Ebola virus (EBOV) causes acute hemorrhagic fever that is fatal in up to 90% of cases in both humans and nonhuman primates. No vaccines or treatments are available for human use. We evaluated the effects in nonhuman primates of vaccine strategies that had protected mice or guinea pigs from lethal EBOV infection. The following immunogens were used: RNA replicon particles derived from an attenuated strain of Venezuelan equine encephalitis virus (VEEV) expressing EBOV glycoprotein and nucleoprotein; recombinant Vaccinia virus expressing EBOV glycoprotein; liposomes containing lipid A and inactivated EBOV; and a concentrated, inactivated whole-virion preparation. None of these strategies successfully protected nonhuman primates from robust challenge with EBOV. The disease observed in primates differed from that in rodents, suggesting that rodent models of EBOV may not predict the efficacy of candidate vaccines in primates and that protection of primates may require different mechanisms.

Inhibition of HSV-1 DNA synthesis with either ~abinosyladenine plus the adenosine deaminase inhibitor pentostatin, or with arabinosylthymine, showed a viral mRNA population identical to that seen prior to viral DNA replication (early) by the criteria of quantitative hybridization, specific mRNA species identifiable by Southern blot hybridization of size-~aetionated RNA, and migration of polypeptides resolved by in vitro translation of purified viral mRNA. The amount of viral mRNA associated with infected cell polyribosomes was determined by quantitative DNA excess solution hybridization. At 2 hr postinfection (p.i.) (before viral DNA synthesis) and in drug-treated cells at 6 hr p.i., the majority of the polyadenylated RNA was cell specific with some virus-specific RNA detectable. In contrast, at 6 hr p.i., in the absence of drugs (during m~imum viral DNA synthesis), nearly all the polyadenylated polyribosomal RNA was viral. Blot hybridization of size-fractionated viral RNA confirmed several specific differences between the viral mRNA species occurring before and after HSV-1 DNA synthesis, which have been reported previously from this laboratory. These differences also were reflected in the in &ro translation products encoded by the viral mRNAs. The mRNA species and the eneoded polypeptides that were present in the absence of viral DNA synthesis are a subset of those viral mRNA species and polypeptides expressed in the presence of viral DNA synthesis. The viral mRNA species fall into several groups based on their relative abundance at various times of infection. These data suggest that, in the normal virus infection cycle, the onset of viral DNA synthesis is necessary for normal expression of later viral genes.

Phosphorothioate oligonucleotides complementary to mRNA of the human cytomegalovirus (HCMV) DNA polymerase gene or to RNA transcripts of the major immediate-early regions 1 and 2 (IE1 and IE2) of HCMV were evaluated for antiviral activity in a 96-well immunoassay with primary human dermal fibroblasts as host cells. Oligonucleotides complementary to RNA of the IE2 region exhibited the most potent antiviral activity.One of these oligonucleotides, ISIS 2922, was at least 30-fold more potent than the nucleoside analog, ganciclovir, with a 50%v effective concentration of 0.37 ,uM in the 96-well immunoassay. In an infectious virus yield reduction assay, ISIS 2922 and ganciclovir reduced production of infectious virus by 2 log units at concentrations of 2.2 and 36 ,M, respectively. A control oligonucleotide showed no inhibition of virus production at concentrations as high as 3 ,uM. ISIS 2922 reduced IE protein synthesis in HCMV-infected cells in a dose-dependent manner which correlated with antiviral activity. The antiviral activity of ISIS 2922 was not due to oligonucleotide-induced cytotoxicity since effects on cell viability or proliferation were observed only at concentrations well in excess of effective antiviral concentrations. The specificity and potency of ISIS 2922 suggest that it may be useful for the treatment of cytomegalovirus disease in humans.

The herpes simplex virus type 1 latency-associated transcript (LAT) is expressed as a major species 2,100 to 2,200 bases in length and a less abundant one ca. 730 bases shorter in latently infected mouse and rabbit neurons. RNA blot hybridization experiments using 20to 22-base synthetic oligonucleotides and mung bean nuclease protection assays have demonstrated that the smaller LAT species is colinear with the larger one, except for a 730-base intron. On the basis of Northern blot analysis, the spliced species which comprises as much as 50% of the total LAT in latent infections of mice with several strains of herpes simplex virus type 1 and latent infections of rabbits with either the McKrae or the KOS(M) strains of virus is not present in the acute phase of infection. Further and rather surprisingly, in mice latently infected with the KOS(M) strain of virus, the spliced LAT species is considerably less abundant. This suggests that both the strain of virus and the animal in which the latent infection occurs are important in either the processing or stability of spliced LAT. Finally, an exhaustive series of experiments failed to provide convincing evidence that a unique, poly(A)+ species of LAT exists in the latent phase of infection.

We report the complete nucleotide sequence of the M and the S genome segments and a portion of the L segments of two hantavirus isolates from Peromyscus maniculatus trapped in eastern California. The isolates, Convict Creek 107 and 74 (CC107 and CC74) are genetically similar to viruses known to cause hantavirus pulmonary syndrome in New Mexico. CC107 and CC74 each have an M segment consisting of 3696 nucleotides with a coding potential of 1140 amino acids in the virus complementary-sense RNA (cRNA). The S segments of CC107 and CC74 are 2083 and 2047 nucleotides long, respectively, and each has an ORF in the cRNA capable of encoding a protein of 428 amino acids. Unusually long 3' noncoding regions of 757 and 721 nucleotides follow the S segment ORF of CC107 and CC74, respectively, and include numerous imperfect repetitive sequences. Whereas the M and S segments of any given hantavirus typically appear to diverge at comparable rates from homologous genes of any other hantavirus, CC107 and CC74 have M segments that differ by only 1% from one another but S segments that differ by 13%. After trivial explanations are rendered improbable, i.e., by consideration of the genetics of closely and distantly related hantaviruses, the most likely explanation for our data is that hantavirus genome segment reassortment occurred within rodent populations in California.