Immunological reactivity of a human immunodeficiency virus type I derived peptide representing a consensus sequence of the GP120 major neutralizing region V3

To reduce the opportunities for human immunodeficiency virus type 1 (HIV-1) to evade vaccine induced immunity, the development of subunit vaccines must focus on the characterization of immunogenic epitopes, which are major targets for the immune system. The most dominant site for elicitation of neutralising immune response is located on the external envelope glycoprotein gp120 within the third variable domain (V3). To overcome virus type specificity of antibodies directed to the V3-domain we designed a 36 amino acids long gp120/V3-consensus peptide (V3-C36) based on published biological data and sequence comparisons of various HIV-1 virus isolates. This peptide contains a conserved core sequence which is suggested to form a surface-exposed beta-turn. This peptide also includes T-cell epitopes defined in mice and humans, an ADCC-epitope and two highly conserved cysteine residues which were oxidized to form a cystine derivate, thus allowing correct peptide folding. In ELISA-tests, this peptide reacts with at least 90% of randomly selected sera of European and African patients infected with HIV-1 and is recognized by three different HIV-1/V3 "type-specific" antisera (MN, RF, IIIB-strain). Using this peptide as immunogen in rabbits, antisera could be raised with highly cross-reactive and HIV-1/IIIB strain neutralizing properties. Moreover, HTLV/HIV-1/IIIB specific cytotoxic T-lymphocytes (CTLs) of BALB/c mice infected with a gp120 recombinant vaccinia virus recognized the central 16- and 12-mer peptides of the V3-C36 consensus peptide in cytolytic assays, indicating perfect compatibility of the consensus peptide with the IIIB-primed CTLs. The DNA-sequence encoding the V3-consensus loop region might be an important component in newly designed recombinant subunit vaccines. In addition, due to its broad serological reactivity, the V3-consensus peptide might play an important role in special diagnostic purposes

Identification of a protein encoded in the EB-viral open reading frame BMRF2

Using monospecific rabbit sera against a peptide derived from a potential antigenic region of the Epstein-Barr viral amino acid sequence encoded in the open reading frame BMRF2 we could identify a protein-complex of 53/55 kDa in chemically induced B95-8, P3HR1 and Raji cell lines. This protein could be shown to be membrane-associated, as predicted by previous computer analysis of the secondary structure and hydrophilicity pattern, and may be a member of EBV-induced membrane proteins in lytically infected cells

Prevalence and clinical aspects of human bocavirus infection in children

AbstractHuman bocavirus (HBoV) was recently described as a new member of the Parvoviridae. In order to investigate the suggested association of HBoV with respiratory and gastric disease in infants and young children, sera of 357 paediatric patients hospitalized with infectious and non-infectious diseases were retrospectively analyzed for the presence of HBoV DNA and virus-specific antibodies using quantitative PCR and ELISA, respectively. HBoV seroprevalence was determined to range from 25% in infants younger than 1 year of age to 93% in children aged more than 3 years. Viral loads between 1 × 102 1.2 × 106 geq/mL were observed in 6.7% (20/297) of sera obtained preferentially from young children suffering from infectious diseases. HBoV genomes were furthermore detected in 5% (3/60) of sera collected from individuals with non-infectious illnesses. HBoV DNA was present most frequently in patients with respiratory disease (9.6%). Whereas only 5.2% of patients with upper respiratory tract disease were viraemic, HBoV DNA was found in 14.6% and 10.0% of patients with lower respiratory tract illness and pneumonia, respectively. Acute HBoV infections were also observed in 7.5% of patients with gastroenteritis and in one child with inflammatory bowel disease. None of 77 patients hospitalized for various other infectious diseases (e.g. rash, urinary tract infection, meningitis) displayed viraemia. In 60.9% and 47.8% of DNA–positive children, HBoV-specific IgM and IgG was observed, respectively. The present prospective study provides comprehensive data on the clinical association of acute HBoV infection with respiratory illness and on the seroprevalence of virus-specific antibodies in children

NS1 Specific CD8(+) T-Cells with Effector Function and TRBV11 Dominance in a Patient with Parvovirus B19 Associated Inflammatory Cardiomyopathy

Background: Parvovirus B19 (B19V) is the most commonly detected virus in endomyocardial biopsies (EMBs) from patients with inflammatory cardiomyopathy (DCMi). Despite the importance of T-cells in antiviral defense, little is known about the role of B19V specific T-cells in this entity. Methodology and Principal Findings: An exceptionally high B19V viral load in EMBs (115,091 viral copies/mg nucleic acids), peripheral blood mononuclear cells (PBMCs) and serum was measured in a DCMi patient at initial presentation, suggesting B19V viremia. The B19V viral load in EMBs had decreased substantially 6 and 12 months afterwards, and was not traceable in PBMCs and the serum at these times. Using pools of overlapping peptides spanning the whole B19V proteome, strong CD8(+) T-cell responses were elicited to the 10-amico-acid peptides SALKLAIYKA (19.7% of all CD8(+) cells) and QSALKLAIYK (10%) and additional weaker responses to GLCPHCINVG (0.71%) and LLHTDFEQVM (0.06%). Real-time RT-PCR of IFN gamma secretion-assay-enriched T-cells responding to the peptides, SALKLAIYKA and GLCPHCINVG, revealed a disproportionately high T-cell receptor Vbeta (TRBV) 11 expression in this population. Furthermore, dominant expression of type-1 (IFN gamma, IL2, IL27 and Tbet) and of cytotoxic T-cell markers (Perforin and Granzyme B) was found, whereas gene expression indicating type-2 (IL4, GATA3) and regulatory T-cells (FoxP3) was low. Conclusions: Our results indicate that B19V Ag-specific CD8(+) T-cells with effector function are involved in B19V associated DCMi. In particular, a dominant role of TRBV11 and type-1/CTL effector cells in the T-cell mediated antiviral immune response is suggested. The persistence of B19V in the endomyocardium is a likely antigen source for the maintenance of CD8(+) T-cell responses to the identified epitopes

Deep Sequencing Reveals Direct Targets of Gammaherpesvirus-Induced mRNA Decay and Suggests That Multiple Mechanisms Govern Cellular Transcript Escape

One characteristic of lytic infection with gammaherpesviruses, including Kaposi's sarcoma-associated herpesvirus (KSHV), Epstein-Barr virus (EBV) and murine herpesvirus 68 (MHV68), is the dramatic suppression of cellular gene expression in a process known as host shutoff. The alkaline exonuclease proteins (KSHV SOX, MHV-68 muSOX and EBV BGLF5) have been shown to induce shutoff by destabilizing cellular mRNAs. Here we extend previous analyses of cellular mRNA abundance during lytic infection to characterize the effects of SOX and muSOX, in the absence of other viral genes, utilizing deep sequencing technology (RNA-seq). Consistent with previous observations during lytic infection, the majority of transcripts are downregulated in cells expressing either SOX or muSOX, with muSOX acting as a more potent shutoff factor than SOX. Moreover, most cellular messages fall into the same expression class in both SOX- and muSOX-expressing cells, indicating that both factors target similar pools of mRNAs. More abundant mRNAs are more efficiently downregulated, suggesting a concentration effect in transcript targeting. However, even among highly expressed genes there are mRNAs that escape host shutoff. Further characterization of select escapees reveals multiple mechanisms by which cellular genes can evade downregulation. While some mRNAs are directly refractory to SOX, the steady state levels of others remain unchanged, presumably as a consequence of downstream effects on mRNA biogenesis. Collectively, these studies lay the framework for dissecting the mechanisms underlying the susceptibility of mRNA to destruction during lytic gammaherpesvirus infection

Stabilization of Scandium Terephthalate MOFs against Reversible Amorphization and Structural Phase Transition by Guest Uptake at Extreme Pressure

Previous high-pressure experiments have shown that pressure-transmitting fluids composed of small molecules can be forced inside the pores of metal organic framework materials, where they can cause phase transitions and amorphization and can even induce porosity in conventionally nonporous materials. Here we report a combined high-pressure diffraction and computational study of the structural response to methanol uptake at high pressure on a scandium terephthalate MOF (Sc2BDC3, BDC = 1,4-benzenedicarboxylate) and its nitro-functionalized derivative (Sc2(NO2–BDC)3) and compare it to direct compression behavior in a nonpenetrative hydrostatic fluid, Fluorinert-77. In Fluorinert-77, Sc2BDC3 displays amorphization above 0.1 GPa, reversible upon pressure release, whereas Sc2(NO2–BDC)3 undergoes a phase transition (C2/c to Fdd2) to a denser but topologically identical polymorph. In the presence of methanol, the reversible amorphization of Sc2BDC3 and the displacive phase transition of the nitro-form are completely inhibited (at least up to 3 GPa). Upon uptake of methanol on Sc2BDC3, the methanol molecules are found by diffraction to occupy two sites, with preferential relative filling of one site compared to the other: grand canonical Monte Carlo simulations support these experimental observations, and molecular dynamics simulations reveal the likely orientations of the methanol molecules, which are controlled at least in part by H-bonding interactions between guests. As well as revealing the atomistic origin of the stabilization of these MOFs against nonpenetrative hydrostatic fluids at high pressure, this study demonstrates a novel high-pressure approach to study adsorption within a porous framework as a function of increasing guest content, and so to determine the most energetically favorable adsorption sites