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.

Live attenuated varicella vaccine use in immunocompromised children and adults.

Live attenuated varicella vaccine has been administered to 307 children with leukemia in remission and to 86 healthy adults. The vaccine was well tolerated and immunogenic. The major side effect in leukemic children receiving maintenance chemotherapy was development of a vaccine-associated rash. Vaccinees in whom a rash developed were potentially somewhat infectious to others about 1 month after immunization. Vaccination was not associated with an increase in the incidence of herpes zoster or in relapse of leukemia. Vaccination provided excellent protection against severe varicella. It was associated with a significant decrease in the attack rate of chickenpox following an intimate exposure to varicella-zoster virus, conferring about 80% protection in leukemic children. The cases of breakthrough varicella that occurred were mild. Thus, the vaccine may either prevent or modify varicella in high-risk individuals. It may also have use for prevention of nosocomial varicella.

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.

Varicella-zoster virus infections in children infected with human immunodeficiency virus.

Primary varicella-zoster (VZ) infection in eight children with perinatally acquired human immunodeficiency virus infection tended to be severe, prolonged, complicated by bacterial infections and in one case fatal. Depletion of CD4-lymphocytes was associated with chronic and recurrent VZ infection. In some patients convalescent VZ antibody titers were low and did not correlate with recurrence of VZ lesions. Administration of acyclovir appeared to be beneficial in suppressing VZ in human immunodeficiency virus-infected children with primary or recurrent VZ infection.

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.

Antibody responses to varicella-zoster virus and the role of antibody in host defense.

Antibody titers to varicella-zoster (VZ) virus in persons aged 1-85 years were measured. Through age 50, the percent of seropostitive individuals continued to rise. There was no fall in geometric mean titer with aging, and there may have been an increase in VZ antibody titer in older persons. Thus, a fall in VZ antibody with increasing age does not seem likely to account for the increased incidence of zoster in the elderly. Similarly, immunocompromised patients who are more likely to develop zoster than normals did not have lower VZ antibody titers than normal persons. Finally, immunocompromised persons with disseminated zoster had antibody titers that were somewhat higher that those of immunocompromised persons with localized zoster. It appears that humoral immunity has little or no influence on either the development or the course of herpes zoster.

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.

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.

Measurement of antibodies to varicella-zoster virus by using a latex agglutination test.

We evaluated a rapid, simple-to-perform assay for detecting antibodies to varicella-zoster virus by latex agglutination (LA); dilutions of test sera were added to latex particles coated with varicella-zoster virus antigen. We tested 878 serum specimens by LA and with fluorescent antibody to membrane antigen; of these, 227 were also tested by a commercially marketed enzyme-linked immunosorbent assay (ELISA). LA was almost as sensitive as the fluorescent antibody to membrane antigen test and more sensitive than ELISA. No cross-reacting antibodies were detected by LA, and false-positive reactions were rare.

Live attenuated varicella vaccine: protection in healthy adults compared with leukemic children. National Institute of Allergy and Infectious Diseases Varicella Vaccine Collaborative Study Group.

Protection against varicella infection was assessed in leukemic children and healthy young adults who were immunized with live attenuated varicella vaccine. Attack rates of breakthrough infection following household exposure to varicella in 102 children and 26 adults were similar whether one or two doses of vaccine had been given. The mild breakthrough illness was also similar after one or more doses. Specific antibody titers were similar 1 year after immunization whether individuals had received one or two doses. Humoral and cell-mediated immunity to varicella-zoster virus (VZV) was lower in these vaccinees than in persons who had experienced natural varicella infection. Protection after natural infection in adult family members exposed to varicella was superior to that in vaccinees; none developed varicella infection. These observations suggest that immunization induces less protection than does natural disease in leukemic children and young adults. This may be partly due to the nature of the vaccine virus, but because responses of adults were similar to those of leukemic children, it suggests also that both of these groups have impaired immune responses to VZV. Boosting of humoral immunity after exposure to VZV was common and was observed in healthy adults with past natural infection and in vaccinated adults and leukemic children.

Persistence of immunity to varicella in children with leukemia immunized with live attenuated varicella vaccine.

To determine the safety and efficacy of varicella vaccine, we studied 437 children in remission from leukemia who were immunized with live attenuated varicella virus. Three hundred one of the patients received two doses of vaccine and 136 received a single dose of vaccine from 1 of 10 lots from two manufacturers. The patients have been followed for an average of three years (range, one to six). Seroconversion occurred in 88 percent of the 437 children after the first dose of vaccine and in 98 percent after one or two doses. The proportions of patients who were seronegative after one, three, and five years were 20, 25, and 30 percent, respectively, with little change over time in the geometric mean titers of specific antibody (6.3, 6.5, and 5.7, respectively). Chickenpox has been documented in 36 vaccinated patients (8 percent) who had 3 to 640 vesicles (mean, 100), mild illness, and no complications. Of the 83 vaccinated patients exposed to varicella within their families, 11 had chickenpox; the attack rate was 14 percent (8 percent among seropositive patients, 29 percent among seronegative patients). There was no relation between the time since vaccination and either the attack rate or the severity of the breakthrough illness. Two doses of vaccine appeared to be no more effective than a single dose. Of the 372 patients receiving maintenance chemotherapy when immunized, 149 (40 percent) had a rash, which was treated with acyclovir in 16 children (4 percent) and became a severe febrile illness in 4. These reactions were not fatal and were all associated with vaccine lots, the use of which has since been discontinued. We conclude that in children in remission from leukemia, varicella vaccine is safe and induces an immunity to chickenpox that persists for more than three years.

Inactivation of varicella zoster virus in vitro: effect of leukocytes and specific antibody.

Inactivation of varicella zoster virus in vitro by nonadherent, mononuclear peripheral blood leukocytes and antibody is described. When leukocytes and specific antibody were incubated with this virus, marked inactivation of the virus occurred. In contrast, leukocytes alone or with serum devoid of varicella zoster antibody caused only a small degree of inactivation of varicella zoster virus. The leukocytes involved appeared not to be monocyte-macrophages or T or B lymphocytes, and only minute amounts of specific antibody were required. We had found previously that leukocytes from unsensitized (varicella susceptible) as well as sensitized (varicella immune) donors could cause this reaction. We therefore propose that the reaction may be a form of antibody-dependent cellular cytotoxicity, as has been described for 51Cr release by lymphoid (K) cells for other herpesviruses.

Cellular and humoral immune responses to varicella-zoster virus in immunocompromised patients during and after varicella-zoster infections.

Humoral and cell-mediated immune responses to varicella-zoster (V-Z) virus were assessed in patients during and after V-Z infections. Ongoing V-Z infections was associated with minimal cellular immunity but not necessarily with poor humoral immunity. Recovery from V-Z infection was associated with a vigorous cellular immune response. Cell-mediated immunity to V-Z virus was demonstrable for years after varicella, but responses were lower in immunocompromised patients than in normal individuals.

Cell-mediated immunity to varicella-zoster virus measured by virus inactivation: mechanism and blocking of the reaction by specific antibody.

The process whereby varicella-zoster (V-Z) virus is inactivated in vitro by immune human peripheral blood leukocytes stimulated with V-Z antigen was examined. It was found that stimulation of leukocytes by V-Z antigen, but not by other viral antigens, was required for inactivation of V-Z virus to occur. Viral inactivation could be blocked by addition of V-Z antiserum to either the stimulation phase of the reaction or the inactivation phase, further demonstrating the specificity of the reaction. In addition these blocking experiments suggested that modulation of V-Z membrane antigen by antiserum occurred with an accompanying loss of immunological recognition of virus-infected cells. Inactivation of V-Z virus in vitro in this study appeared not to be dependent upon the secretion of interferon or upon antibody-dependent cellular cytotoxicity. The specific cells required for V-Z inactivation were T lymphocytes and monocytes (macrophage precursors).

Rapid diagnosis of varicella-zoster virus infections by countercurrent immunoelectrophoresis.

A simple, accurate, and rapid method for laboratory diagnosis of varicella-zoster virus (VZV) infections using countercurrent immunoelectrophoresis (CIE) is described. CIE uses zoster convalescent-phase serum to detect VZV antigen in vesticular fluid. Eighty-six patients were studied, 58 with VZV infections and 28 with vesicular or bullous rashes due to causes other than VZV infection. All patients with documented VZV infection had positive CIE tests for VZV, and the controls were uniformly negative. VZV antigen could be detected up to 15 (varicella) or 16 (zoster) days after the onset of rash. The sensitivity of CIE for detection of VZV antigen was compared with results of viral isolation, immune adherence hemmagglutination, and an indirect enzyme-linked immunosorbent assay.

Clinical reinfection with varicella-zoster virus.

Eight patients became clinically reinfected with varicella-zoster virus despite the presence of specific antibody in the blood three days to six months before the onset of illness. One patient had had varicella previously; a second had been immunized with live, attenuated varicella vaccine 10 months earlier. While it was suspected that these patients experienced a reactivation of latent virus that caused atypical disseminated zoster rather than varicella, detailed study of the vaccinated child suggests that this was not the case; by restriction-endonuclease techniques, this vaccinee was shown to have been infected with wild-type varicella-zoster virus despite the presence of specific antibody and cellular immunity to the virus. All cases clinically resembled chickenpox. Thus, not only subclinical varicella (manifested by a rise in antibody titer after close exposure) but also clinical reinfection with the virus can occur. Clinical reinfection probably develops more frequently in immunocompromised than in immunocompetent individuals. Reinfections are usually mild.