Herpesvirus type 2 in the male genitourinary tract.

A population study of 190 randomly selected male patients with no history of genital herpesvirus infection revealed a high incidence of herpesvirus type 2 in genitourinary specimens. This indicates that men serve as a reservoir of genital herpesvirus.

In vitro transformation of hamster cells by herpes simplex virus type 2 from human prostatic cancer cells.

A cell-associated herpes simplex virus type 2 found in a human prostatic carcinoma induced in vitro transformation of hamster embryo cells. The transformed cells (YW-74) have been shown to be hamster cells by karyotype analysis. Their epithelial morphology and growth pattern, which are different from the parental cell, have remained stable through cell passages. The presence of herpesvirus antigens in the transformed cells was determined by specific immunofluorescence and colony inhibition tests. Immunofluorescence staining with specific anti-herpes simplex virus type 2 serum showed an intense and distinctive nuclear and perinuclear fluorescence in about 95% of the transformed cells. In addition, exposure of these transformed cells to herpes simplex virus type 2-sensitized lymphocytes resulted in inhibition of growth and colony formation, while no effect was seen with nonsensitized lymphocytes. Both observations are consistent with the involvement of herpesvirus type 2 in the transformation event. This virus, which does not produce a lytic infection and is not found either in extracellular spaces or supernatant fluid of the transformed cell cultures, is unique in the fact that it is cell associated, noncytopathogenic, and capable of transforming cells in vitro, and its antigens are clearly demonstrated in the transformed cells.

Dextran flux in M-K medium-stored human corneas.

Dextran, with a minimal molecular weight of 40,000, can pass in and out of the corneal endothelium during storage in M-K medium. Results suggest that the degree of penetration of dextran depends on the length of storage and the condition of the endothelium.

Leukocyte migration inhibitory factor in HSV infections.

The cell-mediated immune (CMI) response as measured by a direct assay of leukocyte migration inhibition factor (LMIF) was determined in a population of patients with recurrent herpes simplex virus (HSV) infections in the quiescent stage as well as in healthy volunteers. The migration of leukocytes incubated in the presence of HSV antigens was compared to that without viral antigens for the calculation of the migration index (MI). Eleven of 41 control subjects (16.8%) had a MI below 0.8, indicating a positive CMI response. In contrast, all the herpes patients tested had a MI above 0.8, suggesting an impairment in the production of LMIF at this stage of their disease. This difference was statistically significant (t = 4.296; p less than 0.001) and was not dependent on the age of the population. This study indicates that individuals with recurrent HSV infections have impaired CMI response betweeen attacks which may be associated with the stage of the disease.

Strain specificity of clinical isolates of herpes simplex virus.

Herpes simplex virus (HSV) produces a wide variety of ocular disease in man. Although host factors are important in determining this variation, it is possible that the different clinical patterns of herpetic ocular disease may be attributed at least partially to the differing biological behavior of specific strains of HSV. To test this theory, we compared the anterior segment disease produced by infecting rabbit corneas with seven different strains of HSV. We found that these seven different strains produced different patterns of ocular disease in the rabbit eye. This also may occur in humans, and we hope to define the biological differences that cause one strain to produce disease more severe than that produced by another strain.

Prevention of surface bacterial contamination of donor corneas.

A simple method has been developed to reduce contamination in postmortem donor human eyes in anticipation of corneal transplantation. In vivo investigation of albino rabbits demonstrates that vigorous saline solution irrigation is extremely effective in decreasing the surface bacterial counts of the postmortem eye. In vitro and in vivo studies show that Neosporin kills bacteria at room temperature and further show that a tenfold increase in the thimerosal concentration of the Neosporin will kill fungus. Postmortem eyes contaminated by pathogenic organisms can be effectively cleaned by a combination of saline solution irrigation and the new Neosporin-thimerosal solution. No substantial damage of the donor tissue was noted by scanning electron microscopy. Human eyes cultured before this procedure were all contaminated, but after cleansing and immersion, no bacterial or fungal growth occurred.

Epidermal growth factor receptors on corneal endothelium.

If a growth factor could bind to and stimulate human endothelial healing, corneal disease could be minimized. To this end, primary cultures of feline and human corneal endothelium were tested in receptor binding assays for radiolabeled epidermal growth factor (EGF). Both of these cells bound ten times as much 125I-EGF as did the negative control cell lines. The time course of association of 125I-EGF to cat corneal endothelium was found to be complete after approximately 120 minutes at 22 degrees C. The 125I-EGF was shown not to dissociate greatly when fresh binding buffer was added to endothelial cultures that had bound the radiolabeled peptide. The pH optimum for binding was determined to be approximately 6.4. The receptor number per cell and the affinity constant for binding were determined to be 40,000 receptors per cell and 1.1 x 10(9) L/mole, respectively, using a Scatchard plot. Parallel cultures of human fetal corneal endothelium grew in vitro only when the growth medium was supplemented with low concentrations of EGF. These studies provide evidence that EGF is specifically bound to the corneal endothelium.

Herpesvirus type 2: isolation from seminal vesicle and testes.

Reproductive tissues from 10 recent male cadavers were examined. Herpesvirus type 2 was isolated from testes, seminal vesicle, or both in 4 cases. This is the first report of the isolation of herpesvirus type 2 from human seminal vesicular tissue. The data support previous evidence that herpesvirus type 2 can be isolated from the reproductive tissues of males without active or prior infection and suggest that these tissues may serve as a reservoir for transmission of this virus. The study also documents, for the first time, the ability to culture herpesvirus type 2 in organ explants from cadaveric reproductive tissues with the subsequent release of infectious virus.

Failure of systemically administered adenine arabinoside to affect humoral and cell-mediated immunity.

The effect of a high dosage (250 mg/kg of body weight) of adenine arabinoside or ara-A (9-beta-D-arabinofuranosyladenine) on humoral immunity was studied in New Zealand white rabbits infected with the McKrae strain of herpes simplex virus. The rabbits were treated daily for 14 days with subcutaneous injections of ara-A. The primary and secondary humoral responses, as measured by neutralizing antibody titers, developed similarly in control and treated groups. Similar drug treatment was used on guinea pigs before or after sensitization with BCG vaccine. Subsequent challenge of the sensitized animals with Old tuberculin solution indicated that ara-A treatment had no effect on the induction or previously established cell-mediated immunity. The lack of immunosuppressive activity of ara-A at dosage levels higher than those used in primates makes this drug a potentially effective agent in the systemic treatment of herpetic infections.

Effect of the herpes simplex virus genome on the response of infection to corticosteroids.

The type and severity of ocular herpetic disease, as well as the pattern of recurrence, have been shown to be determined by the virus genome. We infected rabbit eyes with two closely related recombinant strains of herpes simplex virus type 1 and treated one half of the eyes in each group with corticosteroids before or immediately after virus inoculation. The severity of disease in the first week was similar in the treated and untreated eyes infected with the F(MP)F strain; however, with F(MP)E infection, the disease in the treated eyes was significantly worse than the disease in the untreated eyes. Cultures of corneal virus showed similar titers in all of the groups, but cultures of trigeminal ganglia indicated that increased severity of disease did not result in an increased tendency toward ganglionic colonization. The results suggest that the response to corticosteroids is another factor that is determined by the genetics of the infecting virus, but that there is no correlation between worsening of disease with corticosteroid treatment and the establishment of virus latency.

The effect of modulating the synthesis of arachidonic acid cascade products on HSV lesion recurrence.

Induction of HSV lesion recurrence may be achieved by a variety of stimuli. Trauma of almost any kind (physical, chemical, electromagnetic and thermal) to the healed primary lesion site has been successful for induction of recurrence. In common with each of these mechanisms is the release of inflammatory mediators (arachidonic acid (AA), complement, kinins, etc.) following trauma. Because blockade of the AA cascade with steroids has been noted to abort HSV skin lesions, and because steroids have numerous side effects making them a poor therapeutic choice in ocular lesions, we decided to test several relatively different types of AA cascade inhibitory drugs in mouse ear HSV recurrence models. In this series of experiments, it was found that topical steroids gave the greatest initial decrease in lesion number (80% fewer than control on day 3 post recurrence induction (PRI), while meclofenamate resulted in the greatest reduction of lesions by day 5 PRI (85% fewer lesions than control and 60% fewer than the steroid treated group). The NDGA treated group exhibited the least reduction in recurrence severity (27% fewer lesions than control on day 5 PRI and 200% more lesions than the steroid group. Chlorpromazine (thorazine) acted roughly equivalent to the steroid treated group by day 5 PRI (70% fewer lesions than the untreated control group). Relative efficacy in lesion reduction between groups by day 5 PRI is: meclofenamate greater than steroid = chlorpromazine greater than NDGA greater than control. Meclofenamate, steroid and chlorpromazine significantly reduced lesions (p less than .05) when compared with the saline treated control mice. NDGA did not significantly reduce lesions by day 5 PRI.

Antiviral agent from lambda-infected Escherichia coli K-12. I. Isolation.

Phagicin, which is an antiviral agent active against deoxyribonucleic acid (DNA) viruses such as vaccinia and herpes simplex, has been identified as a phage internal protein. It was found in infected Escherichia coli lysates, but could also be obtained by disruption of the purified infective particles after incubation with LiCl at 46 C for 15 min or by sonic treatment. After centrifugation at high speed, the antiviral activity was found in the DNA phase and could be separated by chromatography on Sephadex gels with 0.2 M phosphate buffer (pH 7.5) as the eluent. Phagicin present in lysates after removal of infective particles was nondialyzable and was bound to nucleic acids. It could be released during precipitation of nucleic acids by streptomycin sulfate, and in this form it could be easily dialyzed. The antiviral activity of phagicin was specific for herpes simplex and vaccinia viruses.

Heterogeneity within an HSV-1 wild-type strain and its importance in pathogenesis.

A herpes simplex virus-type 1 low passage, clinical eye isolate, E-43 at P2, was compared with its variant progeny, SLi-43 at P8, in terms of ocular disease, cytopathic effects, and genomic variation. In New Zealand White (NZW) rabbits, E-43 produced mild epithelial defects and punctate lesions with full recovery by Day 10 postinfection (pi). SLi-43 caused dendritic lesions, progressing to geographic ulceration and death from herpes simplex virus encephalitis in 10 days postinfection. In RK, Hep-2, and Vero cells, E-43 displayed the syn+ phenotype (aggregation and cell rounding); SLi-43 showed the syn phenotype (syncytium formation). DNA digestion profiles of E-43, SLi-43, and isolates from the brains of infected animals showed that the genomic differences map within the terminal repeat of the unique long segment and the internal joint region, specifically in bands B, E, N, and S (Bam HI) and bands M and N (Hind III). Analysis of the DNA of virus recovered from the brain stem of SLi-43-infected, encephalitic rabbits demonstrated that an in vivo selection for neurotropic virions had taken place. Plaque purification of 20 clones from the original E-43 strain showed that one of 20 was the syn phenotype, indicating that the SLi-43 variant was present in the original E-43 isolate and did not develop de novo by rapid mutation. The parent-progeny relationship between E-43 and SLi-43 forms an ideal model in which to compare differences in pathogenicity at the genomic level, and underscores the importance of heterogeneity within a single herpes simplex virus-type 1 wild-type population in terms of variations in ocular disease.

Secretory immunoglobulin a and herpes keratitis.

The sensitization of herpes simplex virus by immunoglobulin G (IgG) antibodies was studied. Although herpes simplex virus sensitized by IgG could not be neutralized by IgA antibodies known to neutralize a similar number of unsensitized viruses, this protection could be overcome by larger amounts of IgA antibodies. This suggests that the effective neutralization of herpes simplex virus in ocular tissue is dependent on the relative concentrations of IgA and IgG antibodies. In vivo sensitized virus was isolated from chronically infected animals, and this may be the mechanism by which herpes simplex virus persists in animals with high titers of circulating antibodies in their serum.

9-(2-hydroxyethoxymethyl)guanine as an inhibitor of herpes simplex virus replication.

The drug 9-(2-hydroxyethoxymethyl)guanine effectively inhibits herpes simplex virus replication. It is selectively phosphorylated by the virus-induced thymidine kinase but not by normal cellular thymidine kinase.