Risk of herpes zoster according to past history in the general population: The Japanese Shozu herpes zoster study.

Immunity is known to persist after vaccination for varicella zoster virus, but the duration of immunity in patients who develop herpes zoster (HZ) remains unknown. To investigate the association between a past history of HZ and its occurrence in the general population. The Shozu HZ (SHEZ) cohort study included data for 12 299 individuals aged ≥50 years with information on their HZ history. Cross-sectional and 3-year follow-up studies were carried out to analyze the associations between a history of HZ (yes <10 years, yes ≥10 years, no) and the proportion of positive varicella zoster virus skin test results (erythema diameter ≥5 mm) and the risk of HZ after adjusting for potential confounding factors including age, sex, body mass index, smoking status, sleep duration, and mental stress. The incidences of positive skin test results were 87.7% (470/536) for individuals with a history of HZ <10 years ago, 82.2% (396/482) for those with a history of HZ ≥10 years, and 80.2% (3614/4509) for those with no history of HZ. The multivariable odds ratios (95% confidence intervals) of erythema diameter ≥5 mm were 2.07 (1.57-2.73) and 1. 39 (1.08-1.80) for individuals with a history <10 years and ≥10 years ago, respectively, compared with no history. The corresponding multivariable hazard ratios of HZ were 0.54 (0.34-0.85) and 1.16 (0.83-1.61), respectively. A past history of HZ <10 years ago may reduce the occurrence of HZ.

Predictors of herpes zoster severity and immune responses according to pain trajectories: A community-based prospective cohort study.

The authors aimed to identify determinants of the clinical course of herpes zoster and immunological responses, focusing on pain trajectories. This prospective community-based cohort study involved the analysis of responses to a valid pain survey provided by 375 patients diagnosed with herpes zoster based on clinical symptoms and virus identification by polymerase chain reaction. The authors analyzed most patients for humoral/cell-mediated immune response against varicella-zoster virus at the onset and 3 months post-onset. Six months post-initial visit, patients self-reported pain on a scale of 0 (no pain) to 5 (extremely strong pain) at up to 18 time points. Moreover, the pain trajectories were traced using group-based trajectory modeling. Subsequently, the authors used analysis of covariance to explore predictors and the humoral/cell-mediated immune response according to the pain trajectories. In addition, humoral/cell-mediated immune responses were assessed among each trajectory using paired t tests. Amon the five identified trajectories, two were isolated that particularly developed postherpetic neuralgia, with or without severe acute pain. Cancer therapy and corticosteroid use before herpes zoster onset specifically predicted postherpetic neuralgia without severe acute pain. In contrast, prescription of nonsteroidal anti-inflammatory drugs was uniquely associated with postherpetic neuralgia accompanied by severe acute pain. The aforementioned trajectories with postherpetic neuralgia showed increased antibodies and decreased cell-mediated immunity compared with those without postherpetic neuralgia. The authors could successfully distinguish between postherpetic neuralgia trajectories with and without severe acute pain. The identified key predictors and immunological responses against varicella-herpes zoster contribute further evidence to our understanding of the clinical features of herpes zoster and postherpetic neuralgia.

Associations of Perceived Mental Stress, Sense of Purpose in Life, and Negative Life Events With the Risk of Incident Herpes Zoster and Postherpetic Neuralgia: The SHEZ Study.

In the present population-based prospective study, we examined the associations of psychosocial factors with the incidence of herpes zoster (HZ) and postherpetic neuralgia (PHN). Data were collected from 12,359 participants (≥50 years of age) who answered a self-completed health questionnaire in the Shozu County of Kagawa Prefecture in Japan. During a 3-year follow-up between December 2008 and November 2012, HZ and PHN were diagnosed in 400 and 79 subjects, respectively. We used Cox regression analysis to estimate hazard ratios of incident HZ and PHN according to psychosocial factors, adjusting for age, sex, histories of HZ, cancer, and diabetes, smoking and drinking habits, and time from disease onset to treatment. Men with high levels of mental stress were twice as likely to be at risk for incident HZ. The risk of incident HZ was approximately 60% lower among men and women who reported a high sense of purpose in life. Women who experienced negative life events-particularly changes in their work, living environment, and relationships-had a 2- to 3-fold higher risk of incident PHN. Psychosocial factors such as perceived mental stress, sense of purpose in life, and negative life events may contribute to the development of HZ and PHN in the general population.

Clinical and immunologic features of recurrent herpes zoster (HZ).

BACKGROUND: Recurrent herpes zoster (HZ) is thought to be rare, but there have been few large-scale studies of recurrent HZ. OBJECTIVE: We conducted a large-scale prospective cohort study to characterize recurrent HZ. METHODS: We examined 12,522 participants aged 50 years or older in Shozu County and followed them up for 3 years. We compared the incidence of HZ and postherpetic neuralgia, severity of skin lesions and acute pain, cell-mediated immunity, and varicella-zoster virus-specific antibody titer between primary and recurrent HZ. RESULTS: A total of 401 participants developed HZ: 341 with primary HZ and 60 with recurrent HZ. Skin lesions and acute pain were significantly milder and the incidence of postherpetic neuralgia was lower in patients aged 50 to 79 years with recurrent HZ than in those with primary HZ. Varicella-zoster virus skin test induced a stronger reaction in patients aged 50 to 79 years with recurrent HZ than in those with primary HZ. LIMITATIONS: Information on previous HZ episodes was self-reported by participants, so it could not be confirmed that they actually had a history of HZ. CONCLUSION: Recurrent HZ was associated with milder clinical symptoms than primary HZ, probably because of stronger varicella-zoster virus-specific cell-mediated immunity in the patients with recurrence.

A recombinant varicella vaccine harboring a respiratory syncytial virus gene induces humoral immunity.

The varicella-zoster virus (VZV) Oka vaccine strain (vOka) is highly efficient and causes few adverse events; therefore, it is used worldwide. We previously constructed recombinant vOka (rvOka) harboring the mumps virus gene. Immunizing guinea pigs with rvOka induced the production of neutralizing antibodies against the mumps virus and VZV. Here, we constructed recombinant vOka viruses containing either the respiratory syncytial virus (RSV) subgroup A fusion glycoprotein (RSV A-F) gene or RSV subgroup B fusion glycoprotein (RSV B-F) gene (rvOka-RSV A-F or rvOka-RSV B-F). Indirect immunofluorescence and Western blot analyses confirmed the expression of each recombinant RSV protein in virus-infected cells. Immunizing guinea pigs with rvOka-RSV A-F or rvOka-RSV B-F led to the induction of antibodies against RSV proteins. These results suggest that the current varicella vaccine genome can be used to generate custom-made vaccine vectors to develop the next generation of live vaccines.

Neuraminidase gene homology contributes to the protective activity of influenza vaccines prepared from the influenza virus library.

Whole-virus (WV) vaccines from influenza A/duck/Hokkaido/77 (H3N2), and its reassortant strains H3N4, H3N5 and H3N7, which have the same haemagglutinin (HA) gene but different neuraminidase (NA) genes, were prepared from our influenza virus library. Mice were intranasally immunized with equivalent doses of each vaccine (1-0.01 µg per mouse). All of the mice that received the highest dose of each vaccine (1 µg per mouse) showed equivalent high HA-inhibiting (HI) antibody titres and survived the H3N2 challenge viruses. However, mice that received lower doses of vaccine (0.1 or 0.01 µg per mouse) containing a heterologous NA had lower survival rates than those given the H3N2-based vaccine. The lungs of mice challenged with H3N2 virus showed a significantly higher virus clearance rate when the vaccine contained the homologous NA (N2) versus a heterologous NA, suggesting that NA contributed to the protection, especially when the HI antibody level was low. These results suggested that, even if vaccines prepared for a possible upcoming pandemic do not induce sufficient HI antibodies, WV vaccines can still be effective through other matched proteins such as NA.

Novel polyvalent live vaccine against varicella-zoster and mumps virus infections.

The varicella-zoster virus (VZV) Oka vaccine strain (vOka) is a highly immunogenic and safe live vaccine that has long been used worldwide. Because its genome is large, making it suitable for inserting foreign genes, vOka is considered a candidate vector for novel polyvalent vaccines. Previously, a recombinant vOka, rvOka-HN, that expresses mumps virus (MuV) hemagglutinin-neuraminidase (HN) was generated by the present team. rvOka-HN induces production of neutralizing antibodies against MuV in guinea pigs. MuV also expresses fusion (F) protein, which is important for inducing neutralizing antibodies, in its viral envelope. To induce a more robust immune response against MuV than that obtained with rvOka-HN, here an rvOka expressing both HN and F (rvOka-HN-F) was generated. However, co-expression of HN and F caused the infected cells to form syncytia, which reduced virus titers. To reduce the amount of cell fusion, an rvOka expressing HN and a mutant F, F(S195Y) were generated. Almost no syncytia formed among the rvOka-HN-F(S195Y)-infected cells and the growth of rvOka-HN-F(S195Y) was similar to that of the original vOka clone. Moreover, replacement of serine 195 with tyrosine had no effect on the immunogenicity of F in mice and guinea pigs. Although obvious augmentation of neutralizing antibody production was not observed after adding F protein to vOka-HN, the anti-F antibodies did have neutralizing activity. These data suggest that F protein contributes to induction of immune protection against MuV. Therefore this recombinant virus is a promising candidate vaccine for polyvalent protection against both VZV and MuV.

CD134 is a cellular receptor specific for human herpesvirus-6B entry.

Human herpesvirus-6B (HHV-6B) is a T lymphotropic ß-herpesvirus that is clearly distinct from human herpesvirus-6A (HHV-6A) according to molecular biological features. The International Committee on Taxonomy of Viruses recently classified HHV-6B as a separate species. The primary HHV-6B infection causes exanthem subitum and is sometimes associated with severe encephalopathy. More than 90% of the general population is infected with HHV-6B during childhood, and the virus remains throughout life as a latent infection. HHV-6B reactivation causes encephalitis in immunosuppressed patients. The cellular receptor for HHV-6A entry was identified as human CD46, but the receptor for HHV-6B has not been clear. Here we found that CD134, a member of the TNF receptor superfamily, functions as a specific entry receptor for HHV-6B. A T-cell line that is normally nonpermissive for HHV-6B infection became highly susceptible to infection when CD134 was overexpressed. CD134 was down-regulated in HHV-6B-infected T cells. Soluble CD134 interacted with the HHV-6B glycoprotein complex that serves as a viral ligand for cellular receptor, which inhibited HHV-6B but not HHV-6A infection in target cells. The identification of CD134 as an HHV-6B specific entry receptor provides important insight into understanding HHV-6B entry and its pathogenesis.

Identification of the human herpesvirus 6A gQ1 domain essential for its functional conformation.

Human herpesvirus 6 is a T lymphotropic herpesvirus, long classified into variants A and B (HHV-6A and HHV-6B) based on differences in sequence and pathogenicity. Recently, however, HHV-6A and HHV-6B were reclassified as different species. Here, we isolated a neutralizing monoclonal antibody (Mab) named AgQ 1-1 that was specific for HHV-6A glycoprotein Q1 (AgQ1), and we showed that amino acid residues 494 to 497 of AgQ1 were critical for its recognition by this Mab. This region was also essential for AgQ1's complex formation with gH, gL, and gQ2, which might be important for viral binding to the cellular receptor, CD46. In addition, amino acid residues 494 to 497 are essential for viral replication. Interestingly, this sequence corresponds to the domain on HHV-6B gQ1 that is critical for recognition by an HHV-6B-specific neutralizing Mab. Within this domain, only Q at position 496 of HHV-6A is distinct from the HHV-6B sequence; however, the mutant AgQ1(Q496E) was still clearly recognized by the Mab AgQ 1-1. Surprisingly, replacement of the adjacent amino acid, in mutant AgQ1(C495A), resulted in poor recognition by Mab AgQ 1-1, and AgQ1(C495A) could not form the gH/gL/gQ1/gQ2 complex. Furthermore, the binding ability of mutant AgQ1(L494A) with CD46 decreased, although it could form the gH/gL/gQ1/gQ2 complex and it showed clear reactivity to Mab AgQ 1-1. These data indicated that amino acid residues 494 to 497 of AgQ1 were critical for the recognition by Mab AgQ 1-1 and essential for AgQ1's functional conformation.

Complementation of the function of glycoprotein H of human herpesvirus 6 variant A by glycoprotein H of variant B in the virus life cycle.

Human herpesvirus 6 (HHV-6) is a T-cell-tropic betaherpesvirus. HHV-6 can be classified into two variants, HHV-6 variant A (HHV-6A) and HHV-6B, based on genetic, antigenic, and cell tropisms, although the homology of their entire genomic sequences is nearly 90%. The HHV-6A glycoprotein complex gH/gL/gQ1/gQ2 is a viral ligand that binds to the cellular receptor human CD46. Because gH has 94.3% amino acid identity between the variants, here we examined whether gH from one variant could complement its loss in the other. Recently, we successfully reconstituted HHV-6A from its cloned genome in a bacterial artificial chromosome (BAC) (rHHV-6ABAC). Using this system, we constructed HHV-6ABAC DNA containing the HHV-6B gH (BgH) gene instead of the HHV-6A gH (AgH) gene in Escherichia coli. Recombinant HHV-6ABAC expressing BgH (rHHV-6ABAC-BgH) was successfully reconstituted. In addition, a monoclonal antibody that blocks HHV-6B but not HHV-6A infection neutralized rHHV-6ABAC-BgH but not rHHV-6ABAC. These results indicate that HHV-6B gH can complement the function of HHV-6A gH in the viral infectious cycle.

Human herpesvirus 6 glycoprotein M is essential for virus growth and requires glycoprotein N for its maturation.

Human herpesvirus 6 (HHV-6) is a T-lymphotropic virus belonging to the betaherpesvirus family. Several HHV-6-encoded glycoproteins are required for cell entry and virion maturation. Glycoprotein M (gM) is conserved among all herpesviruses, and therefore thought to have important functions; however, the HHV-6 g has not been characterized. Here, we examined the expression of HHV-6 g, and examined its function in viral replication, using a mutant and revertant gM. HHV-6 g was expressed on virions as a glycoprotein modified with complex N-linked oligosaccharides. As in other herpesviruses, HHV-6 g formed a complex with glycoprotein N (gN), and was transported from the endoplasmic reticulum to the trans-Golgi network only when part of this complex. Finally, a gM mutant virus in which the gM start codon was destroyed was not reconstituted, although its revertant was, indicating that HHV-6 g is essential for virus production, unlike the gM of alphaherpesviruses.

Novel monoclonal antibodies for identification of multicentric Castleman's disease; Kaposi's sarcoma-associated herpesvirus-encoded vMIP-I and vMIP-II.

Recent studies have indicated that vMIP-I and vMIP-II play important roles in the pathogenesis of Kaposi's sarcoma-associated herpesvirus (KSHV)-related diseases due to the effects of these proteins on vascularization. We developed monoclonal antibodies against KSHV-encoded viral macrophage inflammatory protein-I (vMIP-I) and vMIP-II to study these expression profiles and reveal the pathogenesis of KSHV-related diseases. The MAbs against vMIP-I and vMIP-II reacted to KSHV-infected cell lines after lytic induction. Both vMIP-I and the vMIP-II gene products were detected 24 h post-induction with 12-O-tetradecanoylphorbol-13-acetate until 60 h in the cytoplasm of primary effusion lymphoma cell lines. In clinical specimens, both vMIP-I and vMIP-II gene products were detected in the tissues of patients with multicentric Castleman's disease. On the other hand, only vMIP-II was detected in a subset of Kaposi's sarcoma. We concluded that these antibodies might be powerful tools to elucidate the pathogenesis of KSHV-related diseases.

Poly-γ-glutamic acid nanoparticles and aluminum adjuvant used as an adjuvant with a single dose of Japanese encephalitis virus-like particles provide effective protection from Japanese encephalitis virus.

To maintain immunity against Japanese encephalitis virus (JEV), a formalin-inactivated Japanese encephalitis (JE) vaccine should be administered several times. The repeated vaccination is not helpful in the case of a sudden outbreak of JEV or when urgent travel to a high-JEV-risk region is required; however, there are few single-injection JE vaccine options. In the present study, we investigated the efficacy of a single dose of a new effective JE virus-like particle preparation containing the JE envelope protein (JE-VLP). Although single administration with JE-VLP protected less than 50% of mice against lethal JEV infection, adding poly(γ-glutamic acid) nanoparticles (γ-PGA-NPs) or aluminum adjuvant (alum) to JE-VLP significantly protected more than 90% of the mice. A single injection of JE-VLP with either γ-PGA-NPs or alum induced a significantly greater anti-JEV neutralizing antibody titer than JE-VLP alone. The enhanced titers were maintained for more than 6 months, resulting in long-lasting protection of 90% of the immunized mice. Although the vaccine design needs further modification to reach 100% protection, a single dose of JE-VLP with γ-PGA-NPs may be a useful step in developing a next-generation vaccine to stop a JE outbreak or to immunize travelers or military personnel.

[Perspective of development of vaccine against infectious diseases].

It is really expected to develop vaccines which are based on newly developed technologies, since we are facing to various emerging infectious diseases such as new influenza epidemic. Various vaccines for infants or children have been developed to use, but vaccines for adolescent such as human papilloma vaccine, and those for adults such as zoster vaccine have been recently developed in USA and Europe. Vaccines based on new concept such as recombinant vaccine, DNA vaccine, polypeptide vaccine and mucosal vaccine will be on market in near future. Furthermore, development of new adjuvants inducing high immunity but having few side-reaction is urgently needed.

Human herpesvirus 6 major immediate early promoter has strong activity in T cells and is useful for heterologous gene expression.

BACKGROUND: Human herpesvirus-6 (HHV-6) is a beta-herpesvirus. HHV-6 infects and replicates in T cells. The HHV-6-encoded major immediate early gene (MIE) is expressed at the immediate-early infection phase. Human cytomegalovirus major immediate early promoter (CMV MIEp) is commercially available for the expression of various heterologous genes. Here we identified the HHV-6 MIE promoter (MIEp) and compared its activity with that of CMV MIEp in various cell lines. METHODS: The HHV-6 MIEp and some HHV-6 MIEp variants were amplified by PCR from HHV-6B strain HST. These fragments and CMV MIEp were subcloned into the pGL-3 luciferase reporter plasmid and subjected to luciferase reporter assay. In addition, to investigate whether the HHV-6 MIEp could be used as the promoter for expression of foreign genes in a recombinant varicella-zoster virus, we inserted HHV-6 MIEp-DsRed expression casette into the varicella-zoster virus genome. RESULTS: HHV-6 MIEp showed strong activity in T cells compared with CMV MIEp, and the presence of intron 1 of the MIE gene increased its activity. The NF-κB-binding site, which lies within the R3 repeat, was critical for this activity. Moreover, the HHV-6 MIEp drove heterologous gene expression in recombinant varicella-zoster virus-infected cells. CONCLUSIONS: These data suggest that HHV-6 MIEp functions more strongly than CMV MIEp in various T-cell lines.

Human herpesvirus 6 encoded glycoprotein Q1 gene is essential for virus growth.

Human herpesvirus 6 (HHV-6) glycoprotein Q1 (gQ1), a unique gene in HHV-6, forms a complex with glycoproteinH (gH) and gL, which is the viral ligand for its cellular receptor, CD46. However, whether gQ1 is essential for virus growth is unknown, because a system is lacking for making gene knockouts for HHV-6. Recently, bacterial artificial chromosome (BAC) and E. coli mutagenesis techniques have been applied to herpesvirus investigation. Here we successfully inserted the HHV-6A genome into a BAC, and obtained reconstituted infectious virus from the HHV-6A-containing BAC DNA. Using this system, we generated a gQ1 mutant virus genome, which failed to yield reconstituted infectious virus, whereas its revertant virus could be produced, indicating that the HHV-6 gQ1 gene is essential for virus growth. Therefore, we successfully applied BAC and E. coli mutagenesis techniques to the study of HHV-6, and discovered that HHV-6 gQ1 is an essential gene for virus growth.

Characterization of varicella-zoster virus-encoded ORF0 gene--comparison of parental and vaccine strains.

The varicella-zoster virus (VZV) Oka vaccine strain (vOka) differs from the parental strain (pOka) at several amino acid positions, but the mutations responsible for the attenuation of vOka have not been clearly defined. The ORF0 of vOka carries some of the mutations. Although we found that the ORF0 of both strains was incorporated into virus particles, the C-terminal region of vOka ORF0 was presented on the virion surface and was N-glycosylated, suggesting that the mutation in vOka ORF0 changes it into a novel envelope glycoprotein. In a mutant virus in which pOka ORF0 was replaced by vOka ORF0, the molecular weight of ORF0 was altered, but the plaque size was not. In addition, a pOka recombinant virus lacking the hydrophobic domain of ORF0 grew equally well as the wild-type virus, indicating that the mutation in ORF0 is not by itself sufficient for the attenuation of the vOka virus.

Rapid and efficient introduction of a foreign gene into bacterial artificial chromosome-cloned varicella vaccine by Tn7-mediated site-specific transposition.

Using a rapid and reliable system based on Tn7-mediated site-specific transposition, we have successfully constructed a recombinant Oka varicella vaccine (vOka) expressing the mumps virus (MuV) fusion protein (F). The backbone of the vector was our previously reported vOka-BAC (bacterial artificial chromosome) genome. We inserted the transposon Tn7 attachment sequence, LacZalpha-mini-attTn7, into the region between ORF12 and ORF13 to generate a vOka-BAC-Tn genome. The MuV-F expressing cassette was transposed into the vOka-BAC genome at the mini-attTn7 transposition site. MuV-F protein was expressed in recombinant virus, rvOka-F infected cells. In addition, the MuV-F protein was cleaved in the rvOka-F infected cells as in MuV-infected cells. The growth of rvOka-F was similar to that of the original recombinant vOka without the F gene. Thus, we show that Tn7-mediated transposition is an efficient method for introducing a foreign gene expression cassette into the vOka-BAC genome as a live virus vector.

Characterization of the varicella-zoster virus ORF50 gene, which encodes glycoprotein M.

The ORF50 gene of the varicella-zoster virus (VZV) encodes glycoprotein M (gM), which is conserved among all herpesviruses and is important for the cell-to-cell spread of VZV. However, few analyses of ORF50 gene expression or its posttranscriptional and translational modifications have been published. Here we found that in VZV-infected cells, ORF50 encoded four transcripts: a full-size transcript, which was translated into the gM, and three alternatively spliced transcripts, which were not translated. Using a splicing-negative mutant virus, we showed that the alternative transcripts were nonessential for viral growth in cell culture. In addition, we found that two amino acid mutations of gM, V42P and G301M, blocked gM's maturation and transport to the trans-Golgi network, which is generally recognized as the viral assembly complex. We also found that the mutations disrupted gM's interaction with glycoprotein N (gN), revealing their interaction through a bond that is otherwise unreported for herpesviruses. Using this gM maturation-negative virus, we found that immature gM and gN were incorporated into intracellularly isolated virus particles and that mature gM was required for efficient viral growth via cell-to-cell spread but not for virion morphogenesis. The virus particles were more abundant at the abnormally enlarged perinuclear cisternae than those of the parental virus, but they were also found at the cell surface and in the culture medium. Additionally, in the gM maturation-negative mutant virus-infected melanoma cells, typical syncytium formation was rarely seen, again indicating that mature gM functions in cell-to-cell spread via enhancement of syncytium formation.

y Human herpesvirus 6 envelope components enriched in lipid rafts: evidence for virion-associated lipid rafts.

In general, enveloped viruses are highly dependent on their lipid envelope for entry into host cells. Here, we demonstrated that during the course of virus maturation, a significant proportion of human herpesvirus 6 (HHV-6) envelope proteins were selectively concentrated in the detergent-resistant glycosphingolipid- and cholesterol-rich membranes (rafts) in HHV-6-infected cells. In addition, the ganglioside GM1, which is known to partition preferentially into lipid rafts, was detected in purified virions, along with viral envelope glycoproteins, gH, gL, gB, gQ1, gQ2 and gO indicating that at least one raft component was included in the viral particle during the assembly process.