Herpes Simplex Virus Type 1 KOS Research Articles

Efficacy of oral treatment with BV-araU against cutaneous infection with herpes simplex type 1 in shaved mice

The effect of oral BV-araU was tested in cutaneous model infections of shaved Balb/c mice with herpes simplex virus type 1 (HSV-1). Progression of cutaneous symptoms associated with cutaneous infection with HSV-1 F strain was inhibited by BV-araU at doses of 20 and 50 mg/kg twice daily, beginning one day post-infection, resulting in significant increase in the survival rate. Onset of disease was suppressed in most animals receiving 100 mg of BV-araU per kg. BV-araU (20 mg/kg or more) also significantly increased the survival rate of mice infected with HSV-1 WT-51 strain. The efficacy of BV-araU was not affected by gender or age (6–9 weeks) of the mice. BV-araU was effective even when the treatment was started 2.5 days post-infection. The efficacy of BV-araU against F strain infection was comparable to that of acyclovir, but acyclovir showed therapeutic effects at lower doses compared with BV-araU against WT-51 strain infection. Against infection of cyclophosphamide-treated immunosuppressed mice with HSV-1 KOS(S) strain, BV-araU decreased the morbidity rate and severity of symptoms at doses of 200 and 100 mg/kg, respectively, and all mice given 50 mg of BV-araU or more per kg survived, suggesting oral efficacy can be achieved against HSV-1 infections in immunosuppressed individuals.

Effect of 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil (brovavir) on experimental infections in mice with herpes simplex virus type 1 strains of different degrees of virulence

In vivo antiherpesvirus effects of 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil (brovavir) were tested in two mouse model infections with herpes simplex virus type 1 (HSV-1) strains which showed different degrees of virulence in mice. Successful efficacies of oral and intraperitoneal (i.p.) treatments with brovavir were demonstrated in both intracerebral and i.p. infections with the HSV-1 WT-51 strain of moderate virulence. However, only weak or modest effects of brovavir were observed against the two model infections with a highly virulent strain, HSV-1 VR-3. Brovavir was not effective in reducing mortality of mice infected i.p. with HSV-1 KOS, which exhibited the highest virulence in mice among HSV-1 strains used when inoculated i.p. However, the drug had a significant effect on intracerebral infection with the KOS strain. Efficacies of oral treatment with brovavir were almost equal to those of i.p. administration in the model infections. After intracerebral inoculation, the VR-3 strain grew in brains of mice at a higher rate than the WT-51 strain. By oral treatment with 50 mg of brovavir per kg twice daily, replication of the WT-51 strain in the brains was markedly suppressed and was eliminated after transient elevation of the titer. Growth of the VR-3 strain in the brains was simply delayed by the drug treatment. Thus, the antiviral efficacy of brovavir in mice was affected by the degree of virulence of the challenge virus strain used for infection.

Mild Acidic pH Inhibition of the Major Pathway of Herpes Simplex Virus Entry into HEp-2 Cells

Penetration of the KOS strain of herpes simplex virus type 1 (HSV-1) and the MS and 333 strains of herpes simplex virus type 2 (HSV-2) into HEp-2 cells at pH 6.3 was at least 100-fold less efficient than at pH 7.4. Penetration of two low passage clinical isolates was completely blocked at pH 6.3. The syncytium-forming HSV-1 strains GC and MP were less sensitive than KOS to the mild acidic conditions. The inhibition was completely reversed upon neutralization of the medium. Penetration was assayed by plaque production following protection from acid inactivation upon virus entry. Penetration of HSV-1 KOS into Vero and HEL diploid fibroblast cells was similarly inhibited. HSV-1 KOS grown in 2-deoxy-D-glucose and monensin was also extensively inhibited at pH 6.3 but virus grown in 2-deoxy-D-glucose penetrated more slowly than normal virus at pH 7.4. Electron microscopy of HSV-1 KOS infection indicated that fusion and endocytosis occur at both pH 7.4 and 6.3 but that fusion predominates at pH 7.4 and endocytosis predominates at pH 6.3. These results indicate that fusion at the plasma membrane is the major route of productive entry for HSV, that strains of HSV can differ in their pH dependence for penetration and this may determine whether virus infection can occur following endocytic uptake.

Detection and preliminary characterization of herpes simplex virus type 1 transcripts in latently infected human trigeminal ganglia.

RNA extracted from human trigeminal ganglia was examined for the presence of herpes simplex virus type 1 (HSV-1) transcripts by Northern hybridization (RNA blot) analysis. By using cloned DNA and single-stranded RNA probes, two abundant colinear HSV-1 transcripts (1.85 and 1.35 kilobases) were detected in ganglia from 9 of 17 individuals. These RNAs overlap the 3' end of the transcript for immediate-early gene ICP0 but are transcribed from the opposite strand; thus, they are antisense relative to the ICP0 mRNA. We also report evidence by in situ hybridization that these latently infected ganglia contain HSV-1 RNA homologous to the BamHI SP region of the genome which is transcribed in the same direction as the other latency transcripts. In Vero cells productively infected with laboratory strain HSV-1 KOS and in cultures infected with different low-passage clinical isolates, only the 1.85-kilobase transcript was detected, and there was variation in the size of this larger latency transcript. These novel transcripts may play a role in maintaining HSV-1 latency in human ganglia.

Establishment of latency in vitro with herpes simplex virus temperature-sensitive mutants at nonpermissive temperature

This report describes a latency model using human embryo lung cells that were infected with herpes simplex virus type 1 (HSV-1) temperature-sensitive (ts) mutants and cultivated at nonpermissive temperature (40.5 °C). ts mutants tsG8 (parental strain HSV-1 KOS) and tsG5 (parental strain HSV-1 13) could be maintained in a latent state at 40.5 °C for at least 40 days without exhibiting virus infectivity. During this time, viable virus could be reactivated by reducing the incubation temperature to the permissive level (34 °C). Virus replication could be detected 2 to 6 days after temperature reduction and the virus reactivated from the latent state seemed to retain the same ts phenotype as the input virus for at least 14 days.

The pseudorabies virus gII gene is closely related to the gB glycoprotein gene of herpes simplex virus.

We have looked for conserved DNA sequences between four herpes simplex virus type 1 (HSV-1) glycoprotein genes encoding gB, gC, gD, and gE and pseudorabies virus (PRV) DNA, HSV-1 DNA fragments representing these four glycoprotein-coding sequences were hybridized to restriction enzyme fragments of PRV DNA by the Southern blot procedure. Specific hybridization was observed only when HSV-1 gB DNA was used as probe. This region of hybridization was localized to a 5.2-kilobase (kb) region mapping at approximately 0.15 map units on the PRV genome. Northern blot (RNA blot) analysis, with a 1.2-kb probe derived from this segment, revealed a predominant hybridizing RNA species of approximately 3 kb in PRV-infected PK15 cells. DNA sequence analysis of the region corresponding to this RNA revealed a single large open reading frame with significant nucleotide homology with the gB gene of HSV-1 KOS 321. In addition, the beginning of the sequenced PRV region also contained the end of an open reading frame with amino acid homology to HSV-1 ICP 18.5, a protein that may be involved in viral glycoprotein transport. This sequence partially overlaps the PRV gB homolog coding sequence. We have shown that the PRV gene with homology to HSV-1 gB encoded the gII glycoprotein gene by expressing a 765-base-pair segment of the PRV open reading frame in Escherichia coli as a protein fused to beta-galactosidase. Antiserum, raised in rabbits, against this fusion protein immunoprecipitated a specific family of PRV glycoproteins of apparent molecular mass 110, 68, and 55 kilodaltons that have been identified as the gII family of glycoproteins. Analysis of the predicted amino acid sequence indicated that the PRV gII protein shares 50% amino acid homology with the aligned HSV-1 gB protein. All 10 cysteine residues located outside of the signal sequence, as well as 4 of 6 potential N-linked glycosylation sites, were conserved between the two proteins. The primary protein sequence for HSV-1 gB regions known to be involved in the rate of virus entry into the cells and cell-cell fusion, as well as regions known to be associated with monoclonal antibody resistance, were highly homologous with the PRV protein sequence. Furthermore, monospecific antibody made against PRV gII immunoprecipitated HSV-1 gB from infected cells. Taken together, these findings suggest significant conservation of structure and function between the two proteins and may indicate a common evolutionary history.

Replacement of glycoprotein B gene sequences in herpes simplex virus type 1 strain ANG by corresponding sequences of the strain KOS causes changes of plaque morphology and neuropathogenicity.

DNA sequences encoding glycoprotein B (gB) derived from herpes simplex virus type 1 (HSV-1) strain KOS321 were transferred to HSV-1 ANG. In cotransfection experiments the cloned HSV-1 KOS BamHI G fragment served as donor, and genomic DNA of two ANG variants as recipients. One of these variants, HSV-1 ANG path, expresses gC and the other, C18, was a spontaneous gC-negative mutant. Both ANG strains are of the syncytial (syn) phenotype whereas HSV-1 KOS321 is non-syncytial (syn+). Recombinants were identified by means of a monoclonal antibody which selectively recognizes gBKOS. Among the HSV-1 ANG path/gBKOS recombinants, the majority displayed an altered plaque morphology, i.e. they were of the syn+ phenotype. In contrast all of the C18/gBKOS recombinants were of the syn phenotype. The possibility that the mutant C18 carries a syn mutation not present in the parental strain could be excluded. Marker transfer experiments involving subfragments of the gB gene mapped the syn mutation of HSV-1 ANG path to a locus within the gene that has been previously termed syn 3. Subclones of HSV-1 ANG path were established either directly or after intermittent transfection or cotransfection with the KOS BamHI G fragment. The pathogenicity in mice of these clones was compared. The data obtained indicated that at high frequency, the BamHI G fragment confers apathogenicity.

Two avirulent herpes simplex viruses generate lethal recombinants in vivo.

While it is widely appreciated that infection with a virulent virus can produce disease in an animal, the ability of a mixture of avirulent viruses to produce disease by means of complementation or recombination in vivo has not been established. In this study, two weakly neuroinvasive herpes simplex virus type 1 (HSV-1) strains were simultaneously inoculated onto the footpads of mice. Many (62%) of the animals that received a 1:1 mixture of the viruses died, whereas the animals that received a similar or 100-fold higher dose of each agent alone survived. Of fourteen viruses isolated from the brains of ten mice that died after receiving the mixture of the two weakly neuroinvasive viruses, eleven were recombinants; three of these recombinants were lethal when reapplied to the footpads of mice. These results show that two avirulent HSV-1 variants may interact in vivo to produce virulent recombinants and a lethal infection. They also suggest that different genetic lesions account for the weakly neuroinvasive character of the HSV-1 strains ANG and KOS after footpad inoculation.

2-Deoxy- d-glucose inhibition of herpes simplex virus type-1 receptor expression

Growth of HEp-2 cells in 2-deoxy- d-glucose (2-DOG) supplemented media decreased the cells' binding capacity for herpes simplex virus type-1 KOS (HSV-1) but not vesicular stomatitis virus. HEp-2 cells tolerated up to 30 mM 2-DOG, but 2-DOG was toxic for Vero cells over 2 mM. The reduction in binding was maintained for at least 24 h even after careful removal of the inhibitor and growth in normal media. Complete regeneration of the receptor sites on HEp-2 cells was observed 8 h after mild trypsinization of cells grown in either normal or the 2-DOG supplemented media. Specific glycoprotein characteristics of the HSV-1 binding site were indicated by its inactivation upon trypsinization (0.1 mg per 5 × 10 5 cells for 30 s) and blocking by wheat germ agglutinin but not limulin. These results suggest that 2-DOG inhibits the proper expression of cell surface glycoprotein HSV-1 receptor sites on HEp-2 cells.

Virion component of herpes simplex virus type 1 KOS interferes with early shutoff of host protein synthesis induced by herpes simplex virus type 2 186.

Herpes simplex virus (HSV) strains HSV type 1 (HSV-1) KOS and HSV-2 186 are representative of delayed and early shutoff strains, respectively, with regard to their ability to inhibit protein synthesis in Friend erythroleukemia cells. When these cells were simultaneously infected with HSV-1 KOS and HSV-2 186, HSV-1 KOS interfered with the rapid suppression of globin synthesis induced by HSV-2 186. The observed interference was competitive and not due to exclusion of HSV-2 by HSV-1 at the level of adsorption. Furthermore, UV-irradiated HSV-1 KOS was also effective at interfering with the early shutoff function of HSV-2 186, indicating that a virion component is responsible for the observed interference.

Intratypic and intertypic specificity of lymphocytes involved in the recognition of herpes simplex virus glycoproteins

Cytotoxic T lymphocytes (CTL) were generated in C57BL/6 mice with herpes simplex virus type 1 (HSV-1) (strains KOS, 17, HFEM, and mP) and HSV-2 (strains 186, G, and GP6). Effector lymphocytes were tested for cytotoxicity against syngeneic HSV-1- and HSV-2-infected cells in a 5-h 51Cr release assay. HSV-1 strain HFEM was found to induce CTL efficiently only when 100-fold more virus was used as compared with HSV-1 strains KOS, 17, and mP. All HSV-1 and HSV-2 strains induced cross-reactive populations of CTL. CTL generated by HSV-1 KOS and HSV-2 186 also demonstrated cross-reactivity in an ear-swelling model for delayed-type hypersensitivity. Lymphocytes generated by all HSV-2 strains were highly efficient at lysing HSV-1-infected target cells. However, HSV-2-infected target cells were found to be less susceptible to lysis by either HSV-1 or HSV-2 CTL than were HSV-1-infected target cells. The lowered susceptibility of HSV-2-infected cells was not due to an inefficient infection of BL/6 WT-3 cells as measured by standard growth assays and infectious center assays. Varying the multiplicity of infection or the time of infection did not increase the susceptibility of HSV-2-infected target cells to lysis by CTL. Increasing the effector-to-target-cell ratio resulted in an increased lysis of both HSV-1- and HSV-2-infected target cells by CTL, but the level of HSV-2-infected target cell lysis still did not approach the level of HSV-1-infected target cell lysis. HSV-2-infected cells were as efficient as HSV-1-infected cells in the cold cell competition assay employed in reducing the lysis of 51Cr-labeled, HSV-1-infected target cells. In addition, HSV-2-infected cells were susceptible to lysis by HSV-immune serum and complement.