antigens from infected cell extracts. Disulphide-linked preparations derived from unrelated sera adsorbed neither host nor virus-specific antigens.

A previous report (Breese, 1969) dealt with the direct reaction between type A1 foot-and-mouth disease virus (FMDV) and antibody tagged with ferritin. In the same study, ferritin-tagged anti-O2 globulin and normal guinea-pig globulin gave no reaction. This report describes the indirect test with ferritin in which infected cells are treated first with an antibody globulin and then with an antibody to globulin which has been tagged with ferritin. In the case of types A1 and O2 FMDV, globulins were used which were antibody to the whole virus, while in the A-119 experiments, specific antibodies to separate antigens (Cowan & Graves, 1966) were used. The indirect test allows the study of several antigen + antibody reactions while requiring only a single ferritin conjugation.

Tissue cultures of primary swine kidney cells were infected as previously reported (Breese, 1969; Breese & Graves, 1967).