Immunogenicity of meningococcal polysaccharide ACWY vaccine in primary immunized or revaccinated adults.

Meningococcal polysaccharide (Men-Ps) vaccine immunogenicity following either primary immunization or revaccination in adults was evaluated. The study population consisted of subjects who have received tetravalent Men-Ps vaccine once (group 1) or at least twice, with a 2-6 dose range (group 2). Human leucocyte antigen (HLA)-typing was performed by polymerase chain reaction and specific immunoglobulin (Ig)G was measured by enzyme-linked immunosorbent assay. Nine months post-immunization, the percentages of individuals with levels of anti-Men-Ps IgG ≥ 2 µg/ml were comparable in both groups, with the exception of anti-Men-PsW135 IgG, which were significantly higher in group 2. The percentage of subjects doubling IgG levels at 9 months was significantly higher in group 1. The high baseline anti-Men-Ps antibody levels negatively influenced the response to revaccination, suggesting a feedback control of specific IgG. The calculated durability of anti-Men-Ps IgG was 2·5-4·5 years, depending on the Men-Ps, following a single vaccine dose. No interference by other vaccinations nor HLA alleles association with immune response were observed. This study confirms that Men-Ps vaccine in adults is immunogenic, even when administered repeatedly, and underlines the vaccine suitability for large-scale adult immunization programmes that the higher costs of conjugate vaccines may limit in developing countries.

Safety and immunogenicity of co-administered MF59-adjuvanted 2009 pandemic and plain 2009-10 seasonal influenza vaccines in rheumatoid arthritis patients on biologicals.

Rheumatoid arthritis (RA) patients under immunosuppressive therapy are particularly susceptible to infections, mainly of the respiratory tract, thus vaccination may represent a strategy to reduce their incidence in this vulnerable population. In the 2009-10 influenza season, the safety and immunogenicity of co-administered non-adjuvanted seasonal and MF59-adjuvanted pandemic influenza vaccines were evaluated in this study in 30 RA patients under therapy with anti-tumour necrosis factor (TNF)-α agents or Abatacept and in 13 healthy controls (HC). Patients and HC underwent clinical and laboratory evaluation before (T0), 1 (T1) and 6 months (T2) after vaccinations. No severe adverse reactions, but a significant increase in total mild side effects in patients versus HC were observed. Both influenza vaccines fulfilled the three criteria of the Committee for Proprietary Medicinal Products (CPMP). Seroconversion rate for any viral strain in patients and HC was, respectively, 68 versus 45 for H1-A/Brisbane/59/07, 72 versus 81 for H3-A/Brisbane/10/07, 68 versus 54 for B/Brisbane/60/08 and 81 versus 54 for A/California/7/2009. A slight increase in activated interferon (IFN)-γ-, TNF-α- or interleukin (IL)-17A-secreting T cells at T1 compared to T0, followed by a reduction at T2 in both patients and HC, was registered. In conclusion, simultaneous administration of adjuvanted pandemic and non-adjuvanted seasonal influenza vaccines is safe and highly immunogenic. The largely overlapping results between patients and HC, in terms of antibody response and cytokine-producing T cells, may represent further evidence for vaccine safety and immunogenicity in RA patients on biologicals.

Strong CD8+ T cell antigenicity and immunogenicity of large foreign proteins incorporated in HIV-1 VLPs able to induce a Nef-dependent activation/maturation of dendritic cells.

Virus-like particles (VLPs) are excellent tools for vaccines against pathogens and tumors. They can accommodate foreign polypeptides whose incorporation efficiency and immunogenicity however decrease strongly with the increase of their size. We recently described the CD8(+) T cell immune response against a small foreign antigen (i.e., the 98 amino acid long human papilloma virus E7 protein) incorporated in human immunodeficiency virus (HIV)-1 based VLPs as product of fusion with an HIV-1 Nef mutant (Nef(mut)). Here, we extended our previous investigations by testing the antigenic/immunogenic properties of Nef(mut)-based VLPs incorporating much larger heterologous products, i.e., human hepatitis C virus (HCV) NS3 and influenza virus NP proteins, which are composed of 630 and 498 amino acids, respectively. We observed a remarkable cross-presentation of HCV NS3 in dendritic cells challenged with Nef(mut)-NS3 VLPs, as detected using a NS3 specific CD8(+) T cell clone as well as PBMCs from HCV infected patients. On the other hand, when injected in mice, Nef(mut)-NP VLPs elicited strong anti-NP CD8(+) T cell and CTL immune responses. In addition, we revealed the ability of Nef(mut) incorporated in VLPs to activate and mature primary human immature dendritic cells (iDCs). This phenomenon correlated with the activation of Src tyrosine kinase-related intracellular signaling, and can be transmitted from VLP-challenged to bystander iDCs. Overall, these results prove that Nef(mut)-based VLPs represent a rather flexible platform for the design of innovative CD8(+) T cell vaccines.

Influenza vaccine administration in rheumatoid arthritis patients under treatment with TNFalpha blockers: safety and immunogenicity.

Twenty-eight patients with low-moderate, stable rheumatoid arthritis (RA), under treatment with tumor necrosis factor (TNF) alpha blockers, were immunized at least once with non-adjuvanted trivalent influenza vaccine during three consecutive influenza seasons. Antibodies toward A influenza antigens significantly increased and reached protective levels, still detectable 6 months after vaccination, both in RA patients and healthy controls. Response to B antigen instead was only observed from the second year for healthy controls and in the third year for patients. No significant difference in disease activity and anti-nuclear antibodies was observed as a consequence of vaccine administration, whereas T regulatory cells showed a significant increase 30 days after immunization in RA patients. This study confirms safety of influenza vaccine administration in RA patients treated with TNFalpha blockers. The cohort follow-up revealed the overcoming of poor B vaccine antigen immunogenicity via repeated vaccinations. Finally, protective antibody response was still observed 6 months after vaccination.

Detection of interleukin-2 in addition to interferon-gamma discriminates active tuberculosis patients, latently infected individuals, and controls.

Effective control of tuberculosis (TB) includes discrimination of subjects with active TB from individuals with latent TB infection (LTBI). As distinct interferon (IFN)-gamma and interleukin (IL)-2 profiles of antigen-specific T-cells have been associated with different clinical stages and antigen loads in several viral and bacterial diseases, we analysed these cytokines in TB using a modified QuantiFERON-TB Gold In Tube test. Detection of IL-2 in addition to IFN-gamma distinguishes not only Mycobacterium tuberculosis-infected subjects from healthy controls, but also individuals with LTBI from active TB patients. This may help to improve diagnostic tests for TB.

Influenza vaccine administration in patients with systemic lupus erythematosus and rheumatoid arthritis. Safety and immunogenicity.

OBJECTIVE: To evaluate immunological safety and immunogenicity of influenza vaccine administration in patients with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). PATIENTS AND METHODS: Twenty-four patients with low and/or stable disease activity 14 with SLE (mean age 43.42+/-12.18 years; 13 women) and 10 with RA (mean age 51+/-14.57 years; 9 women), diagnosed on the basis of the American College of Rheumatology criteria, have been immunized with trivalent split influenza vaccine without adjuvant. Further 24 non-vaccinated patients, 14 with SLE and 10 with RA, and 10 vaccinated healthy subjects, all age- and sex-matched, were used as controls. The patients underwent clinical and laboratory (specific anti-influenzavirus antibodies, auto-antibodies, peripheral blood lymphocyte subpopulations) evaluation before and 30 days after vaccination; auto-antibodies were also assessed at 90 days and disease activity at 90 and 180 days. RESULTS: The specific antibody response towards the three used antigens (A/New Caledonia/20/99, A/Moscow/10/99, and B/Shangdong/7/97) significantly increased in both patients and healthy controls, without any significant difference between them. No significant difference could instead be observed on the clinical activity, auto-antibodies, and peripheral blood lymphocyte subpopulations before and after vaccination, and between patients and controls. CONCLUSIONS: Trivalent split influenza vaccine without adjuvant seems to be safe and immunogenic in patients with SLE and RA, provided that only patients with low and/or stable disease activity are selected.

Unilineage monocytopoiesis in hematopoietic progenitor culture: switching cytokine treatment at all Mo developmental stages induces differentiation into dendritic cells.

We have developed a new culture system whereby human hematopoietic progenitors purified from adult peripheral blood extensively proliferate and gradually differentiate into >95% pure monocytic (Mo) cells. At all developmental stages treatment with interleukin (IL)-4+granulocyte-macrophage colony-stimulating factor or IL-4+c-Kit-ligand+FLT-3 ligand switched the Mo precursors into dendritic cells (DCs). The switching capacity decreased only at the end of the culture, when most Mo cells matured to macrophages. Moreover, the Mo precursors were highly susceptible to transduction with lentiviral vectors: once switched to DCs, they maintained the transgene expression, as well as the phenotype and function of the DC lineage. Our results provide new insight into the potential role of the Mo lineage as a reservoir of DCs in vivo. Furthermore, the methodology for transduction of Mo precursors provides a tool to generate genetically modified, normally functioning DCs potentially useful for immunotherapy.

Antibody classes & subclasses induced by mucosal immunization of mice with Streptococcus pyogenes M6 protein & oligodeoxynucleotides containing CpG motifs.

BACKGROUND & OBJECTIVES: Type-specific antibodies against M protein are critical for human protection as they enhance phagocytosis and are protective. An ideal vaccine for the protection against Streptococcus pyogenes would warrant mucosal immunity, but mucosally administered M-protein has been shown to be poorly immunogenic in animals. We used a recombinant M type 6 protein to immunize mice in the presence of synthetic oligodeoxynucleotides containing CpG motifs (immunostimulatory sequences: ISS) or cholera toxin (CT) to explore its possible usage in a mucosal vaccine. METHODS: Mice were immunized by intranasal (in) or intradermal (id) administration with four doses at weekly intervals of M6-protein (10 microg/mouse) with or without adjuvant (ISS, 10 microg/mouse or CT, 0,5 microg/mouse). M6 specific antibodies were measured by enzyme linked immunosorbent assay using class and subclass specific monoclonal antibodies. RESULTS: The use of ISS induced an impressive anti M-protein serum IgG response but when id administered was not detectable in the absence of adjuvant. When used in, M-protein in the presence of both ISS and CT induced anti M-protein IgA in the bronchoalveolar lavage, as well as specific IgG in the serum. IgG were able to react with serotype M6 strains of S. pyogenes. The level of antibodies obtained by immunizing mice in with M-protein and CT was higher in comparison to M-protein and ISS. The analysis of anti-M protein specific IgG subclasses showed high levels of IgG1, IgG2a and IgG2b, and low levels of IgG3 when ISS were used as adjuvant. Thus, in the presence of ISS, the ratio IgG2a/IgG1 and (IgG2a+IgG3)/IgG1 >1 indicated a type 1-like response obtained both in mucosally or systemically vaccinated mice. INTERPRETATION & CONCLUSION: Our study offers a reproducible model of anti-M protein vaccination that could be applied to test new antigenic formulations to induce an anti-group A Streptococcus (GAS) vaccination suitable for protection against the different diseases caused by this bacterium.

The interaction of human dendritic cells with yeast and germ-tube forms of Candida albicans leads to efficient fungal processing, dendritic cell maturation, and acquisition of a Th1 response-promoting function.

T helper cell type 1 (Th1) cell-mediated immunity plays a critical role in protection against the opportunistic pathogen Candida albicans. Virulence of the fungus is closely associated with its ability to form germ-tubes (GT), the early phase of the dimorphic transition from the commensal yeast (Y) to the more invasive hyphal (H) form. In this study, we examined the functional outcome of the interaction of Y or GT forms with human dendritic cells (DCs), professional antigen-presenting cells, which are pivotal for initiation and modulation of T cell responses. DCs phagocytosed and killed Y and GT cells with a comparable efficiency, becoming able to trigger strong proliferative responses by Candida-specific, autologous T cell clones. Both fungal forms induced DC maturation, as indicated by up-regulation of CD83, CD80, CD86, CD40, and major histocompatibility complex classes I and II surface antigens. Chemokine receptors were also modulated in Candida-DCs, which showed increased CCR7/CXCR4 and decreased CCR5 expression. Y- and GT-activated DCs differed in the pattern of cytokine expression. In particular, GT cells, in common with fully differentiated H cells, induced significantly more elevated levels of interleukin (IL)-10 than Y cells. Nevertheless, Y-, GT-, or H-pulsed DCs secreted comparable amounts of IL-12p70. In addition, irrespective of the fungal form triggering DC activation, Candida-DCs acquired the ability to prime naive T lymphocytes with a defined Th1 phenotype. Overall, our findings highlight the induction of substantially similar functional patterns in human DCs encountering the different forms of growth of C. albicans, both seemingly activating the Th1-type immunity which is characteristic of the healthy human subjects, naturally immunized and protected against the fungus.

Antigenic properties and processing requirements of 65-kilodalton mannoprotein, a major antigen target of anti-Candida human T-cell response, as disclosed by specific human T-cell clones.

T-cell-mediated immunity is known to play a central role in the host response to Candida albicans. T-cell clones are useful tools for the exact identification of fungal T-cell epitopes and the processing requirements of C. albicans antigens. We isolated human T-cell clones from an HLA-DRB1*1101 healthy donor by using an antigenic extract (MP-F2) of the fungus. Specific clones were T-cell receptor alpha/beta and CD4(+)/CD8(-) and showed a T-helper type 1 cytokine profile (production of gamma interferon and not interleukin-4). The large majority of these clones recognized both the natural (highly glycosylated) and the recombinant (nonglycosylated) 65-kDa mannoprotein (MP65), an MP-F2 minor constituent that was confirmed to be an immunodominant antigen of the human T-cell response. Surprisingly, most of the clones recognized two synthetic peptides of different MP65 regions. However, the peptides shared the amino acid motif IXSXIXXL, which may be envisaged as a motif sequence representing the minimal epitope recognized by these clones. Three clones recognized natural and pronase-treated MP65 but did not detect nonglycosylated, recombinant MP65 or the peptides, suggesting a possible role for polysaccharides in T-cell recognition of C. albicans. Finally, lymphoblastoid B-cell lines were efficient antigen-presenting cells (APC) for recombinant MP65 and peptides but failed to present natural, glycosylated antigens, suggesting that nonprofessional APC might be defective in processing highly glycosylated yeast proteins. In conclusion, this study provides the first characterization of C. albicans-specific human T-cell clones and provides new clues for the definition of the cellular immune response against C. albicans.

The adjuvant effect of synthetic oligodeoxynucleotide containing CpG motif converts the anti-Haemophilus influenzae type b glycoconjugates into efficient anti-polysaccharide and anti-carrier polyvalent vaccines.

Synthetic oligodeoxynucleotides containing CpG immunostimulatory sequences (ISS) have been shown to act as potent adjuvants of type 1 immune responses when co-administered with protein or peptide vaccines. We have recently shown that ISS can increase the anti-polysaccharide (CHO) and anti-tetanus toxoid (TT) or anti-diphtheria (CRM) toxoid antibody levels if used as adjuvant of anti-Haemophilus influenzae type b (Hib) CHO vaccine conjugated with TT or CRM. The analysis of anti-TT and anti-CRM IgG subclasses showed a significant increase in IgG2a, IgG2b and/or IgG3 in the presence of ISS. Anti-TT and anti-CRM antibodies were shown to neutralize the activity of both the tetanus and diphtheria toxin in vivo or in vitro tests respectively. These data show that ISS have the potential to increase host antibody response against both the CHO and the protein component of a conjugated vaccine, and encourage the investigation to identify strategies of vaccination with schedules aimed at the valuation of protein carriers as protective immunogens.

Generation of a recombinant 65-kilodalton mannoprotein, a major antigen target of cell-mediated immune response to Candida albicans.

A 65-kDa mannoprotein (CaMp65) has long been studied as a major, immunodominant antigen of the human opportunistic pathogen Candida albicans. An expression library of C. albicans was screened with serum from mice immunized with ScMp65 (ScW10), a Saccharomyces cerevisiae recombinant protein of about 48 kDa. This serum recognized the CaMp65 from a cell wall extract of C. albicans. After cloning and sequencing of the relevant C. albicans cDNA, an open reading frame encoding a protein of 379 amino acids was identified. Its deduced amino acid sequence showed regions of identity with all previously characterized tryptic fragments of CaMp65, as well as with the corresponding regions of ScMp65. A prepeptide of 32 amino acids with signal peptidase and Kex2 cleavage sites as well as a high number of potential O-glycosylation sites but no N-glycosylation sites or GPI anchor were observed in sequence studies of CaMp65. A putative adhesin RGD sequence was also present in the C-terminal region of the molecule. This triplet was absent in the ScMp65. The relevant gene (designated CaMP65) was localized to chromosome R of C. albicans as determined by pulse-field gel electrophoresis. Northern blot analysis demonstrated that gene transcription was heat inducible and associated with germ-tube formation by the fungus. A recombinant, His(6)-tagged protein (rCaMp65) was expressed in Escherichia coli under an inducible promoter. After purification by nickel-chelate affinity chromatography, the recombinant product was detected as a 47-kDa protein band in immunoblots with the anti-ScMp65 serum, as well as with CaMp65-specific monoclonal antibodies. Both ScMp65 and CaMp65 were assayed for antigenic stimulation in cultures of peripheral blood mononuclear cells (PBMC) from 10 unselected human donors. While ScMp65 was substantially nonstimulatory, both rCaMp65 and the native CaMp65 were equally able to induce lymphoproliferation of the PBMC from all the donors. In addition, a number of CD4(+) T-cell clones were generated using a C. albicans mannoprotein fraction as an antigenic stimulant. Several of these clones specifically responded to both the native and the recombinant C. albicans Mp65 but not to ScMp65. Thus, the recombinant Mp65 of C. albicans retains antigenicity and, as such, could be a valid, standardized reagent for serodiagnostic and immunological studies.

Synthetic oligodeoxynucleotide containing CpG motif induces an anti-polysaccharide type 1-like immune response after immunization of mice with Haemophilus influenzae type b conjugate vaccine.

Synthetic oligodeoxynucleotides containing CpG motifs [immunostimulatory sequences (ISS)] have been described as potent adjuvants of type 1 immune responses when co-administered with protein or peptide vaccines. To investigate their role in the immune response to polysaccharides (CHO), different preparations of anti-Haemophilus influenzae type b (Hib) conjugate vaccine were administered to mice. The unconjugated CHO did not induce the synthesis of specific antibodies even in the presence of ISS. On the other hand, anti-CHO-specific antibodies significantly increased in the presence of ISS, when tetanus (TT) or diphtheria [cross-reacting material (CRM)] toxoid-conjugated CHO were used to immunize mice. The adjuvant effect was also observed for the immune response against the carrier protein (TT and CRM). ISS insured an early and long-lasting specific IgG production. The effects of ISS on the anti-CHO immune response could be attributed to the amplification of the T help provided by the carrier. The analysis of anti-CHO IgG subclasses showed a significant increase of IgG2a and IgG3 in the presence of ISS. ISS caused a rapid release of IL-12 and IFN-gamma in sera from treated mice. This data provide a first evidence for the ability of ISS to induce an anti-CHO type 1-like immune response and demonstrate that ISS have the potential to increase host antibody response against both the CHO and the protein component of a conjugated vaccine.

Generation of an MHC class II-restricted T cell epitope by extracellular processing of hepatitis delta antigen.

Hepatitis delta virus is a human pathogen that is responsible for a severe form of hepatitis affecting hepatitis B envelope Ag carriers. We have previously identified a series of hepatitis delta Ag (HDAg) epitopes that are recognized by CD4+ T cell clones isolated from infected subjects. Herein, we show that the presentation of soluble HDAg to CD4+ T cell clones that are specific for the HDAg(106-121) epitope was exceptionally unaffected by the inhibition of the APC-processing machinery when APCs were fixed with glutaraldehyde before Ag pulsing or treated with chloroquine or brefeldin A. Interestingly, 5 h of pulsing was strictly required for the efficient presentation of the HDAg(106-21) epitope by fixed APCs, suggesting that some form of extracellular processing had occurred. Indeed, fixed APCs were able to present HDAg after only 1 h of pulsing when HDAg was preincubated with serum for 5 h. More important, presentation was completely abrogated when Ag was previously incubated in medium containing serum in the presence of a potent inhibitor of trypsin activity such as the secretory leukoprotease inhibitor. These results show that HDAg may undergo extracellular processing and suggest that the generation of immunogenic epitopes directly by serum proteases could play a role in the immune response against hepatitis delta virus during infection.

Autoimmune T-cell response to the CD4 molecule in HIV-infected patients.

In a previous study, we demonstrated that by downregulating plasma membrane CD4 and increasing its processing, human immunodeficiency (HIV)-1-gp120 unveils hidden CD4 epitopes, inducing an in vitro anti-CD4-specific T-cell response. We report herein that this mechanism may potentially have important implications in HIV immunopathogenesis, because it could take part in the severe depletion of CD4+ cells that characterizes acquired immune deficiency syndrome (AIDS) and be related to disease progression. Freshly isolated peripheral blood lymphocytes (PBMC) from about 1/4 of a conspicuous cohort of HIV-infected patients responded to CD4 and this response was correlated with beta2-microglobulin levels, widely recognized as marker for progression of HIV infection. Moreover, we provide evidence that a CD4-specific T cell priming can occur in vivo, following a gp120 or anti-CD4 monoclonal antibody (mAb)-mediated CD4 molecule downregulation on antigen-presenting cells (APC). To our knowledge, this is the first study indicating that an autoimmune T-cell response is linked to HIV infection and that it could have an important impact on the immunopathogenesis of this disease.

Isoelectricfocusing and reverse blotting as a diagnostic tool in pediatric HIV infection.

BACKGROUND: Early diagnosis of perinatally acquired HIV-infection is based on either direct HIV detection--by means of viral culture and/or PCR--or anti-HIV antibody detection. However, due to the passive, transplacental passage of maternal immunoglobulin G, antibody detection is nor reliable until 15-18 months of age. In this regard, clonotypic analysis of specific antibodies performed by isoelectricfocusing and reverse blotting (IEF-RB) can be very helpful, as it recognizes possibly different patterns between mother and infant. OBJECTIVES: We used IEF-RB in order to analyze the kinetics of development of anti-HIV antibodies in infants born to seropositive mothers. STUDY DESIGN: Sera from ten mother/infant pairs (all mothers were HIV-infected) were retrospectively analyzed in order to detect different patterns, between mother and infant, in anti-gp120 V3-loop clonotype. RESULTS: We diagnosed the real HIV status of the examined infants no later than month 6 and in one case as early as month 2. CONCLUSIONS: Considering the small size of sample number, these data are preliminary and should be confirmed by larger scale studies. However, they show IEF-RB, when applied to infants born to seropositive mothers, may be useful in evaluating the infants' dynamics of anti-HIV humoral immune response.

Fas/Apo1 mutations and autoimmune lymphoproliferative syndrome in a patient with type 2 autoimmune hepatitis.

Inherited mutations of the Fas/Apo1/CD95 gene, a cell-surface receptor involved in cell death signaling and in the control of self-reactivity, characterize the recently identified autoimmune lymphoproliferative syndromes. A patient with type 2 autoimmune hepatitis with the immunologic and genetic features of autoimmune lymphoproliferative syndrome is described. The clinical picture was dominated by liver disease with hepatosplenomegaly and positivity for anti-liver-kidney microsome 1 and anti-liver-cytosol 1 antibodies. A marked increase in CD3+CD4-CD8-T lymphocytes and inherited mutations in Fas alleles that led to the expression of a soluble form of the protein were also found. Fas-mediated apoptosis was deficient in the patient as it was in her mother and her sister, who carried the same allele 2 mutation. This observation links type 2 autoimmune hepatitis, an organ-specific disease, with a genetically determined defect in peripheral tolerance control.

Cross sectional retrospective study of prevalence of atopy among Italian military students with antibodies against hepatitis A virus.

OBJECTIVE: To investigate the working hypothesis that common infections occurring early in life prevent atopy. DESIGN: Cross sectional, retrospective study of young Italian men with results for hepatitis A serology and atopy. SETTING: Air force school of military students in Caserta, Italy. SUBJECTS: 1659 male students aged 17-24, most of whom (90%) were from central and southern Italy. MAIN OUTCOME MEASURES: Skin sensitisation and specific IgE antibodies to locally relevant airborne allergens; diagnosis of respiratory allergy (asthma or rhinitis, or both); hepatitis A seropositivity. RESULTS: 443 of the 1659 subjects (26.7%) were positive for hepatitis A virus antibody. Atopy was less common among seropositive than seronegative subjects according to skin sensitization (weal reaction > or = 3 mm) to one or more allergens (21.9% (97/443) v 30.2% (367/1216), P < 0.001); polysensitisation (sensitive to three or more allergens) (2.7% (12/443) v 6.4% (78/1216), P < 0.01); high specific IgF concentration (9.7% (43/443) v 18.4% (224/1216), P < 0.00005); and lifetime prevalence of allergic rhinitis or asthma, or both (8.4% (37/443) v 16.7% (203/1216), P < 0.001). Hepatitis A seropositivity remained inversely associated with atopy after adjusting for father's education, the number of older siblings, and the area of residence (based on the number of inhabitants). The prevalence of atopy was constantly low among seropositive subjects, whatever the number of older siblings; by contrast, it increased with a decreasing number of older siblings among seronegative subjects. CONCLUSION: Indirect but important evidence is added to the working hypothesis as common infections acquired early in life because of the presence of many older siblings (among seronegative subjects) or because of unhygienic living conditions (among seropositive subjects) may have reduced the risk of developing atopy.

Human CD4+ T-cell response to hepatitis delta virus: identification of multiple epitopes and characterization of T-helper cytokine profiles.

The T-cell-mediated immune response plays a crucial role in defense against hepatotropic viruses as well as in the pathogenesis of viral chronic hepatitides. However, very little is known about the role of specific T cells during hepatitis delta virus (HDV) infection in humans. In this study, the T-cell response to HDV in chronic hepatitis B virus (HBV) carriers with HDV superinfection was investigated at different levels. Analysis of peripheral blood mononuclear cell (PBMC) proliferation in response to a recombinant form of large hepatitis delta antigen (HDAg) revealed that 8 of 30 patients studied (27%) specifically responded to HDAg. By employing synthetic peptides spanning the entire HDAg sequence, we found that T-cell recognition was directed against different antigenic determinants, with patient-to-patient variation in the pattern of response to peptides. Interestingly, all responders had signs of inactive HDV-induced disease, while none of the patients with active disease and none of the control subjects showed any significant proliferation. More accurate information about the specific T-cell response was obtained at the clonal level. A panel of HDAg-specific CD4+ T-cell clones from three HDV-infected individuals and fine-specificity analysis revealed that the clones tested individually recognized four epitopes corresponding to amino acids (aa) 26 to 41, 50 to 65, 66 to 81, or 106 to 121 of HDAg sequence. The study of human leukocyte antigen (HLA) restriction revealed that peptides 50 to 65 and 106 to 121 were presented to specific T cells in association with multiple class II molecules. In addition, peptide 26 to 41 was efficiently generated after processing of HDAg through the endogenous processing pathway. Cytokine secretion analysis showed that all the CD4+ T-cell clones assayed were able to produce high levels of gamma interferon (IFN-gamma), belonging either to T helper-1 (Th1) or Th0 subsets and that some of them were cytotoxic in a specific assay. This study provides the first evidence that detection of a specific T-cell response to HDAg in the peripheral blood of individuals with hepatitis delta is related to the decrease of HDV-induced disease activity. The HDAg epitopes identified here and particularly those recognized by CD4+ T cells in association with multiple major histocompatibility complex class II molecules may be potentially exploited for the preparation of a vaccine for prophylaxis and therapy of HDV infection.