Noncytocidal persistent infections at 37 C of mouse L cells (LX X) with infective B particles of vesicular stomatitis virus (VSV) could be established only in the presence of large numbers of defective interfering (DI) particles. Under these conditions, there was a rapid spontaneous selection of temperature-sensitive (ts) virus. At 10 days there was an increase to 17.8% in the frequency of ts clones in the virus population; by 17 days this frequency had reached 85.2%, and by 63 days 100% of the clones isolated were ts at 39.5 C, the nonpermissive temperature used. All 34 of the clones isolated from the 84-day fluid had an RNA-phenotype, and 8 clones that were tested all belonged to VSV complementation group I. When tested by an interference assay, L,,fluids did not contain significant numbers of DI particles (<1 DI/PFU). Furthermore, persistent infection of L cells at 37 C could be initiated under conditions in which few, if any, DI particles were present by using low input multiplicities (10-4 and 10-5) of a clonal isolate of an RNA group I mutant obtained from LX ,\ cells. On the basis of these and other results, a mechanism is proposed to explain the role of ts mutants in both the establishment and maintenance of the persistently infected state. MATERIALS AND METHODS Cells. Primary chicken embryo (CE) cells, mouse L cells (clone 929), and a line (BHK-21) of hamster kidney cells were propagated in Eagle minimal essential medium plus 4% calf serum. Viruses. The large-plaque mutant of VSVINI) (L,VSV) described by Wertz and Youngner (22) was grown in BHK-21 cells and assayed in CE cells. Monolayers in 32-ounce (ca. 950-ml) culture bottles were infected with less than 0.01 PFU/cell to avoid production of DI particles. This wild-type virus stock is referred to in the text as VSVe. Analysis of [PH1uridine-labeled viral particles by sucrose gradients failed to detect DI particles in lysates produced under these conditions. ts mutants of VSVINI) 90

Although two deoxyribonucleic acid (DNA) viruses, pseudorabies (PsRV) and vaccinia, are as susceptible as a ribonucleic acid (RNA) virus, vesicular stomatitis (VSV), to interferon when tested in chicken or mouse cells, they are refractory to inhibition in interferon-treated primary rabbit kidney cells and in a continuous line (RK-13) of rabbit kidney cells. Superinfection with VSV of RK-13 cells first infected with PsRV completely blocks the replication of PsRV with no effect on VSV yield. When the same experiment is carried out in RK-1 3 cells pretreated with 1,000 units of interferon, VSV replication is inhibited, which permits PsRV to replicate normally. These findings demonstrate that in the same cell one virus (PsRV) can be refractory to interferon and a second virus (VSV) can be susceptible. These experiments show that rabbit kidney cell cultures are deficient in the synthesis of resistance factors active against the DNA viruses tested and raise the possibility that separate resistance factors may exist for RNA and DNA viruses. In the case of sequential infection of interferon-treated RK-13 cells with vaccinia and VSV, it was found that not only was vaccinia replication refractory to inhibition by interferon, but also that prior infection with vaccinia was able to partially reverse the effect of the inhibitor on the replication of the VSV used for superinfection. On the basis of these and other data it is postulated that a vaccinia virion component or a replication product of vaccinia virus, or both, enables VSV to escape the inhibiting action of interferoninduced resistance factors.

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YOUNGNER, JULIUS S. (University of Pittsburgh, Pittsburgh, Pa.), AND MIARION E. KELLY. Inhibition by exogenous interferon of replication of poliovirus ribonucleic acid in chick brain. J. Bacteriol. 90:443-445. 1965.-The replication of poliovirus was studied after the intracerebral inoculation of infectious ribonucleic acid (RNA) into the brains of 2-day-old chicks; these animals are not susceptible to intact virus. Single-cycle replication of virus, which reached a peak in about 12 hr, was completely inhibited by prior intraperitoneal injection of interferon prepared from the allantoic fluid of chick embryos infected with influenza virus. A single dose of as little as 500 units of interferon, measured by a plaque-reduction method, completely suppressed viral replication when injected 24 hr prior to infectious RNA. This system provides a model for the study of the protection of target organs by passively transferred interferon injected at a distance.