Studies have been reported concerning the relationships between virus materials found in the allantoic membranes and media of eggs deembryonated after injection of Standard (ST), heat-inactivated (37°C.) standard (ΔST), and undiluted passage (UP) seeds.
It was found that the membranes always contained relatively more non-infectious hemagglutinins (NIHA) than the media and, correspondingly, the ratios between infectious virus and hemagglutinin units (ID50/HA) in the tissues were up to 1.5 log10 units lower than in the liberated progeny. These differences were seen not only following inoculation of undiluted ST, ΔST, and UP seeds, the progenies of which always contain considerable proportions of NIHA, but also when dilute ST inocula were employed which lead to the liberation of only infectious virus. Essentially similar differences in the ID50/HA ratios were observed also in the allantoic membranes and fluids obtained from growth curve experiments in the intact chick embryo employing the various types of seeds.
In correlating the liberated virus materials in the media of deembryonated eggs to those in the membranes it was noted that in any given 2 hour interval during the phase of nearly constant production and release up to 10 times the quantity of infectious virus was shed as was present in the tissues at the onset of that period. In contrast, only about ¼ of the hemagglutinins were released during the same time. The viral (V) and soluble (S) complement-fixing antigens were found in the tissues but no detectable quantities were released during any 2 hour interval.
The NIHA in the membranes apparently is located within the cells since it could not be released by the action of RDE. Intracellular inhibitors of hemagglutination were readily inactivated following inoculation of undiluted ST, ΔST, or UP seeds but not when ultraviolet-inactivated virus was used. The inhibitor activity decreased in proportion to the hemagglutinins produced.
Transfer of infected deembryonated eggs to the cold room after production and liberation of progeny were well under way immediately halted further release but in the tissues the status quo was maintained and release was resumed on return to the 37°C. incubator. The addition of potassium cyanide to the medium of deembryonated eggs at 37°C. during the period of nearly constant production and release of virus material reduced immediately and to comparable extents the ID50 and HA titers in the tissues and liberation decreased in proportion. On removal of the cyanide 2 hours later, both titers in the tissues gradually returned to those of the untreated control eggs with a corresponding increase in liberation. The ID50/HA ratios were not affected by these manipulations. It is concluded that the NIHA in the membranes forms part of a dynamic process.
An attempt has been made in the discussion to integrate the present results with previous observations concerning the formation of incomplete forms of virus and their nature and role in the infectious process.