Varicella zoster antibody titers after the administration of intravenous immune serum globulin or varicella zoster immune globulin.

Varicella is a serious infection in the immunocompromised patient. Prophylaxis with varicella zoster immune globulin is known to decrease the incidence of severe varicella infection. The titers of antibody to varicella zoster virus were compared in patients who received either varicella zoster immune globulin or intravenous immune globulin, 4 ml or 6 ml/kg per dose. The titers of antibody to varicella zoster virus were comparable in each group.

The synthesis and immunogenicity of varicella-zoster virus glycoprotein E and immediate-early protein (IE62) expressed in recombinant herpes simplex virus-1.

In order to evaluate the conditions for optimal expression and immunogenicity of varicella-zoster virus (VZV) proteins in a herpes simplex virus-1 (HSV-1) vector, we selected the VZV glycoprotein E (gE), encoded by ORF 68 and the VZV product of ORF 62, an immediate-early major tegument protein (IE62). Three HSV/VZV recombinants were generated: (1) VZV gE protein coding sequences along with the promoter region were inserted into the thymidine kinase (TK) gene of HSV-1 strain KOS; (2) VZV gE expressed from the HSV-1 ICP4 promoter was inserted into the glycoprotein C (gC) gene of HSV-1 strain F; and (3) VZV IE62 protein coding sequences under the control of the HSV-1 ICP4 promoter were inserted into the gC gene of HSV-1 strain F. Immunoblot analysis and immunoperoxidase staining of infected cell monolayers demonstrated vector expression of VZV proteins. Following intracranial inoculation in mice, both VZV gE-HSV (TK) and VZV IE62-HSV (gC) induced an IgG response against VZV gE or VZV IE62. When tested in cytotoxicity assays using T-lymphocytes from VZV immune human donors, the range of precursor frequencies for T-lymphocytes that recognized VZV gE or VZV IE62 was similar whether these proteins were expressed by HSV-1 or a vaccinia vector. These experiments demonstrate that HSV-1 is a competent vector for expression of these VZV proteins and support the feasibility of engineering a combined vaccine for these closely related alpha-herpesviruses.

Immunoglobulins M and G to varicella-zoster virus measured by solid-phase radioimmunoassay: antibody responses to varicella and herpes zoster infections.

Both immunoglobulin M (IgM) and IgG antibodies to varicella-zoster virus (VZV) were detectable in a solid-phase radioimmunoassay with 125I-labeled goat antisera to human immunoglobulins. Primary infection with VZV was associated with early production of IgM and IgG antibodies and rapid development of lymphocyte transformation to VZV antigen. Among eight subjects with varicella tested 1 to 4 days after onset, seven patients had IgG and six patients had IgM antibodies; all patients had both IgG and IgM antibodies within 7 days. An IgM response was documented by radioimmunoassay in 18 of 26 patients with herpes zoster. VZV antibodies could be assayed by radioimmunoassay in unfractionated serum with commercial goat antisera to human immunoglobulins and commercial VZV antigen. VZV-specific IgG binding was present in all sera from 42 subjects with a VZB antibody titer of greater than or equal to 1:8 as determined by indirect immunofluorescence and cellular immunity to VZV as determined by lymphocyte transformation and who had had varicella at least 20 years before testing. The geometric mean titer was 1:6,309, and titers were greater than or equal to 1:16,384 in 20 subjects. Antibody was present as determined by radioimmunoassay in 14 samples negative by complement fixation and in five samples negative by complement fixation and immune adherence hemagglutination. No specific binding was observed in 21 sera from subjects who were not immune to VZV as determined by indirect immunofluorescence or lymphocyte transformation despite the presence of herpes simplex or cytomegalovirus antibody indicated by complement fixation in 15 sera. High titers of VZV IgM antibody were detected in unfractionated sera despite the presence of high titers of VZV IgG antibody. The VZV radioimmunoassay provided a sensitive and practical method for measuring VZV IgG and IgM antibodies.

Comparison of WI-38, MRC-5, and IMR-90 cell strains for isolation of viruses from clinical specimens.

With the diminishing supply of the human fetal lung WI-38 cell strain, a replacement for viral isolation is needed. Two candidates are the human fetal lung strains MRC-5 and IMR-90. A comparison of WI-38, MRC-5, and IMR-90 was performed to evaluate efficiency and speed of viral isolation, clarity of cytophatic effect, and ease of growing the cells. The inocula were clinical specimens rather than tissue culture-adapted isolates. Frozen samples of 46 specimens that had previously yielded an isolate on WI-38 were thawed and inoculated onto WI-38, MRC-5, and IMR-90 cells. In addition, 95 freshly taken clinical specimens uf undetermined infectivity were inoculated onto the cell strains. Viral recovery rates were similar on all three strains, as were the appearance and speed of onset of the cytophatic effect. MRC-5 and WI-38 cells remained healthy until generation 36, whereas IMR-90 cells went into crisis by generation 20. The longer life span of the MRC-5 cells makes them more suitable than IMR-90 cells to replace the WI-38 strain for routine use in viral diagnosis.

Immunologic evidence of reinfection with varicella-zoster virus.

Resistance to reinfection with varicella-zoster virus (VZV) was evaluated in immune adults who had household exposure to varicella. Sixty-four percent of 25 adults exposed to varicella had a fourfold or greater rise in IgG antibody to VZV or had a high initial IgG antibody titer to VZV that declined by fourfold. IgM antibody was detected in only 12% of 25 VZV-immune subjects. Seventy percent of 23 subjects exposed to varicella had IgA antibody to VZV compared with 13% of 23 subjects with antibody to VZV who had no recent exposure (P less than 0.001, chi 2 test). Enhanced cellular immunity was documented by an increase in lymphocyte transformation to VZV antigen from a mean +/- SE index of 7.8 +/- 1.30 to 15.3 +/- 2.56 (P = 0.01, paired t-test). The increase in immunity to VZV in many immune subjects exposed to VZV suggests the occurrence of subclinical reinfection.

Serum immunoglobulin A antibody to varicella-zoster virus in subjects with primary varicella and herpes zoster infections and in immune subjects.

Immunoglobulin A (IgA) antibodies to varicella-zoster virus (VZV) were measured in sera from subjects with acute varicella and herpes zoster, VZV-immune subjects remote from infection, and recipients of a live attenuated varicella vaccine, using a solid-phase radioimmunoassay. Primary infection with VZV was associated with early production of IgA antibodies. Among 36 subjects with varicella tested 1 to 5 days after onset, 22 had detectable IgA, and all of the negative sera were obtained before day 3 of the varicella exanthem. VZV IgA was detected in one of three sera obtained more than 60 days after onset of the illness. Four of five sera obtained from subjects within 1 week of the onset of herpes zoster had measurable levels of IgA. Between 1 and 4 weeks after onset of zoster, all 10 subjects tested had detectable IgA to VZV. VZV IgA was detected as late as 63 days after the onset of herpes zoster. Of 10 vaccine recipients, 5 developed VZV IgA which was detected as early as 4 weeks and persisted for as long as 16 weeks after vaccination. VZV IgA was not detected in sera from 42 children who had no detectable IgG antibody to VZV. VZV IgA was found on only 3 of 23 sera from adults who had varicella more than 20 years before.

Subclinical varicella-zoster virus viremia, herpes zoster, and T lymphocyte immunity to varicella-zoster viral antigens after bone marrow transplantation.

Bone marrow transplant (BMT) recipients were evaluated for subclinical varicella-zoster virus (VZV) viremia and symptoms of herpes zoster after transplantation. Viremia was demonstrated by testing peripheral blood mononuclear cells using polymerase chain reaction and was documented in 19% of 37 patients. When reactivation was defined as herpes zoster and/or subclinical VZV viremia, 41% of VZV-seropositive BMT recipients experienced VZV reactivation. None of 12 patients tested before VZV reactivation had T lymphocyte proliferation to VZV antigen (mean stimulation index, 1.0 +/- 0.42 [SD] at less than 100 days; 12.0 +/- 6.03 at greater than 100 days [P = .003]). Among patients tested at greater than 100 days, 5 (63%) of 8 with detectable T cell proliferation had subclinical or clinical VZV reactivation compared with none of 6 who lacked VZV T cell responses. Recovery of VZV-specific cytotoxic T lymphocyte function was observed in 50% of BMT patients, but BMT recipients had significantly fewer circulating cytotoxic T lymphocytes that recognized VZV immediate early protein (P = .03) or glycoprotein I (P = .004) than did healthy VZV immune subjects. In vivo reexposure to VZV antigens due to subclinical VZV viremia or symptomatic VZV reactivation may explain the recovery of virus-specific T cell immunity after BMT.

Investigation of varicella-zoster virus infection of lymphocytes by in situ hybridization.

Peripheral blood mononuclear cells harboring viral gene sequences were detected during primary varicella-zoster virus (VZV) infection of the human host and the strain 2 guinea pig by in situ hybridization with a 3H-labeled VZV DNA probe. Activated T lymphocytes were permissive for VZV infection at low frequency in vitro.

Detection of type-specific antibody to herpes simplex virus type 1 by radioimmunoassay with herpes simplex virus type 1 glycoprotein C purified with monoclonal antibody.

Herpes simplex virus type 1 (HSV-1) and HSV-2 specify at least four glycoproteins designated gA/gB, gC, gD, and gE. Previous studies have shown that gC produced by HSV-1 is antigenically distinct from the corresponding HSV-2 glycoprotein. With the exception of gC, the glycoproteins of both serotypes share antigenic sites. Standard serological assays fail to differentiate the antibody to the shared antigenic determinants from the type-specific antibody. In this paper, we describe a procedure for purifying gC from HSV-1-infected cell extracts with an immunoadsorbent prepared with an HCL monoclonal antibody. When used in a solid-phase radioimmunoassay, gC proved to be a type-specific antigen for quantitation of antibody to HSV-1. Among individuals who had no antibody to HSV at the onset of infection, all of those with primary HSV-1 infection developed antibody to gC. Subjects with primary HSV-2 infection failed to develop antibody reactive with gC of HSV-1 (P less than 0.01). Both immunoglobulin G and M antibodies against gC were detected in sera from subjects with either primary or recurrent HSV-1 infection. Higher antibody titers to gC were found in sera from individuals with recurrent infection than in sera from those with primary HSV-1 infection.

Investigation of varicella-zoster virus-infected cell proteins that elicit antibody production during primary varicella using the immune transfer method.

The varicella-zoster virus-infected cell proteins (VZV-ICPs) against which IgG, IgM and IgA antibodies were made in the course of primary varicella-zoster virus (VZV) infection were analysed by the immune transfer method. IgG antibodies were made against one or more of 18 VZV-ICPs by patients with varicella. IgM antibodies were produced which reacted with 21 VZV-ICPs. The spectrum of IgG antibody production during the first week after the onset of infection was limited to an average of three VZV-ICPs while IgM antibodies which reacted with an average of seven VZV-ICPs were detectable in the acute phase of varicella. Equivalent VZV IgG or IgM antibody titres by radioimmunoassay did not correlate with a similar pattern of antibody specificity for VZV-ICPs by immune transfer. A detectable immune response to all VZV-ICPs was not required for the recovery of individual patients from primary VZV infection.

Early immune response in healthy and immunocompromised subjects with primary varicella-zoster virus infection.

Events in pathogenesis and immunity during primary varicella-zoster virus (VZV) infection were examined in 64 healthy subjects and 21 immunocompromised patients. Activation of the interferon system and activation of circulating T lymphocytes were early immune responses that occurred during the incubation period in some healthy subjects. Elevated levels of 2-5A synthetase in peripheral blood mononuclear cells and detection of serum alpha interferon (IFN-alpha) and gamma interferon (IFN-gamma) were present in the majority of healthy subjects who had acute primary VZV infection. Expression of HLA-DR antigen occurred on circulating T lymphocytes from subjects with acute VZV infection. The early production of VZV-specific IgG or IgM antibodies did not correlate with the severity of the clinical infection, but the detection of T lymphocyte proliferation to VZV antigen within three days after the appearance of the varicella exanthem was associated with milder illness. The mean VZV-specific lymphocyte transformation for subjects with less than 100 lesions/m2 was 7.5 +/- 10.43 SD compared with 1.4 +/- 1.85 SD for those with greater than 400 lesions/m2 (P less than .05). Only one (7.7%) of 13 immunocompromised patients had early VZV-specific lymphocyte transformation compared with 19 (42%) of 45 healthy subjects (P less than .05). The rapid host response to primary VZV infection was associated with rapid termination of viremia in healthy subjects; VZV was isolated from only 11% of peripheral blood mononuclear cell samples cultured within 48 hr after the appearance of the exanthem.

Humoral and cellular immunity to varicella-zoster virus glycoprotein gpI and to a non-glycosylated protein, p170, in the strain 2 guinea-pig.

Strain 2 guinea-pigs were inoculated with infectious varicella-zoster virus (VZV) or with immunoaffinity-purified proteins of VZV. Monoclonal antibodies to the VZV gpI (90,000/58,000 complex) and to a non-glycosylated protein, p170, were used to prepare the polypeptide antigens. Humoral and cell-mediated immune responses to the infectious virus were compared with those elicited by the gpI and p170 proteins. Both VZV IgG antibody production and T lymphocyte proliferation to VZV were detected after immunization with infectious VZV and with VZV proteins. The antibody and T lymphocyte responses waned after protein immunization in comparison with the responses induced by infectious VZV but were detected again immediately after reimmunization with gpI or p170.

C-terminal deletion of human tissue transglutaminase enhances magnesium-dependent GTP/ATPase activity.

Tissue transglutaminase (tTG) exhibits a magnesium-dependent GTP/ATPase activity that is involved in the regulation of the cell cycle and cell receptor signaling. The portion of the molecule involved in GTP/ATP hydrolysis is unknown. We expressed and purified a series of C-terminal truncation mutants of human tTG as glutathione S-transferase fusion proteins (DeltaS538, DeltaE447, DeltaP345, DeltaC290, DeltaV228, and DeltaF185) to determine the effect on GTP/ATPase activity. The truncation of the C terminus did not change significantly the apparent Km value for either GTP or ATP. In contrast, the Kcat value for GTP was increased by 4.6- and 3-fold for the DeltaS538 and DeltaE447 mutants, respectively. The DeltaP345 mutant had the highest hydrolysis activity with a 34-fold increase. The hydrolysis activity then declined to 8.1-, 8.7-, and 1. 9-fold for the DeltaC290, DeltaV228, and DeltaF185 mutants, respectively. The Kcat for ATP changed in parallel with the GTPase results. Thin layer chromatography analysis of the hydrolysis reaction products revealed that ATP was rapidly converted to ADP followed by a much slower conversion of ADP to AMP when incubated with wild type tTG or the DeltaP345 mutant. There was a substantial decrease in the calcium-dependent TGase activity when the last 149 amino acid residues were deleted from the C terminus. Less than 5% of the TGase activity was detected for the DeltaS538 and DeltaE447 mutants. In conclusion, we have located the ATP and GTP hydrolytic domain to amino acid residues 1-185. The C terminus functions to inhibit the expression of endogenous GTP/ATPase activity of tTG, and the potential role of the C terminus in modulating this activity is discussed.

Effects of (E)-5-(2-bromovinyl)-2'-deoxyuridine on proliferation of human fibroblasts, peripheral blood mononuclear cells, and granulocyte-monocyte progenitor cells in vitro.

Inhibition of human fibroblasts, granulocyte-monocyte progenitor cells, and lymphocytes was observed at (E)-5-(2-bromovinyl)-2'-deoxyuridine concentrations ranging from 21 to 197 micrograms/ml. These concentrations were 10- to 100-fold above usual serum concentrations after oral administration. (E)-5-(2-Bromovinyl)-2'-deoxyuridine compares favorably with currently used antivirals in terms of in vitro myelotoxicity and immunotoxicity.

Equivalent recognition of a varicella-zoster virus immediate early protein (IE62) and glycoprotein I by cytotoxic T lymphocytes of either CD4+ or CD8+ phenotype.

Immunity to varicella-zoster virus (VZV), a member of the alpha-herpes virus family, exemplifies the host response to an ubiquitous human viral pathogen. In this investigation of the cytotoxic T lymphocyte (CTL) response to VZV, the depletion of CD4+ T lymphocytes made it possible to demonstrate CD8(+)-mediated cytotoxic function against autologous VZV-infected lymphoblastoid cells targets. CTL recognition of two major VZV proteins, the immediate early protein (IE62) and gp I, was demonstrated in limiting dilution cultures of T lymphocytes obtained from immune donors, stimulated with inactivated VZV Ag, and tested against lymphoblastoid cells infected with vaccinia recombinants expressing these VZV proteins. Among 11 VZV donors tested at least 20 y after primary infection, the mean precursor frequency for T lymphocytes that recognized the IE62 protein was 1:105,000 +/- 85,000 SD, with a range of 1:13,000 to 1:231,000. The mean frequency of CTL precursors specific for gp I in 11 subjects was equivalent, with a mean of 1:121,000 +/- 86,000 SD (range 1:15,000 to 1:228,000) (p = 0.68). Limiting dilution cultures were also prepared using purified CD4+ or CD8+ T lymphocyte populations recovered from PBMC by sterile fluorescence-activated cell sorting. CTL precursors that recognized the IE62 protein or gp I were derived from each of the major T lymphocyte populations by stimulation with inactivated VZV Ag; CD4+ and CD8+ CTL precursor frequencies for the IE62 protein and gp I were equivalent (p = 0.2). We conclude that antiviral CTL activity against targets expressing VZV proteins was mediated equally well by T lymphocytes of the CD4+ or CD8+ phenotype and that antiviral CTL function could be elicited in each subpopulation by exposure to non-infectious viral Ag.

Immunity to varicella-zoster viral glycoproteins, gp I (gp 90/58) and gp III (gp 118), and to a nonglycosylated protein, p 170.

Humoral and cellular immunity against two major glycoproteins (gp) of varicella-zoster virus (VZV), gp I (gp 90/58) and gp III (gp 118), and against a nonglycosylated phosphoprotein (p 170) was demonstrated in human subjects. Primary VZV infection was accompanied by the development of IgG to gp I (mean titer 1:200), gp III (mean titer 1:132), and p 170 (mean titer 1:331). Increased IgG antibody production to each of the VZV proteins occurred during recurrent VZV infection with mean titers to gp I of 1:29512, to gp III of 1:15848, and to p 170 of 1:15848. Persistent high titers to gp III (mean titer 1:891) and to p 170 (mean titer 1:2238) were observed in 75% and 88% of VZV-immune subjects, respectively. T lymphocytes which proliferated on stimulation with gp I, gp III, and p 170 developed with primary VZV infection. VZV-immune subjects had mean transformation indices of 4.2 +/- 0.70 SE to gp I, 4.7 +/- 1 SE to gp III, and 3 +/- 0.39 SE to p 170. Among individual subjects, humoral and cellular immunity was not always detected to all three of the VZV proteins. Resolution of primary VZV infection and maintenance of VZV latency did not require a host response to each of these major viral proteins.

Human monoclonal antibodies neutralizing varicella-zoster virus.

Hybridomas secreting human monoclonal antibodies to varicella-zoster virus were produced by fusing B cells of a patient recovering from acute varicella infection with a human-mouse cell line. Two hybrid lines have continued to secrete IgG1, one with kappa and the other with lambda chains, for at least 12 months. Each antibody neutralizes virus infectivity between 1-5 micrograms of partially purified immunoglobulin/ml, each shows a different pattern of immunofluorescent staining of virus-infected cells, and one identifies three viral proteins with molecular weights of 60,000, 95,000, and 97,000.

Investigation of the pathogenesis of varicella-zoster virus infection in guinea pigs by using polymerase chain reaction.

The polymerase chain reaction method (PCR) was used to investigate events in the pathogenesis of varicella-zoster virus (VZV) infection in strain 2, Hartley, and euthymic hairless guinea pigs. VZV was detected in peripheral blood mononuclear cells (PBMC) obtained 2-5 days after infection in 8 (50%) of 16 strain 2, 4 (40%) of 10 hairless, and 10 (34%) of 29 Hartley guinea pigs. The frequency of VZV-infected PBMC was estimated to be at least 1/200,000, which is comparable to that observed in human infection. When VZV PCR was used to test ganglia from hairless guinea pigs, samples from 6 of 8 animals were positive. Of 45 VZV-infected guinea pigs that were tested for cellular immunity by VZV T lymphocyte proliferation assay, 44 developed a stimulation index > 2.0. Control animals had no detectable virus by PCR and did not develop cellular immunity to VZV. These experiments showed that viremia was detectable by PCR during primary VZV infection of guinea pigs in about half of the animals regardless of the strain of guinea pig. Acquisition of cellular immunity provided a consistent marker of infection in all guinea pig strains. PCR was also useful for demonstrating VZV in guinea pig ganglia tissue, with VZV gene sequences being detectable for at least 80 days after infection. With the combination of PCR and immunologic assays, various guinea pig strains should be useful for studies of VZV pathogenesis and for the evaluation of antiviral agents and vaccine strategies.

Immunization with the immediate-early tegument protein (open reading frame 62) of varicella-zoster virus protects guinea pigs against virus challenge.

The IE62 protein, the primary regulatory protein of varicella-zoster virus (VZV) and the major component of the virion tegument, was an effective immunogen in the guinea pig model of VZV infection, whereas the ORF 29 gene product, a nonstructural DNA replication protein, did not elicit protection. All animals immunized with the ORF 29 protein had cell-associated viremia compared with 2 of 11 guinea pigs given the IE62 protein (P = 0.005). VZV was detected in ganglia from 38% of the animals given the ORF 29 protein and 44% of the control animals compared with 9% of the animals immunized with the IE62 protein (P = 0.04). In contrast to the IE62 protein, immunization with the ORF 29 protein did not prime the animals for an enhanced T-cell response upon challenge with infectious virus. The VZV IE62 protein has potential value as a vaccine component.