Previous work has shown that while typhus rickettsiae grow freely in the yolk sacs of embryonated eggs incubated at 34 ° or 37.5°C., their growth is almost completely inhibited when the temperature of incubation is raised to 40°C. (1). The introduction of potassium cyanide in low concentrations, however, resulted in excellent growth of rickettsiae in eggs incubated at 40°C. This work suggested that the determining factor might be the rate of intracellular respiration, rather than the temperature of incubation per se (2).In view of evidence that certain small viruses, in contrast to rickettsiae, grow best when the rate of intracellular metabolism is relatively high (3) it seemed that a study of the influenza virus, by methods previously used for typhus rickettsiae (1, 2, 4), might lead to results of interest. When this study was begun, it was believed that the hemagglutination titre could be used as an index of the rate of multiplication of the virus. It was thought necessary, however, in the initial experiments, to check the titre of hemagglutination by the more laborious process of infectivity titration. This study has brought out anomalous relationships between the hemagglutination and infectivity titre curves.
Material and MethodsEleven day old eggs of the White Rock variety, incubated with air sacs uppermost, were used throughout these experiments. Previous to injection, all eggs were incubated at 37.5°C. The eggs were then candled and a pencil mark placed at the base of the air space directly over the embryo. This portion of the egg shell was painted with an alcoholic solution of iodine and a hole was made directly above the pencil mark. A 10 -s dilution of pooled allantoie fluid from chick embryos infected with an egg-adapted PR8 strain of influenza A virus was used as a source of virus in the majority of experiments. Physiological saline was used as the diluting fluid. Several preliminary experiments showed this solution to work as well as the horse serum broth originally utilized by Hirst (5). Infected allantoic fluids were stored at --40°C.