1. A method is described for propagation of bacteria-free vaccinia virus in the rabbit fetus.
2. For the production of severe generalized infection, affording a high yield of virus, the following conditions were found to be favorable: the use of 24 day fetuses, a virus dosage of 10,000 adult rabbit skin units, injection into multiple sites, and an incubation period of 4 days in utero.
3. Under these conditions the virus was found to disseminate widely in the infected fetus, being recoverable particularly from liver, lungs, brain, skin, placenta, and kidney.
4. Under varied conditions of virus dosage, fetal age, and incubation period, all grades of reaction were observed in the fetus, ranging from the mildest infection, with only an occasional small lesion demonstrable, to generalized vaccinia with pocks in nearly every organ of the body, and often death.
5. It was shown that the virus could be carried in serial passage through fetuses. A series of 27 such passages was made by using fetal skin for subinoculation at each transfer. Similarly, two other strains of the virus were evolved in shorter series of transfers by using respectively fetal brain and fetal liver as the subinocula.
6. The passage strains maintained their identity and titer, as judged by the intradermal inoculation of adult rabbits. However, these strains underwent a significant reduction in their capacity to infect by the scarification method, and the lesions induced by intradermal inoculation were much milder than those produced by the parent strain. None of the passage strains manifested properties characteristic of the so called neurovaccine strain, despite the frequent use of intracerebral inoculation in the fetuses.
7. In explanation of the reduction of virulence of the passage strains, it is suggested that a given suspension of virus may be considered as consisting of particles of varying degrees of virulence, and that when cultivation is carried out in highly susceptible tissues a relative overgrowth of the less virulent elements is permitted, so that apparent attenuation results.