The herpes simplex virus 1 (HSV-1) genome contains three origins of DNA synthesis (Ori) utilized by viral DNA synthesis proteins. One sequence (OriL) maps in the L component, whereas two sequences (Oris) map in the S component. We report the construction of a recombinant virus, R7711, from which both Oris sequences have been deleted, and show that the Oris sequences are not essential for the replication of HSV-1 in cultured cells. In addition to the deletions of Oris in R7711, the a47 gene and the 5' untranscribed and transcribed noncoding regions of the Us11 gene were deleted, one of the a4 promoter-regulatory regions was replaced with the simian virus 40 promoter, and the at22 promoter was substituted with the av27 promoter. The total amount of viral DNA synthesized in Vero cells infected with the Oris-negative (Oris-) virus was approximately that seen in cells infected with the Oris-positive virus. However, cells infected with the Orisvirus accumulated viral DNA more slowly than those infected with the wild-type virus during the first few hours after the onset of DNA synthesis. In single-step growth experiments, the yield of Orins progeny virus was reduced at most fourfold. Although a single Oris (R. Longnecker and B.

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Herpes simplex viruses maintained in a latent state in sensory neurons in mice do not reactivate spontaneously, and therefore the factors or procedures which cause the virus to reactivate serve as a clue to the mechanisms by which the virus is maintained in a latent state. We report that cadmium sulfate induces latent virus to reactivate in 75 to 100% of mice tested. The following specific findings are reported. (i) The highest frequency of induction was observed after two to four daily administrations of 100 micrograms of cadmium sulfate. (ii) Zinc, copper, manganese, or nickel sulfate administered in equimolar amounts under the same regimen did not induce viral reactivation; however, zinc sulfate in molar ratios 25-fold greater than those of cadmium induced viral replication in 2 of 16 ganglia tested. (iii) Administration of zinc, nickel, or manganese prior to the cadmium sulfate reduced the incidence of ganglia containing infectious virus. (iv) Administration of cadmium daily during the first week after infection and at 2-day intervals to 13 days after infection resulted in the recovery from ganglia of infectious virus in titers 10- to 100-fold higher than those obtained from animals given saline. Moreover, infectious virus was recovered as late as 11 days after infection compared with 6 days in mice administered saline. (v) Administration of cadmium immediately after infection or repeatedly after establishment of latency did not exhaust the latent virus harbored by sensory neurons, inasmuch as the fraction of ganglia of mice administered cadmium and yielding infectious virus was similar to that observed in mice treated with saline. We conclude that induction of cadmium tolerance precludes reactivation of latent virus. If the induction of metallothionein genes was the sole factor required to cause reactivation of latent virus, it would have been expected that all metals which induce metallothioneins would also induce reactivation, which was not observed. The results therefore raise the possibility that in addition to inducing the metallothionein genes, cadmium inactivates the factors which maintain the virus in latent state.