Persistent reactivable latent herpes simplex virus infection in trigeminal ganglia of mice treated with antiviral drugs

Summary Latent reactivable infection was established with HSV in mouse trigeminal ganglion. A number of antiviral drugs (IUDR, acycloguanosine, Ara-A, PAA and PFA), effective against acute CNS infection with HSV, failed to influence the latent infection once established. Neither lipophilic properties of PFA-derivatives nor the combination of Ara-A and acyloguanosine improved the drug-effects on HSV latency.With 1 Figure     

The proportion of galanin-immunoreactive neurons in mouse trigeminal ganglia is transiently increased following corneal inoculation of herpes simplex virus type-1

To investigate whether neurobiological functions are modified by viral infection, we inoculated mouse corneas with herpes simplex virus type 1 (HSV-1) and examined neuronal galanin content in trigeminal ganglia at selected survival times. HSV-1 antigen appeared in neurons at day 3, peaked at day 6 and disappeared by day 11. Increased galanin positivity was first seen at day 6, peaked at day 10 and approached control values by day 21. This result provides further evidence that the biological program of peripheral sensory neurons is modified by herpes-virus infection.     

Effect of immune serum on the establishment of herpes simplex virus infection in trigeminal ganglia of hairless mice

Administration of immune serum to herpes simplex virus (HSV)-infected hairless mice could not prevent acute infection in the trigeminal ganglia and the eventual establishment of latency. However, immune serum reduced the amount of free virus in the ganglion during the acute phase of the infection. It appears also that the amount of virus that can be reactivated in the latently infected ganglion is decreased. This was indicated by a prolonged reactivation time and by a reduced virus content of ganglion homogenates prepared after various periods of cocultivation.     

Induction of 2',5'-oligoadenylate synthetase and interferon in mouse trigeminal ganglia infected with herpes simplex virus.

The activity of 2',5'-oligoadenylate synthetase, which is induced by interferon action, was detected in mouse trigeminal ganglia infected with herpes simplex virus (HSV) type 1. When HSV was inoculated on the left cheek of mice, the virus began to appear in the ipsilateral trigeminal ganglia on day 2, reached its maximum accumulation on day 4, declined thereafter, and was no longer detected in the tissue homogenate after 11 days. After this short acute productive phase, the virus entered into the latent phase of infection. 2',5'-Oligoadenylate synthetase appeard in the trigeminal ganglia shortly after the beginning of virus multiplication; the synthetase activity began to rise on day 3, reached a maxium level on day 4, and then declined. Interferon activity (type I) also appeared in the infected ganglia, but diminished more rapidly than the synthetase. The antibody against HSV in the sera began to appear at the time when the virus and synthetase were declining. From these results it may be hypothesized, although not concluded, that in the acute productive phase of HSV infection in mouse trigeminal ganglia, virus multiplication is suppressed by interferon action, including 2',5'-oligoadenylate synthetase induction.     

In vivo reactivation of herpes simplex virus in rabbit trigeminal ganglia: electrode model.

The rabbit provides an excellent model for the study of ocular herpes because herpetic keratitis in the rabbit eye resembles human disease in its clinical features and in its propensity for spontaneous recurrence. This paper presents a method for the electrical induction of multiple episodes of in vivo reactivation of latent HSV-1 infection with peripheral shedding of virus. Physiological levels of current delivered via an electrode implanted over the trigeminal ganglion of latently infected animals has enabled us to modify and synchronize virus shedding in preocular tear film and to cause multiple episodes of reactivation in a single animal. For this reason, the model is well suited for antiviral efficacy testing and provides an excellent opportunity for investigation of virus-host cell interactions in latent and recurring herpetic disease.     

Neutralising antibody in mice with primary and recurrent herpes simplex virus infection

Summary Neutralising antibody was studied in mice infected in the ear with herpes simplex virus type 1. Antibody was detected six days after infection and after one month it had reached titres which subsequently varied little. Geometric group mean titres were doubled by reinoculation of virus whereas recurrent disease induced by trauma did not change them (although titres did change in some individual animals).Mean titres in mice which had had unequivocal signs of primary disease were twice as high as in those which had not, and the frequency of recurrent disease in response to trauma in the former animals was greater than in the latter. A reduction in the frequency of induced recurrent disease was seen if the application of trauma was delayed for some time after the establishment of latency.Hyperimmunisation increased mean titres tenfold. If given soon after primary infection it reduced the frequency of induced recurrent disease but if its administration was delayed then the frequency was increased.     

Viable herpes simplex virus type 1 and varicella‐zoster virus in the trigeminal ganglia of human cadavers

Human herpes simplex virus type 1 (HSV‐1) and varicella‐zoster virus (VZV) were isolated in the bilateral trigeminal ganglia of 12 human cadavers with no history of herpes‐related symptoms within 1–5 days of death. Sixteen trigeminal ganglia were subjected to explant culture by using Vero cells, but no cytopathogenic effects (CPE) were observed. However, when another eight trigeminal ganglia were placed in a cell strainer and kept from direct contact with Vero cells during culture, CPE were clearly apparent in all cultures. The amount of DNA in the culture supernatants of 16 trigeminal ganglia decreased over time; 12 and 9 of these samples were PCR‐positive for HSV‐1 and VZV, respectively. In new Vero cells inoculated with supernatants collected 2 days after culture initiation, immunofluorescence staining revealed HSV‐1 and VZV in 6 and 5 of 8 trigeminal ganglia, respectively. HSV‐1 and VZV DNA was detected in supernatants collected 3 and 7 days after culture initiation and in Vero cells collected after culture completion, but real‐time PCR revealed the DNA amounts decreased over time. There was less VZV DNA than HSV‐1 DNA. These results demonstrate that infective HSV‐1 and VZV can be isolated in culture, and confirm that viable HSV‐1 and VZV persist in human trigeminal ganglia for some time after death. J. Med. Virol. 85:833–838, 2013. © 2013 Wiley Periodicals, Inc.     

Antibody activity to herpes simplex virus in mouse Ig classes and IgG subclasses

Summary Recent studies indicate that Ig class and IgG subclass induction varies for different proteins and further that some Ig subclasses, like IgG2a, are more efficient in important biologic processes such as antibody-dependant cell-mediated cytotoxicity (ADCC). Many proteins of herpes simplex virus type 1 (HSV-1) are immunogenic and induce immunoglobulin responses. To determine the distribution of immunoglobulins induced by HSV-1 proteins, we studied immune mouse serum using an Ig isotype specific Elisa assay for antiviral activity. We found by endpoint analysis that the antiviral titer was 1:12,903 for IgG1, 1:5141 for IgG2a, 1:2140 for IgG2b and 1:229 for IgG3. To identify which isotypes were induced by individual glycoproteins and other viral proteins, Western blots containing HSV-1 proteins were probed with immune serum and isotype specific second antibodies. gB, gC, gD and the 42/44KDa nucleocapsid complex induced strong IgG1, IgG2a, IgG2b responses. IgG3 reactivity with viral proteins appeared weaker. Among the IgG3 reactivities detected on immunoblots, gB and gC were the most intense. Other proteins which elicited IgG1, IgG2a and IgG2b responses were 170KDa, 154KDa and gE. IgA responses were induced by 154KDa, gC, gB, gE and gD. Prominent IgM responses included gB, gC, gD and the 42KDa protein. These results indicate that HSV-1 glycoproteins induce prominent responses in all IgG isotypes except IgG3. The biologic implications of the data are discussed.     

Effects of long-term acyclovir chemosuppression on serum IgG antibody to herpes simplex virus

Patients with severe or frequent recurrent genital herpes simplex virus (HSV) infection can be managed either by treating each recurrence with acyclovir or by suppressing recurrences with daily administration of the drug. To determine the effects of long-term acyclovir therapy on the immune response to HSV, we studied the change in IgG antibody concentration to HSV in 46 individuals with recurrent genital HSV-2 infection who received acyclovir for 1 year, Twenty-seven subjects received daily acyclovir chemosuppression, while 19 subjects received daily placebo (with acyclovir administered intermittently only during recurrences). Immunoglobulin G (IgG) antibody to HSV was determined before medication began, at completion of 1 year of therapy, and 22 weeks following the first untreated HSV recurrence. Daily acyclovir chemosuppression for 1 year reduced mean IgG antibody concentration by 10% from baseline values (P less than 0.01), whereas in patients receiving intermittent therapy no significant decline was observed. In both groups, however, the first untreated recurrence produced a rise in mean antibody concentrations. We conclude that prolonged daily acyclovir chemosuppression reduces humoral immunity to HSV, but antibody concentrations increase following the first untreated recurrence.     

Detection of antibody to viral proteins following primary infection with herpes simplex virus

Sera from patients with primary genital infection with herpes simplex virus (HSV) were tested by immunoblotting (IB) and radioimmunoprecipitation (RIP) analysis to determine the protein targets of antibody elicited by infection. The two tests detected antibody to different antigens: IB primarily detected reactivity with p40, a phosphorylated capsid protein, and RIP detected antibody to glycoprotein B, a viral envelope component. Furthermore, RIP detected antibody at an earlier stage of infection then IB. A nondenaturing version of IB was developed and used to investigate the role that the solubilisation of antigen plays in the sensitivity of each test for antibodies with different specificities.     

Efficacy of phosphonoacetic acid on herpes simplex virus infection of sensory ganglia.

Treatment of mice with phosphonoacetic acid markedly reduced the incidence of latent ganglionic infection with herpes simplex virus when administered within 24 h after viral inoculation, but had no effect on an already established ganglionic infection.     

Detection of latent thymidine kinase-deficient herpes simplex virus in trigeminal ganglia of mice using the polymerase chain reaction

Summary Latency of thymidine kinase-negative mutants of herpes simplex virus (TK^– HSV) could not be detected by reactivating the virus from the ganglia of infected mice. Because Southern blot hybridization was not sensitive enough to detect viral DNA, positive results obtained by dot blot hybridization were ascertained by the highly specific and sensitive polymerase chain reaction (PCR), which detected both latent TK^– HSV type 1 and 2 DNA from the trigeminal ganglia of infected mice.     

Acute and latent infection of sensory ganglia with herpes simplex virus: immune control and virus reactivation.

The role of antiviral antibody in controlling the acute and latent phases of herpes simplex virus (HSV) infection in sensory ganglia of mice was studied in vitro and in vivo. Organ cultures of ganglia inoculated in vitro with HSV produced infectious virus for at least 3 weeks. In the presence of antiviral antibody, the titre of virus was markedly reduced, but the infection was not eliminated. Similarly, passive administration of antibody to HSV-infected immunodeficient (nude) mice reduced the virus titre but did not eliminate the acute phase of the ganglionic infection. Suppression of the cell-mediated immune response in latently infected immunocompetent mice by treatment with cyclophosphamide and/or X-irradiation resulted in reactivation of HSV in up to 70% of the animals. Reactivation was demonstrated by recovering infectious virus in cell-free homogenates of ganglia and eye globes and by finding virus antigens in ganglia by immunofluorescent staining. Reactivation occurred both in vitro and in vivo in the presence of high concentrations of neutralizing antibody. It is concluded that antibody alone is not sufficient to eliminate the acute phase of the ganglionic infection and that cytotoxic agents known to suppress the host's cellular immune response can reactivate virus in the presence of neutralizing antibody.     

Peripheral infection in culture of rat sensory neurons by herpes simplex virus.

The peripheral infection of neurons from dissociated rat sensory ganglia by herpes simplex virus was studied with a newly developed tissue culture system. The results indicated that the virus arrives at the neuronal soma by an axoplasmic route and that at least some of these neurons are productively infected since eventually all cells in the culture appeared to be destroyed by the virus. This system showed promise in elucidating the possible roles of various effector cells, antiviral antibody, or antiviral compounds in the mechanism of herpes simplex virus latency.     

Immunological mechanisms giving rise to latency of herpes simplex virus in the spinal ganglia of the mouse

In the model of genital herpes simplex virus (HSV)-infection of mice, early latency could be induced by passive immunization with HSV-specific antibodies and, to a lesser degree, by adoptive transfer of immune lymphocytes prepared from spleen and draining lymph nodes of genitally infected syngeneic mice. Conversely, spontaneously occurring latency was inhibited by treatment of the animals with cyclophosphamide (Cph) and, to a lesser degree, with cyclosporin A (CyA). Whereas the effect of CyA could be compensated by passively administered HSV-specific antibodies, that of Cph could not. Apparently specific antibodies cooperate with a non-specific proliferating cell type, probably macrophages and/or NK-cells, as could be demonstrated by significantly reduced antibody effect in silica-treated mice. Moreover, F(ab)₂ fragments, in contrast to complete antibody molecules, were inactive. HSV-specific antibodies and also immune lymphocytes had little effect on virus production in the mucous membranes, immune lymphocytes being at least as active as antibodies. It is therefore not probable that latency is induced by attenuation of the peripheral disease. It can rather be concluded that the neuron itself is the target for the action of specific antibodies, cooperating in turn with macrophages and/or NK cells.With support of the Deutsche Forschungsgemeinschaft, Schn 174/6-3     

Immunological mechanisms giving rise to latency of herpes simplex virus in the spinal ganglia of the mouse

In the model of genital herpes simplex virus (HSV)-infection of mice, early latency could be induced by passive immunization with HSV-specific antibodies and, to a lesser degree, by adoptive transfer of immune lymphocytes prepared from spleen and draining lymph nodes of genitally infected syngeneic mice. Conversely, spontaneously occurring latency was inhibited by treatment of the animals with cyclophosphamide (Cph) and, to a lesser degree, with cyclosporin A (CyA). Whereas the effect of CyA could be compensated by passively administered HSV-specific antibodies, that of Cph could not. Apparently specific antibodies cooperate with a non-specific proliferating cell type, probably macrophages and/or NK-cells, as could be demonstrated by significantly reduced antibody effect in silica-treated mice. Moreover, F(ab)2 fragments, in contrast to complete antibody molecules, were inactive. HSV-specific antibodies and also immune lymphocytes had little effect on virus production in the mucous membranes, immune lymphocytes being at least as active as antibodies. It is therefore not probable that latency is induced by attenuation of the peripheral disease. It can rather be concluded that the neuron itself is the target for the action of specific antibodies, cooperating in turn with macrophages and/or NK cells.