The agent of infectious bovine rhinotracheitis ( I B R ) an acute upper respiratory disease of cattle, has been isolated from nasal washings of infected cattle on bovine kidney tissue culture. The viral nature of the etiologic agent has recently been established by extensive serologic and transmission studies ( 1-5 ) .The virus is cytopathogenic to cells of both bovine kidney and human amnion cultures in which it induces development of intranuclear inclusions. readily observed in stained preparations(6). The agent is relatively stable and can be held at 4°C over 30 days and at 2 2 3 C for 3 days with no measurable loss of titer (7). This study was undertaken to demonstrate the entity associated with the infectious unit and to determine its physical charac teristics.Materials and methods. Tissue czrlturc. Bovine kidney cell cultures were prepared by the method of JIadin ct a l . ( 8 ) with a medium of 0.5% lactalbumin hydrolysate in Hanks balanced salt solution containing .01 J I Tris buffer. with addition of 5% lamb serum. Human amnion cell cultures were prepared by the method of Zitcer ct d ( 9 ) with a medium of bovine amniotic fluid containing 5 9 horse serum. Virus. Pritchard and American strains of IRR virus as described by Cheatham and Crandell(6) were used. 17irus pools of Pritchard strain were prepared by inoculating the agent in bovine kidney cultures into 32 oz. prescription bottles. When the cytopathogenic effect was complete at about 72 hours. the fluid was frozen and thawed several times to disrupt the infected cells. then centrifuged in refrigerated centrifuge at 2.000 rpm for
Summary
The occurrence of infection with the Venezuelan equine encephalomyelitis virus in two members of our laboratory staff and the reports of 24 other laboratory infections with the same virus indicate the exigency for the protective immunization of personnel having contact with this agent.
A formalinized vaccine prepared from chick embryos infected with the Venezuelan virus and partially purified by differential high-speed centrifugation was used to immunize 20 laboratory workers who had not previously had contact with the agent. Following two injections of 1.0 ml amounts of vaccine, the majority developed appreciable amounts of specific neutralizing antibody and one-half of them developed detectable amounts of specific complement-fixing antibodies. A third injection of vaccine resulted in increased amounts of both neutralizing and complement-fixing antibodies in the majority of the group. The neutralization indices now ranged from 150 to 1,585,000 and the complement-fixation titers from 0 to 1/32. While there was a general correlation between the results of neutralization and complement-fixation tests there was no close relation between the levels of antibody as determined by the two methods.
None of the 20 persons who received the present vaccine developed clinical signs of disease when they subsequently engaged in the large scale manufacture of Venezuelan equine encephalomyelitis vaccine.
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