The differences in short- and long-term varicella-zoster virus (VZV) immunoglobulin G levels following varicella vaccination of healthcare workers measured by VZV fluorescent-antibody-to-membrane-antigen assay (FAMA), VZV time-resolved fluorescence immunoassay and a VZV purified glycoprotein enzyme immunoassay.

Healthcare workers (HCWs) reporting no history of varicella frequently receive varicella vaccination (vOka) if they test varicella-zoster virus (VZV) immunoglobulin G (IgG) negative. In this study, the utilities of VZV-IgG time-resolved fluorescence immunoassay (VZV-TRFIA) and a commercial VZV-IgG purified glycoprotein enzyme immunoassay (gpEIA) currently used in England for confirming VZV immunity have been compared to the fluorescent-antibody-to-membrane-antigen assay (FAMA). A total of 110 HCWs received two doses of vOka vaccine spaced 6 weeks apart and sera collected pre-vaccination (n = 100), at 6 weeks post-completion of vaccination (n = 86) and at 12-18 months follow-up (n = 73) were analysed. Pre-vaccination, by FAMA, 61·0% sera were VZV IgG negative, and compared to FAMA the sensitivities of VZV-TRFIA and gpEIA were 74·4% [95% confidence interval (CI) 57·9-87·0] and 46·2% (95% CI 30·1-62·8), respectively. Post-completion of vaccination the seroconversion rate by FAMA was 93·7% compared to rates of 95·8% and 70·8% determined by VZV-TRFIA and gpEIA, respectively. At 12-18 months follow-up seropositivity rates by FAMA, VZV-TRFIA and gpEIA were 78·1%, 74·0% and 47·9%, respectively. Compared to FAMA the sensitivities of VZV-TRFIA and gpEIA for measuring VZV IgG following vaccination were 96·4% (95% CI 91·7-98·8) and 74·6% (95% CI 66·5-81·6), respectively. Using both FAMA and VZV-TRFIA to identify healthy adult VZV susceptibles and measure seroconversion showed that vOka vaccination of HCWs is highly immunogenic.

Comparative study of the standard fluorescent antibody to membrane antigen (FAMA) assay and a flow cytometry-adapted FAMA assay to assess immunity to varicella-zoster virus.

A flow cytometry-adapted fluorescent antibody to membrane antigen (FAMA) assay to detect IgG antibodies against varicella-zoster virus (VZV) was developed and tested in 62 serum samples, showing 90.32% accuracy obtained from a receiver operating characteristic (ROC) curve with a 0.9125 (95% confidence interval [CI], 0.829 to 1.00) area below the curve compared to the result with standard FAMA.

Population diversity in batches of the varicella Oka vaccine.

The Oka vaccine is a live attenuated vaccine for the prevention of varicella. Although the vaccine differs from the progenitor virus by over 40 mutations, only three of these are fixed, the rest being a mixture of the wildtype and the vaccine allele. To examine the extent of this variability between two of the three commercially available vaccine preparations, we analysed the vaccine/wildtype allele frequencies present at fifteen vaccine loci in five preparations each from two different manufacturers of the vOka vaccine. Our results suggest that differences in manufacturing processes between the two companies have resulted in significant variation in the frequencies of the vaccine/wildtype alleles in their vaccines. Yet despite these differences, the allele frequencies in the vaccines from the two companies are strongly correlated. We discuss the significance of these findings and the role of evolutionary processes that influence the production of this live attenuated vaccine.

Evaluation of the time resolved fluorescence immunoassay (TRFIA) for the detection of varicella zoster virus (VZV) antibodies following vaccination of healthcare workers.

Determination of varicella zoster virus (VZV) immunity in healthcare workers without a history of chickenpox is important for identifying those in need of vOka vaccination. Post immunisation, healthcare workers in the UK who work with high risk patients are tested for seroconversion. To assess the performance of the time-resolved fluorescence immunoassay (TRFIA) for the detection of antibody in vaccinated as well as unvaccinated individuals, a cut-off was first calculated. VZV-IgG specific avidity and titres six weeks after the first dose of vaccine were used to identify subjects with pre-existing immunity among a cohort of 110 healthcare workers. Those with high avidity (≥ 60%) were considered to have previous immunity to VZV and those with low or equivocal avidity (<60%) were considered naive. The former had antibody levels ≥ 400 mIU/mL and latter had levels < 400 mIU/mL. Comparison of the baseline values of the naive and immune groups allowed the estimation of a TRFIA cut-off value of > 130 mIU/mL which best discriminated between the two groups and this was confirmed by ROC analysis. Using this value, the sensitivity and specificity of TRFIA cut-off were 90% (95% CI 79-96), and 78% (95% CI 61-90) respectively in this population. A subset of samples tested by the gold standard Fluorescence Antibody to Membrane Antigen (FAMA) test showed 84% (54/64) agreement with TRFIA.

Cholesterol dependence of varicella-zoster virion entry into target cells.

The entry of inhaled virions into airway cells is presumably the initiating step of varicella-zoster infection. In order to characterize viral entry, we studied the relative roles played by lipid rafts and clathrin-mediated transport. Virus and target cells were pretreated with agents designed to perturb selected aspects of endocytosis and membrane composition, and the effects of these perturbations on infectious focus formation were monitored. Infectivity was exquisitely sensitive to methyl-beta-cyclodextrin (M beta CD) and nystatin, which disrupt lipid rafts by removing cholesterol. These agents inhibited infection by enveloped, but not cell-associated, varicella-zoster virus (VZV) in a dose-dependent manner and exerted these effects on both target cell and viral membranes. Inhibition by M beta CD, which could be reversed by cholesterol replenishment, rapidly declined as a function of time after exposure of target cells to VZV, suggesting that an early step in viral infection requires cholesterol. No effect of cholesterol depletion, however, was seen on viral binding; moreover, there was no reduction in the surface expression or internalization of mannose 6-phosphate receptors, which are required for VZV entry. Viral entry was energy dependent and showed concentration-dependent inhibition by chlorpromazine, which, among other actions, blocks clathrin-mediated endocytosis. These data suggest that both membrane lipid composition and clathrin-mediated transport are critical for VZV entry. Lipid rafts are likely to contribute directly to viral envelope integrity and, in the host membrane, may influence endocytosis, evoke downstream signaling, and/or facilitate membrane fusion.

A vaccine to prevent herpes zoster and postherpetic neuralgia in older adults.

BACKGROUND: The incidence and severity of herpes zoster and postherpetic neuralgia increase with age in association with a progressive decline in cell-mediated immunity to varicella-zoster virus (VZV). We tested the hypothesis that vaccination against VZV would decrease the incidence, severity, or both of herpes zoster and postherpetic neuralgia among older adults. METHODS: We enrolled 38,546 adults 60 years of age or older in a randomized, double-blind, placebo-controlled trial of an investigational live attenuated Oka/Merck VZV vaccine ("zoster vaccine"). Herpes zoster was diagnosed according to clinical and laboratory criteria. The pain and discomfort associated with herpes zoster were measured repeatedly for six months. The primary end point was the burden of illness due to herpes zoster, a measure affected by the incidence, severity, and duration of the associated pain and discomfort. The secondary end point was the incidence of postherpetic neuralgia. RESULTS: More than 95 percent of the subjects continued in the study to its completion, with a median of 3.12 years of surveillance for herpes zoster. A total of 957 confirmed cases of herpes zoster (315 among vaccine recipients and 642 among placebo recipients) and 107 cases of postherpetic neuralgia (27 among vaccine recipients and 80 among placebo recipients) were included in the efficacy analysis. The use of the zoster vaccine reduced the burden of illness due to herpes zoster by 61.1 percent (P<0.001), reduced the incidence of postherpetic neuralgia by 66.5 percent (P<0.001), and reduced the incidence of herpes zoster by 51.3 percent (P<0.001). Reactions at the injection site were more frequent among vaccine recipients but were generally mild. CONCLUSIONS: The zoster vaccine markedly reduced morbidity from herpes zoster and postherpetic neuralgia among older adults.

The role of the trans-Golgi network in varicella zoster virus biology.

The task of assembling nascent virions presents a formidable challenge to large, enveloped DNA viruses such as varicella zoster virus (VZV). After parasitising the host cell's compartmentalised biosynthetic machinery, viral constituents must be brought together in appropriate proportions for packaging and export. Recent evidence places the trans-Golgi network (TGN) in an orchestrating role with respect to the assembly, envelopment and egress of herpesviruses. This role accords with known functions of the TGN in the uninfected cell. The targeting of viral glycoproteins to the TGN appears to provide a crucial platform for viral assembly. Tegument proteins, interacting with the cytoplasmic domains of glycoproteins, in turn recruit nucleocapsids to the developing supramolecular array. Molecular studies are continually refining understanding of these processes, building upon elegant electron microscopic data. Knowledge of VZV's use of endogenous trafficking pathways from the TGN sheds light on important aspects of viral behaviour in vitro and in vivo.

Immunization of HIV-infected children with varicella vaccine.

OBJECTIVE: To determine the safety and immunogenicity of varicella vaccine in children with human immunodeficiency virus (HIV) infection. Children (n = 41) who were mildly affected by HIV (Centers for Disease Control and Prevention stage N1 or A1) and had no history or serum antibody indicative of prior varicella infection were immunized with two doses of live attenuated varicella vaccine. RESULTS: A minority of the vaccine recipients had mild local or systemic reactions. Vaccination had no effect on the clinical stage of HIV or the HIV RNA plasma load. CD4 cell percentage and CD4 cell count were marginally decreased at week 4 after the first vaccination; this effect was no longer present at week 8 after vaccination. Two months after the second dose of vaccine, 60% of vaccine recipients had anti-varicella antibody in their serum, and 83% had a positive lymphocyte proliferation assay response to varicella antigen. CONCLUSION: On the basis of its safety and immunogenicity, varicella vaccine should be considered in the childhood vaccines given to mildly affected HIV-infected children.

Persistence of immunity to varicella-zoster virus after vaccination of healthcare workers.

OBJECTIVE: Varicella-zoster virus (VZV) vaccine is recommended to protect susceptible healthcare workers (HCWs) from serious disease and to prevent nosocomial spread of VZV. We evaluated clinical outcomes and serological responses in HCWs after immunization with live attenuated VZV vaccine. DESIGN: Vaccinees were immunized from 1979 to 1998 during VZV vaccine trials, as well as after licensure, and followed prospectively for 1 month to 20.6 (mean 4.6) years after vaccination. Sera were tested by fluorescent antibody to membrane antigen (FAMA), latex agglutination (LA), and enzyme-linked immunoassay (EIA) to detect VZV-specific antibodies. STUDY PARTICIPANTS: The median age of the 120 HCWs was 26 years; 51 (42%) were males. INTERVENTIONS: Ninety eight (82%) of these study subjects received vaccine prepared by Merck and 22 (18%) by SmithKline Beecham; 25, 81, and 14 vaccinees received one dose, two doses, and three doses, respectively. RESULTS: The crude attack rate was 10%; 12 of 120 HCWs developed chickenpox 6 months to 8.4 years after vaccination. The attack rates following household and hospital exposures were 18% (4/22) and 8% (6/72), respectively. All resulting illness was mild to moderate (mean 40 vesicles). Seroconversion after vaccination was documented by FAMA in 96% of HCWs, although 31% lost detectable antibodies. Compared with FAMA, LA and EIA were 82% and 74% sensitive and 94% and 89% specific, respectively. CONCLUSIONS: The VZV vaccine effectively protected HCWs from varicella, particularly from serious disease. Currently available serological tests are not optimal, and improved assays are needed.

The current status of live attenuated varicella vaccine.

This manuscript reviews the means by which live attenuated varicella vaccine offers protection against varicella and zoster. It is accepted that although varicella is usually a mild illness, complications leading to morbidity and mortality are significant and the disease is worth preventing. The vaccine offers close to 100% protection from severe chickenpox and 90% protection from illness. Waning of immunity after vaccination, particularly in children, has not been a significant problem. Ways in which vaccination may decrease the incidence and severity of zoster include the following. Vaccine virus may be less likely to establish latency and to be able to reactivate than wild type virus. In addition, by selective immunization of certain hosts such as HIV-infected children whose, immune systems are still relatively intact and individuals with latency due to wild type virus to boost the cell-mediated immune response to the virus, zoster may be decreased. Varicella vaccine is predicted to have a major impact on the epidemiology of varicella and zoster in countries with high vaccine uptake.

Biologic and geographic differences between vaccine and clinical varicella-zoster virus isolates.

Vaccine and wild-type strains of varicella-zoster virus differ both in their biologic characteristics and in the clinical manifestations of infection caused by each strain. The biologic differences described for the vaccine strain (temperature sensitivity and host cell preference) probably reflect the methods used to adapt the wild-type strain to the in vitro growth conditions imposed during the attenuation process in cell culture. In addition, restriction fragment polymorphisms have been described that reflect geographic strain variations between the parental virus used to develop the vaccine strain and other wild-type strains. These polymorphisms have been exploited as tools for the identification and differentiation of vaccine and wild-type strains in clinical studies. Infection with the wild-type strain results in the typical extensive rash of varicella, frequent transmission to other susceptible contacts, establishment of latency, and in some individuals, reactivation with the clinical picture of zoster. Infection with the vaccine strain results in the development of a protective immune response, minimal rash in a minority of individuals, rare transmission to other susceptible contacts, and a greatly reduced risk of zoster.

Live-attenuated varicella vaccine.

This article reviews the history and development of live attenuated varicella vaccine from its early days in Japan to its widespread use throughout the world. The vaccine has proven extremely safe after immunization of as many as 10 million healthy children and adults in the United States alone. The vaccine is also highly immunogenic and offers close to 100% protection from severe chickenpox and 90% protection from illness. It is expected to have a major impact on the epidemiology of varicella and zoster in countries with high vaccine uptake.

The effectiveness of the varicella vaccine in clinical practice.

BACKGROUND: A live attenuated varicella vaccine was approved for use in the United States in March 1995 and is recommended for all susceptible persons 12 months of age or older. METHODS: To assess the effectiveness of the varicella vaccine, we conducted a case-control study with two controls per child with chickenpox, matched according to both age and pediatric practice. Children with potential cases of chickenpox were identified by active surveillance of pediatric practices in the New Haven, Connecticut, area. Research assistants visited the children on day 3, 4, or 5 of the illness, assessed the severity of the illness, and collected samples from lesions to test for varicella-zoster virus by polymerase chain reaction (PCR). RESULTS: From March 1997 through November 2000, data collection was completed for 330 potential cases, of which 243 (74 percent) were in children who had positive PCR tests for varicella-zoster virus. Of the 56 vaccinated children with chickenpox, 86 percent had mild disease, whereas only 48 percent of the 187 unvaccinated children with chickenpox had mild disease (P<0.001). Among the 202 children with PCR-confirmed varicella-zoster virus and their 389 matched controls, 23 percent of the children with chickenpox and 61 percent of the matched controls had received the vaccine (vaccine effectiveness, 85 percent; 95 percent confidence interval, 78 to 90 percent; P<0.001). Against moderately severe and severe disease the vaccine was 97 percent effective (95 percent confidence interval, 93 to 99 percent). The effectiveness of the vaccine was virtually unchanged (87 percent) after adjustment for potential confounders by means of conditional logistic regression. CONCLUSIONS: Varicella vaccine is highly effective as used in clinical practice.

Essential role played by the C-terminal domain of glycoprotein I in envelopment of varicella-zoster virus in the trans-Golgi network: interactions of glycoproteins with tegument.

Varicella-zoster virus (VZV) is enveloped in the trans-Golgi network (TGN). Here we report that glycoprotein I (gI) is required within the TGN for VZV envelopment. Enveloping membranous TGN cisternae were microscopically identified in cells infected with intact VZV. These sacs curved around, and ultimately enclosed, nucleocapsids. Tegument coated the concave face of these sacs, which formed the viral envelope, but the convex surface was tegument-free. TGN cisternae of cells infected with VZV mutants lacking gI (gI(Delta)) or its C (gI(DeltaC))- or N-terminal (gI(DeltaN))-terminal domains were uniformly tegument coated and adhered to one another, forming bizarre membranous stacks. Viral envelopment was compromised, and no virions were delivered to post-Golgi structures. The TGN was not gI-immunoreactive in cells infected with the gI(Delta) or gI(DeltaN) mutants, but it was in cells infected with gI(DeltaC) (because the ectodomains of gI and gE interact). The presence in the TGN of gI lacking a C-terminal domain, therefore, was not sufficient to maintain enveloping cisternae. In cells infected with intact VZV or with gI(Delta), gI(DeltaN), or gI(DeltaC) mutants, ORF10p immunoreactivity was concentrated on the cytosolic face of TGN membranes, suggesting that it interacts with the cytosolic domains of glycoproteins. Because of the gE-gI interaction, cotransfected cells that expressed gE or gI were able to target truncated forms of the other to the TGN. Our data suggest that the C-terminal domain of gI is required to segregate viral and cellular proteins in enveloping TGN cisternae.

Prevention and treatment of VZV infections in patients with HIV.

Varicella zoster virus (VZV) infections in human immunodeficiency virus (HIV)-infected patients are known to have a different disease spectrum from that seen in other types of patients. Varicella in children with HIV infection is likely to be more serious than in otherwise healthy children and routine antiviral therapy is recommended. There is evidence that the development of varicella in HIV-infected children is not associated with progression to AIDS, suggesting that it may be safe to immunize HIV-infected children with live attenuated varicella vaccine. There are no published data on varicella in HIV-infected adults, however, probably because most adults have already experienced varicella prior to HIV infection. Zoster in HIV-infected children differs somewhat from that in HIV-infected adults. In particular, HIV-infected children who develop varicella in the setting of severe immunodeficiency are at an especially high risk to develop zoster. Given the low rate of toxicity of aciclovir as well as its ease of administration and its efficacy in hastening the healing of VZV infections, prompt treatment with this antiviral agent is recommended for both HIV-infected children and adults. Foscarnet should be used for zoster that is strongly suspected or proven to be caused by aciclovir-resistant VZV. Patients with HIV for whom there is no evidence of significant immunosuppression and who have not had varicella should be immunized with live attenuated varicella vaccine as a preventative measure for both varicella and zoster. It is hoped that immunization of VZV seropositive HIV-infected patients will decrease the incidence of zoster. Studies to determine this are under way.

The postmarketing safety profile of varicella vaccine.

The postmarketing safety profile of varicella vaccine was evaluated by analyzing selected adverse experience reports temporally associated with the administration of the vaccine. There were 7963 reports voluntarily submitted to Merck for an overall reporting rate of 5.0 per 10000 doses of vaccine distributed. A varicella zoster virus (VZV) identification program detected the presence of the Oka vaccine strain in three individuals with an immune deficiency - two with pneumonia and one with hepatitis - and in three instances of secondary transmission from vaccinees with vesicular lesions to susceptible household contacts. The Oka vaccine strain was present in 23 patients and wild-type VZV was present in 15 patients with herpes zoster. Vesicular rashes that occurred within 2 weeks of vaccination were more likely to contain the presence of wild-type VZV, while vesicular rashes that occurred more than 2 weeks post-vaccination were more likely to contain the Oka vaccine strain. Eleven patients were hospitalized with complications of breakthrough varicella infection.

Viral strain identification in varicella vaccinees with disseminated rashes.

BACKGROUND: Approximately 15% of recipients of live attenuated varicella vaccine may develop mild breakthrough varicella months to years after immunization. Although some vaccinees will develop zoster, it is less common in recipients of vaccine than in those who have had natural varicella. OBJECTIVE: To determine the varicella-zoster virus (VZV) strain responsible for breakthrough varicella and zoster in recipients of varicella vaccine. METHODS: A PCR assay capable of distinguishing wild-type from vaccine strain VZV was performed on samples from skin lesions from vaccinees with breakthrough varicella and zoster. RESULTS: All of 57 vaccinees with breakthrough varicella, clinically diagnosed on the basis of a generalized maculopapular or vesicular rash, in which there was amplifiable DNA [corrected], had wild-type VZV infection based on analysis of viral DNA. The Oka vaccine strain of VZV was not identified in any of these cases. In contrast, in 32 patients with zosteriform rashes, the vaccine strain was identified in 22 samples, and the wild-type strain was identified in 10 samples. CONCLUSIONS: Wild-type virus was identified in all generalized rashes occurring after the immediate 6-week postvaccination period. When reactivation of vaccine strain occurred, it presented as typical zoster. We find no evidence that reactivation of vaccine virus occurs with the clinical picture of generalized rash.