Placental expression of asialoglycoprotein receptor associated with Hepatitis B virus transmission from mother to child.

PubMed

Vyas, Ashish Kumar; Ramakrishna, Usha; Sen, Bijoya; Islam, Mojahidul; Ramakrishna, Gayatri; Patra, Sharda; Rastogi, Archana; Sarin, Shiv Kumar; Trehanpati, Nirupma

2018-04-30

Asialoglycoprotein receptor expression on hepatocytes has been associated with endocytosis, binding and uptake of hepatitis B virus. The role of asialoglycoprotein receptor in hepatitis B virus vertical transmission and its expression on placenta has not yet been studied. Thirty-four HBsAg+ve and 13 healthy pregnant mothers along with their newborns were enrolled. The former were categorized into transmitting and non-transmitting mothers based on their newborns being hepatitis B surface antigen and hepatitis B virus DNA positive. Expression of asialoglycoprotein receptor and hepatitis B surface antigen in placenta and isoform of asialoglycoprotein receptor on dendritic cell in peripheral and cord blood dendritic cells were analysed using flowcytometry, immune histochemistry, immune florescence and qRT-PCR. Twelve HBsAg+ve mothers transmitted hepatitis B virus to their newborns whereas the rest (n = 22) did not. Hepatitis B virus-transmitting mothers showed increased expression of asialoglycoprotein receptor in trophoblasts of placenta. Immunofluorescence microscopy revealed colocalization of hepatitis B surface antigen and asialoglycoprotein receptor in placenta as well as in DCs of transmitting mothers. There was no significant difference in the expression of asialoglycoprotein receptor on peripheral blood mononuclear cells or chord blood mononuclear cells between the 2 groups. However, hepatitis B virus-transmitting mothers and their HBsAg+ve newborns showed increased mRNA levels of isoform of asialoglycoprotein receptor on dendritic cell in peripheral blood mononuclear cells. Hepatitis B virus-transmitting mothers and their HBsAg+ve newborns showed an increased expression of isoform of asialoglycoprotein receptor on dendritic cell on circulating dendritic cells compared to hepatitis B virus non-transmitting mothers and their negative newborns. This study revealed that increased expression of asialoglycoprotein receptor in placenta and colocalization with hepatitis B surface antigen strongly indicates its role in intrauterine transmission of hepatitis B virus. Asialoglycoprotein receptor-blocking strategy can be used for therapeutic intervention of vertical transmission. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The molecular determinants of antibody recognition and antigenic drift in the H3 hemagglutinin of swine influenza A virus

USDA-ARS?s Scientific Manuscript database

Influenza A virus (IAV) of the H3 subtype is an important pathogen that affects both humans and swine. The main intervention strategy for preventing infection is vaccination to induce neutralizing antibodies against the surface glycoprotein hemagglutinin (HA). However, due to antigenic drift, vaccin...

Inference of Genotype–Phenotype Relationships in the Antigenic Evolution of Human Influenza A (H3N2) Viruses

PubMed Central

Steinbrück, Lars; McHardy, Alice Carolyn

2012-01-01

Distinguishing mutations that determine an organism's phenotype from (near-) neutral ‘hitchhikers’ is a fundamental challenge in genome research, and is relevant for numerous medical and biotechnological applications. For human influenza viruses, recognizing changes in the antigenic phenotype and a strains' capability to evade pre-existing host immunity is important for the production of efficient vaccines. We have developed a method for inferring ‘antigenic trees’ for the major viral surface protein hemagglutinin. In the antigenic tree, antigenic weights are assigned to all tree branches, which allows us to resolve the antigenic impact of the associated amino acid changes. Our technique predicted antigenic distances with comparable accuracy to antigenic cartography. Additionally, it identified both known and novel sites, and amino acid changes with antigenic impact in the evolution of influenza A (H3N2) viruses from 1968 to 2003. The technique can also be applied for inference of ‘phenotype trees’ and genotype–phenotype relationships from other types of pairwise phenotype distances. PMID:22532796

Pseudorabies virus-induced suppression of major histocompatibility complex class I antigen expression.

PubMed Central

Mellencamp, M W; O'Brien, P C; Stevenson, J R

1991-01-01

The ability of pseudorabies virus (PrV) to down-modulate expression of major histocompatibility complex class I antigens in murine and porcine cells was investigated. When quantified by flow cytometry, surface expression of class I Kk and Dk antigens on PrV-infected cells decreased by 60% or more. Down-modulation was associated with a decrease in total cellular class I antigens, indicating regulation at the transcriptional or posttranscriptional level. PrV did not suppress expression of transferrin receptor, suggesting a selective regulatory mechanism. Images PMID:1851884

Expression of the human hepatitis B virus large surface antigen gene in transgenic tomato plants.

PubMed

Lou, Xiao-Ming; Yao, Quan-Hong; Zhang, Zhen; Peng, Ri-He; Xiong, Ai-Sheng; Wang, Hua-Kun

2007-04-01

The original hepatitis B virus (HBV) large surface antigen gene was synthesized. In order to optimize the expression of this gene in tomato plants, the tobacco pathogenesis-related protein S signal peptide was fused to the 5' end of the modified gene and the sequence encoding amino acids S, E, K, D, E, and L was placed at the 3' end. The gene encoding the modified HBV large surface antigen under the control of a fruit-specific promoter was constructed and expressed in transgenic tomato plants. The expression of the antigen from transgenic plants was confirmed by PCR and reverse transcriptase PCR. Enzyme-linked immunoassays using a monoclonal antibody directed against human serum-derived HBsAg revealed that the maximal level of HBsAg was about 0.02% of the soluble protein in transgenic tomato fruit. The amount of HBsAg in mature fruits was found to be 65- to 171-fold larger than in small or medium fruits and leaf tissues. Examination of transgenic plant samples by transmission electron microscopy proved that HBsAg had been expressed and had accumulated. The HBsAg protein was capable of assembling into capsomers and virus-like particles. To our knowledge, this is the first time the HBV large surface antigen has been expressed in plants. This work suggests the possibility of producing a new alternative vaccine for human HBV.

Novel reassortant H9N2 viruses in pigeons and evidence for antigenic diversity of H9N2 viruses isolated from quails in Egypt

PubMed Central

Kandeil, Ahmed; El-Shesheny, Rabeh; Maatouq, Asmaa; Moatasim, Yassmin; Cai, Zhipeng; McKenzie, Pamela; Webby, Richard

2017-01-01

The endemicity of avian influenza viruses (AIVs) among Egyptian poultry represents a public health risk. Co-circulation of low pathogenic AIV H9N2 subtype with highly pathogenic AIV H5N1 subtype in Egyptian farms provides a possibility to generate novel reassortant viruses. Here, the genetic characteristics of surface glycoproteins of 59 Egyptian H9N2 viruses, isolated between 2013 and 2015, were analysed. To elucidate the potential of genetic reassortment, 10 H9N2 isolates were selected based on different avian hosts (chickens, ducks, pigeons and quails) and phylogenetic analyses of their full genome sequences were conducted. Additionally, we performed antigenic analysis to further investigate the antigenic evolution of H9N2 viruses isolated during 2011–2015. Different viral characteristics including receptor-binding affinity and drug resistance of representative Egyptian H9N2 viruses were further investigated. The surface glycoproteins of current Egyptian H9N2 viruses were closely related to viruses of the G1-like lineage isolated from Egypt. Several genetic markers that enhance virulence in poultry and transmission to humans were detected. Analysis of the full genome of 10 H9N2 isolates indicated that two pigeon isolates inherited five internal genes from Eurasian AIVs circulating in wild birds. Antigenic conservation of different Egyptian H9N2 isolates from chickens, pigeons and ducks was observed, whereas quail isolates showed antigenic drift. The Egyptian H9N2 viruses preferentially bound to the human-like receptor rather than to the avian-like receptor. Our results suggest that the endemic H9N2 viruses in Egypt contain elements that may favour avian-to-human transmission and thus represent a public health risk. PMID:27902350

Kinetics of antibody-induced modulation of respiratory syncytial virus antigens in a human epithelial cell line

PubMed Central

Sarmiento, Rosa E; Tirado, Rocio G; Valverde, Laura E; Gómez-Garcia, Beatriz

2007-01-01

Background The binding of viral-specific antibodies to cell-surface antigens usually results in down modulation of the antigen through redistribution of antigens into patches that subsequently may be internalized by endocytosis or may form caps that can be expelled to the extracellular space. Here, by use of confocal-laser-scanning microscopy we investigated the kinetics of the modulation of respiratory syncytial virus (RSV) antigen by RSV-specific IgG. RSV-infected human epithelial cells (HEp-2) were incubated with anti-RSV polyclonal IgG and, at various incubation times, the RSV-cell-surface-antigen-antibody complexes (RSV Ag-Abs) and intracellular viral proteins were detected by indirect immunoflourescence. Results Interaction of anti-RSV polyclonal IgG with RSV HEp-2 infected cells induced relocalization and aggregation of viral glycoproteins in the plasma membrane formed patches that subsequently produced caps or were internalized through clathrin-mediated endocytosis participation. Moreover, the concentration of cell surface RSV Ag-Abs and intracellular viral proteins showed a time dependent cyclic variation and that anti-RSV IgG protected HEp-2 cells from viral-induced death. Conclusion The results from this study indicate that interaction between RSV cell surface proteins and specific viral antibodies alter the expression of viral antigens expressed on the cells surface and intracellular viral proteins; furthermore, interfere with viral induced destruction of the cell. PMID:17608950

The Molecular Determinants of Antibody Recognition and Antigenic Drift in the H3 Hemagglutinin of Swine Influenza A Virus

PubMed Central

Abente, Eugenio J.; Santos, Jefferson; Lewis, Nicola S.; Gauger, Phillip C.; Stratton, Jered; Skepner, Eugene; Rajao, Daniela S.

2016-01-01

ABSTRACT Influenza A virus (IAV) of the H3 subtype is an important respiratory pathogen that affects both humans and swine. Vaccination to induce neutralizing antibodies against the surface glycoprotein hemagglutinin (HA) is the primary method used to control disease. However, due to antigenic drift, vaccine strains must be periodically updated. Six of the 7 positions previously identified in human seasonal H3 (positions 145, 155, 156, 158, 159, 189, and 193) were also indicated in swine H3 antigenic evolution. To experimentally test the effect on virus antigenicity of these 7 positions, substitutions were introduced into the HA of an isogenic swine lineage virus. We tested the antigenic effect of these introduced substitutions by using hemagglutination inhibition (HI) data with monovalent swine antisera and antigenic cartography to evaluate the antigenic phenotype of the mutant viruses. Combinations of substitutions within the antigenic motif caused significant changes in antigenicity. One virus mutant that varied at only two positions relative to the wild type had a >4-fold reduction in HI titers compared to homologous antisera. Potential changes in pathogenesis and transmission of the double mutant were evaluated in pigs. Although the double mutant had virus shedding titers and transmissibility comparable to those of the wild type, it caused a significantly lower percentage of lung lesions. Elucidating the antigenic effects of specific amino acid substitutions at these sites in swine H3 IAV has important implications for understanding IAV evolution within pigs as well as for improved vaccine development and control strategies in swine. IMPORTANCE A key component of influenza virus evolution is antigenic drift mediated by the accumulation of amino acid substitutions in the hemagglutinin (HA) protein, resulting in escape from prior immunity generated by natural infection or vaccination. Understanding which amino acid positions of the HA contribute to the ability of the virus to avoid prior immunity is important for understanding antigenic evolution and informs vaccine efficacy predictions based on the genetic sequence data from currently circulating strains. Following our previous work characterizing antigenic phenotypes of contemporary wild-type swine H3 influenza viruses, we experimentally validated that substitutions at 6 amino acid positions in the HA protein have major effects on antigenicity. An improved understanding of the antigenic diversity of swine influenza will facilitate a rational approach for selecting more effective vaccine components to control the circulation of influenza in pigs and reduce the potential for zoonotic viruses to emerge. PMID:27384658

Prediction and characterization of novel epitopes of serotype A foot-and-mouth disease viruses circulating in East Africa using site-directed mutagenesis

PubMed Central

Bari, Fufa Dawo; Parida, Satya; Asfor, Amin S.; Haydon, Daniel T.; Reeve, Richard; Paton, David J.

2015-01-01

Epitopes on the surface of the foot-and-mouth disease virus (FMDV) capsid have been identified by monoclonal antibody (mAb) escape mutant studies leading to the designation of four antigenic sites in serotype A FMDV. Previous work focused on viruses isolated mainly from Asia, Europe and Latin America. In this study we report on the prediction of epitopes in African serotype A FMDVs and testing of selected epitopes using reverse genetics. Twenty-four capsid amino acid residues were predicted to be of antigenic significance by analysing the capsid sequences (n = 56) using in silico methods, and six residues by correlating capsid sequence with serum–virus neutralization data. The predicted residues were distributed on the surface-exposed capsid regions, VP1–VP3. The significance of residue changes at eight of the predicted epitopes was tested by site-directed mutagenesis using a cDNA clone resulting in the generation of 12 mutant viruses involving seven sites. The effect of the amino acid substitutions on the antigenic nature of the virus was assessed by virus neutralization (VN) test. Mutations at four different positions, namely VP1-43, VP1-45, VP2-191 and VP3-132, led to significant reduction in VN titre (P value = 0.05, 0.05, 0.001 and 0.05, respectively). This is the first time, to our knowledge, that the antigenic regions encompassing amino acids VP1-43 to -45 (equivalent to antigenic site 3 in serotype O), VP2-191 and VP3-132 have been predicted as epitopes and evaluated serologically for serotype A FMDVs. This identifies novel capsid epitopes of recently circulating serotype A FMDVs in East Africa. PMID:25614587

Isolation of a High Affinity Neutralizing Monoclonal Antibody against 2009 Pandemic H1N1 Virus That Binds at the ‘Sa’ Antigenic Site

PubMed Central

Mishra, Arpita; Yeolekar, Leena; Dhere, Rajeev; Kapre, Subhash; Varadarajan, Raghavan; Gupta, Satish Kumar

2013-01-01

Influenza virus evades host immunity through antigenic drift and shift, and continues to circulate in the human population causing periodic outbreaks including the recent 2009 pandemic. A large segment of the population was potentially susceptible to this novel strain of virus. Historically, monoclonal antibodies (MAbs) have been fundamental tools for diagnosis and epitope mapping of influenza viruses and their importance as an alternate treatment option is also being realized. The current study describes isolation of a high affinity (K D = 2.1±0.4 pM) murine MAb, MA2077 that binds specifically to the hemagglutinin (HA) surface glycoprotein of the pandemic virus. The antibody neutralized the 2009 pandemic H1N1 virus in an in vitro microneutralization assay (IC50 = 0.08 µg/ml). MA2077 also showed hemagglutination inhibition activity (HI titre of 0.50 µg/ml) against the pandemic virus. In a competition ELISA, MA2077 competed with the binding site of the human MAb, 2D1 (isolated from a survivor of the 1918 Spanish flu pandemic) on pandemic H1N1 HA. Epitope mapping studies using yeast cell-surface display of a stable HA1 fragment, wherein ‘Sa’ and ‘Sb’ sites were independently mutated, localized the binding site of MA2077 within the ‘Sa’ antigenic site. These studies will facilitate our understanding of antigen antibody interaction in the context of neutralization of the pandemic influenza virus. PMID:23383214

Cell-mediated immunity to herpes simplex virus: recognition of type-specific and type-common surface antigens by cytotoxic T cell populations.

PubMed Central

Eberle, R; Russell, R G; Rouse, B T

1981-01-01

In this communication, we examine the specificity of anti-herpes simplex virus (HSV) cytotoxic T lymphocytes (CTL). Serological studies of the two related HSV serotypes (HSV-1 and HSV-2) have revealed both type-specific and cross-reactive antigenic determinants in the viral envelope and on the surface of infected cells. By analysis of cytotoxicity of CTL, generated in vitro by restimulation of splenocytes from mice primed with one or the other HSV serotype, the recognition of both type-specific and cross-reactive determinants on infected target cells by anti-HSV CTL was detectable. Thus, effector cells generated by priming and restimulating with the same virus recognized both type-specific and cross-reactive determinants on target cells infected with the homologous virus, but only cross-reactive determinants on target cells infected with the heterologous HSV serotype. CTL generated by restimulation with the heterologous virus were capable of recognizing only the cross-reactive determinants on either HSV-1- or HSV-2-infected target cells. These results indicate that two subpopulations of CTL exist in a population of anti-HSV immune spleen cells--those which recognize type-specific determinants and those specific for cross-reactive antigenic determinants present on the surface of HSV infected cells. The type-specific subset of anti-HSV CTL was shown to recognize the gC glycoprotein of HSV-1 infected target cells. In addition to the gC glycoprotein, at least one other type-specific surface antigen was also recognized by anti-HSV CTL in addition to the cross-reactive determinants recognized by anti-HSV CTL. PMID:6277790

Rules of co-occurring mutations characterize the antigenic evolution of human influenza A/H3N2, A/H1N1 and B viruses.

PubMed

Chen, Haifen; Zhou, Xinrui; Zheng, Jie; Kwoh, Chee-Keong

2016-12-05

The human influenza viruses undergo rapid evolution (especially in hemagglutinin (HA), a glycoprotein on the surface of the virus), which enables the virus population to constantly evade the human immune system. Therefore, the vaccine has to be updated every year to stay effective. There is a need to characterize the evolution of influenza viruses for better selection of vaccine candidates and the prediction of pandemic strains. Studies have shown that the influenza hemagglutinin evolution is driven by the simultaneous mutations at antigenic sites. Here, we analyze simultaneous or co-occurring mutations in the HA protein of human influenza A/H3N2, A/H1N1 and B viruses to predict potential mutations, characterizing the antigenic evolution. We obtain the rules of mutation co-occurrence using association rule mining after extracting HA1 sequences and detect co-mutation sites under strong selective pressure. Then we predict the potential drifts with specific mutations of the viruses based on the rules and compare the results with the "observed" mutations in different years. The sites under frequent mutations are in antigenic regions (epitopes) or receptor binding sites. Our study demonstrates the co-occurring site mutations obtained by rule mining can capture the evolution of influenza viruses, and confirms that cooperative interactions among sites of HA1 protein drive the influenza antigenic evolution.

Antigen vehiculization particles based on the Z protein of Junin virus.

PubMed

Borio, Cristina S; Bilen, Marcos F; Argüelles, Marcelo H; Goñi, Sandra E; Iserte, Javier A; Glikmann, Graciela; Lozano, Mario E

2012-11-02

Arenavirus matrix protein Z plays an important role in virus budding and is able to generate enveloped virus-like-particles (VLPs) in absence of any other viral proteins. In these VLPs, Z protein is associated to the plasma membrane inner surface by its myristoyl residue. Budding induction and vesicle formation properties can be exploited to generate enveloped VLPs platform. These structures can be designed to carry specific antigen in the inner side or on the surface of VLPs.Vaccines based on VLPs are a highly effective type of subunit vaccines that mimic the overall structure of virus particles in absence of viral nucleic acid, being noninfectious.In this work we assayed the capacity of Junin Z protein to produce VLPs carrying the green fluorescent protein (eGFP), as a model antigen. In this report the Junin Z protein ability to produce VLPs from 293T cells and its capacity to deliver a specific antigen (eGFP) fused to Z was evaluated. Confocal microscopy showed a particular membrane bending in cells expressing Z and a spot welded distribution in the cytoplasm. VLPs were detected by TEM (transmission electron microscopy) and were purified from cell supernatant. The proteinase protection assay demonstrated the VLPs integrity and the absence of degradation of the fused antigen, thus indicating its internal localization. Finally, immunization of mice with purified VLPs produced high titres of anti-eGFP antibodies compared to the controls. It was proved that VLPs can be generated from cells transfected with a fusion Junin virus Z-eGFP protein in absence of any other viral protein, and the capacity of Z protein to support fusions at the C-terminal, without impairing its budding activity, allowing vehiculization of specific antigens into VLPs.

Temperature-sensitive mutants of influenza A virus. XIV. Production and evaluation of influenza A/Georgia/74-ts-1[E] recombinant viruses in human adults.

PubMed

Richman, D D; Murphy, B R; Belshe, R B; Rusten, H M; Chanock, R M; Blacklow, N R; Parrino, T A; Rose, F B; Levine, M M; Caplan, E

1977-08-01

The two temperature-sensitive (ts) lesions present in influenza A/Hong Kong/68-ts-1[E] (H3N2 68) virus were transferred via genetic reassortment to influenza A/Georgia/74 (H3N2 74) wild-type virus. A recombinant clone possessing both ts lesions and the shutoff temperature of 38 C of the Hong Kong/68 ts donor and the two surface antigens of the Georgia/74 wild-type virus was administered to 32 seronegative adult volunteers. Thirty-one volunteers were infected, of whom only five experienced mild afebrile upper respiratory tract illness. The wild-type recipient virus was a cloned population that induced illness in five of six infected volunteers. Therfore, the attenuation exhibited by the Georgia/74-ts-1[E] virus could reasonably be assumed to be due to the acquisition of the two ts-1[E] lesions by the Georgia/74 wild-type virus. The serum and nasal wash antibody responses of the ts-1[E] vaccinees were equivalent to those of the volunteers who received wild-type virus. The two ts lesions present in the Hong Kong/68-ts-1[E] virus have now been transferred three times to a wild-type virus bearing a new hemagglutinin, and in each instance the new ts recombination exhibited a similar, satisfactory level of attenuation and antigenicity for adults. It seems likely that the transfer of the ts-1[E] lesions to any new influenza virus will regularly result in attenuation of a recombinat virus possessing the new surface antigens.

The Epstein-Barr Virus Glycoprotein gp150 Forms an Immune-Evasive Glycan Shield at the Surface of Infected Cells

PubMed Central

Gram, Anna M.; Oosenbrug, Timo; Lindenbergh, Marthe F. S.; Büll, Christian; Comvalius, Anouskha; Dickson, Kathryn J. I.; Wiegant, Joop; Vrolijk, Hans; Lebbink, Robert Jan; Wolterbeek, Ron; Adema, Gosse J.; Griffioen, Marieke; Heemskerk, Mirjam H. M.; Tscharke, David C.; Hutt-Fletcher, Lindsey M.; Ressing, Maaike E.

2016-01-01

Cell-mediated immunity plays a key role in host control of viral infection. This is exemplified by life-threatening reactivations of e.g. herpesviruses in individuals with impaired T-cell and/or iNKT cell responses. To allow lifelong persistence and virus production in the face of primed immunity, herpesviruses exploit immune evasion strategies. These include a reduction in viral antigen expression during latency and a number of escape mechanisms that target antigen presentation pathways. Given the plethora of foreign antigens expressed in virus-producing cells, herpesviruses are conceivably most vulnerable to elimination by cell-mediated immunity during the replicative phase of infection. Here, we show that a prototypic herpesvirus, Epstein-Barr virus (EBV), encodes a novel, broadly acting immunoevasin, gp150, that is expressed during the late phase of viral replication. In particular, EBV gp150 inhibits antigen presentation by HLA class I, HLA class II, and the non-classical, lipid-presenting CD1d molecules. The mechanism of gp150-mediated T-cell escape does not depend on degradation of the antigen-presenting molecules nor does it require gp150’s cytoplasmic tail. Through its abundant glycosylation, gp150 creates a shield that impedes surface presentation of antigen. This is an unprecedented immune evasion mechanism for herpesviruses. In view of its likely broader target range, gp150 could additionally have an impact beyond escape of T cell activation. Importantly, B cells infected with a gp150-null mutant EBV displayed rescued levels of surface antigen presentation by HLA class I, HLA class II, and CD1d, supporting an important role for iNKT cells next to classical T cells in fighting EBV infection. At the same time, our results indicate that EBV gp150 prolongs the timespan for producing viral offspring at the most vulnerable stage of the viral life cycle. PMID:27077376

Towards a universal vaccine for avian influenza: Protective efficacy of modified Vaccinia virus Ankara and Adenovirus vaccines expressing conserved influenza antigens in chickens challenged with low pathogenic avian influenza virus

PubMed Central

Boyd, Amy C.; Ruiz-Hernandez, Raul; Peroval, Marylene Y.; Carson, Connor; Balkissoon, Devanand; Staines, Karen; Turner, Alison V.; Hill, Adrian V.S.; Gilbert, Sarah C.; Butter, Colin

2013-01-01

Current vaccines targeting surface proteins can drive antigenic variation resulting either in the emergence of more highly pathogenic viruses or of antigenically distinct viruses that escape control by vaccination and thereby persist in the host population. Influenza vaccines typically target the highly mutable surface proteins and do not provide protection against heterologous challenge. Vaccines which induce immune responses against conserved influenza epitopes may confer protection against heterologous challenge. We report here the results of vaccination with recombinant modified Vaccinia virus Ankara (MVA) and Adenovirus (Ad) expressing a fusion construct of nucleoprotein and matrix protein (NP + M1). Prime and boost vaccination regimes were trialled in different ages of chicken and were found to be safe and immunogenic. Interferon-γ (IFN-γ) ELISpot was used to assess the cellular immune response post secondary vaccination. In ovo Ad prime followed by a 4 week post hatch MVA boost was identified as the most immunogenic regime in one outbred and two inbred lines of chicken. Following vaccination, one inbred line (C15I) was challenged with low pathogenic avian influenza (LPAI) H7N7 (A/Turkey/England/1977). Birds receiving a primary vaccination with Ad-NP + M1 and a secondary vaccination with MVA-NP + M1 exhibited reduced cloacal shedding as measured by plaque assay at 7 days post infection compared with birds vaccinated with recombinant viruses containing irrelevant antigen. This preliminary indication of efficacy demonstrates proof of concept in birds; induction of T cell responses in chickens by viral vectors containing internal influenza antigens may be a productive strategy for the development of vaccines to induce heterologous protection against influenza in poultry. PMID:23200938

Detection of Multiple Pathogens in Serum Using Silica-Encapsulated Nanotags in a Surface-Enhanced Raman Scattering-Based Immunoassay.

PubMed

Neng, Jing; Li, Yina; Driscoll, Ashley J; Wilson, William C; Johnson, Patrick A

2018-06-06

A robust immunoassay based on surface-enhanced Raman scattering (SERS) has been developed to simultaneously detect trace quantities of multiple pathogenic antigens from West Nile virus, Rift Valley fever virus, and Yersinia pestis in fetal bovine serum. Antigens were detected by capture with silica-encapsulated nanotags and magnetic nanoparticles conjugated with polyclonal antibodies. The magnetic pull-down resulted in aggregation of the immune complexes, and the silica-encapsulated nanotags provided distinct spectra corresponding to each antigen captured. The limit of detection was ∼10 pg/mL in 20% fetal bovine serum, a significant improvement over previous studies in terms of sensitivity, level of multiplexing, and medium complexity. This highly sensitive multiplex immunoassay platform provides a promising method to detect various antigens directly in crude serum samples without the tedious process of sample preparation, which is desirable for on-site diagnostic testing and real-time disease monitoring.

Antigenic properties of the envelope of influenza virus rendered soluble by surfactant-solvent systems

PubMed Central

Larin, N. M.; Gallimore, P. H.

1971-01-01

Dissociating chemical treatments employing surfactant-solvent systems were applied to purified influenza A and B viruses to obtain viral preparations possessing a significantly higher or lower haemagglutinating activity than the intact virus. All preparations, whether with high or low haemagglutinating activity, with the exception of envelope protein solubilized by Triton X-100, were significantly lacking in the ability to excite the formation of haemagglutination-inhibiting and virus-neutralizing antibodies in inoculated ferrets. In contrast to other treatments, Triton X-100 treatment of virus significantly enhanced the antigenicity of viral protein as judged by virus neutralization and haemagglutination inhibition tests. Yet the haemagglutinating activity of the envelope protein solubilized with Triton X-100 was about 1% that of the intact virus. Results suggest that the correlation assumed to exist between the haemagglutinating activity of influenza virus and its ability to excite the formation of humoral antibodies is coincidental. Another important point is that the specific antigenicity of viral protein may be lost or enhanced owing to effects, other than solubilization, by surface-active agents. PMID:5291750

Clonal analysis of T-cell responses to herpes simplex virus: isolation, characterization and antiviral properties of an antigen-specific helper T-cell clone.

PubMed

Leung, K N; Nash, A A; Sia, D Y; Wildy, P

1984-12-01

A herpes simplex virus (HSV)-specific long-term T-cell clone has been established from the draining lymph node cells of BALB/c mice; the cells required repeated in vitro restimulation with UV-irradiated virus. The established T-cell clone expresses the Thy-1 and Lyt-1+2,3- surface antigens. For optimal proliferation of the cloned cells, both the presence of specific antigen and an exogenous source of T-cell growth factor are required. The proliferative response of the cloned T cells was found to be virus-specific but it did not distinguish between HSV-1 and HSV-2. Adoptive cell transfer of the cloned T cells helped primed B cells to produce anti-herpes antibodies: the response was antigen-specific and cell dose-dependent. The clone failed to produce a significant DTH reaction in vivo, but did produce high levels of macrophage-activating factor. Furthermore, the T-cell clone could protect from HSV infection, as measured by a reduction in local virus growth, and by enhanced survival following the challenge of mice with a lethal dose of virus. The mechanism(s) whereby this clone protects in vivo is discussed.

Immunogenicity of a novel, bivalent, plant-based oral vaccine against hepatitis B and human immunodeficiency viruses.

PubMed

Shchelkunov, Sergei N; Salyaev, Rurik K; Pozdnyakov, Sergei G; Rekoslavskaya, Natalia I; Nesterov, Andrei E; Ryzhova, Tatiana S; Sumtsova, Valentina M; Pakova, Natalia V; Mishutina, Uliana O; Kopytina, Tatiana V; Hammond, Rosemarie W

2006-07-01

A synthetic chimeric gene, TBI-HBS, encoding the immunogenic ENV and GAG epitopes of human immunodeficiency virus (HIV-1) and the surface protein antigen (HBsAg) of hepatitis B virus (HBV), was expressed in tomato plants. Tomato fruits containing the TBI-HBS antigen were fed to experimental mice and, on days 14 and 28 post-feeding, high levels of HIV- and HBV-specific antibodies were present in the serum and feces of the test animals. Intraperitoneal injection of a DNA vaccine directing synthesis of the same TBI-HBsAg antigen boosted the antibody response to HIV in the blood serum; however, it had no effect on the high level of antibodies produced to HBV.

The detection of hepatitis c virus core antigen using afm chips with immobolized aptamers.

PubMed

Pleshakova, T O; Kaysheva, A L; Bayzyanova, J М; Anashkina, А S; Uchaikin, V F; Ziborov, V S; Konev, V A; Archakov, A I; Ivanov, Y D

2018-01-01

In the present study, the possibility of hepatitis C virus core antigen (HCVcoreAg) detection in buffer solution, using atomic force microscope chip (AFM-chip) with immobilized aptamers, has been demonstrated. The target protein was detected in 1mL of solution at concentrations from 10 -10 М to 10 -13 М. The registration of aptamer/antigen complexes on the chip surface was carried out by atomic force microscopy (AFM). The further mass-spectrometric (MS) identification of AFM-registered objects on the chip surface allowed reliable identification of HCVcoreAg target protein in the complexes. Aptamers, which were designed for therapeutic purposes, have been shown to be effective in HCVcoreAg detection as probe molecules. Copyright © 2017 Elsevier B.V. All rights reserved.

Determining the Epitope Dominance on the Capsid of a Serotype SAT2 Foot-and-Mouth Disease Virus by Mutational Analyses

PubMed Central

Opperman, Pamela A.; Rotherham, Lia S.; Esterhuysen, Jan; Charleston, Bryan; Juleff, Nicholas; Capozzo, Alejandra V.; Theron, Jacques

2014-01-01

ABSTRACT Monoclonal-antibody (MAb)-resistant mutants were used to map antigenic sites on foot-and-mouth disease virus (FMDV), which resulted in the identification of neutralizing epitopes in the flexible βG-βH loop in VP1. For FMDV SAT2 viruses, studies have shown that at least two antigenic sites exist. By use of an infectious SAT2 cDNA clone, 10 structurally exposed and highly variable loops were identified as putative antigenic sites on the VP1, VP2, and VP3 capsid proteins of SAT2/Zimbabwe (ZIM)/7/83 (topotype II) and replaced with the corresponding regions of SAT2/Kruger National Park (KNP)/19/89 (topotype I). Virus neutralization assays using convalescent-phase antisera raised against the parental virus, SAT2/ZIM/7/83, indicated that the mutant virus containing the TQQS-to-ETPV mutation in the N-terminal part of the βG-βH loop of VP1 showed not only a significant increase in the neutralization titer but also an increase in the index of avidity to the convalescent-phase antisera. Furthermore, antigenic profiling of the epitope-replaced and parental viruses with nonneutralizing SAT2-specific MAbs led to the identification of two nonneutralizing antigenic regions. Both regions were mapped to incorporate residues 71 to 72 of VP2 as the major contact point. The binding footprint of one of the antigenic regions encompasses residues 71 to 72 and 133 to 134 of VP2 and residues 48 to 50 of VP1, and the second antigenic region encompasses residues 71 to 72 and 133 to 134 of VP2 and residues 84 to 86 and 109 to 11 of VP1. This is the first time that antigenic regions encompassing residues 71 to 72 of VP2 have been identified on the capsid of a SAT2 FMDV. IMPORTANCE Monoclonal-antibody-resistant mutants have traditionally been used to map antigenic sites on foot-and-mouth disease virus (FMDV). However, for SAT2-type viruses, which are responsible for most of the FMD outbreaks in Africa and are the most varied of all seven serotypes, only two antigenic sites have been identified. We have followed a unique approach using an infectious SAT2 cDNA genome-length clone. Ten structurally surface-exposed, highly varied loops were identified as putative antigenic sites on the VP1, VP2, and VP3 capsid proteins of the SAT2/ZIM/7/83 virus. These regions were replaced with the corresponding regions of an antigenically disparate virus, SAT2/KNP/19/89. Antigenic profiling of the epitope-replaced and parental viruses with SAT2-specific MAbs led to the identification of two unique antibody-binding footprints on the SAT2 capsid. In this report, evidence for the structural engineering of antigenic sites of a SAT2 capsid to broaden cross-reactivity with antisera is provided. PMID:24829347

The cell-mediated immune response to ectromelia virus infection. Secondary response in vitro: specificity, nature of effector and responder cells and requirements for induction of antigenic changes in stimulator cells.

PubMed

Pang, T; Blanden, R V

1976-06-01

An in vitro culture method was used to study secondary cell-mediated responses to ectromelia virus infection in mice. Infected, syngeneic spleen cells or peritoneal cells were efficient "stimulator" cells when cultured with "responder" cells obtained from mice infected with ectromelia 4-6 weeks previously. The kinetics of generation of cytotoxic cells in cultures were determined; a peak occurred on days 4-5. A separation procedure performed on the cytotoxic cells showed that activity was associated mainly with the Ig-negative subpopulation (T cell-rich) and that H-2 compatibility between cytotoxic cells and target cells was required. The secondary response was virus-specific, at the level of both induction and target cell lysis, at least so far as ectromelia and lymphocytic choriomeningitis (LCM) viruses are concerned. Seperation of responder cells prior to culture showed that a potent secondary response was generated with the Ig-negative (T cell-rich) subpopulation and only a weak response was observed when the responder cells were Ig-positive (rich in B cells). Infected stimulator cells did not appear to secrete significant amounts of soluble antigen into the medium over 4 days of culture. Thus, antigenic patterns effective in memory T cell stimulation may be largely associated with the surfaces of infected cells.Pretreatment of ectromelia virus with UV- or gamma-irradiation did not impair its ability to induce antigenic changes in stimulator cells. Stimulator cells treated with UV-or gamma-irradiated virus for 1 h and then immediately with pactamycin to inhibit further viral protein synthesis and replication were efficient stimulators, thus indicating that antigenic changes are induced very rapidly on the surface of stimulator cells after uptake of virus. These treatments are being used to further characterize the cellular requirements in the stimulator population.

Amino Acids in Hemagglutinin Antigenic Site B Determine Antigenic and Receptor Binding Differences between A(H3N2)v and Ancestral Seasonal H3N2 Influenza Viruses

PubMed Central

Wang, Xiaoquan; Ilyushina, Natalia A.; Lugovtsev, Vladimir Y.; Bovin, Nicolai V.; Couzens, Laura K.; Gao, Jin

2016-01-01

ABSTRACT Influenza A H3N2 variant [A(H3N2)v] viruses, which have caused human infections in the United States in recent years, originated from human seasonal H3N2 viruses that were introduced into North American swine in the mid-1990s, but they are antigenically distinct from both the ancestral and current circulating H3N2 strains. A reference A(H3N2)v virus, A/Minnesota/11/2010 (MN/10), and a seasonal H3N2 strain, A/Beijing/32/1992 (BJ/92), were chosen to determine the molecular basis for the antigenic difference between A(H3N2)v and the ancestral viruses. Viruses containing wild-type and mutant MN/10 or BJ/92 hemagglutinins (HAs) were constructed and probed for reactivity with ferret antisera against MN/10 and BJ/92 in hemagglutination inhibition assays. Among the amino acids that differ between the MN/10 and BJ/92 HAs, those in antigenic site A had little impact on the antigenic phenotype. Within antigenic site B, mutations at residues 156, 158, 189, and 193 of MN/10 HA to those in BJ/92 switched the MN/10 antigenic phenotype to that of BJ/92. Mutations at residues 156, 157, 158, 189, and 193 of BJ/92 HA to amino acids present in MN/10 were necessary for BJ/92 to become antigenically similar to MN/10. The HA amino acid substitutions responsible for switching the antigenic phenotype also impacted HA binding to sialyl receptors that are usually present in the human respiratory tract. Our study demonstrates that antigenic site B residues play a critical role in determining both the unique antigenic phenotype and receptor specificity of A(H3N2)v viruses, a finding that may facilitate future surveillance and risk assessment of novel influenza viruses. IMPORTANCE Influenza A H3N2 variant [A(H3N2)v] viruses have caused hundreds of human infections in multiple states in the United States since 2009. Most cases have been children who had contact with swine in agricultural fairs. These viruses originated from human seasonal H3N2 viruses that were introduced into the U.S. swine population in the mid-1990s, but they are different from both these ancestral viruses and current circulating human seasonal H3N2 strains in terms of their antigenic characteristics as measured by hemagglutination inhibition (HI) assay. In this study, we identified amino acids in antigenic site B of the surface glycoprotein hemagglutinin (HA) that explain the antigenic difference between A(H3N2)v and the ancestral H3N2 strains. These amino acid mutations also alter binding to minor human-type glycans, suggesting that host adaptation may contribute to the selection of antigenically distinct H3N2 variants which pose a threat to public health. PMID:27807224

Universal influenza vaccines, a dream to be realized soon.

PubMed

Zhang, Han; Wang, Li; Compans, Richard W; Wang, Bao-Zhong

2014-04-29

Due to frequent viral antigenic change, current influenza vaccines need to be re-formulated annually to match the circulating strains for battling seasonal influenza epidemics. These vaccines are also ineffective in preventing occasional outbreaks of new influenza pandemic viruses. All these challenges call for the development of universal influenza vaccines capable of conferring broad cross-protection against multiple subtypes of influenza A viruses. Facilitated by the advancement in modern molecular biology, delicate antigen design becomes one of the most effective factors for fulfilling such goals. Conserved epitopes residing in virus surface proteins including influenza matrix protein 2 and the stalk domain of the hemagglutinin draw general interest for improved antigen design. The present review summarizes the recent progress in such endeavors and also covers the encouraging progress in integrated antigen/adjuvant delivery and controlled release technology that facilitate the development of an affordable universal influenza vaccine.

Universal Influenza Vaccines, a Dream to Be Realized Soon

PubMed Central

Zhang, Han; Wang, Li; Compans, Richard W.; Wang, Bao-Zhong

2014-01-01

Due to frequent viral antigenic change, current influenza vaccines need to be re-formulated annually to match the circulating strains for battling seasonal influenza epidemics. These vaccines are also ineffective in preventing occasional outbreaks of new influenza pandemic viruses. All these challenges call for the development of universal influenza vaccines capable of conferring broad cross-protection against multiple subtypes of influenza A viruses. Facilitated by the advancement in modern molecular biology, delicate antigen design becomes one of the most effective factors for fulfilling such goals. Conserved epitopes residing in virus surface proteins including influenza matrix protein 2 and the stalk domain of the hemagglutinin draw general interest for improved antigen design. The present review summarizes the recent progress in such endeavors and also covers the encouraging progress in integrated antigen/adjuvant delivery and controlled release technology that facilitate the development of an affordable universal influenza vaccine. PMID:24784572

The Density Code for the Development of a Vaccine?

PubMed Central

Cheng, Wei

2016-01-01

The development of prophylactic vaccines remains largely empirical in nature and rarely have general rules been applied in the strategic decision and the formulation of a viral vaccine. Currently there are a total of 15 virus agents from 12 unique virus families with vaccines licensed by the US Food and Drug Administration. Extensive structural information on these viral particles and potential mechanisms of protection are available for the majority of these virus pathogens and their respective vaccines. Here I review the quantitative features of these viral surface antigens in relation to the molecular mechanisms of B cell activation, and point out a potential correlation between the density of immunogenic proteins displayed on the surface of the vaccine antigen carrier and the success of a vaccine. These features help us understand the humoral immunity induced by viral vaccines on a quantitative ground and re-emphasize the importance of antigen density on the activation of the immune system. Although the detailed mechanisms behind this phenomenon remain to be explored, it implies that both the size of antigen carriers and the density of immunogenic proteins displayed on these carriers are important parameters that may need to be optimized for the formulation of a vaccine. PMID:27649885

Genetic and antigenic relationship of foot-and-mouth disease virus serotype O isolates with the vaccine strain O1/BFS.

PubMed

Xu, Wanhong; Zhang, Zhidong; Nfon, Charles; Yang, Ming

2018-05-15

Foot-and-mouth disease serotype O viruses (FMDV/O) are responsible for the most outbreaks in FMD endemic countries. O1/BFS is one of the recommended FMD/O vaccine strains by World Reference Laboratory for FMD. In the current study, FMDV/O1 BFS vaccine strain and serotype O field isolates (45) were analyzed phylogenetically and antigenically to gain more insight into the genetic and antigenic characteristics of the vaccine strain and field isolates. O1/BFS showed similarity with 89% of the field isolates using a virus neutralization test (VNT). The P1 region encoding the FMDV capsid was sequenced and analysed for 46 strains of FMDV/O. Phylogenetic analysis showed these viruses originated from five continents and covered eight of 11 reported topotypes. Five isolates that demonstrated low antigenic similarities with O1/BFS were analyzed for their antigenic variation at the known neutralizing antigenic sites. Three of the five isolates demonstrated unique amino acid substitutions at various antigenic sites. No unique amino acid substitutions were observed for the other two unmatched isolates. Positively selected residues were identified on the surface of the FMD virus capsid supporting that it is important to continuously monitor field isolates for their antigenic and phenotypic changes. In conclusion, the vaccine strain O1/BFS is likely to confer protection against 89% of the 45 FMDV/O isolates based on VNT. Thus O1/BFS vaccine strain is still suitable for use in global FMD serotype O outbreak control. Combining data from phylogenetic, molecular and antigenic analysis can provide improvements in the process of vaccine selection. Crown Copyright © 2018. Published by Elsevier Ltd. All rights reserved.

Evaluation of single-round infectious, chimeric dengue type 1 virus as an antigen for dengue functional antibody assays.

PubMed

Yamanaka, Atsushi; Suzuki, Ryosuke; Konishi, Eiji

2014-07-23

Dengue fever and dengue hemorrhagic fever are endemic throughout tropical and subtropical countries. Four serotypes of dengue viruses (DENV-1 to DENV-4), each with several genotypes including various subclades, are co-distributed in most endemic areas. Infection-neutralizing and -enhancing antibodies are believed to play protective and pathogenic roles, respectively. Measurement of these functional antibodies against a variety of viral strains is thus important for evaluating coverage and safety of dengue vaccine candidates. Although transportation of live virus materials beyond national borders is increasingly limited, this difficulty may be overcome using biotechnology that enables generation of an antibody-assay antigen equivalent to authentic virus based on viral sequence information. A rapid system to produce flavivirus single-round infectious particles (SRIPs) was recently developed using a Japanese encephalitis virus (JEV) subgenomic replicon plasmid. This system allows production of chimeric SRIPs that have surface proteins of other flaviviruses. In the present study, SRIPs of DENV-1 (D1-SRIPs) were evaluated as an antigen for functional antibody assays. Inclusion of the whole mature capsid gene of JEV into the replicon plasmid provided higher D1-SRIP yields than did its exclusion in cases where a DENV-1 surface-protein-expressing plasmid was used for co-transfection of 293T cells with the replicon plasmid. In an assay to measure the balance between neutralizing and enhancing activities, dose (antibody dilution)-dependent activity curves in dengue-immune human sera or mouse monoclonal antibodies obtained using D1-SRIP antigen were equivalent to those obtained using DENV-1 antigen. Similar results were obtained using additional DENV-2 and DENV-3 systems. In a conventional Vero-cell neutralization test, a significant correlation was shown between antibody titers obtained using D1-SRIP and DENV-1 antigens. These results demonstrate the utility of D1-SRIPs as an alternative antigen to authentic DENV-1 in functional antibody assays. SRIP antigens may contribute to dengue vaccine candidate evaluation, understanding of dengue pathogenesis, and development of serodiagnostic systems. Copyright © 2014 Elsevier Ltd. All rights reserved.

PROPAGATION OF INFLUENZA VIRUS IN "IMMUNE" ENVIRONMENTS

PubMed Central

Magill, Thomas P.

1955-01-01

Influenza virus can survive, and can be propagated in immunological environments induced in mice by vaccination with the homologous strain of virus: survival was associated with the emergence of variants which differed from the parent strain in antigenic characteristics. The data concerning hemagglutinating activity of the variants, on the one hand, and of the antigenicity, on the other, are compatible with the concept that the structure of the influenza virus includes a surface arrangement which is distinct from the inner virus bulk. The points (a) that propagation was accomplished with difficulty whenever the immunological environment was altered, and (b) that once established, passage was continued without difficulty, are interpreted to indicate that the mechanism of variation may involve a rearrangement of the basic hereditary mechanism. PMID:13252183

Dynamically correlated mutations drive human Influenza A evolution.

PubMed

Tria, F; Pompei, S; Loreto, V

2013-01-01

Human Influenza A virus undergoes recurrent changes in the hemagglutinin (HA) surface protein, primarily involved in the human antibody recognition. Relevant antigenic changes, enabling the virus to evade host immune response, have been recognized to occur in parallel to multiple mutations at antigenic sites in HA. Yet, the role of correlated mutations (epistasis) in driving the molecular evolution of the virus still represents a challenging puzzle. Further, though circulation at a global geographic level is key for the survival of Influenza A, its role in shaping the viral phylodynamics remains largely unexplored. Here we show, through a sequence based epidemiological model, that epistatic effects between amino acids substitutions, coupled with a reservoir that mimics worldwide circulating viruses, are key determinants that drive human Influenza A evolution. Our approach explains all the up-to-date observations characterizing the evolution of H3N2 subtype, including phylogenetic properties, nucleotide fixation patterns, and composition of antigenic clusters.

Localization and force analysis at the single virus particle level using atomic force microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Chih-Hao; Horng, Jim-Tong; Chang, Jeng-Shian

2012-01-06

Highlights: Black-Right-Pointing-Pointer Localization of single virus particle. Black-Right-Pointing-Pointer Force measurements. Black-Right-Pointing-Pointer Force mapping. -- Abstract: Atomic force microscopy (AFM) is a vital instrument in nanobiotechnology. In this study, we developed a method that enables AFM to simultaneously measure specific unbinding force and map the viral glycoprotein at the single virus particle level. The average diameter of virus particles from AFM images and the specificity between the viral surface antigen and antibody probe were integrated to design a three-stage method that sets the measuring area to a single virus particle before obtaining the force measurements, where the influenza virus was usedmore » as the object of measurements. Based on the purposed method and performed analysis, several findings can be derived from the results. The mean unbinding force of a single virus particle can be quantified, and no significant difference exists in this value among virus particles. Furthermore, the repeatability of the proposed method is demonstrated. The force mapping images reveal that the distributions of surface viral antigens recognized by antibody probe were dispersed on the whole surface of individual virus particles under the proposed method and experimental criteria; meanwhile, the binding probabilities are similar among particles. This approach can be easily applied to most AFM systems without specific components or configurations. These results help understand the force-based analysis at the single virus particle level, and therefore, can reinforce the capability of AFM to investigate a specific type of viral surface protein and its distributions.« less

Ultrastructural localization of human HL-A membrane antigens by use of hybrid antibodies

PubMed Central

Neauport-Sautes, Catherine; Silvestre, Daniele; Niccolai, Marie-Gabrielle; Kourilsky, F. M.; Levy, J. P.

1972-01-01

The localization of HL-A histocompatibility antigens at the surface of human lymphocytes in electron microscopy has been studied using hybrid antibodies to bind electron-dense particles (ferritin and plant viruses) to anti-HL-A antibody. A discontinuous distribution of the markers is observed at the cell surface, which is identical with that described for H-2 antigens on mouse lymphocytes with the same technique. Double labelling experiments suggest that the areas of the cell surface where HL-A antigens are detected contain also the heterologous lymphocyte antigens detected by an anti-thymocyte serum and that HL-A antigens are not renewed at a detectable level during the period of the labelling procedure in the areas of the cell surface which are not labelled primarily with ferritin-anti-IgG-anti-HL-A complexes. The interpretation of the discontinuous labelling of HL-A antigens with direct immunoferritin techniques is discussed. ImagesFIG. 2FIG. 3FIG. 4FIG. 5 PMID:5063188

Rapid detection of hepatitis B virus surface antigen by an agglutination assay mediated by a bispecific diabody against both human erythrocytes and hepatitis B virus surface antigen.

PubMed

Chen, Yu-Ping; Qiao, Yuan-Yuan; Zhao, Xiao-Hang; Chen, Hong-Song; Wang, Yan; Wang, Zhuozhi

2007-06-01

Bispecific antibodies have immense potential for use in clinical applications. In the present study, a bispecific diabody against human red blood cells (RBCs) and hepatitis B virus surface antigen (HBsAg) was used to detect HBsAg in blood specimens. The bispecific diabody was constructed by crossing over the variable region of the heavy chains and the light chains of anti-RBC and anti-HBsAg antibodies with a short linker, SRGGGS. In enzyme-linked immunosorbent assays, this bispecific diabody showed specific binding to both RBCs and HBsAg. When this bispecific diabody was mixed with human blood specimens in the presence of HBsAg, the dual binding sites of the diabody caused agglutination of human RBCs. This diabody-mediated agglutination assay was then used to test 712 clinical blood specimens and showed 97.7% sensitivity and 100% specificity when the results were compared with those of the conventional immunoassay, which was used as a reference. This autologous RBC agglutination assay provides a simple approach for rapid screening for HBsAg in blood specimens.

Tomato bushy stunt virus (TBSV), a versatile platform for polyvalent display of antigenic epitopes and vaccine design.

PubMed

Kumar, Shantanu; Ochoa, Wendy; Singh, Pratik; Hsu, Catherine; Schneemann, Anette; Manchester, Marianne; Olson, Mark; Reddy, Vijay

2009-05-25

Viruses-like particles (VLPs) are frequently being used as platforms for polyvalent display of foreign epitopes of interest on their capsid surface to improve their presentation enhancing the antigenicity and host immune response. In the present study, we used the VLPs of Tomato bushy stunt virus (TBSV), an icosahedral plant virus, as a platform to display 180 copies of 16 amino acid epitopes of ricin toxin fused to the C-terminal end of a modified TBSV capsid protein (NDelta52). Expression of the chimeric recombinant protein in insect cells resulted in spontaneous assembly of VLPs displaying the ricin epitope. Cryo-electron microscopy and image reconstruction of the chimeric VLPs at 22 A resolution revealed the locations and orientation of the ricin epitope exposed on the TBSV capsid surface. Furthermore, injection of chimeric VLPs into mice generated antisera that detected the native ricin toxin. The ease of fusing of short peptides of 15-20 residues and their ability to form two kinds (T=1, T=3) of bio-nanoparticles that result in the display of 60 or 180 copies of less constrained and highly exposed antigenic epitopes makes TBSV an attractive and versatile display platform for vaccine design.

Clonal analysis of T-cell responses to herpes simplex virus: isolation, characterization and antiviral properties of an antigen-specific helper T-cell clone.

PubMed Central

Leung, K N; Nash, A A; Sia, D Y; Wildy, P

1984-01-01

A herpes simplex virus (HSV)-specific long-term T-cell clone has been established from the draining lymph node cells of BALB/c mice; the cells required repeated in vitro restimulation with UV-irradiated virus. The established T-cell clone expresses the Thy-1 and Lyt-1+2,3- surface antigens. For optimal proliferation of the cloned cells, both the presence of specific antigen and an exogenous source of T-cell growth factor are required. The proliferative response of the cloned T cells was found to be virus-specific but it did not distinguish between HSV-1 and HSV-2. Adoptive cell transfer of the cloned T cells helped primed B cells to produce anti-herpes antibodies: the response was antigen-specific and cell dose-dependent. The clone failed to produce a significant DTH reaction in vivo, but did produce high levels of macrophage-activating factor. Furthermore, the T-cell clone could protect from HSV infection, as measured by a reduction in local virus growth, and by enhanced survival following the challenge of mice with a lethal dose of virus. The mechanism(s) whereby this clone protects in vivo is discussed. PMID:6209206

Stable expression of hepatitis delta virus antigen in a eukaryotic cell line.

PubMed

Macnaughton, T B; Gowans, E J; Reinboth, B; Jilbert, A R; Burrell, C J

1990-06-01

The gene encoding the hepatitis delta virus structural antigen (HDAg) was linked to a neomycin resistance gene in a retrovirus expression vector, and human HepG2 cells were transfected with the recombinant plasmid. A stable cell line was cloned that expressed HDAg in the nuclei of 100% of cells, in a pattern indicating a close relationship with cell nucleoli. Analysis of partially purified recombinant HDAg by HPLC showed an Mr in the range of 7 x 10(5) to 2 x 10(6), which appeared to contain conformation-dependent epitopes, whereas the density of the antigen was 1.19 g/ml by equilibrium centrifugation in caesium chloride, and in rate zonal centrifugation it sedimented with a value of 50S, close to that of particulate hepatitis B virus surface antigen. Immunoblotting demonstrated a single polypeptide with an Mr of 24K which corresponded to the smaller of the two HDAg-specific polypeptides present in infected sera. The recombinant HDAg polypeptide was shown to be a RNA-binding protein with specificity for both genomic and antigenomic species of hepatitis delta virus RNA.

Preventing vaccinia virus class-I epitopes presentation by HSV-ICP47 enhances the immunogenicity of a TAP-independent cancer vaccine epitope.

PubMed

Raafat, Nermin; Sadowski-Cron, Charlotte; Mengus, Chantal; Heberer, Michael; Spagnoli, Giulio C; Zajac, Paul

2012-09-01

Herpes simplex virus protein ICP47, encoded by US12 gene, strongly downregulates major histocompatibility complex (MHC) class-I antigen restricted presentation by blocking transporter associated with antigen processing (TAP) protein. To decrease viral vector antigenic immunodominance and MHC class-I driven clearance, we engineered recombinant vaccinia viruses (rVV) expressing ICP47 alone (rVV-US12) or together with endoplasmic reticulum (ER)-targeted Melan-A/MART-1(27-35) model tumor epitope (rVV-MUS12). In this study, we show that antigen presenting cells (APC), infected with rVV-US12, display a decreased ability to present TAP dependent MHC class-I restricted viral antigens to CD8+ T-cells. While HLA class-I cell surface expression is strongly downregulated, other important immune related molecules such as CD80, CD44 and, most importantly, MHC class-II are unaffected. Characterization of rVV-MUS12 infected cells demonstrates that over-expression of a TAP-independent peptide, partially compensates for ICP47 induced surface MHC class-I downregulation (30% vs. 70% respectively). Most importantly, in conditions where clearance of infected APC by virus-specific CTL represents a limiting factor, a significant enhancement of CTL responses to the tumor epitope can be detected in cultures stimulated with rVV-MUS12, as compared to those stimulated by rVV-MART alone. Such reagents could become of high relevance in multiple boost protocols required for cancer immunotherapy, to limit vector-specific responsiveness. Copyright © 2011 UICC.

Nanoparticle-based flow virometry for the analysis of individual virions

PubMed Central

Arakelyan, Anush; Fitzgerald, Wendy; Margolis, Leonid; Grivel, Jean-Charles

2013-01-01

While flow cytometry has been used to analyze the antigenic composition of individual cells, the antigenic makeup of viral particles is still characterized predominantly in bulk. Here, we describe a technology, “flow virometry,” that can be used for antigen detection on individual virions. The technology is based on binding magnetic nanoparticles to virions, staining the virions with monoclonal antibodies, separating the formed complexes with magnetic columns, and characterizing them with flow cytometers. We used this technology to study the distribution of two antigens (HLA-DR and LFA-1) that HIV-1 acquires from infected cells among individual HIV-1 virions. Flow virometry revealed that the antigenic makeup of virions from a single preparation is heterogeneous. This heterogeneity could not be detected with bulk analysis of viruses. Moreover, in two preparations of the same HIV-1 produced by different cells, the distribution of antigens among virions was different. In contrast, HIV-1 of two different HIV-1 genotypes replicating in the same cells became somewhat antigenically similar. This nanotechnology allows the study of virions in bodily fluids without virus propagation and in principle is not restricted to the analysis of HIV, but can be applied to the analysis of the individual surface antigenic makeup of any virus. PMID:23925291

Boosting immunity to small tumor-associated carbohydrates with bacteriophage qβ capsids.

PubMed

Yin, Zhaojun; Comellas-Aragones, Marta; Chowdhury, Sudipa; Bentley, Philip; Kaczanowska, Katarzyna; Benmohamed, Lbachir; Gildersleeve, Jeffrey C; Finn, M G; Huang, Xuefei

2013-01-01

The development of an effective immunotherapy is an attractive strategy toward cancer treatment. Tumor associated carbohydrate antigens (TACAs) are overexpressed on a variety of cancer cell surfaces, which present tempting targets for anticancer vaccine development. However, such carbohydrates are often poorly immunogenic. To overcome this challenge, we show here that the display of a very weak TACA, the monomeric Tn antigen, on bacteriophage Qβ virus-like particles elicits powerful humoral responses to the carbohydrate. The effects of adjuvants, antigen display pattern, and vaccine dose on the strength and subclasses of antibody responses were established. The local density of antigen rather than the total amount of antigen administered was found to be crucial for induction of high Tn-specific IgG titers. The ability to display antigens in an organized and high density manner is a key advantage of virus-like particles such as Qβ as vaccine carriers. Glycan microarray analysis showed that the antibodies generated were highly selective toward Tn antigens. Furthermore, Qβ elicited much higher levels of IgG antibodies than other types of virus-like particles, and the IgG antibodies produced reacted strongly with the native Tn antigens on human leukemia cells. Thus, Qβ presents a highly attractive platform for the development of carbohydrate-based anticancer vaccines.

Sialidase-Inhibiting Antibody Titers Correlate with Protection from Heterologous Influenza Virus Strains of the Same Neuraminidase Subtype.

PubMed

Walz, Lisa; Kays, Sarah-Katharina; Zimmer, Gert; von Messling, Veronika

2018-06-20

Immune responses induced by currently licensed inactivated influenza vaccines are mainly directed against the hemagglutinin (HA) glycoprotein, the immunodominant antigen of influenza viruses. The resulting antigenic drift of HA requires frequent updating of the vaccine composition and annual revaccination. On the other hand, the level of antibodies directed against the neuraminidase (NA) glycoprotein, the second major influenza virus antigen, vary greatly. To investigate the potential of the more conserved NA protein for the induction of a subtype-specific protection, vesicular stomatitis virus-based replicons expressing a panel of N1 proteins from prototypic seasonal and pandemic H1N1 strain and human H5N1 and H7N9 isolates were generated. Immunization of mice and ferrets with the replicon carrying the matched N1 protein resulted in robust humoral and cellular immune responses and protected against challenge with the homologous influenza virus with similar efficacy as the matched HA protein, illustrating the potential of the NA protein as vaccine antigen. The extent of protection after immunization with mismatched N1 proteins correlated with the level of cross-reactive sialidase-inhibiting antibody titers. Passive serum transfer experiments in mice confirmed that these functional antibodies determine subtype-specific cross-protection. Our findings illustrate the potential of NA-specific immunity for achieving broader protection against antigenic drift variants or newly emerging viruses carrying the same NA but a different HA subtype. IMPORTANCE Despite the availability of vaccines, annual influenza virus epidemics cause 250,000 to 500,000 deaths worldwide. Currently licensed inactivated vaccines, which are standardized for the amount of the hemagglutinin (HA) antigen, primarily induce strain-specific antibodies whereas the immune response to the neuraminidase (NA) antigen, which is also present on the viral surface, is usually low. Using NA-expressing single-cycle vesicular stomatitis virus replicons, we show that the NA antigen not only conferred protection of mice and ferrets to the matched influenza strains, but also against viruses carrying NA proteins from other strains of the same subtype. The extent of protection correlated with the level of cross-reactive NA-inhibiting antibodies. This highlights the potential of the NA antigen for the development of more broadly protective influenza vaccines. Such vaccines may also provide partial protection against newly emerging strains with the same NA but a different HA subtype. Copyright © 2018 American Society for Microbiology.

Novel antibody binding determinants on the capsid surface of serotype O foot-and-mouth disease virus

PubMed Central

Asfor, Amin S.; Upadhyaya, Sasmita; Knowles, Nick J.; King, Donald P.; Paton, David J.

2014-01-01

Five neutralizing antigenic sites have been described for serotype O foot-and-mouth disease viruses (FMDV) based on monoclonal antibody (mAb) escape mutant studies. However, a mutant virus selected to escape neutralization of mAb binding at all five sites was previously shown to confer complete cross-protection with the parental virus in guinea pig challenge studies, suggesting that amino acid residues outside the mAb binding sites contribute to antibody-mediated in vivo neutralization of FMDV. Comparison of the ability of bovine antisera to neutralize a panel of serotype O FMDV identified three novel putative sites at VP2-74, VP2-191 and VP3-85, where amino acid substitutions correlated with changes in sero-reactivity. The impact of these positions was tested using site-directed mutagenesis to effect substitutions at critical amino acid residues within an infectious copy of FMDV O1 Kaufbeuren (O1K). Recovered viruses containing additional mutations at VP2-74 and VP2-191 exhibited greater resistance to neutralization with both O1K guinea pig and O BFS bovine antisera than a virus that was engineered to include only mutations at the five known antigenic sites. The changes at VP2-74 and VP3-85 are adjacent to critical amino acids that define antigenic sites 2 and 4, respectively. However VP2-191 (17 Å away from VP2-72), located at the threefold axis and more distant from previously identified antigenic sites, exhibited the most profound effect. These findings extend our knowledge of the surface features of the FMDV capsid known to elicit neutralizing antibodies, and will improve our strategies for vaccine strain selection and rational vaccine design. PMID:24584474

Gene constellation of influenza A virus reassortants with high growth phenotype prepared as seed candidates for vaccine production.

PubMed

Fulvini, Andrew A; Ramanunninair, Manojkumar; Le, Jianhua; Pokorny, Barbara A; Arroyo, Jennifer Minieri; Silverman, Jeanmarie; Devis, Rene; Bucher, Doris

2011-01-01

Influenza A virus vaccines undergo yearly reformulations due to the antigenic variability of the virus caused by antigenic drift and shift. It is critical to the vaccine manufacturing process to obtain influenza A seed virus that is antigenically identical to circulating wild type (wt) virus and grows to high titers in embryonated chicken eggs. Inactivated influenza A seasonal vaccines are generated by classical reassortment. The classical method takes advantage of the ability of the influenza virus to reassort based on the segmented nature of its genome. In ovo co-inoculation of a high growth or yield (hy) donor virus and a low yield wt virus with antibody selection against the donor surface antigens results in progeny viruses that grow to high titers in ovo with wt origin hemagglutinin (HA) and neuraminidase (NA) glycoproteins. In this report we determined the parental origin of the remaining six genes encoding the internal proteins that contribute to the hy phenotype in ovo. The genetic analysis was conducted using reverse transcription-polymerase chain reaction (RT-PCR) and restriction fragment length polymorphism (RFLP). The characterization was conducted to determine the parental origin of the gene segments (hy donor virus or wt virus), gene segment ratios and constellations. Fold increase in growth of reassortant viruses compared to respective parent wt viruses was determined by hemagglutination assay titers. In this study fifty-seven influenza A vaccine candidate reassortants were analyzed for the presence or absence of correlations between specific gene segment ratios, gene constellations and hy reassortant phenotype. We found two gene ratios, 6:2 and 5:3, to be the most prevalent among the hy reassortants analyzed, although other gene ratios also conferred hy in certain reassortants.

Gene Constellation of Influenza A Virus Reassortants with High Growth Phenotype Prepared as Seed Candidates for Vaccine Production

PubMed Central

Fulvini, Andrew A.; Ramanunninair, Manojkumar; Le, Jianhua; Pokorny, Barbara A.; Arroyo, Jennifer Minieri; Silverman, Jeanmarie; Devis, Rene; Bucher, Doris

2011-01-01

Background Influenza A virus vaccines undergo yearly reformulations due to the antigenic variability of the virus caused by antigenic drift and shift. It is critical to the vaccine manufacturing process to obtain influenza A seed virus that is antigenically identical to circulating wild type (wt) virus and grows to high titers in embryonated chicken eggs. Inactivated influenza A seasonal vaccines are generated by classical reassortment. The classical method takes advantage of the ability of the influenza virus to reassort based on the segmented nature of its genome. In ovo co-inoculation of a high growth or yield (hy) donor virus and a low yield wt virus with antibody selection against the donor surface antigens results in progeny viruses that grow to high titers in ovo with wt origin hemagglutinin (HA) and neuraminidase (NA) glycoproteins. In this report we determined the parental origin of the remaining six genes encoding the internal proteins that contribute to the hy phenotype in ovo. Methodology The genetic analysis was conducted using reverse transcription-polymerase chain reaction (RT-PCR) and restriction fragment length polymorphism (RFLP). The characterization was conducted to determine the parental origin of the gene segments (hy donor virus or wt virus), gene segment ratios and constellations. Fold increase in growth of reassortant viruses compared to respective parent wt viruses was determined by hemagglutination assay titers. Significance In this study fifty-seven influenza A vaccine candidate reassortants were analyzed for the presence or absence of correlations between specific gene segment ratios, gene constellations and hy reassortant phenotype. We found two gene ratios, 6∶2 and 5∶3, to be the most prevalent among the hy reassortants analyzed, although other gene ratios also conferred hy in certain reassortants. PMID:21695145

Mucosal and systemic adjuvant activity of alphavirus replicon particles

NASA Astrophysics Data System (ADS)

Thompson, Joseph M.; Whitmore, Alan C.; Konopka, Jennifer L.; Collier, Martha L.; Richmond, Erin M. B.; Davis, Nancy L.; Staats, Herman F.; Johnston, Robert E.

2006-03-01

Vaccination represents the most effective control measure in the fight against infectious diseases. Local mucosal immune responses are critical for protection from, and resolution of, infection by numerous mucosal pathogens. Antigen processing across mucosal surfaces is the natural route by which mucosal immunity is generated, as peripheral antigen delivery typically fails to induce mucosal immune responses. However, we demonstrate in this article that mucosal immune responses are evident at multiple mucosal surfaces after parenteral delivery of Venezuelan equine encephalitis virus replicon particles (VRP). Moreover, coinoculation of null VRP (not expressing any transgene) with inactivated influenza virions, or ovalbumin, resulted in a significant increase in antigen-specific systemic IgG and fecal IgA antibodies, compared with antigen alone. Pretreatment of VRP with UV light largely abrogated this adjuvant effect. These results demonstrate that alphavirus replicon particles possess intrinsic systemic and mucosal adjuvant activity and suggest that VRP RNA replication is the trigger for this activity. We feel that these observations and the continued experimentation they stimulate will ultimately define the specific components of an alternative pathway for the induction of mucosal immunity, and if the activity is evident in humans, will enable new possibilities for safe and inexpensive subunit and inactivated vaccines. vaccine vector | Venezuelan equine encephalitis virus | viral immunology | RNA virus

[Production of marker-free plants expressing the gene of the hepatitis B virus surface antigen].

PubMed

Rukavtsova, E B; Gaiazova, A R; Chebotareva, E N; Bur'ianova, Ia I

2009-08-01

The pBM plasmid, carrying the gene of hepatitis B virus surface antigen (HBsAg) and free of any selection markers of antibiotic or herbicide resistance, was constructed for genetic transformation of plants. A method for screening transformed plant seedlings on nonselective media was developed. Enzyme immunoassay was used for selecting transgenic plants with HBsAg gene among the produced regenerants; this method provides for a high sensitivity detection of HBsAg in plant extracts. Tobacco and tomato transgenic lines synthesizing this antigen at a level of 0.01-0.05% of the total soluble protein were obtained. The achieved level of HBsAg synthesis is sufficient for preclinical trials of the produced plants as a new generation safe edible vaccine. The developed method for selecting transformants can be used for producing safe plants free of selection markers.

Recognition of Typhus Group Rickettsia-Infected Targets by Human Lymphokine-Activated Killer Cells

DTIC Science & Technology

1988-09-01

rick- Similar problems in detection of antigens of Theileria parva ettsia-specific cell surface antigens by performing polyacryl- (7) or influenza virus...infected with the protozoan parasite Theileria parva: workers in our laboratory are now in the process of cloning parasite strain specificity and class I

Recombinant poxviruses as mucosal vaccine vectors.

PubMed

Gherardi, M Magdalena; Esteban, Mariano

2005-11-01

The majority of infections initiate their departure from a mucosal surface, such as Human immunodeficiency virus (HIV), a sexually transmitted virus. Therefore, the induction of mucosal immunity is a high priority in the development of vaccines against mucosal pathogens. The selection of an appropriate antigen delivery system is necessary to induce an efficient mucosal immune response. Poxvirus vectors have been the most intensively studied live recombinant vector, and numerous studies have demonstrated their ability to induce mucosal immune responses against foreign expressed antigens. Previous studies have demonstrated that recombinants based on the attenuated modified vaccinia virus Ankara (MVA) vector were effective in inducing protective responses against different respiratory viruses, such as influenza and respiratory syncytial virus, following immunization via mucosal routes. Recent studies performed in the murine and macaque models have shown that recombinant MVA (rMVA) does not only stimulate HIV-specific immunity in the genital and rectal tracts following mucosal delivery, but can also control simian/human immunodeficiency viraemia and disease progression. In addition, a prime-boost vaccination approach against tuberculosis emphasized the importance of the intranasal rMVA antigen delivery to induce protective immunity against Mycobacterium tuberculosis. The aim of this review is to summarize the studies employing recombinant poxviruses, specifically rMVA as a mucosal delivery vector. The results demonstrate that rMVAs can activate specific immune responses at mucosal surfaces, and encourage further studies to characterize and improve the MVA mucosal immunogenicity of poxvirus vectors.

A Nano-Au/C-MWCNT based label free amperometric immunosensor for the detection of capsicum chlorosis virus in bell pepper.

PubMed

Sharma, Anshul; Kaushal, Ankur; Kulshrestha, Saurabh

2017-07-01

Accurate and on time diagnosis of plant viruses is an essential prerequisite for efficient control in field conditions. A number of diagnostic methods have been reported with the required level of sensitivity. Here, we propose a label free immunosensor for efficient and sensitive detection of capsicum chlorosis virus (CaCV) in bell pepper. Antigen was immobilized over the surface of gold nanoparticle/multi-walled carbon nanotube (Nano-Au/C-MWCNT) screen printed electrodes using 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC)/N-hydroxysuccinimide (NHS) cross linking chemistry followed by interaction with groundnut bud necrosis virus (GBNV)/CaCV specific polyclonal antibody. The electrochemical response was measured by cyclic voltammetry (CV), differential pulse voltammetry (DPV) using the redox indicator. Electrode surface characterization was done by performing scanning electron microscopy (SEM). Electrochemical studies showed positive results at different antigenic dilutions ranging from 10 -2 - 8x10 -5 . The sensitivity of the immunosensor developed has been compared with direct antigen coated enzyme-linked immunosorbent assay (DAC-ELISA) and the results showed that the immunosensor developed was 800-1000 times more sensitive, when compared to DAC-ELISA for CaCV detection. The immunosensor we have developed is economical and sensitive and could be used for immediate determination of the presence of virus in extracts from bell pepper leaves.

Effects of egg-adaptation on receptor-binding and antigenic properties of recent influenza A (H3N2) vaccine viruses.

PubMed

Parker, Lauren; Wharton, Stephen A; Martin, Stephen R; Cross, Karen; Lin, Yipu; Liu, Yan; Feizi, Ten; Daniels, Rodney S; McCauley, John W

2016-06-01

Influenza A virus (subtype H3N2) causes seasonal human influenza and is included as a component of influenza vaccines. The majority of vaccine viruses are isolated and propagated in eggs, which commonly results in amino acid substitutions in the haemagglutinin (HA) glycoprotein. These substitutions can affect virus receptor-binding and alter virus antigenicity, thereby, obfuscating the choice of egg-propagated viruses for development into candidate vaccine viruses. To evaluate the effects of egg-adaptive substitutions seen in H3N2 vaccine viruses on sialic acid receptor-binding, we carried out quantitative measurement of virus receptor-binding using surface biolayer interferometry with haemagglutination inhibition (HI) assays to correlate changes in receptor avidity with antigenic properties. Included in these studies was a panel of H3N2 viruses generated by reverse genetics containing substitutions seen in recent egg-propagated vaccine viruses and corresponding cell culture-propagated wild-type viruses. These assays provide a quantitative approach to investigating the importance of individual amino acid substitutions in influenza receptor-binding. Results show that viruses with egg-adaptive HA substitutions R156Q, S219Y, and I226N, have increased binding avidity to α2,3-linked receptor-analogues and decreased binding avidity to α2,6-linked receptor-analogues. No measurable binding was detected for the viruses with amino acid substitution combination 156Q+219Y and receptor-binding increased in viruses where egg-adaptation mutations were introduced into cell culture-propagated virus. Substitutions at positions 156 and 190 appeared to be primarily responsible for low reactivity in HI assays with post-infection ferret antisera raised against 2012-2013 season H3N2 viruses. Egg-adaptive substitutions at position 186 caused substantial differences in binding avidity with an insignificant effect on antigenicity.

Clinical and virological improvement of hepatitis B virus-related or hepatitis C virus-related chronic hepatitis with concomitant hepatitis A virus infection.

PubMed

Sagnelli, Evangelista; Coppola, Nicola; Pisaturo, Mariantonietta; Pisapia, Raffaella; Onofrio, Mirella; Sagnelli, Caterina; Catuogno, Antonio; Scolastico, Carlo; Piccinino, Felice; Filippini, Pietro

2006-06-01

We evaluated the clinical and virological characteristics of hepatitis A virus infection in persons concomitantly infected with hepatitis B virus (HBV) or hepatitis C virus (HCV). We enrolled 21 patients with acute hepatitis A and chronic hepatitis with no sign of liver cirrhosis, 13 patients who were positive for hepatitis B surface antigen (case B group), 8 patients who were anti-HCV positive (case C group), and 21 patients with acute hepatitis A without a preexisting liver disease (control A group). Two control groups of patients with chronic hepatitis B (control B group) or C (control C group) were also chosen. All control groups were pair-matched by age and sex with the corresponding case group. Fulminant hepatitis A was never observed, and hepatitis A had a severe course in 1 patient in the case B group and in 1 patient in the control A group. Both patients recovered. On admission, HBV DNA was detected in 1 patient in the case B group (7.7%) and in 13 patients (50%) in the control B group; HCV RNA was found in no patient in the case C group and in 16 patients (81.2%) in the control C group. Of 9 patients in the case B group who were followed up for 6 months, 3 became negative for hepatitis B surface antigen and positive for hepatitis B surface antibody, 2 remained positive for hepatitis B surface antigen and negative for HBV DNA, and 4 became positive for HBV DNA with a low viral load [corrected] Of 6 patients in the case C group who were followed up for 6 months, 3 remained negative for HCV RNA, and 3 had persistently low viral loads. Concomitant hepatitis A was always self-limited, associated with a marked inhibition of HBV and HCV genomes, and possibly had a good prognosis for the underlying chronic hepatitis.

A Modular Vaccine Development Platform Based on Sortase-Mediated Site-Specific Tagging of Antigens onto Virus-Like Particles

PubMed Central

Tang, Shubing; Xuan, Baoqin; Ye, Xiaohua; Huang, Zhong; Qian, Zhikang

2016-01-01

Virus-like particles (VLPs) can be used as powerful nanoscale weapons to fight against virus infection. In addition to direct use as vaccines, VLPs have been extensively exploited as platforms on which to display foreign antigens for prophylactic vaccination and immunotherapeutic treatment. Unfortunately, fabrication of new chimeric VLP vaccines in a versatile, site-specific and highly efficient manner is beyond the capability of traditional VLP vaccine design approaches, genetic insertion and chemical conjugation. In this study, we described a greatly improved VLP display strategy by chemoenzymatic site-specific tailoring antigens on VLPs surface with high efficiency. Through the transpeptidation mediated by sortase A, one protein and two epitopes containing N-terminal oligoglycine were conjugated to the LPET motif on the surface of hepatitis B virus core protein (HBc) VLPs with high density. All of the new chimeric VLPs induced strong specific IgG responses. Furthermore, the chimeric VLPs with sortase A tagged enterovirus 71 (EV71) SP70 epitope could elicit effective antibodies against EV71 lethal challenging as well as the genetic insertion chimeric VLPs. The sortase A mediated chemoenzymatic site-specific tailoring of the HBc VLP approach shows great potential in new VLP vaccine design for its simplicity, site specificity, high efficiency, and versatility. PMID:27170066

Tomato bushy stunt virus (TBSV), a versatile platform for polyvalent display of antigenic epitopes and vaccine design

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kumar, Shantanu; Ochoa, Wendy; Singh, Pratik

2009-05-25

Viruses-like particles (VLPs) are frequently being used as platforms for polyvalent display of foreign epitopes of interest on their capsid surface to improve their presentation enhancing the antigenicity and host immune response. In the present study, we used the VLPs of Tomato bushy stunt virus (TBSV), an icosahedral plant virus, as a platform to display 180 copies of 16 amino acid epitopes of ricin toxin fused to the C-terminal end of a modified TBSV capsid protein (NDELTA52). Expression of the chimeric recombinant protein in insect cells resulted in spontaneous assembly of VLPs displaying the ricin epitope. Cryo-electron microscopy and imagemore » reconstruction of the chimeric VLPs at 22 A resolution revealed the locations and orientation of the ricin epitope exposed on the TBSV capsid surface. Furthermore, injection of chimeric VLPs into mice generated antisera that detected the native ricin toxin. The ease of fusing of short peptides of 15-20 residues and their ability to form two kinds (T = 1, T = 3) of bio-nanoparticles that result in the display of 60 or 180 copies of less constrained and highly exposed antigenic epitopes makes TBSV an attractive and versatile display platform for vaccine design.« less

Comprehensive Analysis and Characterization of Linear Antigenic Domains on HN Protein from Genotype VII Newcastle Disease Virus Using Yeast Surface Display System

PubMed Central

Li, Tao; Wang, Gaoling; Shi, Bingtian; Liu, Peixin; Si, Wei; Wang, Bin; Jiang, Li; Zhou, Lunjiang; Xiu, Jinsheng; Liu, Henggui

2015-01-01

Circulation of genotype VII Newcastle disease virus (NDV) has posed a great threat for the poultry industry worldwide. Antibodies against Hemagglutinin-neuraminidase (HN), a membrane protein of NDV with critical roles in NDV infection, have been reported to provide chickens protection from NDV infection. In this study, we comprehensively analyzed the in vivo antibody responses against the linear antigenic domains of the HN protein from genotype VII NDV using a yeast surface display system. The results revealed four distinct regions of HN, P1 (1-52aa), P2 (53-192aa), P3 (193-302aa) and P4 (303-571aa), respectively, according to their antigenic potency. Analysis by FACS and ELISA assay indicated P2 to be the dominant linear antigenic domain, with the immunogenic potency to protect the majority of chickens from NDV challenge. In contrast, the P1, P3 and P4 domains showed weak antigenicity in vivo and could not protect chickens from NDV challenge. These results provide important insight into the characteristic of humoral immune responses elicited by HN of NDV in vivo. PMID:26121247

The rescue and evaluation of FLAG and HIS epitope-tagged Asia 1 type foot-and-mouth disease viruses.

PubMed

Yang, Bo; Yang, Fan; Zhang, Yan; Liu, Huanan; Jin, Ye; Cao, Weijun; Zhu, Zixiang; Zheng, Haixue; Yin, Hong

2016-02-02

The VP1 G-H loop of the foot-and-mouth disease virus (FMDV) contains the primary antigenic site, as well as an Arg-Gly-Asp (RGD) binding motif for the αv-integrin family of cell surface receptors. We anticipated that introducing a foreign epitope tag sequence downstream of the RGD motif would be tolerated by the viral capsid and would not destroy the antigenic site of FMDV. In this study, we have designed, generated, and characterized two recombinant FMDVs with a FLAG tag or histidine (HIS) inserted in the VP1 G-H loop downstream of the RGD motif +9 position. The tagged viruses were genetically stable and exhibited similar growth properties with their parental virus. What is more, the recombinant viruses rFMDV-FLAG and rFMDV-HIS showed neutralization sensitivity to FMDV type Asia1-specific mAbs, as well as to polyclonal antibodies. Additionally, the r1 values of the recombinant viruses were similar to that of the parental virus, indicating that the insertion of FLAG or HIS tag sequences downstream of the RGD motif +9 position do not eradicate the antigenic site of FMDV and do not affect its antigenicity. These results indicated that the G-H loop of Asia1 FMDV is able to effectively display the foreign epitopes, making this a potential approach for novel FMDV vaccines development. Copyright © 2015 Elsevier B.V. All rights reserved.

Influence of occult hepatitis B virus infection in chronic hepatitis C outcomes

PubMed Central

Fernandez-Rodriguez, Conrado M; Gutierrez, Maria Luisa; Lledó, José Luis; Casas, Maria Luisa

2011-01-01

Persistence of hepatitis B virus-DNA in the sera, peripheral blood mononuclear cells or in the liver of hepatitis B surface antigen (HBsAg)-negative patients with or without serological markers of previous exposure (antibodies to HBsAg and/or to HB-core antigen) defines the entity called occult hepatitis B infection (OBI). Co-infection with hepatitis B and hepatitis C viruses is frequent in highly endemic areas. While this co-infection increases the risk of liver disease progression, development of cirrhosis and hepatocellular carcinoma and also increases the rate of therapeutic failure to interferon-based treatments than either virus alone, a potentially negative effect of OBI on clinical outcomes and of therapeutic response to current antiviral regimes of patients with chronic hepatitis C remains inconclusive. PMID:21472121

Human immunodeficiency virus (HIV) seropositivity and hepatitis B surface antigenemia (HBSAG) among blood donors in Benin city, Edo state, Nigeria.

PubMed

Umolu, Patience Idia; Okoror, Lawrence Ehis; Orhue, Philip

2005-03-01

Human Immunodeficiency Virus and Hepatitis B virus are blood borne pathogens that can be transmitted through blood transfusion and could pose a huge problem in areas where mechanisms of ensuring blood safety are suspect. This study became necessary in a population where most of the blood for transfusion is from commercial blood donors. A total of 130 donors comprising 120 commercial donors and 10 voluntary donors were tested for antibodies to human immunodeficiency virus and hepatitis B surface antigen in Benin city using Immunocomb HIV - 1 and 2 Biospot kit and Quimica Clinica Aplicada direct latex agglutination method respectively. Thirteen (10%) samples were HIV seropositive and 7(5.8%) were HBsAg positive. The age bracket 18 - 25years had the highest numbers of donors and also had the highest number of HBsAg positive cases (7.8%) while the age group 29 - 38years had highest number of HIV seropositive cases. High prevalence of HIV antibodies and Hepatitis B surface antigen was found among commercial blood donors. Appropriate and compulsory screening of blood donors using sensitive methods, must be ensured to prevent post transfusion hepatitis and HIV.

Adsorption of virus-like particles on ion exchange surface: Conformational changes at different pH detected by dual polarization interferometry.

PubMed

Yang, Yanli; Mengran Yu; Zhang, Songping; Ma, Guanghui; Su, Zhiguo

2015-08-21

Disassembling of virus-like particles (VLPs) like hepatitis B virus surface antigen (HB-VLPs) during chromatographic process has been identified as a major cause of loss of antigen activity. In this study, dual polarization interferometry (DPI) measurement, together with chromatography experiments, were performed to study the adsorption and conformational change of HB-VLPs on ion exchange surface at three different pHs. Changes in pH values of buffer solution showed only minimal effect on the HB-VLPs assembly and antigen activity, while significantly different degree of HB-VLPs disassembling was observed after ion exchange chromatography (IEC) at different pHs, indicating the conformational change of HB-VLPs caused mainly by its interactions with the adsorbent surface. By creating an ion exchange surface on chip surface, the conformational changes of HB-VLPs during adsorption to the surface were monitored in real time by DPI for the first time. As pH increased from 7.0 to 9.0, strong electrostatic interactions between oppositely charged HB-VLPs and the ion exchange surface make the HB-VLPs spread thinly or even adsorbed in disassembled formation on the surface as revealed by significant decrease in thickness of the adsorbed layer measured by DPI. Such findings were consistent with the results of IEC experiments operated at different pHs, that more disassembled HB-VLPs were detected in the eluted proteins at pH 9.0. At low pH like pH 5.0, however, possible bi-layer adsorption was involved as evidenced by an adsorbed layer thickness higher than average diameter of the HB-VLPs. The "lateral" protein-protein interactions might be unfavorable and would make additional contribution to the disassembling of HB-VLPs besides the primary mechanism related to the protein-surface interactions; therefore, the lowest antigen activity was observed after IEC at pH 5.0. Such real-time information on conformational change of VLPs is helpful for better understanding the real mechanism for the disassembling of VLPs on the solid-liquid interface. Copyright © 2015 Elsevier B.V. All rights reserved.

Macromolecular Assemblage in the Design of a Synthetic AIDS Vaccine

NASA Astrophysics Data System (ADS)

Defoort, Jean-Philippe; Nardelli, Bernardetta; Huang, Wolin; Ho, David D.; Tam, James P.

1992-05-01

We describe a peptide vaccine model based on the mimicry of surface coat protein of a pathogen. This model used a macromolecular assemblage approach to amplify peptide antigens in liposomes or micelles. The key components of the model consisted of an oligomeric lysine scaffolding to amplify peptide antigens covalently 4-fold and a lipophilic membrane-anchoring group to further amplify noncovalently the antigens many-fold in liposomal or micellar form. A peptide antigen derived from the third variable domain of glycoprotein gp120 of human immunodeficiency virus type 1 (HIV-1), consisting of neutralizing, T-helper, and T-cytotoxic epitopes, was used in a macromolecular assemblage model (HIV-1 linear peptide amino acid sequence 308-331 in a tetravalent multiple antigen peptide system linked to tripalmitoyl-S-glycerylcysteine). The latter complex, in liposome or micelle, was used to immunize mice and guinea pigs without any adjuvant and found to induce gp120-specific antibodies that neutralize virus infectivity in vitro, elicit cytokine production, and prime CD8^+ cytotoxic T lymphocytes in vivo. Our results show that the macromolecular assemblage approach bears immunological mimicry of the gp120 of HIV virus and may lead to useful vaccines against HIV infection.

[Molecular characterization of human influenza viruses--a look back on the last 10 years].

PubMed

Schweiger, Brunhilde

2006-01-01

Influenza A (H3N2) viruses and influenza B viruses have caused more than 90% of influenza infections in Germany during the last then years. Continuous and extensive antigenic variation was evident for both the hemagglutinin (HA) and neuraminidase (NA) surface proteins of H3N2 and influenza B viruses. Molecular characterisation revealed an ongoing genetic drift even in years when the antigenic profiles of circulating strains were indistinguishable from those of the previous season. Retrospective phylogenetic studies showed that viruses similar to vaccine strains circulated one or two years before a given strain was recommended as vaccine strain. New drift variants of H3N2 viruses with significantly changed antigenic features appeared during the seasons 1997/1998 and 2002/2003. Most influenza seasons were characterised by a co-circulation of at least two different lineages of H3N2 viruses. Genetic reassortment between H3N2 viruses belonging to separate lineages caused the different evolutionary pathways of the HA and viruses was responsible for the occurrence of H1N2 viruses during the season 2001/02. This new subtype has been detected only sporadically in Germany. The evolution of influenza B viruses was characterised by the re-emergence of B/Victoria/2/87-lineage viruses and their co-circulation with viruses of the B/Yamagata/16/88-lineage. Reassortant B viruses possessing a Victoria/87-lineage HA and a Yamagata/88-like NA were predominant in Germany during 2002/03 and 2004/05.

USSR Report, Life Sciences, Biomedical and Behavioral Sciences

DTIC Science & Technology

1987-01-20

Varnek, P. D. Brezhestovskiy; DOKLADY AKADEMII NAUK SSSR, No 6, Aug 86) 13 Effects of Selenomethionine on Proton Magnetic Relaxation in Hepatic ...Virus Expressing Surface Antigen of B Hepatitis Virus and Thymidinekinase of Herpes Simplex Virus (A. D. Altshteyn, 0. G. Andzhaparidze, et al...Influence of Dimetpramide and Metoclopramide on Catecholamine Turnover Rates in Rat Brain Subcortical-Brainstem Structures (V. I. Legeza, M. F. Kamynina

Antigenic Variation of Clade 2.1 H5N1 Virus Is Determined by a Few Amino Acid Substitutions Immediately Adjacent to the Receptor Binding Site

PubMed Central

Koel, Björn F.; van der Vliet, Stefan; Burke, David F.; Bestebroer, Theo M.; Bharoto, Eny E.; Yasa, I. Wayan W.; Herliana, Inna; Laksono, Brigitta M.; Xu, Kemin; Skepner, Eugene; Russell, Colin A.; Rimmelzwaan, Guus F.; Perez, Daniel R.; Osterhaus, Albert D. M. E.; Smith, Derek J.; Prajitno, Teguh Y.

2014-01-01

ABSTRACT Highly pathogenic avian influenza (HPAI) viruses of the H5N1 subtype are genetically highly variable and have diversified into multiple phylogenetic clades over the past decade. Antigenic drift is a well-studied phenomenon for seasonal human influenza viruses, but much less is known about the antigenic evolution of HPAI H5N1 viruses that circulate in poultry. In this study, we focused on HPAI H5N1 viruses that are enzootic to Indonesia. We selected representative viruses from genetically distinct lineages that are currently circulating and determined their antigenic properties by hemagglutination inhibition assays. At least six antigenic variants have circulated between 2003, when H5N1 clade 2.1 viruses were first detected in Indonesia, and 2011. During this period, multiple antigenic variants cocirculated in the same geographic regions. Mutant viruses were constructed by site-directed mutagenesis to represent each of the circulating antigenic variants, revealing that antigenic differences between clade 2.1 viruses were due to only one or very few amino acid substitutions immediately adjacent to the receptor binding site. Antigenic variants of H5N1 virus evaded recognition by both ferret and chicken antibodies. The molecular basis for antigenic change in clade 2.1 viruses closely resembled that of seasonal human influenza viruses, indicating that the hemagglutinin of influenza viruses from different hosts and subtypes may be similarly restricted to evade antibody recognition. PMID:24917596

Silent killers: Transfusion Transmissible Infections-TTI, among asymptomatic population of Pakistan.

PubMed

Saeed, Muhammad; Hussain, Shahida; Rasheed, Farhan; Ahmad, Maqsood; Arif, Mizna; Hamid Rahmani, Muhammad Tariq

2017-03-01

To analyse transfusion transmissible infections in asymptomatic population. This study was conducted at the Allama Iqbal Medical College and Jinnah Hospital, Lahore, Pakistan, from December 2014 to November 2015, and comprised healthy asymptomatic blood donors.Every sample was screened for the presence of antibodies/antigens of hepatitis C virus, human immunodeficiency virus, treponemapallidum, hepatitis B virus and malaria parasite through rapid immunochromatographic technique. Of the 18,274 blood donors, 17,276(94.53%) were found healthy and 998(5.46%) were infected. Besides, 71(0.38%) had multiple infections. The overall frequency of anti-hepatitis C virus, treponemapallidum (syphilis), hepatitis B surface antigen, malaria parasite and anti-human immunodeficiency virus was 480(2.62%), 284(1.55%), 210(1.10%), 20(0.10%) and 4(0.02%), respectively. Blood transfusion was found to be a significant but preventable mode of spread of transfusion transmissible infections.

[The experimental evaluation of the possibility of the penetration of enteric viruses from the surface into the pulp of contaminated fruits and vegetables].

PubMed

Sergevnin, V I; Ladeyshchikova, Yu I; Sarmometov, E V; Podgorunskaya, I L; Kudrevatykh, E V

2014-01-01

According to the results of complex microbiological examination of samples of vegetables, fruits and grapes there was established significant contamination of them with opportunistic bacteria, antigens of intestinal viruses and cysts of intestinal Protozoa, that confirms the epidemiological role of these products as factors in transmission of acute intestinal infections. There was revealed ribonucleic acid of enteric viruses in experimentally infected pulp from the surface of tomatoes and apples, that indicates to the possibility of penetration of these pathogens into the fruits and vegetables through intact (having no visible damages) surface.

Complex antigen presentation pathway for an HLA-A*0201-restricted epitope from Chikungunya 6K protein

PubMed Central

Lemonnier, François A.; Esteban, Mariano

2017-01-01

Background The adaptive cytotoxic T lymphocyte (CTL)-mediated immune response is critical for clearance of many viral infections. These CTL recognize naturally processed short viral antigenic peptides bound to human leukocyte antigen (HLA) class I molecules on the surface of infected cells. This specific recognition allows the killing of virus-infected cells. The T cell immune T cell response to Chikungunya virus (CHIKV), a mosquito-borne Alphavirus of the Togaviridae family responsible for severe musculoskeletal disorders, has not been fully defined; nonetheless, the importance of HLA class I-restricted immune response in this virus has been hypothesized. Methodology/Principal findings By infection of HLA-A*0201-transgenic mice with a recombinant vaccinia virus that encodes the CHIKV structural polyprotein (rVACV-CHIKV), we identified the first human T cell epitopes from CHIKV. These three novel 6K transmembrane protein-derived epitopes are presented by the common HLA class I molecule, HLA-A*0201. One of these epitopes is processed and presented via a complex pathway that involves proteases from different subcellular locations. Specific chemical inhibitors blocked these events in rVACV-CHIKV-infected cells. Conclusions/Significance Our data have implications not only for the identification of novel Alphavirus and Togaviridae antiviral CTL responses, but also for analyzing presentation of antigen from viruses of different families and orders that use host proteinases to generate their mature envelope proteins. PMID:29084215

Complex antigen presentation pathway for an HLA-A*0201-restricted epitope from Chikungunya 6K protein.

PubMed

Lorente, Elena; Barriga, Alejandro; García-Arriaza, Juan; Lemonnier, François A; Esteban, Mariano; López, Daniel

2017-10-01

The adaptive cytotoxic T lymphocyte (CTL)-mediated immune response is critical for clearance of many viral infections. These CTL recognize naturally processed short viral antigenic peptides bound to human leukocyte antigen (HLA) class I molecules on the surface of infected cells. This specific recognition allows the killing of virus-infected cells. The T cell immune T cell response to Chikungunya virus (CHIKV), a mosquito-borne Alphavirus of the Togaviridae family responsible for severe musculoskeletal disorders, has not been fully defined; nonetheless, the importance of HLA class I-restricted immune response in this virus has been hypothesized. By infection of HLA-A*0201-transgenic mice with a recombinant vaccinia virus that encodes the CHIKV structural polyprotein (rVACV-CHIKV), we identified the first human T cell epitopes from CHIKV. These three novel 6K transmembrane protein-derived epitopes are presented by the common HLA class I molecule, HLA-A*0201. One of these epitopes is processed and presented via a complex pathway that involves proteases from different subcellular locations. Specific chemical inhibitors blocked these events in rVACV-CHIKV-infected cells. Our data have implications not only for the identification of novel Alphavirus and Togaviridae antiviral CTL responses, but also for analyzing presentation of antigen from viruses of different families and orders that use host proteinases to generate their mature envelope proteins.

Broadly neutralizing antibodies from human survivors target a conserved site in the Ebola virus glycoprotein HR2-MPER region.

PubMed

Flyak, Andrew I; Kuzmina, Natalia; Murin, Charles D; Bryan, Christopher; Davidson, Edgar; Gilchuk, Pavlo; Gulka, Christopher P; Ilinykh, Philipp A; Shen, Xiaoli; Huang, Kai; Ramanathan, Palaniappan; Turner, Hannah; Fusco, Marnie L; Lampley, Rebecca; Kose, Nurgun; King, Hannah; Sapparapu, Gopal; Doranz, Benjamin J; Ksiazek, Thomas G; Wright, David W; Saphire, Erica Ollmann; Ward, Andrew B; Bukreyev, Alexander; Crowe, James E

2018-05-07

Ebola virus (EBOV) in humans causes a severe illness with high mortality rates. Several strategies have been developed in the past to treat EBOV infection, including the antibody cocktail ZMapp, which has been shown to be effective in nonhuman primate models of infection 1 and has been used under compassionate-treatment protocols in humans 2 . ZMapp is a mixture of three chimerized murine monoclonal antibodies (mAbs) 3-6 that target EBOV-specific epitopes on the surface glycoprotein 7,8 . However, ZMapp mAbs do not neutralize other species from the genus Ebolavirus, such as Bundibugyo virus (BDBV), Reston virus (RESTV) or Sudan virus (SUDV). Here, we describe three naturally occurring human cross-neutralizing mAbs, from BDBV survivors, that target an antigenic site in the canonical heptad repeat 2 (HR2) region near the membrane-proximal external region (MPER) of the glycoprotein. The identification of a conserved neutralizing antigenic site in the glycoprotein suggests that these mAbs could be used to design universal antibody therapeutics against diverse ebolavirus species. Furthermore, we found that immunization with a peptide comprising the HR2-MPER antigenic site elicits neutralizing antibodies in rabbits. Structural features determined by conserved residues in the antigenic site described here could inform an epitope-based vaccine design against infection caused by diverse ebolavirus species.

Detection of viral antigens in renal tissue of glomerulonephritis patients without serological evidence of hepatitis B virus and hepatitis C virus infection.

PubMed

Kong, Dan; Wu, Di; Wang, Tianzhen; Li, Tianzhu; Xu, Shengjie; Chen, Fulai; Jin, Xiaoming; Lou, Ge

2013-07-01

Glomerulonephritis is an important extrahepatic manifestation of hepatitis B virus (HBV) and hepatitis C virus (HCV) infection. HBV and HCV infection may be occult, and they are often overlooked by both patients and doctors. The aim of this study was to assess the importance of HBV and HCV infection in glomerulonephritis patients with undetectable HBV surface antigen (HBsAg) and HCV antibody in serum. The HBsAg, the HBV core antigen (HBcAg), and the HCV antigen were detected using immunohistochemistry in frozen renal tissues of 500 glomerulonephritis patients without serological evidence of HBV and HCV infection. Electron microscopy was used to trace the virus particles, and clinicopathological features were also reviewed. HBsAg or HBcAg was positive in nine out of 500 cases (9/500, 1.8%). Three cases were HBsAg-positive and another six cases were HBcAg-positive. The HCV antigen was found in eight cases (8/500, 1.6%). There was one case of HBV and HCV co-infection (1/500, 0.2%). Under electron microscopy, virus particles were found in the base membrane and cytoplasm of endotheliocytes in the glomerulus. The most common clinical manifestation was nephrotic syndrome (9/18), followed by nephritic syndrome (7/18). Membranous nephropathy was the most common pathological diagnosis (5/18), followed by mesangioproliferative glomerulonephritis (4/18) and IgA nephropathy (4/18). Occult HBV and HCV infection might be implicated in HBV- or HCV-associated glomerulonephritis. More attention should be focused on the underlying cause. Copyright © 2013 International Society for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Differences in antigen presentation to MHC class I-and class II- restricted influenza virus-specific cytolytic T lymphocyte clones

PubMed Central

1986-01-01

We have examined requirements for antigen presentation to a panel of MHC class I-and class II-restricted, influenza virus-specific CTL clones by controlling the form of virus presented on the target cell surface. Both H-2K/D- and I region-restricted CTL recognize target cells exposed to infectious virus, but only the I region-restricted clones efficiently lysed histocompatible target cells pulsed with inactivated virus preparations. The isolated influenza hemagglutinin (HA) polypeptide also could sensitize target cells for recognition by class II-restricted, HA-specific CTL, but not by class I-restricted, HA- specific CTL. Inhibition of nascent viral protein synthesis abrogated the ability of target cells to present viral antigen relevant for class I-restricted CTL recognition. Significantly, presentation for class II- restricted recognition was unaffected in target cells exposed to preparations of either inactivated or infectious virus. This differential sensitivity suggested that these H-2I region-restricted CTL recognized viral polypeptides derived from the exogenously introduced virions, rather than viral polypeptides newly synthesized in the infected cell. In support of this contention, treatment of the target cells with the lysosomotropic agent chloroquine abolished recognition of infected target cells by class II-restricted CTL without diminishing class I-restricted recognition of infected target cells. Furthermore, when the influenza HA gene was introduced into target cells without exogenous HA polypeptide, the target cells that expressed the newly synthesized protein product of the HA gene were recognized only by H-2K/D-restricted CTL. These observations suggest that important differences may exist in requirements for antigen presentation between H-2K/D and H-2I region-restricted CTL. These differences may reflect the nature of the antigenic epitopes recognized by these two CTL subsets. PMID:3485173

Hepatitis B virus surface antigen and anti-hepatitis C virus rapid tests underestimate hepatitis prevalence among HIV-infected patients.

PubMed

Hønge, Bl; Jespersen, S; Medina, C; Té, Ds; da Silva, Zj; Ostergaard, L; Laursen, Al; Wejse, C; Krarup, H; Erikstrup, C

2014-10-01

In the case of coinfection with HIV and hepatitis B virus (HBV) and/or hepatitis C virus (HCV), hepatic disease progression is often accelerated, with higher rates of liver cirrhosis and liver-related mortality. We aimed to evaluate the performance of the rapid tests used routinely to detect HBV surface antigen (HBsAg) and anti-HCV among HIV-infected patients in Guinea-Bissau. Blood samples from HIV-infected patients in Guinea-Bissau were stored after testing for HBsAg and anti-HCV with rapid tests. Samples were subsequently re-tested for HBsAg and anti-HCV in Denmark. Two rapid tests were used in Guinea-Bissau: HBsAg Strip Ref 2034 (VEDA.LAB, Alençon, France; sensitivity 62.3%; specificity 99.2%) and HEPA-SCAN (Bhat Bio-Tech, Bangalore, India; sensitivity 57.1%; specificity 99.7%). In the two tests the ability to obtain the correct outcome depended on the antigen and antibody concentrations, respectively. Sex, age, CD4 cell count and antiretroviral therapy status did not differ between false negative and true positive samples in either of the tests. The study is limited by a low number of anti-HCV positive samples. New diagnostic rapid tests should always be evaluated in the setting in which they will be used before implementation. © 2014 British HIV Association.

Evaluating the Use of Commercial West Nile Virus Antigens as Positive Controls in the Rapid Analyte Measurement Platform West Nile Virus Assay.

PubMed

Burkhalter, Kristen L; Savage, Harry M

2015-12-01

We evaluated the utility of 2 types of commercially available antigens as positive controls in the Rapid Analyte Measurement Platform (RAMP®) West Nile virus (WNV) assay. Purified recombinant WNV envelope antigens and whole killed virus antigens produced positive RAMP results and either type would be useful as a positive control. Killed virus antigens provide operational and economic advantages and we recommend their use over purified recombinant antigens. We also offer practical applications for RAMP positive controls and recommendations for preparing them.

Genetic variation of occult hepatitis B virus infection

PubMed Central

Zhu, Hui-Lan; Li, Xu; Li, Jun; Zhang, Zhen-Hua

2016-01-01

Occult hepatitis B virus infection (OBI), characterized as the persistence of hepatitis B virus (HBV) surface antigen (HBsAg) seronegativity and low viral load in blood or liver, is a special form of HBV infection. OBI may be related mainly to mutations in the HBV genome, although the underlying mechanism of it remains to be clarified. Mutations especially within the immunodominant “α” determinant of S protein are “hot spots” that could contribute to the occurrence of OBI via affecting antigenicity and immunogenicity of HBsAg or replication and secretion of virion. Clinical reports account for a large proportion of previous studies on OBI, while functional analyses, especially those based on full-length HBV genome, are rare. PMID:27053845

Structure, Receptor Binding, and Antigenicity of Influenza Virus Hemagglutinins from the 1957 H2N2 Pandemic

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Rui; McBride, Ryan; Paulson, James C.

2010-03-04

The hemagglutinin (HA) envelope protein of influenza viruses mediates essential viral functions, including receptor binding and membrane fusion, and is the major viral antigen for antibody neutralization. The 1957 H2N2 subtype (Asian flu) was one of the three great influenza pandemics of the last century and caused 1 million deaths globally from 1957 to 1968. Three crystal structures of 1957 H2 HAs have been determined at 1.60 to 1.75 {angstrom} resolutions to investigate the structural basis for their antigenicity and evolution from avian to human binding specificity that contributed to its introduction into the human population. These structures, which representmore » the highest resolutions yet recorded for a complete ectodomain of a glycosylated viral surface antigen, along with the results of glycan microarray binding analysis, suggest that a hydrophobicity switch at residue 226 and elongation of receptor-binding sites were both critical for avian H2 HA to acquire human receptor specificity. H2 influenza viruses continue to circulate in birds and pigs and, therefore, remain a substantial threat for transmission to humans. The H2 HA structure also reveals a highly conserved epitope that could be harnessed in the design of a broader and more universal influenza A virus vaccine.« less

Induction of neutralising antibodies by virus-like particles harbouring surface proteins from highly pathogenic H5N1 and H7N1 influenza viruses

PubMed Central

Szécsi, Judit; Boson, Bertrand; Johnsson, Per; Dupeyrot-Lacas, Pia; Matrosovich, Mikhail; Klenk, Hans-Dieter; Klatzmann, David; Volchkov, Viktor; Cosset, François-Loïc

2006-01-01

There is an urgent need to develop novel approaches to vaccination against the emerging, highly pathogenic avian influenza viruses. Here, we engineered influenza viral-like particles (Flu-VLPs) derived from retroviral core particles that mimic the properties of the viral surface of two highly pathogenic influenza viruses of either H7N1 or H5N1 antigenic subtype. We demonstrate that, upon recovery of viral RNAs from a field strain, one can easily generate expression vectors that encode the HA, NA and M2 surface proteins of either virus and prepare high-titre Flu-VLPs. We characterise these Flu-VLPs incorporating the HA, NA and M2 proteins and we show that they induce high-titre neutralising antibodies in mice. PMID:16948862

Influenza Virus PB1 and Neuraminidase Gene Segments Can Cosegregate during Vaccine Reassortment Driven by Interactions in the PB1 Coding Region

PubMed Central

Cobbin, Joanna C. A.; Ong, Chi; Verity, Erin; Gilbertson, Brad P.; Rockman, Steven P.

2014-01-01

ABSTRACT Egg-grown influenza vaccine yields are maximized by infection with a seed virus produced by “classical reassortment” of a seasonal isolate with a highly egg-adapted strain. Seed viruses are selected based on a high-growth phenotype and the presence of the seasonal hemagglutinin (HA) and neuraminidase (NA) surface antigens. Retrospective analysis of H3N2 vaccine seed viruses indicated that, unlike other internal proteins that were predominantly derived from the high-growth parent A/Puerto Rico/8/34 (PR8), the polymerase subunit PB1 could be derived from either parent depending on the seasonal strain. We have recently shown that A/Udorn/307/72 (Udorn) models a seasonal isolate that yields reassortants bearing the seasonal PB1 gene. This is despite the fact that the reverse genetics-derived virus that includes Udorn PB1 with Udorn HA and NA on a PR8 background has inferior growth compared to the corresponding virus with PR8 PB1. Here we use competitive plasmid transfections to investigate the mechanisms driving selection of a less fit virus and show that the Udorn PB1 gene segment cosegregates with the Udorn NA gene segment. Analysis of chimeric PB1 genes revealed that the coselection of NA and PB1 segments was not directed through the previously identified packaging sequences but through interactions involving the internal coding region of the PB1 gene. This study identifies associations between viral genes that can direct selection in classical reassortment for vaccine production and which may also be of relevance to the gene constellations observed in past antigenic shift events where creation of a pandemic virus has involved reassortment. IMPORTANCE Influenza vaccine must be produced and administered in a timely manner in order to provide protection during the winter season, and poor-growing vaccine seed viruses can compromise this process. To maximize vaccine yields, manufacturers create hybrid influenza viruses with gene segments encoding the surface antigens from a seasonal virus isolate, important for immunity, and others from a virus with high growth properties. This involves coinfection of cells with both parent viruses and selection of dominant progeny bearing the seasonal antigens. We show that this method of creating hybrid viruses does not necessarily select for the best yielding virus because preferential pairing of gene segments when progeny viruses are produced determines the genetic makeup of the hybrids. This not only has implications for how hybrid viruses are selected for vaccine production but also sheds light on what drives and limits hybrid gene combinations that arise in nature, leading to pandemics. PMID:24872588

Influenza virus PB1 and neuraminidase gene segments can cosegregate during vaccine reassortment driven by interactions in the PB1 coding region.

PubMed

Cobbin, Joanna C A; Ong, Chi; Verity, Erin; Gilbertson, Brad P; Rockman, Steven P; Brown, Lorena E

2014-08-01

Egg-grown influenza vaccine yields are maximized by infection with a seed virus produced by "classical reassortment" of a seasonal isolate with a highly egg-adapted strain. Seed viruses are selected based on a high-growth phenotype and the presence of the seasonal hemagglutinin (HA) and neuraminidase (NA) surface antigens. Retrospective analysis of H3N2 vaccine seed viruses indicated that, unlike other internal proteins that were predominantly derived from the high-growth parent A/Puerto Rico/8/34 (PR8), the polymerase subunit PB1 could be derived from either parent depending on the seasonal strain. We have recently shown that A/Udorn/307/72 (Udorn) models a seasonal isolate that yields reassortants bearing the seasonal PB1 gene. This is despite the fact that the reverse genetics-derived virus that includes Udorn PB1 with Udorn HA and NA on a PR8 background has inferior growth compared to the corresponding virus with PR8 PB1. Here we use competitive plasmid transfections to investigate the mechanisms driving selection of a less fit virus and show that the Udorn PB1 gene segment cosegregates with the Udorn NA gene segment. Analysis of chimeric PB1 genes revealed that the coselection of NA and PB1 segments was not directed through the previously identified packaging sequences but through interactions involving the internal coding region of the PB1 gene. This study identifies associations between viral genes that can direct selection in classical reassortment for vaccine production and which may also be of relevance to the gene constellations observed in past antigenic shift events where creation of a pandemic virus has involved reassortment. Influenza vaccine must be produced and administered in a timely manner in order to provide protection during the winter season, and poor-growing vaccine seed viruses can compromise this process. To maximize vaccine yields, manufacturers create hybrid influenza viruses with gene segments encoding the surface antigens from a seasonal virus isolate, important for immunity, and others from a virus with high growth properties. This involves coinfection of cells with both parent viruses and selection of dominant progeny bearing the seasonal antigens. We show that this method of creating hybrid viruses does not necessarily select for the best yielding virus because preferential pairing of gene segments when progeny viruses are produced determines the genetic makeup of the hybrids. This not only has implications for how hybrid viruses are selected for vaccine production but also sheds light on what drives and limits hybrid gene combinations that arise in nature, leading to pandemics. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Rapid screening and identification of dominant B cell epitopes of HBV surface antigen by quantum dot-based fluorescence polarization assay

NASA Astrophysics Data System (ADS)

Meng, Zhongji; Song, Ruihua; Chen, Yue; Zhu, Yang; Tian, Yanhui; Li, Ding; Cui, Daxiang

2013-03-01

A method for quickly screening and identifying dominant B cell epitopes was developed using hepatitis B virus (HBV) surface antigen as a target. Eleven amino acid fragments from HBV surface antigen were synthesized by 9-fluorenylmethoxy carbonyl solid-phase peptide synthesis strategy, and then CdTe quantum dots were used to label the N-terminals of all peptides. After optimizing the factors for fluorescence polarization (FP) immunoassay, the antigenicities of synthetic peptides were determined by analyzing the recognition and combination of peptides and standard antibody samples. The results of FP assays confirmed that 10 of 11 synthetic peptides have distinct antigenicities. In order to screen dominant antigenic peptides, the FP assays were carried out to investigate the antibodies against the 10 synthetic peptides of HBV surface antigen respectively in 159 samples of anti-HBV surface antigen-positive antiserum. The results showed that 3 of the 10 antigenic peptides may be immunodominant because the antibodies against them existed more widely among the samples and their antibody titers were higher than those of other peptides. Using three dominant antigenic peptides, 293 serum samples were detected for HBV infection by FP assays; the results showed that the antibody-positive ratio was 51.9% and the sensitivity and specificity were 84.3% and 98.2%, respectively. In conclusion, a quantum dot-based FP assay is a very simple, rapid, and convenient method for determining immunodominant antigenic peptides and has great potential in applications such as epitope mapping, vaccine designing, or clinical disease diagnosis in the future.

The Dynamics of a SEIR-SIRC Antigenic Drift Influenza Model.

PubMed

Adi-Kusumo, Fajar

2017-06-01

We consider the dynamics of an influenza model with antigenic drift mechanism. Antigenic drift is an antigen mutation on the skin surface of the influenza virus that do not produce a new virus strain. The mutation produces the same virus but with slightly different antigens that cannot be recognized by the immune receptors formed by the previous infection. There are some type of influenza that involve the interaction between two populations such as human and animal. In this paper, we construct an influenza model with antigenic drift mechanism on the human population that has interaction with the animal population. The animal population is assumed to follow the SEIR epidemic model. Our model is motivated by the fact that some of the influenza cases in human come from the animal such as the swine and the avian. The transmission parameter that shows number of contact between the susceptible human and the infectious animals are important to study. The parameter plays an important role to detect the cycle of infection of the disease. The other important parameters are the seasonality degree, which shows the pathogen appearance and disappearance via annual migration, and the infection rate on the human population. We employ the bifurcation theory to analyze the behavior of the system and to detect the cycle of infection types when the parameters values are varied.

Antigenic Drift in H5N1 Avian Influenza Virus in Poultry Is Driven by Mutations in Major Antigenic Sites of the Hemagglutinin Molecule Analogous to Those for Human Influenza Virus▿†

PubMed Central

Cattoli, Giovanni; Milani, Adelaide; Temperton, Nigel; Zecchin, Bianca; Buratin, Alessandra; Molesti, Eleonora; Aly, Mona Meherez; Arafa, Abdel; Capua, Ilaria

2011-01-01

H5N1 highly pathogenic avian influenza virus has been endemic in poultry in Egypt since 2008, notwithstanding the implementation of mass vaccination and culling of infected birds. Extensive circulation of the virus has resulted in a progressive genetic evolution and an antigenic drift. In poultry, the occurrence of antigenic drift in avian influenza viruses is less well documented and the mechanisms remain to be clarified. To test the hypothesis that H5N1 antigenic drift is driven by mechanisms similar to type A influenza viruses in humans, we generated reassortant viruses, by reverse genetics, that harbored molecular changes identified in genetically divergent viruses circulating in the vaccinated population. Parental and reassortant phenotype viruses were antigenically analyzed by hemagglutination inhibition (HI) test and microneutralization (MN) assay. The results of the study indicate that the antigenic drift of H5N1 in poultry is driven by multiple mutations primarily occurring in major antigenic sites at the receptor binding subdomain, similarly to what has been described for human influenza H1 and H3 subtype viruses. PMID:21734057

Classification of infectious bursal disease virus into genogroups.

PubMed

Michel, Linda O; Jackwood, Daral J

2017-12-01

Infectious bursal disease virus (IBDV) causes infectious bursal disease (IBD), an immunosuppressive disease of poultry. The current classification scheme of IBDV is confusing because it is based on antigenic types (variant and classical) as well as pathotypes. Many of the amino acid changes differentiating these various classifications are found in a hypervariable region of the capsid protein VP2 (hvVP2), the major host protective antigen. Data from this study were used to propose a new classification scheme for IBDV based solely on genogroups identified from phylogenetic analysis of the hvVP2 of strains worldwide. Seven major genogroups were identified, some of which are geographically restricted and others that have global dispersion, such as genogroup 1. Genogroup 2 viruses are predominately distributed in North America, while genogroup 3 viruses are most often identified on other continents. Additionally, we have identified a population of genogroup 3 vvIBDV isolates that have an amino acid change from alanine to threonine at position 222 while maintaining other residues conserved in this genogroup (I242, I256 and I294). A222T is an important mutation because amino acid 222 is located in the first of four surface loops of hvVP2. A similar shift from proline to threonine at 222 is believed to play a role in the significant antigenic change of the genogroup 2 IBDV strains, suggesting that antigenic drift may be occurring in genogroup 3, possibly in response to antigenic pressure from vaccination.

Seroprevalence of antibodies and antigens against hepatitis A-E viruses in refugees and asylum seekers in Germany in 2015.

PubMed

Jablonka, Alexandra; Solbach, Philipp; Wöbse, Michael; Manns, Michael P; Schmidt, Reinhold E; Wedemeyer, Heiner; Cornberg, Markus; Behrens, Georg M N; Hardtke, Svenja

2017-08-01

Migration because of miscellaneous political crises in countries in the Middle East and Africa is a global challenge for whole Europe from an economic, social, and public health view. There is an urgent need to generate comprehensive, evidence-based data to expedite further screening and vaccination strategies. A total of 604 individuals ranging in age from 2 to 68 years who enrolled at a single reception center were tested for the prevalence of serologic markers for hepatitis virus types A, B, C, D, and E (HAV, HBV, HCV, HDV, HEV), respectively. Anti-HAV antibody prevalence was 91.2 and 70.3% in children younger than 18 years of age. The prevalence of anti-HEV antibodies was 20.1% among the individuals. 3.0% were positive for hepatitis B surface antigen, whereas 15.2% tested positive for anti-hepatitis B core antigen. None of the refugees tested positive for anti-HDV. 14.1% of refugees were vaccinated against hepatitis B and had a protective anti-hepatitis B surface level of at least 10 mIU/ml. Significant differences in vaccination status were found between the regions (Eastern Mediterranean Region with 77/482 (16.0%; 95% confidence interval=12.7-19.3%) versus African Region with 1/55 (1.8%; 95% confidence interval=0-5.0%). The prevalence of anti-HCV antibodies was 1.2% (n=7), with 0.7% HCV RNA positivity; 16.7% of hepatitis B surface antigen-positive individuals were HCV coinfected (n=3). The prevalence of refugees with previous exposure to hepatitis viruses was higher than that in the general German population, but lower than in other migrant populations in Germany. The vaccination status against hepatitis B was poor.

Selection of antigenically advanced variants of seasonal influenza viruses.

PubMed

Li, Chengjun; Hatta, Masato; Burke, David F; Ping, Jihui; Zhang, Ying; Ozawa, Makoto; Taft, Andrew S; Das, Subash C; Hanson, Anthony P; Song, Jiasheng; Imai, Masaki; Wilker, Peter R; Watanabe, Tokiko; Watanabe, Shinji; Ito, Mutsumi; Iwatsuki-Horimoto, Kiyoko; Russell, Colin A; James, Sarah L; Skepner, Eugene; Maher, Eileen A; Neumann, Gabriele; Klimov, Alexander I; Kelso, Anne; McCauley, John; Wang, Dayan; Shu, Yuelong; Odagiri, Takato; Tashiro, Masato; Xu, Xiyan; Wentworth, David E; Katz, Jacqueline M; Cox, Nancy J; Smith, Derek J; Kawaoka, Yoshihiro

2016-05-23

Influenza viruses mutate frequently, necessitating constant updates of vaccine viruses. To establish experimental approaches that may complement the current vaccine strain selection process, we selected antigenic variants from human H1N1 and H3N2 influenza virus libraries possessing random mutations in the globular head of the haemagglutinin protein (which includes the antigenic sites) by incubating them with human and/or ferret convalescent sera to human H1N1 and H3N2 viruses. We also selected antigenic escape variants from human viruses treated with convalescent sera and from mice that had been previously immunized against human influenza viruses. Our pilot studies with past influenza viruses identified escape mutants that were antigenically similar to variants that emerged in nature, establishing the feasibility of our approach. Our studies with contemporary human influenza viruses identified escape mutants before they caused an epidemic in 2014-2015. This approach may aid in the prediction of potential antigenic escape variants and the selection of future vaccine candidates before they become widespread in nature.

Antigenic evolution of H9N2 chicken influenza viruses isolated in China during 2009–2013 and selection of a candidate vaccine strain with broad cross-reactivity

PubMed Central

Wei, Yandi; Xu, Guanlong; Zhang, Guozhong; Wen, Chu; Anwar, Furkat; Wang, Shuoguo; Lemmon, Gordon; Wang, Jinliang; Carter, Robert; Wang, Min; Sun, Honglei; Sun, Yipeng; Zhao, Jixun; Wu, Gang; Webster, Robert G.; Liu, Jinhua; Pu, Juan

2016-01-01

We previously demonstrated that H9N2 subtype avian influenza viruses (AIVs) isolated from 1994 to 2008 evolved into distinct antigenic groups (C, D, and E) and then underwent antigenic drift from commercial vaccines, causing a country-wide outbreak during 2010–2013. In this study, H9N2 AIVs isolated from chickens during 2009–2013 were antigenically analyzed by performing hemagglutination inhibition and neutralization assays using a panel of polyclonal antibodies. Our findings confirmed the antigenic drift of recent H9N2 viruses from the commercial vaccine and showed that most of these antigenic variants form a novel HI antigenic group, F, with a few belonging to groups D and E. Slight antigenic variation was observed in group F viruses. Genetic analysis of amino acid sequences deduced from hemagglutinin (HA) gene sequences indicated that 9 of 15 mutations predominant in the 2009–2013 viruses can be mapped to known antigenic sites, which might be responsible for the novel antigenicity of group F. These antigenic changes make it necessary to modify the influenza vaccine to ensure efficient protection. A vaccine candidate, Ck/HeB/YT/10, was selected and provided significant protection against viruses from different antigenic groups in terms of reduction in virus shedding, suggesting broad cross-reactivity. Taken together, our results indicate that the H9N2 chicken influenza viruses in China have evolved from distinct antigenic groups into a novel group F that became dominant during the country-wide outbreak and now seems to be undergoing new antigenic divergence. Systematic surveillance and timely updating of vaccine strains are important for viral prevention and control in the future. PMID:26711021

Antigenic evolution of H9N2 chicken influenza viruses isolated in China during 2009-2013 and selection of a candidate vaccine strain with broad cross-reactivity.

PubMed

Wei, Yandi; Xu, Guanlong; Zhang, Guozhong; Wen, Chu; Anwar, Furkat; Wang, Shuoguo; Lemmon, Gordon; Wang, Jinliang; Carter, Robert; Wang, Min; Sun, Honglei; Sun, Yipeng; Zhao, Jixun; Wu, Gang; Webster, Robert G; Liu, Jinhua; Pu, Juan

2016-01-01

We previously demonstrated that H9N2 subtype avian influenza viruses (AIVs) isolated from 1994 to 2008 evolved into distinct antigenic groups (C, D, and E) and then underwent antigenic drift from commercial vaccines, causing a country-wide outbreak during 2010-2013. In this study, H9N2 AIVs isolated from chickens during 2009-2013 were antigenically analyzed by performing hemagglutination inhibition and neutralization assays using a panel of polyclonal antibodies. Our findings confirmed the antigenic drift of recent H9N2 viruses from the commercial vaccine and showed that most of these antigenic variants form a novel HI antigenic group, F, with a few belonging to groups D and E. Slight antigenic variation was observed in group F viruses. Genetic analysis of amino acid sequences deduced from hemagglutinin (HA) gene sequences indicated that 9 of 15 mutations predominant in the 2009-2013 viruses can be mapped to known antigenic sites, which might be responsible for the novel antigenicity of group F. These antigenic changes make it necessary to modify the influenza vaccine to ensure efficient protection. A vaccine candidate, Ck/HeB/YT/10, was selected and provided significant protection against viruses from different antigenic groups in terms of reduction in virus shedding, suggesting broad cross-reactivity. Taken together, our results indicate that the H9N2 chicken influenza viruses in China have evolved from distinct antigenic groups into a novel group F that became dominant during the country-wide outbreak and now seems to be undergoing new antigenic divergence. Systematic surveillance and timely updating of vaccine strains are important for viral prevention and control in the future. Copyright © 2015 Elsevier B.V. All rights reserved.

Identification of Low- and High-Impact Hemagglutinin Amino Acid Substitutions That Drive Antigenic Drift of Influenza A(H1N1) Viruses

PubMed Central

Harvey, William T.; Benton, Donald J.; Gregory, Victoria; Hall, James P. J.; Daniels, Rodney S.; Bedford, Trevor; Haydon, Daniel T.; Hay, Alan J.; McCauley, John W.; Reeve, Richard

2016-01-01

Determining phenotype from genetic data is a fundamental challenge. Identification of emerging antigenic variants among circulating influenza viruses is critical to the vaccine virus selection process, with vaccine effectiveness maximized when constituents are antigenically similar to circulating viruses. Hemagglutination inhibition (HI) assay data are commonly used to assess influenza antigenicity. Here, sequence and 3-D structural information of hemagglutinin (HA) glycoproteins were analyzed together with corresponding HI assay data for former seasonal influenza A(H1N1) virus isolates (1997–2009) and reference viruses. The models developed identify and quantify the impact of eighteen amino acid substitutions on the antigenicity of HA, two of which were responsible for major transitions in antigenic phenotype. We used reverse genetics to demonstrate the causal effect on antigenicity for a subset of these substitutions. Information on the impact of substitutions allowed us to predict antigenic phenotypes of emerging viruses directly from HA gene sequence data and accuracy was doubled by including all substitutions causing antigenic changes over a model incorporating only the substitutions with the largest impact. The ability to quantify the phenotypic impact of specific amino acid substitutions should help refine emerging techniques that predict the evolution of virus populations from one year to the next, leading to stronger theoretical foundations for selection of candidate vaccine viruses. These techniques have great potential to be extended to other antigenically variable pathogens. PMID:27057693

Simian virus 40-related antigens in three human meningiomas with defined chromosome loss.

PubMed Central

Weiss, A F; Portmann, R; Fischer, H; Simon, J; Zang, K D

1975-01-01

Two out of seven meningiomas tested in early cell cultures by indirect immunofluorescence staining showed simian virus 40 (SV40)-related tumor (T) antigen. In one tumor 90% of the cells were positive. An additional SV40-related antigen (U) was found in 10% of cells of a third tumor. These findings indicate that the meningioma cells showing a positive reaction are transformed by a papova virus that has at least partly the same antigenic properties as SV40 virus. SV40-related viral capsid (V) antigen was absent in all the meningiomas tested. No virus infectious for African green monkey kidney (AGMK) cells could be isolated. The tumors positive for T and U antigens showed the chromosome aberration typical for human meningiomas, i.e., the loss of one chromosome, G-22. The T-antigen-positive tumors showed further hypodiploidization. Experiments to rescue virus from the T-antigen-positive tumors showed further hypodiploidization. Experiments to rescue virus from the T-antigen-positive meningioma cells were performed: fusion of cells pretreated with 8-azaguanine with cells premissive for SV40 led to a low percentage (0.01-0.05%) of V-antigen-positive nuclei in heterokaryon cultures. On the basis of these results, the possibility of a correlation between the meningioma, a relatively common intracranial tumor in man, and an SV40-related papova virus must be considered. It remains to be shown whether this virus is a causative agent for human meningiomas. Images PMID:164660

Immunizations with chimeric hepatitis B virus-like particles to induce potential anti-hepatitis C virus neutralizing antibodies.

PubMed

Vietheer, Patricia T K; Boo, Irene; Drummer, Heidi E; Netter, Hans-Jürgen

2007-01-01

Virus-like particles (VLPs) are highly immunogenic and proven to induce protective immunity. The small surface antigen (HBsAg-S) of hepatitis B virus (HBV) self-assembles into VLPs and its use as a vaccine results in protective antiviral immunity against HBV infections. Chimeric HBsAg-S proteins carrying foreign epitopes allow particle formation and have the ability to induce anti-foreign humoral and cellular immune responses. The insertion of the hypervariable region 1 (HVR1) sequence derived from the envelope protein 2 (E2) of hepatitis C virus (HCV) into the major antigenic site of HBsAg-S ('a'-determinant) resulted in the formation of highly immunogenic VLPs that retained the antigenicity of the inserted HVR1 sequence. BALB/c mice were immunized with chimeric VLPs, which resulted in antisera with anti-HCV activity. The antisera were able to immunoprecipitate native HCV envelope complexes (E1E2) containing homologous or heterologous HVR1 sequences. HCV E1E2 pseudotyped HIV-1 particles (HCVpp) were used to measure entry into HuH-7 target cells in the presence or absence of antisera that were raised against chimeric VLPs. Anti-HVR1 VLP sera interfered with entry of entry-competent HCVpps containing either homologous or heterologous HVR1 sequences. Also, immunizations with chimeric VLPs induced antisurface antigen (HBsAg) antibodies, indicating that HBV-specific antigenicity and immunogenicity of the 'a'-determinant region is retained. A multivalent vaccine against different pathogens based on the HBsAg delivery platform should be possible. We hypothesize that custom design of VLPs with an appropriate set of HCV-neutralizing epitopes will induce antibodies that would serve to decrease the viral load at the initial infecting inoculum.

Peripheral B cells latently infected with Epstein–Barr virus display molecular hallmarks of classical antigen-selected memory B cells

PubMed Central

Souza, Tatyana A.; Stollar, B. David; Sullivan, John L.; Luzuriaga, Katherine; Thorley-Lawson, David A.

2005-01-01

Epstein–Barr virus (EBV) establishes a lifelong persistent infection within peripheral blood B cells with the surface phenotype of memory cells. To date there is no proof that these cells have the genotype of true germinal-center-derived memory B cells. It is critical to understand the relative contribution of viral mimicry versus antigen signaling to the production of these cells because EBV encodes proteins that can affect the surface phenotype of infected cells and provide both T cell help and B cell receptor signals in the absence of cognate antigen. To address these questions we have developed a technique to identify single EBV-infected cells in the peripheral blood and examine their expressed Ig genes. The genes were all isotype-switched and somatically mutated. Furthermore, the mutations do not cause stop codons and display the pattern expected for antigen-selected memory cells based on their frequency, type, and location within the Ig gene. We conclude that latently infected peripheral blood B cells display the molecular hallmarks of classical antigen-selected memory B cells. Therefore, EBV does not disrupt the normal processing of latently infected cells into memory, and deviations from normal B cell biology are not tolerated in the infected cells. This article provides definitive evidence that EBV in the peripheral blood persists in true memory B cells. PMID:16330748

Persistent Lymphadenopathy due to IgG4-Related Disease

DTIC Science & Technology

2012-10-01

worsened, she was referred to Infectious Disease who entertained a broad differential including Kikuchi’s syndrome, Epstein - Barr virus (EBV...anti-Smith; EBV = Epstein - Barr virus , CMV: cytomegalovirus, HBs Ag = hepatitis B surface antigen, and HBc Ab: hepatitis B core antibody IgG, HBs Ab...plasmosis, HIV, and mycobacterium), lymphoma/leukemia, metastatic neoplasia, systemic lupus erythematous, Castle- man’s disease , autoimmune

Multiplex detection of IgG and IgM to Rift Valley fever virus nucleoprotein, nonstructural proteins, and glycoprotein in ovine and bovine

USDA-ARS?s Scientific Manuscript database

A multiplex fluorescence microsphere immunoassay (FMIA) was used to detect bovine and ovine IgM and IgG antibodies to several Rift Valley fever virus (RVFV) proteins, including the major surface glycoprotein, Gn; the nonstructural proteins, NSs and NSm; and the nucleoprotein, N. Target antigens were...

Rapid Detection of Hepatitis B Virus Surface Antigen by an Agglutination Assay Mediated by a Bispecific Diabody against Both Human Erythrocytes and Hepatitis B Virus Surface Antigen▿

PubMed Central

Chen, Yu-Ping; Qiao, Yuan-Yuan; Zhao, Xiao-Hang; Chen, Hong-Song; Wang, Yan; Wang, Zhuozhi

2007-01-01

Bispecific antibodies have immense potential for use in clinical applications. In the present study, a bispecific diabody against human red blood cells (RBCs) and hepatitis B virus surface antigen (HBsAg) was used to detect HBsAg in blood specimens. The bispecific diabody was constructed by crossing over the variable region of the heavy chains and the light chains of anti-RBC and anti-HBsAg antibodies with a short linker, SRGGGS. In enzyme-linked immunosorbent assays, this bispecific diabody showed specific binding to both RBCs and HBsAg. When this bispecific diabody was mixed with human blood specimens in the presence of HBsAg, the dual binding sites of the diabody caused agglutination of human RBCs. This diabody-mediated agglutination assay was then used to test 712 clinical blood specimens and showed 97.7% sensitivity and 100% specificity when the results were compared with those of the conventional immunoassay, which was used as a reference. This autologous RBC agglutination assay provides a simple approach for rapid screening for HBsAg in blood specimens. PMID:17442848

Membranous glomerulonephritis in a child asymptomatic for hepatitis B virus. Concomitant seropositivity for HBsAG and anti-HBs.

PubMed

Hirsch, H Z; Ainsworth, S K; DeBeukelaer, M; Brissie, R M; Hennigar, G R

1981-04-01

The presence of hepatitis B surface antigen (HBsAg) in association with immunoglobulins and complement components within the glomerular basement membranes of adults having chronic active hepatitis has been well documented. In addition, investigators in Poland have demonstrated HBsAg immune complexes in glomeruli of children who did not have clinical evidence of hepatitis. More recently, a single case of childhood membranous glomerulonephritis in an asymptomatic carrier of hepatitis B virus was cited by observers in Canada. Reported here is the deposition of HBsAg immune complexes in the glomerular basement membranes of a 13-year-old black boy who had membranous glomerulopathy but not clinical evidence of hepatitis. This may be the first reported case in the United States of HbsAg-associated membranous glomerulonephritis in a child asymptomatic for hepatitis B virus, and only the second such case in North America. However, unlike previous studies of childhood glomerulopathy in association with hepatitis B virus, this patient is seropositive for both HBsAg and anti-HBs (antibody for hepatitis B surface antigen). Similar "rare" serologic findings were found for the patient's eldest male sib.

Membrane and envelope virus proteins co-expressed as lysosome associated membrane protein (LAMP) fused antigens: a potential tool to develop DNA vaccines against flaviviruses.

PubMed

Dhalia, Rafael; Maciel, Milton; Cruz, Fábia S P; Viana, Isabelle F T; Palma, Mariana L; August, Thomas; Marques, Ernesto T A

2009-12-01

Vaccination is the most practical and cost-effective strategy to prevent the majority of the flavivirus infection to which there is an available vaccine. However, vaccines based on attenuated virus can potentially promote collateral side effects and even rare fatal reactions. Given this scenario, the development of alternative vaccination strategies such as DNA-based vaccines encoding specific flavivirus sequences are being considered. Endogenous cytoplasmic antigens, characteristically plasmid DNA-vaccine encoded, are mainly presented to the immune system through Major Histocompatibility Complex class I - MHC I molecules. The MHC I presentation via is mostly associated with a cellular cytotoxic response and often do not elicit a satisfactory humoral response. One of the main strategies to target DNA-encoded antigens to the MHC II compartment is expressing the antigen within the Lysosome-Associated Membrane Protein (LAMP). The flavivirus envelope protein is recognized as the major virus surface protein and the main target for neutralizing antibodies. Different groups have demonstrated that co-expression of flavivirus membrane and envelope proteins in mammalian cells, fused with the carboxyl-terminal of LAMP, is able to induce satisfactory levels of neutralizing antibodies. Here we reviewed the use of the envelope flavivirus protein co-expression strategy as LAMP chimeras with the aim of developing DNA vaccines for dengue, West Nile and yellow fever viruses.

Binding of a neutralizing antibody to dengue virus alters the arrangement of surface glycoproteins

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lok, Shee-Mei; Kostyuchenko, Victor; Nybakken, Grant E.

The monoclonal antibody 1A1D-2 has been shown to strongly neutralize dengue virus serotypes 1, 2 and 3, primarily by inhibiting attachment to host cells. A crystal structure of its antigen binding fragment (Fab) complexed with domain III of the viral envelope glycoprotein, E, showed that the epitope would be partially occluded in the known structure of the mature dengue virus. Nevertheless, antibody could bind to the virus at 37 degrees C, suggesting that the virus is in dynamic motion making hidden epitopes briefly available. A cryo-electron microscope image reconstruction of the virus:Fab complex showed large changes in the organization ofmore » the E protein that exposed the epitopes on two of the three E molecules in each of the 60 icosahedral asymmetric units of the virus. The changes in the structure of the viral surface are presumably responsible for inhibiting attachment to cells.« less

The global antigenic diversity of swine influenza A viruses

PubMed Central

Lewis, Nicola S; Russell, Colin A; Langat, Pinky; Anderson, Tavis K; Berger, Kathryn; Bielejec, Filip; Burke, David F; Dudas, Gytis; Fonville, Judith M; Fouchier, Ron AM; Kellam, Paul; Koel, Bjorn F; Lemey, Philippe; Nguyen, Tung; Nuansrichy, Bundit; Peiris, JS Malik; Saito, Takehiko; Simon, Gaelle; Skepner, Eugene; Takemae, Nobuhiro; Webby, Richard J; Van Reeth, Kristien; Brookes, Sharon M; Larsen, Lars; Watson, Simon J; Brown, Ian H; Vincent, Amy L

2016-01-01

Swine influenza presents a substantial disease burden for pig populations worldwide and poses a potential pandemic threat to humans. There is considerable diversity in both H1 and H3 influenza viruses circulating in swine due to the frequent introductions of viruses from humans and birds coupled with geographic segregation of global swine populations. Much of this diversity is characterized genetically but the antigenic diversity of these viruses is poorly understood. Critically, the antigenic diversity shapes the risk profile of swine influenza viruses in terms of their epizootic and pandemic potential. Here, using the most comprehensive set of swine influenza virus antigenic data compiled to date, we quantify the antigenic diversity of swine influenza viruses on a multi-continental scale. The substantial antigenic diversity of recently circulating viruses in different parts of the world adds complexity to the risk profiles for the movement of swine and the potential for swine-derived infections in humans. DOI: http://dx.doi.org/10.7554/eLife.12217.001 PMID:27113719

Virus-based nanoparticles as platform technologies for modern vaccines

PubMed Central

Lee, Karin L.; Twyman, Richard M.; Fiering, Steven

2017-01-01

Nanoscale engineering is revolutionizing the development of vaccines and immunotherapies. Viruses have played a key role in this field because they can function as prefabricated nanoscaffolds with unique properties that are easy to modify. Viruses are immunogenic through multiple pathways, and antigens displayed naturally or by engineering on the surface can be used to create vaccines against the cognate virus, other pathogens, specific molecules or cellular targets such as tumors. This review focuses on the development of virus-based nanoparticle systems as vaccines indicated for the prevention or treatment of infectious diseases, chronic diseases, cancer, and addiction. PMID:26782096

Overview of expression of hepatitis B surface antigen in transgenic plants.

PubMed

Guan, Zheng-jun; Guo, Bin; Huo, Yan-lin; Guan, Zheng-ping; Wei, Ya-hui

2010-10-28

Hepatitis B virus (HBV), a pathogen for chronic liver infection, afflicts more than 350 million people world-wide. The effective way to control the virus is to take HBV vaccine. Hepatitis B surface antigen (HBsAg) is an effective protective antigen suitable for vaccine development. At present, "edible" vaccine based on transgenic plants is one of the most promising directions in novel types of vaccines. HBsAg production from transgenic plants has been carried out, and the transgenic plant expression systems have developed from model plants (such as tobacco, potato and tomato) to other various plant platforms. Crude or purified extracts of transformed plants have been found to conduct immunological responses and clinical trials for hepatitis B, which gave the researches of plant-based HBsAg production a big boost. The aim of this review was to summarize the recent data about plant-based HBsAg development including molecular biology of HBsAg gene, selection of expression vector, the expression of HBsAg gene in plants, as well as corresponding immunological responses in animal models or human. Copyright © 2010 Elsevier Ltd. All rights reserved.

Antigenic variation: Molecular and genetic mechanisms of relapsing disease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cruse, J.M.; Lewis, R.E.

1987-01-01

This book contains 10 chapters. They are: Contemporary Concepts of Antigenic Variation; Antigenic Variation in the Influenza Viruses; Mechanisms of Escape of Visna Lentiviruses from Immunological Control; A Review of Antigenic Variation by the Equine Infectious Anemia Virus; Biologic and Molecular Variations in AIDS Retrovirus Isolates; Rabies Virus Infection: Genetic Mutations and the Impact on Viral Pathogenicity and Immunity; Immunobiology of Relapsing Fever; Antigenic Variation in African Trypanosomes; Antigenic Variation and Antigenic Diversity in Malaria; and Mechanisms of Immune Evasion in Schistosomiasis.

Selection of antigenically advanced variants of seasonal influenza viruses

PubMed Central

Ozawa, Makoto; Taft, Andrew S.; Das, Subash C.; Hanson, Anthony P.; Song, Jiasheng; Imai, Masaki; Wilker, Peter R.; Watanabe, Tokiko; Watanabe, Shinji; Ito, Mutsumi; Iwatsuki-Horimoto, Kiyoko; Russell, Colin A.; James, Sarah L.; Skepner, Eugene; Maher, Eileen A.; Neumann, Gabriele; Kelso, Anne; McCauley, John; Wang, Dayan; Shu, Yuelong; Odagiri, Takato; Tashiro, Masato; Xu, Xiyan; Wentworth, David E.; Katz, Jacqueline M.; Cox, Nancy J.; Smith, Derek J.; Kawaoka, Yoshihiro

2016-01-01

Influenza viruses mutate frequently, necessitating constant updates of vaccine viruses. To establish experimental approaches that may complement the current vaccine strain selection process, we selected antigenic variants from human H1N1 and H3N2 influenza virus libraries possessing random mutations in the globular head of the haemagglutinin protein (which includes the antigenic sites) by incubating them with human and/or ferret convalescent sera to human H1N1 and H3N2 viruses. Further, we selected antigenic escape variants from human viruses treated with convalescent sera and from mice that had been previously immunized against human influenza viruses. Our pilot studies with past influenza viruses identified escape mutants that were antigenically similar to variants that emerged in nature, establishing the feasibility of our approach. Our studies with contemporary human influenza viruses identified escape mutants before they caused an epidemic in 2014–2015. This approach may aid in the prediction of potential antigenic escape variants and the selection of future vaccine candidates before they become widespread in nature. PMID:27572841

Rabies virus glycoprotein as a carrier for anthrax protective antigen

DOE Office of Scientific and Technical Information (OSTI.GOV)

Smith, Mary Ellen; Koser, Martin; Xiao Sa

2006-09-30

Live viral vectors expressing foreign antigens have shown great promise as vaccines against viral diseases. However, safety concerns remain a major problem regarding the use of even highly attenuated viral vectors. Using the rabies virus (RV) envelope protein as a carrier molecule, we show here that inactivated RV particles can be utilized to present Bacillus anthracis protective antigen (PA) domain-4 in the viral membrane. In addition to the RV glycoprotein (G) transmembrane and cytoplasmic domains, a portion of the RV G ectodomain was required to express the chimeric RV G anthrax PA on the cell surface. The novel antigen wasmore » also efficiently incorporated into RV virions. Mice immunized with the inactivated recombinant RV virions exhibited seroconversion against both RV G and anthrax PA, and a second inoculation greatly increased these responses. These data demonstrate that a viral envelope protein can carry a bacterial protein and that a viral carrier can display whole polypeptides compared to the limited epitope presentation of previous viral systems.« less

Immunization against Rabies with Plant-Derived Antigen

NASA Astrophysics Data System (ADS)

Modelska, Anna; Dietzschold, Bernard; Sleysh, N.; Fu, Zhen Fang; Steplewski, Klaudia; Hooper, D. Craig; Koprowski, Hilary; Yusibov, Vidadi

1998-03-01

We previously demonstrated that recombinant plant virus particles containing a chimeric peptide representing two rabies virus epitopes stimulate virus neutralizing antibody synthesis in immunized mice. We show here that mice immunized intraperitoneally or orally (by gastric intubation or by feeding on virus-infected spinach leaves) with engineered plant virus particles containing rabies antigen mount a local and systemic immune response. After the third dose of antigen, given intraperitoneally, 40% of the mice were protected against challenge infection with a lethal dose of rabies virus. Oral administration of the antigen stimulated serum IgG and IgA synthesis and ameliorated the clinical signs caused by intranasal infection with an attenuated rabies virus strain.

Presenting Influenza A M2e Antigen on Recombinant Spores of Bacillus subtilis

PubMed Central

Obuchowski, Michał; Nidzworski, Dawid

2016-01-01

Effective vaccination against influenza virus infection is a serious problem mainly due to antigenic variability of the virus. Among many of investigated antigens, the extracellular domain of the M2 protein (M2e) features high homology in all strains of influenza A viruses and antibodies against M2e and is protective in animal models; this makes it a potential candidate for generation of a universal influenza vaccine. However, due to the low immunogenicity of the M2e, formulation of a vaccine based on this antigen requires some modification to induce effective immune responses. In this work we evaluated the possible use of Bacillus subtilis spores as a carrier of the Influenza A M2e antigen in mucosal vaccination. A tandem repeat of 4 consensus sequences coding for human—avian—swine—human M2e (M2eH-A-S-H) peptide was fused to spore coat proteins and stably exposed on the spore surface, as demonstrated by the immunostaining of intact, recombinant spores. Oral immunization of mice with recombinant endospores carrying M2eH-A-S-H elicited specific antibody production without the addition of adjuvants. Bacillus subtilis endospores can serve as influenza antigen carriers. Recombinant spores constructed in this work showed low immunogenicity although were able to induce antibody production. The System of influenza antigen administration presented in this work is attractive mainly due to the omitting time-consuming and cost-intensive immunogen production and purification. Therefore modification should be made to increase the immunogenicity of the presented system. PMID:27902762

Demonstration of feline corona virus (FCV) antigen in organs of cats suspected of feline infectious peritonitis (FIP) disease.

PubMed

Hök, K

1990-07-01

Cryosections of organs and smears from membrana nicitians from cats suspected of having spontaneous infection with feline infectious peritonitis virus (FIPV), were investigated using an indirect immunofluorescence assay (IIFA) in order to detect the presence of feline corona virus (FCV). In 113 cats, from each of which six organs were screened, virus antigen was found most often in membrana nicitians and lung. Out of these animals an additional six organs from a group of 30 cats were screened. In these cats membrana nicitians, parotid gland, thymus and apex of caecum had the highest incidence of virus antigen (90%). The lowest incidence of virus antigen was found in the spleen (60%). There was a clear demonstration of a higher incidence of antigen present in more than half of the total number of screened organs per cat (P less than 0.0005). No statistical difference was observed between sexes when comparing the incidence of virus antigen in different organs. Virus antigen was present in less organs in cats with no lesions suggestive of FIP disease compared to cats with such lesions (P less than 0.001). A similar distribution of the incidence of FCV antigen in the investigated organs was observed in these two groups.

Surface expression of squamous cell carcinoma antigen (SCCA) can be increased by the preS1(21-47) sequence of hepatitis B virus.

PubMed

Ruvoletto, M G; Tono, N; Carollo, D; Vilei, T; Trentin, L; Muraca, M; Marino, M; Gatta, A; Fassina, G; Pontisso, P

2004-03-01

A variant of the serpin squamous cell carcinoma antigen (SCCA) has been identified as a hepatitis B virus binding protein and high expression of SCCA has recently been found in hepatocarcinoma. Since HBV is involved in liver carcinogenesis, experiments were carried out to examine the effect of HBV preS1 envelope protein on SCCA expression. Surface and intracellular staining for SCCA was assessed by FACS analysis. Preincubation of HepG2 cells and primary human hepatocytes with preS1 protein or with preS1(21-47) tetrameric peptide significantly increased the surface expression of SCCA, without modification of its overall cellular burden, suggesting a surface redistribution of the serpin. An increase in HBV binding and internalization was observed after pre-incubation of the cells with preS1 preparations, compared to cells preincubated with medium alone. Pretreatment of cells with DMSO, while not influencing SCCA basal expression, was responsible for an increase in the efficiency of HBV internalization and this effect was additive to that obtained after incubation with preS1 preparations. In conclusion, the HBV preS1(21-47) sequence is able to induce overexpression of SCCA at the cell surface facilitating virus internalization, while the increased efficiency of HBV entry following DMSO addition is not mediated by SCCA.

Proteins of Vasicular Stomatitis Virus

PubMed Central

Kang, C. Y.; Prevec, L.

1969-01-01

Infection of L cells with vesicular stomatitis virus results in the release, into the cell-free fluid, of four antigenic components separable by rate zonal centrifugation on sucrose gradients. The largest antigens are the infectious (B) particle and a shorter noninfectious, autointerfering (T) particle. The two small antigens are characterized by sedimentation coefficients of approximately 20S and 6S. Treatment of purified B or T particles with sodium deoxycholate results in the release from the particle of a nucleoprotein core which can be purified on sucrose gradient and which has a sedimentation coefficient characteristic of the virus from which it arose. Utilizing purified antigens labeled with 14C-amino acids during growth, we examined the protein constituents of each antigen by acrylamide-gel electrophoresis. The proteins of B and T particles are identical, each containing one minor (virus protein 1) and three major (virus proteins 2, 3, and 4) proteins, numbered in order of increasing mobility. Virus protein 3 originates from the nucleoprotein core, whereas proteins 2 and 4 come from the coat. The origin of virus protein 1 is not known. The 20S antigen contains a single protein equivalent to virus protein 3, whereas the 6S antigen shows a single protein which is similar to, but probably distinct from, virus protein 2. PMID:4306195

Plasticity of a critical antigenic determinant in the West Nile virus NY99 envelope protein domain III.

PubMed

Plante, Jessica A; Torres, Maricela; Huang, Claire Y-H; Beasley, David W C

2016-09-01

West Nile virus (WNV) is a mosquito-borne flavivirus that causes febrile illness, encephalitis, and occasionally death in humans. The envelope protein is the main component of the WNV virion surface, and domain III of the envelope protein (EIII) is both a putative receptor binding domain and a target of highly specific, potently neutralizing antibodies. Envelope E-332 (E-332) is known to have naturally occurring variation and to be a key determinant of neutralization for anti-EIII antibodies. A panel of viruses containing all possible amino acid substitutions at E-332 was constructed. E-332 was found to be highly tolerant of mutation, and almost all of these changes had large impacts on antigenicity of EIII but only limited effects on growth or virulence phenotypes. Copyright © 2016 Elsevier Inc. All rights reserved.

A New Group of Hepadnaviruses Naturally Infecting Orangutans (Pongo pygmaeus)

PubMed Central

Warren, Kristin S.; Heeney, Jonathan L.; Swan, Ralph A.; Heriyanto; Verschoor, Ernst J.

1999-01-01

A high prevalence (42.6%) of hepatitis B virus (HBV) infection was suspected in 195 formerly captive orangutans due to a large number of serum samples which cross-reacted with human HBV antigens. It was assumed that such viral infections were contracted from humans during captivity. However, two wild orangutans were identified which were HBV surface antigen positive, indicating that HBV or related viruses may be occurring naturally in the orangutan populations. Sequence analyses of seven isolates revealed that orangutans were infected with hepadnaviruses but that these were clearly divergent from the six known human HBV genotypes and those of other nonhuman hepadnaviruses reported. Phylogenetic analyses revealed geographic clustering with Southeast Asian genotype C viruses and gibbon ape HBV. This implies a common origin of infection within this geographic region, with cross-species transmission of hepadnaviruses among hominoids. PMID:10438880

[A structural protein study of the influenza A (H1N1) virus by polyacrylamide gel electrophoresis].

PubMed

Pérez Guevara, M T; Savón Valdés, C; Rivas Arjona, M; Goyenechea Hernández, A

1992-01-01

Influenza is an acute respiratory disease typically appearing as an epidemic. Three immunological types of the influenza virus are known: A, B and C. Continually, antigen changes occur, especially in type A. Therefore, a comparative study was carried out on 4 influenza A(H1N1) virus strains in relation to protein structure (surface antigens), by using polyacrylamide gel electrophoresis by the modified Laemmli method. The objective was to compare the structural proteins of the A/Havana/1292/78 (H1N1) national strain with the proteins of 3 international pattern strains. In all the cases, 6 bands were detected by densitometry. In the 4 strains studied the most abundant protein was M. Great differences between the Cuban strain and the 3 international patterns were not seen.

Renal transplantation from hepatitis B surface antigen (HBsAg)-positive donors to HBsAg-negative recipients: a case of post-transplant fulminant hepatitis associated with an extensively mutated hepatitis B virus strain and review of the current literature.

PubMed

Magiorkinis, E; Paraskevis, D; Pavlopoulou, I D; Kantzanou, M; Haida, C; Hatzakis, A; Boletis, I N

2013-08-01

The purpose of this study was to present a fatal case of fulminant hepatitis B (FHB) that developed in a renal transplant recipient, immunized against hepatitis B, 1 year post transplantation. Polymerase chain reaction amplification and full genome sequencing were performed to investigate whether specific mutations were associated with hepatitis B virus (HBV) transmission and FHB. Molecular analysis revealed multiple mutations in various open reading frames of HBV, the most important being the G145R escape mutation and a frameshift mutation-insertion (1838insA) within the pre-C/C reading frame. Our results highlight the possibility of developing FHB, despite previous immunization against HBV or administration of hyperimmune gammaglobulin, because of the selection of escape virus mutants. The current literature and guidelines regarding renal transplantation from hepatitis B surface antigen (HBsAg)-positive to HBsAg-negative patients were also reviewed. © 2013 John Wiley & Sons A/S.

Structure of Adeno-Associated Virus Type 4

PubMed Central

Padron, Eric; Bowman, Valorie; Kaludov, Nikola; Govindasamy, Lakshmanan; Levy, Hazel; Nick, Phillip; McKenna, Robert; Muzyczka, Nicholas; Chiorini, John A.; Baker, Timothy S.; Agbandje-McKenna, Mavis

2005-01-01

Adeno-associated virus (AAV) is a member of the Parvoviridae, belonging to the Dependovirus genus. Currently, several distinct isolates of AAV are in development for use in human gene therapy applications due to their ability to transduce different target cells. The need to manipulate AAV capsids for specific tissue delivery has generated interest in understanding their capsid structures. The structure of AAV type 4 (AAV4), one of the most antigenically distinct serotypes, was determined to 13-Å resolution by cryo-electron microscopy and image reconstruction. A pseudoatomic model was built for the AAV4 capsid by use of a structure-based sequence alignment of its major capsid protein, VP3, with that of AAV2, to which AAV4 is 58% identical and constrained by its reconstructed density envelope. The model showed variations in the surface loops that may account for the differences in receptor binding and antigenicity between AAV2 and AAV4. The AAV4 capsid surface topology also shows an unpredicted structural similarity to that of Aleutian mink disease virus and human parvovirus B19, autonomous members of the genus, despite limited sequence homology. PMID:15795290

Antibody Against Integrin Lymphocyte Function-Associated Antigen 1 Inhibits HIV Type 1 Infection in Primary Cells Through Caspase-8-Mediated Apoptosis

PubMed Central

Walker, Tiffany N.; Cimakasky, Lisa M.; Coleman, Ebony M.; Madison, M. Nia

2013-01-01

Abstract HIV-1 infection induces formation of a virological synapse wherein CD4, chemokine receptors, and cell-adhesion molecules such as lymphocyte function-associated antigen 1 (LFA-1) form localized domains on the cell surface. Studies show that LFA-1 on the surface of HIV-1 particles retains its adhesion function and enhances virus attachment to susceptible cells by binding its counterreceptor intercellular adhesion molecule 1 (ICAM-1). This virus–cell interaction augments virus infectivity by facilitating binding and entry events. In this study, we demonstrate that inhibition of the LFA-1/ICAM-1 interaction by a monoclonal antibody leads to decreased virus production and spread in association with increased apoptosis of HIV-infected primary T cells. The data indicate that the LFA-1/ICAM-1 interaction may limit apoptosis in HIV-1-infected T cells. This phenomenon appears similar to anoikis wherein epithelial cells are protected from apoptosis conferred by ligand-bound integrins. These results have implications for further understanding HIV pathogenesis and replication in peripheral compartments and lymphoid organs. PMID:22697794

Immunofluorescence of bovine virus diarrhea viral antigen in white blood cells from experimentally infected immunocompetent calves.

PubMed Central

Bezek, D M; Baker, J C; Kaneene, J B

1988-01-01

A study to evaluate the detection of bovine virus diarrhea viral antigen using immunofluorescence testing of white blood cells was conducted. Five colostrum-deprived calves were inoculated intravenously with a cytopathic strain of the virus. Lymphocyte and buffy coat smears were prepared daily for direct immunofluorescent staining for detection of antigen. Lymphocytes were separated from heparinized blood using a Ficoll density procedure. Buffy coat smears were prepared from centrifuged blood samples collected using ethylenediaminetetraacetic acid as an anticoagulant. Bovine viral diarrhea virus antigen was detected by immunofluorescence between 3 and 11 days postinfection in lymphocyte smears and 3 to 12 days postinfection in buffy coat smears. Isolation of virus from both lymphocytes and buffy coat preparations correlated with detection of immunofluorescence. Serum neutralizing antibody to bovine virus diarrhea virus was detected on day 10 postinfection. Buffy coat smears were as sensitive as lymphocyte smears for the detection of antigen by immunofluorescence. It appeared that immunofluorescent staining of white blood cells was an effective method of detecting bovine virus diarrhea viral antigen. PMID:2836047

Specific Antibodies Reacting with SV40 Large T Antigen Mimotopes in Serum Samples of Healthy Subjects

PubMed Central

Tognon, Mauro; Corallini, Alfredo; Manfrini, Marco; Taronna, Angelo; Butel, Janet S.; Pietrobon, Silvia; Trevisiol, Lorenzo; Bononi, Ilaria; Vaccher, Emanuela; Barbanti-Brodano, Giuseppe; Martini, Fernanda; Mazzoni, Elisa

2016-01-01

Simian Virus 40, experimentally assayed in vitro in different animal and human cells and in vivo in rodents, was classified as a small DNA tumor virus. In previous studies, many groups identified Simian Virus 40 sequences in healthy individuals and cancer patients using PCR techniques, whereas others failed to detect the viral sequences in human specimens. These conflicting results prompted us to develop a novel indirect ELISA with synthetic peptides, mimicking Simian Virus 40 capsid viral protein antigens, named mimotopes. This immunologic assay allowed us to investigate the presence of serum antibodies against Simian Virus 40 and to verify whether Simian Virus 40 is circulating in humans. In this investigation two mimotopes from Simian Virus 40 large T antigen, the viral replication protein and oncoprotein, were employed to analyze for specific reactions to human sera antibodies. This indirect ELISA with synthetic peptides from Simian Virus 40 large T antigen was used to assay a new collection of serum samples from healthy subjects. This novel assay revealed that serum antibodies against Simian Virus 40 large T antigen mimotopes are detectable, at low titer, in healthy subjects aged from 18–65 years old. The overall prevalence of reactivity with the two Simian Virus 40 large T antigen peptides was 20%. This new ELISA with two mimotopes of the early viral regions is able to detect in a specific manner Simian Virus 40 large T antigen-antibody responses. PMID:26731525

Critical role for the chemokine receptor CXCR6 in NK cell-mediated antigen-specific memory of haptens and viruses.

PubMed

Paust, Silke; Gill, Harvinder S; Wang, Bao-Zhong; Flynn, Michael P; Moseman, E Ashley; Senman, Balimkiz; Szczepanik, Marian; Telenti, Amalio; Askenase, Philip W; Compans, Richard W; von Andrian, Ulrich H

2010-12-01

Hepatic natural killer (NK) cells mediate antigen-specific contact hypersensitivity (CHS) in mice deficient in T cells and B cells. We report here that hepatic NK cells, but not splenic or naive NK cells, also developed specific memory of vaccines containing antigens from influenza, vesicular stomatitis virus (VSV) or human immunodeficiency virus type 1 (HIV-1). Adoptive transfer of virus-sensitized NK cells into naive recipient mice enhanced the survival of the mice after lethal challenge with the sensitizing virus but not after lethal challenge with a different virus. NK cell memory of haptens and viruses depended on CXCR6, a chemokine receptor on hepatic NK cells that was required for the persistence of memory NK cells but not for antigen recognition. Thus, hepatic NK cells can develop adaptive immunity to structurally diverse antigens, an activity that requires NK cell-expressed CXCR6.

[The lysate and recombinant antigens in ELISA-test-systems for diagnostic of herpes simplex].

PubMed

Ganova, L A; Kovtoniuk, G V; Korshun, L N; Kiseleva, E K; Tereshchenko, M I; Vudmaska, M I; Moĭsa, L N; Shevchuk, V A; Spivak, N Ia

2014-08-01

The lysate and recombinant antigens of various production included informula of ELISA-test-systems were analyzed. The ELISA-test-systems are used for detection of IgG to Herpes simplex virus type I and II. For testing the panel of serums PTH 201 (BBI Inc.) were used. The samples of this panel contain antibodies to Herpes simplex virus type I and II in mixed titers. The 69 serums of donors were used too (17 samples had IgG to Herpes simplex virus type I, 23 samples to Herpes simplex virus type II and 29 samples had no antibodies to Herpes simplex virus). The diagnostic capacity of mixture of recombinant antigens gG1 Herpes simplex virus type I and gG2 Herpes simplex virus type II (The research-and-production complex "DiaprofMed") was comparable with mixture of lysate antigen Herpes simplex virus type I and II (Membrane) EIE Antigen ("Virion Ltd."). In the test-systems for differentiation of IgG to Herpes simplex virus type I the recombinant antigen gG1 Herpes simplex virus type I proved to be comparable with commercial analogue Herpes simplex virus-1 gG1M ("Viral Therapeutics Inc."'). At the same time, capacity to detect IgG to Herpes simplex virus type II in recombinant protein gG2 Herpes simplex virus type II is significantly higher than in its analogue Herpes simplex virus-2 gG2c ("Viral Therapeutics Inc.").

Optical fiber immunosensor based on a poly(pyrrole-benzophenone) film for the detection of antibodies to viral antigen.

PubMed

Konry, T; Novoa, A; Shemer-Avni, Y; Hanuka, N; Cosnier, S; Lepellec, Arielle; Marks, R S

2005-03-15

We describe herein a newly developed optical microbiosensor for the diagnosis of hepatitis C virus (HCV) by using a novel photoimmobilization methodology based on a photoactivable electrogenerated polymer film deposited upon surface-conductive fiber optics, which are then used to link a biological receptor to the fiber tip through light mediation. This fiber-optic electroconductive surface modification is done by the deposition of a thin layer of indium tin oxide on the silica surface of the fiber optics. Monomers are then electropolymerized onto the conductive metal oxide surface; thereafter, the fibers are immersed in a solution containing HCV-E2 envelope protein antigen and illuminated with UV light (wavelength approximately 345 nm). As a result of the photochemical reaction, a thin layer of the antigen becomes covalently bound to the benzophenone-modified surface. The photochemically modified fiber optics were tested as immunosensors for the detection of anti-E2 protein antibody analyte that was measured through chemiluminescence reaction. The biosensor was tested for sensitivity, specificity, and overall practicality. Our results suggest that the detection of anti-E2 antibodies with this microbiosensor may enhance significantly HCV serological standard testing especially among patients during dialysis, which were diagnosed as HCV negative, by standard immunological tests, but were known to carry the virus. If transformed into an easy to use procedure, this assay might be used in the future as an important clinical tool for HCV screening in blood banks.

Scaffolded Antigens in Yeast Cell Particle Vaccines Provide Protection against Systemic Polyoma Virus Infection.

PubMed

Tipper, Donald J; Szomolanyi-Tsuda, Eva

2016-01-01

Background. U65, a self-aggregating peptide scaffold, traps fused protein antigens in yeast cells. Conversion to Yeast Cell Particle (YCP) vaccines by partial removal of surface mannoproteins exposes β-glucan, mediating efficient uptake by antigen-presenting cells (APCs). YCP vaccines are inexpensive, capable of rapid large-scale production and have potential for both parenteral and oral use. Results. YCP processing by alkaline hydrolysis exposes up to 20% of the glucan but converts scaffolded antigen and internal yeast proteins into a common aggregate, preventing selective yeast protein removal. For U65-green fluorescent protein (GFP) or U65-Apolipoprotein A1 (ApoA1) subcutaneous vaccines, maximal IgG responses in mice required 10% glucan exposure. IgG responses to yeast proteins were 5-fold lower. Proteolytic mannoprotein removal produced YCPs with only 6% glucan exposure, insufficiently porous for selective removal of even native yeast proteins. Vaccine efficacy was reduced 10-fold. Current YCP formulations, therefore, are not suitable for human use but have considerable potential for use in feed animal vaccines. Significantly, a YCP vaccine expressing a GFP fusion to VP1, the murine polyoma virus major capsid protein, after either oral or subcutaneous administration, protected mice against an intraperitoneal polyoma virus challenge, reducing viral DNA levels in spleen and liver by >98%.

Immunogenic Subviral Particles Displaying Domain III of Dengue 2 Envelope Protein Vectored by Measles Virus

PubMed Central

Harahap-Carrillo, Indira S.; Ceballos-Olvera, Ivonne; Reyes-del Valle, Jorge

2015-01-01

Vaccines against dengue virus (DV) are commercially nonexistent. A subunit vaccination strategy may be of value, especially if a safe viral vector acts as biologically active adjuvant. In this paper, we focus on an immunoglobulin-like, independently folded domain III (DIII) from DV 2 envelope protein (E), which contains epitopes that elicits highly specific neutralizing antibodies. We modified the hepatitis B small surface antigen (HBsAg, S) in order to display DV 2 DIII on a virus-like particle (VLP), thus generating the hybrid antigen DIII-S. Two varieties of measles virus (MV) vectors were developed to express DIII-S. The first expresses the hybrid antigen from an additional transcription unit (ATU) and the second additionally expresses HBsAg from a separate ATU. We found that this second MV vectoring the hybrid VLPs displaying DIII-S on an unmodified HBsAg scaffold were immunogenic in MV-susceptible mice (HuCD46Ge-IFNarko), eliciting robust neutralizing responses (averages) against MV (1:1280 NT90), hepatitis B virus (787 mIU/mL), and DV2 (1:160 NT50) in all of the tested animals. Conversely, the MV vector expressing only DIII-S induced immunity against MV alone. In summary, DV2 neutralizing responses can be generated by displaying E DIII on a scaffold of HBsAg-based VLPs, vectored by MV. PMID:26350592

Immunodominance and Functional Activities of Antibody Responses to Inactivated West Nile Virus and Recombinant Subunit Vaccines in Mice▿

PubMed Central

Zlatkovic, Juergen; Stiasny, Karin; Heinz, Franz X.

2011-01-01

Factors controlling the dominance of antibody responses to specific sites in viruses and/or protein antigens are ill defined but can be of great importance for the induction of potent immune responses to vaccines. West Nile virus and other related important human-pathogenic flaviviruses display the major target of neutralizing antibodies, the E protein, in an icosahedral shell at the virion surface. Potent neutralizing antibodies were shown to react with the upper surface of domain III (DIII) of this protein. Using the West Nile virus system, we conducted a study on the immunodominance and functional quality of E-specific antibody responses after immunization of mice with soluble protein E (sE) and isolated DIII in comparison to those after immunization with inactivated whole virions. With both virion and sE, the neutralizing response was dominated by DIII-specific antibodies, but the functionality of these antibodies was almost four times higher after virion immunization. Antibodies induced by the isolated DIII had an at least 15-fold lower specific neutralizing activity than those induced by the virion, and only 50% of these antibodies were able to bind to virus particles. Our results suggest that immunization with the tightly packed E in virions focuses the DIII antibody response to the externally exposed sites of this domain which are the primary targets for virus neutralization, different from sE and isolated DIII, which also display protein surfaces that are cryptic in the virion. Despite its low potency for priming, DIII was an excellent boosting antigen, suggesting novel vaccination strategies that strengthen and focus the antibody response to critical neutralizing sites in DIII. PMID:21147919

[Herpes simplex virus and malignancies of female genital organs].

PubMed

Cokić-Damjanović, J; Horvat, E; Balog, A

2001-01-01

Primary herpes simplex virus (HSV) infections of female genital tract usually end with remission, while the virus remains in the organism--almost in the sacral ganglion in a latent form, protected from humoral and cellular immunity. Stress induces the virus and the result is recurrent genital infection. Frequent exacerbations damage some parts of vital cellular structures without cytolysis, but stimulate malignant transformations. Vulvar (portio vaginalis uteri) and endometrial tumor tissue samples were analyzed for HSV by direct and indirect fluorescent antibody technique (FAT). Pre and postoperative sera samples were analyzed for presence of anti-HSV antibodies--IgM and IgG by Elisa-Enzygnost method. Acellular filtrates obtained by ultrasonic destruction of malignant tissues were used as inoculum for rabbit corneal scarification. Out of 63 tissue samples, 42 were positive for HSV antigen i.e. 67.3%. According to location 50% of vulvar, 76% PVU and 65% of endometrial tissues were positive. This antigen induces production of virus specific antibodies. Two types of antigens are known: the so-called T-antigen persisting in the cell nucleus and cell-surface antigen--product of the viral genome and can be evidenced by immunofluorescence method. Anti HSV antibodies were present in 63 preoperative serum samples and belonged to IgG group, but not one to IgM, implying a long and chronic course of infection excluding acute primary. Out of 38 postoperative serums the titer of antibodies decreased in 36 evidently, but in two samples remained unchanged. Two samples of endometrial and one from PVU origin contained HSV antigen type one. In the remaining 16 samples HSV 2 antigen was present. Rabbit corneal scarification was the proof of complete infectious virus in malignant tissues. Acellular filtrate of malignant tissues served as inoculum. Corneas of examined rabbits showed a mild inflammation after 24 hours which disappeared in the next 24 hours. We could not isolate the infectious virus by rabbit corneal scarification. Instead of herpetic changes, mild inflammation was evident. This abortive, incomplete symptomatology was probably caused by nonstructural early protein, which is a product of viral genome incorporated in malignant cells. On the basis of our results, we can conclude that HSV can have, beside other factors, a very important, maybe an initial role in development of malignant changes of female genital tract, not only on vulva and PVU, but on endometrium as well. HSV I can cause genital infections and have some role in malignant changes as well as HSV 2. However, complete infective virion couldn't be isolated from malignant tissues.

Intranasal delivery of Norwalk virus-like particles formulated in an in-situ gelling, dry powder vaccine

PubMed Central

Velasquez, Lissette S.; Shira, Samantha; Berta, Alice N.; Kilbourne, Jacquelyn; Medi, Babu M.; Tizard, Ian; Ni, Yawei; Arntzen, Charles J.; Herbst-Kralovetz, Melissa M.

2011-01-01

The development of a vaccine to prevent norovirus infections has been focused on immunization at a mucosal surface, but has been limited by the low immunogenicity of self-assembling Norwalk virus-like particles (NV VLPs) delivered enterically or at nasal surfaces. Nasal immunization, which offers the advantage of ease of immunization, faces obstacles imposed by the normal process of mucociliary clearance, which limits residence time of applied antigens. Herein, we describe the use of a dry powder formulation (GelVac) of an inert in-situ gelling polysaccharide (GelSite) extracted from Aloe vera for nasal delivery of NV VLP antigen. Powder formulations, with or without NV VLP antigen, were similar in structure in dry form or when rehydrated in simulated nasal fluids. Immunogenicity of the dry powder VLP formulation was compared to equivalent antigen/adjuvant liquid formulations in animals. For the GelVac powder, we observed superior NV-specific serum and mucosal (aerodigestive and reproductive tracts) antibody responses relative to liquid formulations. Incorporation of TLR7 agonist gardiquimod in dry powder formulations did not enhance antibody responses, although its inclusion in liquid formulations did enhance VLP immunogenicity irrespective of the presence or absence of GelSite. We interpret these data as showing that GelSite-based dry powder formulations 1.) stabilize the immunogenic structural properties of VLPs and 2.) induce systemic and mucosal antibody titers which are equal or greater than those achieved by VLPs plus adjuvant in a liquid formulation. We conclude that in-situ gelation of the GelVac dry powder formulation at nasal mucosal surfaces delays mucociliary clearance and thereby prolongs VLP antigen exposure to immune effector sites. PMID:21640778

Viral vector-based influenza vaccines

PubMed Central

de Vries, Rory D.; Rimmelzwaan, Guus F.

2016-01-01

ABSTRACT Antigenic drift of seasonal influenza viruses and the occasional introduction of influenza viruses of novel subtypes into the human population complicate the timely production of effective vaccines that antigenically match the virus strains that cause epidemic or pandemic outbreaks. The development of game-changing vaccines that induce broadly protective immunity against a wide variety of influenza viruses is an unmet need, in which recombinant viral vectors may provide. Use of viral vectors allows the delivery of any influenza virus antigen, or derivative thereof, to the immune system, resulting in the optimal induction of virus-specific B- and T-cell responses against this antigen of choice. This systematic review discusses results obtained with vectored influenza virus vaccines and advantages and disadvantages of the currently available viral vectors. PMID:27455345

Viral vector-based influenza vaccines.

PubMed

de Vries, Rory D; Rimmelzwaan, Guus F

2016-11-01

Antigenic drift of seasonal influenza viruses and the occasional introduction of influenza viruses of novel subtypes into the human population complicate the timely production of effective vaccines that antigenically match the virus strains that cause epidemic or pandemic outbreaks. The development of game-changing vaccines that induce broadly protective immunity against a wide variety of influenza viruses is an unmet need, in which recombinant viral vectors may provide. Use of viral vectors allows the delivery of any influenza virus antigen, or derivative thereof, to the immune system, resulting in the optimal induction of virus-specific B- and T-cell responses against this antigen of choice. This systematic review discusses results obtained with vectored influenza virus vaccines and advantages and disadvantages of the currently available viral vectors.

The Missing Link in Epstein-Barr Virus Immune Evasion: the BDLF3 Gene Induces Ubiquitination and Downregulation of Major Histocompatibility Complex Class I (MHC-I) and MHC-II

PubMed Central

Quinn, Laura L.; Williams, Luke R.; White, Claire; Forrest, Calum; Rowe, Martin

2015-01-01

ABSTRACT The ability of Epstein-Barr virus (EBV) to spread and persist in human populations relies on a balance between host immune responses and EBV immune evasion. CD8+ cells specific for EBV late lytic cycle antigens show poor recognition of target cells compared to immediate early and early antigen-specific CD8+ cells. This phenomenon is due in part to the early EBV protein BILF1, whose immunosuppressive activity increases with lytic cycle progression. However, published data suggest the existence of a hitherto unidentified immune evasion protein further enhancing protection against late EBV antigen-specific CD8+ cells. We have now identified the late lytic BDLF3 gene as the missing link accounting for efficient evasion during the late lytic cycle. Interestingly, BDLF3 also contributes to evasion of CD4+ cell responses to EBV. We report that BDLF3 downregulates expression of surface major histocompatibility complex (MHC) class I and class II molecules in the absence of any effect upon other surface molecules screened, including CD54 (ICAM-1) and CD71 (transferrin receptor). BDLF3 both enhanced internalization of surface MHC molecules and reduced the rate of their appearance at the cell surface. The reduced expression of surface MHC molecules correlated with functional protection against CD8+ and CD4+ T cell recognition. The molecular mechanism was identified as BDLF3-induced ubiquitination of MHC molecules and their subsequent downregulation in a proteasome-dependent manner. IMPORTANCE Immune evasion is a necessary feature of viruses that establish lifelong persistent infections in the face of strong immune responses. EBV is an important human pathogen whose immune evasion mechanisms are only partly understood. Of the EBV immune evasion mechanisms identified to date, none could explain why CD8+ T cell responses to late lytic cycle genes are so infrequent and, when present, recognize lytically infected target cells so poorly relative to CD8+ T cells specific for early lytic cycle antigens. The present work identifies an additional immune evasion protein, BDLF3, that is expressed late in the lytic cycle and impairs CD8+ T cell recognition by targeting cell surface MHC class I molecules for ubiquitination and proteasome-dependent downregulation. Interestingly, BDLF3 also targets MHC class II molecules to impair CD4+ T cell recognition. BDLF3 is therefore a rare example of a viral protein that impairs both the MHC class I and class II antigen-presenting pathways. PMID:26468525

The Missing Link in Epstein-Barr Virus Immune Evasion: the BDLF3 Gene Induces Ubiquitination and Downregulation of Major Histocompatibility Complex Class I (MHC-I) and MHC-II.

PubMed

Quinn, Laura L; Williams, Luke R; White, Claire; Forrest, Calum; Zuo, Jianmin; Rowe, Martin

2016-01-01

The ability of Epstein-Barr virus (EBV) to spread and persist in human populations relies on a balance between host immune responses and EBV immune evasion. CD8(+) cells specific for EBV late lytic cycle antigens show poor recognition of target cells compared to immediate early and early antigen-specific CD8(+) cells. This phenomenon is due in part to the early EBV protein BILF1, whose immunosuppressive activity increases with lytic cycle progression. However, published data suggest the existence of a hitherto unidentified immune evasion protein further enhancing protection against late EBV antigen-specific CD8(+) cells. We have now identified the late lytic BDLF3 gene as the missing link accounting for efficient evasion during the late lytic cycle. Interestingly, BDLF3 also contributes to evasion of CD4(+) cell responses to EBV. We report that BDLF3 downregulates expression of surface major histocompatibility complex (MHC) class I and class II molecules in the absence of any effect upon other surface molecules screened, including CD54 (ICAM-1) and CD71 (transferrin receptor). BDLF3 both enhanced internalization of surface MHC molecules and reduced the rate of their appearance at the cell surface. The reduced expression of surface MHC molecules correlated with functional protection against CD8(+) and CD4(+) T cell recognition. The molecular mechanism was identified as BDLF3-induced ubiquitination of MHC molecules and their subsequent downregulation in a proteasome-dependent manner. Immune evasion is a necessary feature of viruses that establish lifelong persistent infections in the face of strong immune responses. EBV is an important human pathogen whose immune evasion mechanisms are only partly understood. Of the EBV immune evasion mechanisms identified to date, none could explain why CD8(+) T cell responses to late lytic cycle genes are so infrequent and, when present, recognize lytically infected target cells so poorly relative to CD8(+) T cells specific for early lytic cycle antigens. The present work identifies an additional immune evasion protein, BDLF3, that is expressed late in the lytic cycle and impairs CD8(+) T cell recognition by targeting cell surface MHC class I molecules for ubiquitination and proteasome-dependent downregulation. Interestingly, BDLF3 also targets MHC class II molecules to impair CD4(+) T cell recognition. BDLF3 is therefore a rare example of a viral protein that impairs both the MHC class I and class II antigen-presenting pathways. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Antigenic Patterns and Evolution of the Human Influenza A (H1N1) Virus

PubMed Central

Liu, Mi; Zhao, Xiang; Hua, Sha; Du, Xiangjun; Peng, Yousong; Li, Xiyan; Lan, Yu; Wang, Dayan; Wu, Aiping; Shu, Yuelong; Jiang, Taijiao

2015-01-01

The influenza A (H1N1) virus causes seasonal epidemics that result in severe illnesses and deaths almost every year. A deep understanding of the antigenic patterns and evolution of human influenza A (H1N1) virus is extremely important for its effective surveillance and prevention. Through development of antigenicity inference method for human influenza A (H1N1), named PREDAC-H1, we systematically mapped the antigenic patterns and evolution of the human influenza A (H1N1) virus. Eight dominant antigenic clusters have been inferred for seasonal H1N1 viruses since 1977, which demonstrated sequential replacements over time with a similar pattern in Asia, Europe and North America. Among them, six clusters emerged first in Asia. As for China, three of the eight antigenic clusters were detected in South China earlier than in North China, indicating the leading role of South China in H1N1 transmission. The comprehensive view of the antigenic evolution of human influenza A (H1N1) virus can help formulate better strategy for its prevention and control. PMID:26412348

Antigenic Patterns and Evolution of the Human Influenza A (H1N1) Virus.

PubMed

Liu, Mi; Zhao, Xiang; Hua, Sha; Du, Xiangjun; Peng, Yousong; Li, Xiyan; Lan, Yu; Wang, Dayan; Wu, Aiping; Shu, Yuelong; Jiang, Taijiao

2015-09-28

The influenza A (H1N1) virus causes seasonal epidemics that result in severe illnesses and deaths almost every year. A deep understanding of the antigenic patterns and evolution of human influenza A (H1N1) virus is extremely important for its effective surveillance and prevention. Through development of antigenicity inference method for human influenza A (H1N1), named PREDAC-H1, we systematically mapped the antigenic patterns and evolution of the human influenza A (H1N1) virus. Eight dominant antigenic clusters have been inferred for seasonal H1N1 viruses since 1977, which demonstrated sequential replacements over time with a similar pattern in Asia, Europe and North America. Among them, six clusters emerged first in Asia. As for China, three of the eight antigenic clusters were detected in South China earlier than in North China, indicating the leading role of South China in H1N1 transmission. The comprehensive view of the antigenic evolution of human influenza A (H1N1) virus can help formulate better strategy for its prevention and control.

Integrating influenza antigenic dynamics with molecular evolution

PubMed Central

Bedford, Trevor; Suchard, Marc A; Lemey, Philippe; Dudas, Gytis; Gregory, Victoria; Hay, Alan J; McCauley, John W; Russell, Colin A; Smith, Derek J; Rambaut, Andrew

2014-01-01

Influenza viruses undergo continual antigenic evolution allowing mutant viruses to evade host immunity acquired to previous virus strains. Antigenic phenotype is often assessed through pairwise measurement of cross-reactivity between influenza strains using the hemagglutination inhibition (HI) assay. Here, we extend previous approaches to antigenic cartography, and simultaneously characterize antigenic and genetic evolution by modeling the diffusion of antigenic phenotype over a shared virus phylogeny. Using HI data from influenza lineages A/H3N2, A/H1N1, B/Victoria and B/Yamagata, we determine patterns of antigenic drift across viral lineages, showing that A/H3N2 evolves faster and in a more punctuated fashion than other influenza lineages. We also show that year-to-year antigenic drift appears to drive incidence patterns within each influenza lineage. This work makes possible substantial future advances in investigating the dynamics of influenza and other antigenically-variable pathogens by providing a model that intimately combines molecular and antigenic evolution. DOI: http://dx.doi.org/10.7554/eLife.01914.001 PMID:24497547

Relationship Among Tau Antigens Isolated from Various Lines of Simian Virus 40-Transformed Cells

PubMed Central

Simmons, Daniel T.; Martin, Malcolm A.; Mora, Peter T.; Chang, Chungming

1980-01-01

In addition to the virus-specified tumor antigens, simian virus 40-transformed cells contain at least one other protein which can be immunoprecipitated with serum from animals bearing simian virus 40-induced tumors. This protein, which is designated Tau antigen, has an apparent molecular weight of 56,000 as determined by electrophoresis on acrylamide gels. The relationship among Tau antigens isolated from different lines of simian virus 40-transformed cells was examined by comparing the methionine-labeled tryptic peptides of these proteins by two-dimensional fingerprinting on thin-layer cellulose plates. In this fashion, we initially determined that the Tau antigens isolated from three different lines of transformed mouse cells were very similar. Second, we found that Tau antigen isolated from a line of rat transformants was closely related, but not identical, to the mouse cell Tau antigens. Approximately 70% of their methionine peptides comigrated in two dimensions. Finally, we showed that Tau antigen isolated from a line of transformed human cells was only partially related to the mouse and rat proteins. About 40% of the methionine peptides of the human protein were also contained in the Tau antigens from the other two species. These results strongly indicate that the Tau antigens isolated from these various simian virus 40-transformed cell lines contain common amino acid sequences. Images PMID:6247503

Routine screening of blood donations at Qingdao central blood bank, China, for hepatitis B virus (HBV) DNA with a real-time, multiplex nucleic acid test for HBV, hepatitis C virus, and human immunodeficiency virus Types 1 and 2.

PubMed

Yang, Zhongsi; Xu, Lei; Liu, Li; Feng, Qiuxia; Zhang, Longmu; Ma, Weijuan; Saldanha, John; Wang, Mingmin; Zhao, Lin

2013-10-01

The Roche cobas TaqScreen MPX test was used to evaluate the rate of hepatitis B surface antigen (HBsAg)-negative donations that were hepatitis B virus (HBV) DNA reactive from June 2010 to January 2011 in Qingdao, China. HBsAg-negative samples from 65,800 voluntary blood donors were tested with the cobas TaqScreen MPX test in pools of 6 on the Roche cobas s 201 blood screening platform. Samples positive for HBV DNA and negative for HBsAg were quantitated with the Roche COBAS AmpliPrep/COBAS TaqMan HBV test. In addition, serologic tests for HBsAg, hepatitis B surface antibody, anti-hepatitis B core antigen (anti-HBc), anti-hepatitis B e antigen (anti-HBe), and hepatitis B e antigen (HBe) were done using the Roche electrochemiluminescence immunoassay. A total of 80 nucleic acid amplification technology (NAT) test-reactive pools were identified and 59 pools (74%) resolved to a reactive sample. All samples were HBV DNA reactive and the viral load in each sample was quantitated. The viral loads of the samples ranged from less than 20 to 34,600 IU/mL; 13 samples (22%) had viral loads of more than 20 IU/mL, 27 samples (45.8%) had viral loads of less than 20 IU/mL, and 19 samples (32.2%) had undetectable viral loads. Of the 59 NAT-reactive samples, 40 (67.8%) were anti-HBc positive. Fifteen of the 59 samples could not be confirmed as NAT reactive either by an alternative NAT test or by serology. The HBV NAT yield in blood donors in Qingdao is 0.06% (38/65,800). This study confirmed the value of NAT for interdicting HBV-positive donations and preventing transfusion-transmitted HBV infections. © 2013 American Association of Blood Banks.

Candidate Medical Countermeasures Targeting Ebola Virus Cell Entry

DTIC Science & Technology

2017-04-03

interface, rather than as expected to the more exposed surface of the 134 GP1,2 trimer [16]. Importantly, KZ52 protected guinea pigs (Cavia porcellus...from death after 135 inoculation with guinea pig -adapted EBOV [64], but failed to have a beneficial effect on EBOV-136 exposed rhesus monkeys... guinea pigs infected with 145 rodent-adapted EBOV or its antigenically distant relative, Sudan virus (SUDV) [68]. 146 Identification of ebolavirus

Candidate Medical Countermeasures Targeting Ebola Virus Cell Entry

DTIC Science & Technology

2017-03-31

interface, rather than as expected to the more exposed surface of the 134 GP1,2 trimer [16]. Importantly, KZ52 protected guinea pigs (Cavia porcellus...from death after 135 inoculation with guinea pig -adapted EBOV [64], but failed to have a beneficial effect on EBOV-136 exposed rhesus monkeys... guinea pigs infected with 145 rodent-adapted EBOV or its antigenically distant relative, Sudan virus (SUDV) [68]. 146 Identification of ebolavirus

THE LYMPHOCYTE RESPONSE TO PRIMARY MOLONEY SARCOMA VIRUS TUMORS IN BALB/c MICE

PubMed Central

Lamon, E. W.; Wigzell, H.; Klein, E.; Andersson, B.; Skurzak, H. M.

1973-01-01

Adult BALB/c mice were injected with Moloney sarcoma virus (MSV) after which the animals' lymphocytes were examined for activity against Moloney leukemia virus (MLV) antigen-bearing target cells at 5-day intervals for 30 days. Lymphocytes from these animals and appropriately matched controls were fractionated into B cell-deficient (primarily T cells) and T cell-deficient (primarily B cells) subpopulations. Macrophages were removed using iron powder and magnetism. The unfractionated lymphocytes, T cells, and non-T cells were then tested in microcytotoxicity tests. Antigen-specific activity was found in the unfractionated lymphocytes from animals that had not yet developed palpable tumors and from regressor animals. The T cells were active just before tumor development and just after regression; however, by day 30 after virus infection (8–10 days after regression) the T cell subpopulation was much less active. The non-T cell subpopulation was also active before tumor development and soon after regression. However, this activity continued to rise after regression and was highest at 30 days. At day 15 (peak tumor size) neither subpopulation was active. The activity was demonstrated to be specific for the MLV-determined cell surface antigen by testing on control target cells that were MLV antigen negative and by comparison of the inhibitory effects with lymphocytes immune to a nonpertinent antigen as well as normal lymphocytes. The non-T cells were tested for activity before and after removal of macrophages with iron powder and magnetism. Such cells were significantly more active after removal of the macrophages. These data demonstrate specific T cell and non-T cell activity in microcytotoxicity tests with a tumor-specific system and strongly suggest that the non-T cell activity described herein is a B cell function. PMID:4709269

Genotypic evolution and antigenicity of H9N2 influenza viruses in Shanghai, China.

PubMed

Ge, Feifei; Li, Xin; Ju, Houbin; Yang, Dequan; Liu, Jian; Qi, Xinyong; Wang, Jian; Yang, Xianchao; Qiu, Yafeng; Liu, Peihong; Zhou, Jinping

2016-06-01

H9N2 influenza viruses have been circulating in China since 1994, but a systematic investigation of H9N2 in Shanghai has not previously been undertaken. Here, using 14 viruses we isolated from poultry and pigs in Shanghai during 2002 and 2006-2014, together with the commercial vaccine A/chicken/Shanghai/F/1998 (Ck/SH/F/98), we analyzed the evolution of H9N2 influenza viruses in Shanghai and showed that all 14 isolates originated from Ck/SH/F/98 antigenically. We evaluated the immune protection efficiency of the vaccine. Our findings demonstrate that H9N2 viruses in Shanghai have undergone extensive reassortment. Various genotypes emerged in 2002, 2006 and 2007, while during 2009-2014 only one genotype was found. Four antigenic groups, A-D, could be identified among the 14 isolates and a variety of antigenically distinct H9N2-virus-derived avian influenza viruses (AIVs) circulated simultaneously in Shanghai during this period. Challenge experiments using vaccinated chickens indicated that the vaccine prevented shedding of antigenic group A and B viruses, but not those of the more recent groups C and D. Genetic analysis showed that compared to the vaccine strain, representative viruses of antigenic groups C and D possess greater numbers of amino acid substitutions in the hemagglutinin (HA) protein than viruses in antigenic groups A and B. Many of these substitutions are located in antigenic sites. Our results indicate that the persistence of H9N2 AIV in China might be due to incomplete vaccine protection and that the avian influenza vaccine should be regularly evaluated and updated to maintain optimal protection.

Structural basis for immunization with postfusion respiratory syncytial virus fusion F glycoprotein (RSV F) to elicit high neutralizing antibody titers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Swanson, Kurt A.; Settembre, Ethan C.; Shaw, Christine A.

2012-02-07

Respiratory syncytial virus (RSV), the main cause of infant bronchiolitis, remains a major unmet vaccine need despite more than 40 years of vaccine research. Vaccine candidates based on a chief RSV neutralization antigen, the fusion (F) glycoprotein, have foundered due to problems with stability, purity, reproducibility, and potency. Crystal structures of related parainfluenza F glycoproteins have revealed a large conformational change between the prefusion and postfusion states, suggesting that postfusion F antigens might not efficiently elicit neutralizing antibodies. We have generated a homogeneous, stable, and reproducible postfusion RSV F immunogen that elicits high titers of neutralizing antibodies in immunized animals.more » The 3.2-{angstrom} X-ray crystal structure of this substantially complete RSV F reveals important differences from homology-based structural models. Specifically, the RSV F crystal structure demonstrates the exposure of key neutralizing antibody binding sites on the surface of the postfusion RSV F trimer. This unanticipated structural feature explains the engineered RSV F antigen's efficiency as an immunogen. This work illustrates how structural-based antigen design can guide the rational optimization of candidate vaccine antigens.« less

Sequential detection of different antigens induced by Epstein-Barr virus and herpes simplex virus in the same Western blot by using dual antibody probes.

PubMed

Lin, J C; Pagano, J S

1986-08-01

A dual antibody probing technique that permitted a color-coded identification of polypeptides representing different classes of Epstein-Barr virus (EBV) antigens as well as differentiation of the polypeptides induced by different herpesviruses in the same Western blot was developed. When the nitrocellulose sheet was probed first with monoclonal antibody against EBV early antigen diffuse component (EA-D) and then stained with 4-chloro-1-naphthol, four polypeptides specific for EA-D were identified by purple bands. Subsequently, the same nitrocellulose sheet was reprobed with human serum containing antibodies against EBV early antigen, viral capsid antigen, and nuclear antigen and stained with 3,3'-diaminobenzidine. Several brown bands corresponding to early, viral capsid, and nuclear antigen polypeptides were detected. The dual antibody probing technique was used in an analysis to differentiate polypeptides resulting from either EBV or herpes simplex virus infection, either in cells infected by individual virus or in a cell line dually infected by both viruses. On the basis of different colored bands in different lanes of the same gel, 20 polypeptides with molecular weights ranging from 31,000 to 165,000 were identified as herpes simplex virus-specific proteins. These results suggested that the dual antibody probing technique may be applicable in clinical diagnosis for detecting antigens and antibodies derived from different pathogens.

World Reference Center for Arboviruses.

DTIC Science & Technology

1983-01-01

fever virus for antibody detection Antigen- detection ELISA was applied to dengue, and in another study o eastern encephalitis and Highland J viruses for...relatedness of RNA viruses ............................................. 60 Detection of eastern equine encephalitis virus and Highland J virus antigens...narrow. This suggests that Palyam group viruses are genetically quite stable. Virus identification. New viruses were identified in the Kemerovo group

Biological and immunological characterization of recombinant Yellow Fever 17D viruses expressing a Trypanosoma cruzi Amastigote Surface Protein-2 CD8+ T cell epitope at two distinct regions of the genome.

PubMed

Nogueira, Raquel T; Nogueira, Alanderson R; Pereira, Mirian C S; Rodrigues, Maurício M; Galler, Ricardo; Bonaldo, Myrna C

2011-03-18

The attenuated Yellow fever (YF) 17D vaccine virus is one of the safest and most effective viral vaccines administered to humans, in which it elicits a polyvalent immune response. Herein, we used the YF 17D backbone to express a Trypanosoma cruzi CD8+ T cell epitope from the Amastigote Surface Protein 2 (ASP-2) to provide further evidence for the potential of this virus to express foreign epitopes. The TEWETGQI CD8+ T cell epitope was cloned and expressed based on two different genomic insertion sites: in the fg loop of the viral Envelope protein and the protease cleavage site between the NS2B and NS3. We investigated whether the site of expression had any influence on immunogenicity of this model epitope. Recombinant viruses replicated similarly to vaccine virus YF 17D in cell culture and remained genetically stable after several serial passages in Vero cells. Immunogenicity studies revealed that both recombinant viruses elicited neutralizing antibodies to the YF virus as well as generated an antigen-specific gamma interferon mediated T-cell response in immunized mice. The recombinant viruses displayed a more attenuated phenotype than the YF 17DD vaccine counterpart in mice. Vaccination of a mouse lineage highly susceptible to infection by T. cruzi with a homologous prime-boost regimen of recombinant YF viruses elicited TEWETGQI specific CD8+ T cells which might be correlated with a delay in mouse mortality after a challenge with a lethal dose of T. cruzi. We conclude that the YF 17D platform is useful to express T. cruzi (Protozoan) antigens at different functional regions of its genome with minimal reduction of vector fitness. In addition, the model T. cruzi epitope expressed at different regions of the YF 17D genome elicited a similar T cell-based immune response, suggesting that both expression sites are useful. However, the epitope as such is not protective and it remains to be seen whether expression of larger domains of ASP-2, which include the TEWETGQI epitope, will elicit better T-CD8+ responses to the latter. It is likely that additional antigens and recombinant virus formulations will be necessary to generate a protective response.

Characterization of the size distribution and aggregation of virus-like nanoparticles used as active ingredients of the HeberNasvac therapeutic vaccine against chronic hepatitis B

NASA Astrophysics Data System (ADS)

Lopez, Matilde; Rodriguez, Elias Nelson; Lobaina, Yadira; Musacchio, Alexis; Falcon, Viviana; Guillen, Gerardo; Aguilar, Julio C.

2017-06-01

The use of virus-like particles (VLPs) as antigens constitutes a well established strategy in preventive vaccination. These non-infective particles have a composition, size, and structure favoring their interaction and processing by the immune system. Recombinant viral nucleocapsids encapsulating bacterial nucleic acids result in potent Th1-driving immunogens. Several antigens have been coadministered with VLPs or conjugated to them to further increase their immunogenicity. In the present work we characterize the size distribution of two different recombinant VLPs obtained as components of HeberNasvac, a novel therapeutic vaccine recently registered to treat chronic hepatitis B. The vaccine ingredients, hepatitis B virus surface and nucleocapsid antigens (HBsAg and HBcAg, respectively) and the vaccine formulation, were evaluated using dynamic light scattering (DLS), transmission electron microscopy (TEM) and light obscuration technology. The results demonstrate that both antigens are nanoparticles with sizes ranging between 20-30 nm, in line with reports in the literature. In addition, DLS studies evidenced the capacity of both antigens to form homologous and heterologous aggregates, both as active ingredients as well as being part of the final product. The evaluation of subvisible particles in HeberNasvac formulation fulfills the requirements in terms of quantity and size established for parenteral pharmaceutical compositions. Invited talk at 8th Int. Workshop on Advanced Materials Science and Nanotechnology (IWAMSN2016) (Ha Long City, Vietnam, 8-12 November 2016)

[Study of the virus carrier state in chicken influenza].

PubMed

Smolenskiĭ, V I; Osidze, N G; Bogautdinov, Z F; Panteleev, Iu V; Siurin, V N

1978-01-01

The problems of virus carrier state in influenza are connected with two aspects of the disease: the duration of virus antigen persistence in convalescents and changes of influenza virus properties in the course of persistence. In the present study, natural influenza infection in chickens caused by influenza A/chicken/USSR/336/74 virus (Hav6H3--N2) was used to determine the duration of virus antigen persistence (up to 60 days) and the entire period of virus isolation from the survivers (up to 30 days). Administration of hydrocortisone on the 50th day of convalescence permitted to obtain from the chickens several influenza A virus isolates antigenically unrelated to the epizootic strain either in hemagglutinin or in neuraminidase. Cultivation of isolate No. 42 (Hav7Neq1) in the presence of the homologous serum yielded strain 42' which was neutralized by the serum to Hav6H3--N2 virus. The isolates differed from the epizootic virus by their biological properties: the eluting activity, pathogenesis and morphology. The above facts of antigenic variability are considered in the light of the antigenic heterogeneity of the natural virus population and the possibility of virus activation by the provoking effect of extreme conditions on the carriers of latent infection.

Detection and Characterization of Clade 1 Reassortant H5N1 Viruses Isolated from Human Cases in Vietnam during 2013

PubMed Central

Balish, Amanda; Hoang, Anh Nguyen; Gustin, Kortney M.; Nhung, Pham Thi; Jones, Joyce; Thu, Ngoc Nguyen; Davis, William; Ngoc, Thao Nguyen Thi; Jang, Yunho; Sleeman, Katrina; Villanueva, Julie; Kile, James; Gubareva, Larisa V.; Lindstrom, Stephen; Tumpey, Terrence M.; Davis, C. Todd; Long, Nguyen Thanh

2015-01-01

Highly pathogenic avian influenza (HPAI) H5N1 is endemic in Vietnamese poultry and has caused sporadic human infection in Vietnam since 2003. Human infections with HPAI H5N1 are of concern due to a high mortality rate and the potential for the emergence of pandemic viruses with sustained human-to-human transmission. Viruses isolated from humans in southern Vietnam have been classified as clade 1 with a single genome constellation (VN3) since their earliest detection in 2003. This is consistent with detection of this clade/genotype in poultry viruses endemic to the Mekong River Delta and surrounding regions. Comparison of H5N1 viruses detected in humans from southern Vietnamese provinces during 2012 and 2013 revealed the emergence of a 2013 reassortant virus with clade 1.1.2 hemagglutinin (HA) and neuraminidase (NA) surface protein genes but internal genes derived from clade 2.3.2.1a viruses (A/Hubei/1/2010-like; VN12). Closer analysis revealed mutations in multiple genes of this novel genotype (referred to as VN49) previously associated with increased virulence in animal models and other markers of adaptation to mammalian hosts. Despite the changes identified between the 2012 and 2013 genotypes analyzed, their virulence in a ferret model was similar. Antigenically, the 2013 viruses were less cross-reactive with ferret antiserum produced to the clade 1 progenitor virus, A/Vietnam/1203/2004, but reacted with antiserum produced against a new clade 1.1.2 WHO candidate vaccine virus (A/Cambodia/W0526301/2012) with comparable hemagglutination inhibition titers as the homologous antigen. Together, these results indicate changes to both surface and internal protein genes of H5N1 viruses circulating in southern Vietnam compared to 2012 and earlier viruses. PMID:26244768

An immuno-biosensor system based on quartz crystal microbalance for avian influenza virus detection

NASA Astrophysics Data System (ADS)

Liu, Shengping; Chen, Guoming; Zhou, Qi; Wei, Yunlong

2007-12-01

For the quick detection of Avian Influenza Virus (AIV), a biosensor based on Quartz Crystal Microbalance (QCM) was fabricated according to the specific bonding principle between antibody and antigen. Staphylococcal Protein A (SPA) was extracted from Staphylococcus and purified. Then SPA was coated on the surface of QCM for immobilizing AIV monoclonal antibodies. The use of AIV monoclonal antibody could enhance the specificity of the immuno-biosensor. A multi-channel piezoelectricity detection system for the immuno-biosensor was developed. The system can work for the quick detection of AIV antigen in the case of the entirely aqueous status owe to one special oscillating circuit designed in this work. The optimum conditions of SPA coating and AIV monoclonal antibody immobilization were investigated utilizing the multi-channel detection system. The preliminary application of the immuno-biosensor system for detection of AIV was evaluated. Results indicate that the immuno-biosensor system can detect the AIV antigens with a linear range of 3-200ng/ml. The system can accomplish the detection of AIV antigens around 40 minutes.

Continuing challenges in influenza

PubMed Central

Webster, Robert G.; Govorkova, Elena A.

2014-01-01

Influenza is an acute respiratory disease in mammals and domestic poultry that emerges from zoonotic reservoirs in aquatic birds and bats. Although influenza viruses are among the most intensively studied pathogens, existing control options require further improvement. Influenza vaccines must be regularly updated because of continuous antigenic drift and sporadic antigenic shifts in the viral surface glycoproteins. Currently, influenza therapeutics are limited to neuraminidase inhibitors; novel drugs and vaccine approaches are therefore urgently needed. Advances in vaccinology and structural analysis have revealed common antigenic epitopes on hemagglutinins across all influenza viruses and suggest that a universal influenza vaccine is possible. In addition, various immunomodulatory agents and signaling pathway inhibitors are undergoing preclinical development. Continuing challenges in influenza include the emergence of pandemic H1N1 influenza in 2009, human infections with avian H7N9 influenza in 2013, and sporadic human cases of highly pathogenic avian H5N1 influenza. Here, we review the challenges facing influenza scientists and veterinary and human public health officials; we also discuss the exciting possibility of achieving the ultimate goal of controlling influenza’s ability to change its antigenicity. PMID:24891213

Comparison of the Abbott Architect i2000 assay, the Roche Modular Analytics E170 assay, and an immunoradiometric assay for serum hepatitis B virus markers.

PubMed

Kim, Hyunjung; Oh, Eun-Jee; Kang, Mi-Sook; Kim, Sung Hoon; Park, Yeon-Joon

2007-01-01

Serum hepatitis B virus (HBV) markers are the most important data for epidemiological screening and clinical diagnosis of HBV infection, especially in endemic areas. We compared the results of the Roche Modular Analytics E170 assay, the Abbott Architect i2000 assay, and an immunoradiometric assay (IRMA) for HBV surface antigen (HBsAg), anti-HBV surface antigen (anti-HBs), HBV e antigen (HBeAg), and anti-HBV e antigen (anti-HBe). A number of serum samples (264, 263, 224, and 202 for HBsAg, anti-HBs, HBeAg, and anti-HBe, respectively) were studied. For samples giving discrepant results for HBeAg between methods, real-time PCR assays were performed. The concordance rates among the three methods were high for HBsAg (100%) and HBeAg (94.6), but low for anti-HBs (91.6%) and anti-HBe (82.2%). For anti-HBs, which could be measured quantitatively by the Modular E170 and Architect i2000 procedures, discrepant results were observed at low levels of anti-HBs. For anti-HBe, the positive rate was highest with Modular E170 (60.9%) followed by the IRMA kit (54.1%) and Architect i2000 (51.0%). This study shows substantial differences between the assay results by the three methods, which should be taken into account in determinations of serum HBV markers.

Antigenic diversity of H5 highly pathogenic avian influenza viruses of clade 2.3.4.4 isolated in Asia.

PubMed

Ohkawara, Ayako; Okamatsu, Masatoshi; Ozawa, Makoto; Chu, Duc-Huy; Nguyen, Lam Thanh; Hiono, Takahiro; Matsuno, Keita; Kida, Hiroshi; Sakoda, Yoshihiro

2017-05-01

H5 highly pathogenic avian influenza viruses (HPAIV) have spread in both poultry and wild birds since late 2003. Continued circulation of HPAIV in poultry in several regions of the world has led to antigenic drift. In the present study, we analyzed the antigenic properties of H5 HPAIV isolated in Asia using four neutralizing mAbs recognizing hemagglutinin, which were established using A/chicken/Kumamoto/1-7/2014 (H5N8), belonging to clade 2.3.4.4 and also using polyclonal antibodies. Viruses of clades 1.1, 2.3.2.1, 2.3.4, and 2.3.4.4 had different reactivity patterns to the panel of mAbs, thereby indicating that the antigenicity of the viruses of clade 2.3.4.4 were similar but differed from the other clades. In particular, the antigenicity of the viruses of clade 2.3.4.4 differed from those of the viruses of clades 2.3.4 and 2.3.2.1, which suggests that the recent H5 HPAIV have further evolved antigenically divergent. In addition, reactivity of antiserum suggests that the antigenicity of viruses of clade 2.3.4.4 differed slightly among groups A, B, and C. Vaccines are still used in poultry in endemic countries, so the antigenicity of H5 HPAIV should be monitored continually to facilitate control of avian influenza. The panel of mAbs established in the present study will be useful for detecting antigenic drift in the H5 viruses that emerge from the current strains. © 2017 The Societies and John Wiley & Sons Australia, Ltd.

Assembly of viral particles in Xenopus oocytes: pre-surface-antigens regulate secretion of the hepatitis B viral surface envelope particle.

PubMed Central

Standring, D N; Ou, J H; Rutter, W J

1986-01-01

Infection with hepatitis B virus (HBV) is associated with the production of a viral envelope particle that contains membrane lipids, surface antigen (S), and two presurface-antigens (pre-S) comprised of the entire S moiety with approximately 55 (pre-S2) and 174 (pre-S1) additional NH2-terminal amino acids. We show here that Xenopus oocytes injected with synthetic S mRNA assemble and secrete characteristic 22-nm viral envelope particles. In contrast, pre-S1 and pre-S2 antigens are synthesized but not secreted. By coinjecting mRNAs, we found that synthesis of high levels of pre-S proteins specifically inhibits S antigen secretion. On the other hand, high levels of S synthesis can drive the secretion of small amounts of either pre-S antigen. These observations are consistent with a model for viral envelope assembly in which both S and pre-S proteins are incorporated into a multimeric particle, presumably via interactions between the S protein domains, while the pre-S amino-terminal moieties regulate the secretion of this structure. Our results indicate that Xenopus oocytes will provide a powerful system for studying the morphogenesis of simple structures of viral or cellular origin. Images PMID:3467308

Recovery of West Nile Virus Envelope Protein Domain III Chimeras with Altered Antigenicity and Mouse Virulence

PubMed Central

McAuley, Alexander J.; Torres, Maricela; Plante, Jessica A.; Huang, Claire Y.-H.; Bente, Dennis A.

2016-01-01

ABSTRACT Flaviviruses are positive-sense, single-stranded RNA viruses responsible for millions of human infections annually. The envelope (E) protein of flaviviruses comprises three structural domains, of which domain III (EIII) represents a discrete subunit. The EIII gene sequence typically encodes epitopes recognized by virus-specific, potently neutralizing antibodies, and EIII is believed to play a major role in receptor binding. In order to assess potential interactions between EIII and the remainder of the E protein and to assess the effects of EIII sequence substitutions on the antigenicity, growth, and virulence of a representative flavivirus, chimeric viruses were generated using the West Nile virus (WNV) infectious clone, into which EIIIs from nine flaviviruses with various levels of genetic diversity from WNV were substituted. Of the constructs tested, chimeras containing EIIIs from Koutango virus (KOUV), Japanese encephalitis virus (JEV), St. Louis encephalitis virus (SLEV), and Bagaza virus (BAGV) were successfully recovered. Characterization of the chimeras in vitro and in vivo revealed differences in growth and virulence between the viruses, with in vivo pathogenesis often not being correlated with in vitro growth. Taken together, the data demonstrate that substitutions of EIII can allow the generation of viable chimeric viruses with significantly altered antigenicity and virulence. IMPORTANCE The envelope (E) glycoprotein is the major protein present on the surface of flavivirus virions and is responsible for mediating virus binding and entry into target cells. Several viable West Nile virus (WNV) variants with chimeric E proteins in which the putative receptor-binding domain (EIII) sequences of other mosquito-borne flaviviruses were substituted in place of the WNV EIII were recovered, although the substitution of several more divergent EIII sequences was not tolerated. The differences in virulence and tissue tropism observed with the chimeric viruses indicate a significant role for this sequence in determining the pathogenesis of the virus within the mammalian host. Our studies demonstrate that these chimeras are viable and suggest that such recombinant viruses may be useful for investigation of domain-specific antibody responses and the more extensive definition of the contributions of EIII to the tropism and pathogenesis of WNV or other flaviviruses. PMID:26912625

Recovery of West Nile Virus Envelope Protein Domain III Chimeras with Altered Antigenicity and Mouse Virulence.

PubMed

McAuley, Alexander J; Torres, Maricela; Plante, Jessica A; Huang, Claire Y-H; Bente, Dennis A; Beasley, David W C

2016-05-01

Flaviviruses are positive-sense, single-stranded RNA viruses responsible for millions of human infections annually. The envelope (E) protein of flaviviruses comprises three structural domains, of which domain III (EIII) represents a discrete subunit. The EIII gene sequence typically encodes epitopes recognized by virus-specific, potently neutralizing antibodies, and EIII is believed to play a major role in receptor binding. In order to assess potential interactions between EIII and the remainder of the E protein and to assess the effects of EIII sequence substitutions on the antigenicity, growth, and virulence of a representative flavivirus, chimeric viruses were generated using the West Nile virus (WNV) infectious clone, into which EIIIs from nine flaviviruses with various levels of genetic diversity from WNV were substituted. Of the constructs tested, chimeras containing EIIIs from Koutango virus (KOUV), Japanese encephalitis virus (JEV), St. Louis encephalitis virus (SLEV), and Bagaza virus (BAGV) were successfully recovered. Characterization of the chimeras in vitro and in vivo revealed differences in growth and virulence between the viruses, within vivo pathogenesis often not being correlated within vitro growth. Taken together, the data demonstrate that substitutions of EIII can allow the generation of viable chimeric viruses with significantly altered antigenicity and virulence. The envelope (E) glycoprotein is the major protein present on the surface of flavivirus virions and is responsible for mediating virus binding and entry into target cells. Several viable West Nile virus (WNV) variants with chimeric E proteins in which the putative receptor-binding domain (EIII) sequences of other mosquito-borne flaviviruses were substituted in place of the WNV EIII were recovered, although the substitution of several more divergent EIII sequences was not tolerated. The differences in virulence and tissue tropism observed with the chimeric viruses indicate a significant role for this sequence in determining the pathogenesis of the virus within the mammalian host. Our studies demonstrate that these chimeras are viable and suggest that such recombinant viruses may be useful for investigation of domain-specific antibody responses and the more extensive definition of the contributions of EIII to the tropism and pathogenesis of WNV or other flaviviruses. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Understanding MHC Class I Presentation of Viral Antigens by Human Dendritic Cells as a Basis for Rational Design of Therapeutic Vaccines

PubMed Central

van Montfoort, Nadine; van der Aa, Evelyn; Woltman, Andrea M.

2014-01-01

Effective viral clearance requires the induction of virus-specific CD8+ cytotoxic T lymphocytes (CTL). Since dendritic cells (DC) have a central role in initiating and shaping virus-specific CTL responses, it is important to understand how DC initiate virus-specific CTL responses. Some viruses can directly infect DC, which theoretically allow direct presentation of viral antigens to CTL, but many viruses target other cells than DC and thus the host depends on the cross-presentation of viral antigens by DC to activate virus-specific CTL. Research in mouse models has highly enhanced our understanding of the mechanisms underlying cross-presentation and the dendritic cells (DC) subsets involved, however, these results cannot be readily translated toward the role of human DC in MHC class I-antigen presentation of human viruses. Here, we summarize the insights gained in the past 20 years on MHC class I presentation of viral antigen by human DC and add to the current debate on the capacities of different human DC subsets herein. Furthermore, possible sources of viral antigens and essential DC characteristics for effective induction of virus-specific CTL are evaluated. We conclude that cross-presentation is not only an efficient mechanism exploited by DC to initiate immunity to viruses that do not infect DC but also to viruses that do infect DC, because cross-presentation has many conceptual advantages and bypasses direct immune modulatory effects of the virus on its infected target cells. Since knowledge on the mechanism of viral antigen presentation and the preferred DC subsets is crucial for rational vaccine design, the obtained insights are very instrumental for the development of effective anti-viral immunotherapy. PMID:24795724

A Longitudinal Hepatitis B Vaccine Cohort Demonstrates Long-lasting Hepatitis B Virus (HBV) Cellular Immunity Despite Loss of Antibody Against HBV Surface Antigen.

PubMed

Simons, Brenna C; Spradling, Philip R; Bruden, Dana J T; Zanis, Carolyn; Case, Samantha; Choromanski, Tammy L; Apodaca, Minjun; Brogdon, Hazel D; Dwyer, Gaelen; Snowball, Mary; Negus, Susan; Bruce, Michael G; Morishima, Chihiro; Knall, Cindy; McMahon, Brian J

2016-07-15

Long-lasting protection resulting from hepatitis B vaccine, despite loss of antibody against hepatitis B virus (HBV) surface antigen (anti-HBs), is undetermined. We recruited persons from a cohort vaccinated with plasma-derived hepatitis B vaccine in 1981 who have been followed periodically since. We performed serological testing for anti-HBs and microRNA-155 and assessed HBV-specific T-cell responses by enzyme-linked immunospot and cytometric bead array. Study subgroups were defined 32 years after vaccination as having an anti-HBs level of either ≥10 mIU/mL (group 1; n = 13) or <10 mIU/mL (group 2; n = 31). All 44 participants, regardless of anti-HBs level, tested positive for tumor necrosis factor α, interleukin 10, or interleukin 6 production by HBV surface antigen-specific T cells. The frequency of natural killer T cells correlated with the level of anti-HBs (P = .008). The proportion of participants who demonstrated T-cell responses to HBV core antigen varied among the cytokines measured, suggesting some natural exposure to HBV in the study group. No participant had evidence of breakthrough HBV infection. Evidence of long-lasting cellular immunity, regardless of anti-HBs level, suggests that protection afforded by primary immunization with plasma-derived hepatitis B vaccine during childhood and adulthood lasts at least 32 years. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Marked differences in the antigenic structure of human respiratory syncytial virus F and G glycoproteins.

PubMed Central

García-Barreno, B; Palomo, C; Peñas, C; Delgado, T; Perez-Breña, P; Melero, J A

1989-01-01

Monoclonal antibodies directed against the glycoproteins of human respiratory syncytial virus were used in competitive enzyme-linked immunosorbent assays for topological mapping of epitopes. Whereas epitopes of the F glycoprotein could be ascribed to five nonoverlapping antigenic sites, anti-G antibodies recognized unique epitopes, many of whose competition profiles overlapped extensively. Variant viruses selected with a neutralizing (47F) anti-F antibody lost the binding for only 47F and 49F antibodies, which mapped in the same antigenic area. In contrast, viruses selected with an anti-G antibody lost the capacity to bind most of the anti-G antibodies, and their G protein was not recognized by an anti-virus antiserum, indicating major changes in the antigenic structure of the G molecule. Finally, we found great antigenic variation of the G protein among viral isolates. This occurred even within viruses of the same subtype with only limited divergence of amino acid sequence between strains. All of these data indicate marked differences in the antigenic organization of the G and F glycoproteins of respiratory syncytial virus; we discuss these differences in terms of the chemical structure of the glycoproteins. Images PMID:2463385

Enhanced activation of human T cell clones specific for virus-like particles expressing the HIV V3 loop in the presence of HIV V3 loop-specific polyclonal antibodies

PubMed Central

Peifang, S.; Pira, G. L.; Fenoglio, D.; Harris, S.; Costa, M. G.; Venturino, V.; Dessì, V.; Layton, G.; Laman, J.; Huisman, J. G.; Manca, F.

1994-01-01

Recombinant virus-like particles (VLP), formed by the yeast Ty p1 protein, carrying the HIV gp120 V3 loop on their surface (V3-VLP) have been tested in vitro for immunogenicity and antigenicity by using VLP p1-specific human CD4+ T cell lines and clones. VLP-specific human T cell lines and clones were generated from normal individuals, indicating that VLP-specific precursor cells present in the peripheral lymphocyte pool can be induced to expand clonally upon antigen challenge in vitro, in the absence of previous immunization. It was also shown that V3-specific polyclonal antibodies enhance V3-VLP-induced activation of VLP-specific T cell clones. Antibody-dependent potentiation has been shown previously in other antigen systems, and it depends on enhanced uptake of complexed antigen by Fc receptor-positive antigen-presenting cells. Since in this case antigen is internalized by presenting cells as a complex, it can be inferred that a similar event of antibody-mediated antigen uptake can take place with V3-specific B cells, resulting in presentation by the B cells of T helper epitopes derived from processing of the VLP p1 moiety. This suggests that T helper cells specific for the carrier VLP p1 protein can be activated to provide help to V3-specific B cells in the presence of the appropriate antigen construct. PMID:7915974

Vaccine approaches conferring cross-protection against influenza viruses

PubMed Central

Vemula, Sai V.; Sayedahmed, Ekramy E; Sambhara, Suryaprakash; Mittal, Suresh K.

2018-01-01

Introduction Annual vaccination is one of the most efficient and cost-effective strategies to prevent and control influenza epidemics. Most of currently available influenza vaccines are strong inducer of antibody responses against viral surface proteins, hemagglutinin (HA) and neuraminidase (NA), but are poor inducers of cell-mediated immune responses against conserved internal proteins. Moreover, due to the high variability of viral surface proteins because of antigenic drift or antigenic shift, many of the currently licensed vaccines confer little or no protection against drift or shift variants. Areas covered Next generation influenza vaccines that can induce humoral immune responses to receptor-binding epitopes as well as broadly neutralizing conserved epitopes, and cell-mediated immune responses against highly conserved internal proteins would be effective against variant viruses as well as a novel pandemic influenza until circulating strain-specific vaccines become available. Here we discuss vaccine approaches that have potential to provide broad spectrum protection against influenza viruses. Expert opinion Based on current progress in defining cross-protective influenza immunity, it seems that the development of a universal influenza vaccine is feasible. It would revolutionize the strategy for influenza pandemic preparedness, and significantly impact the shelf-life and protection efficacy of seasonal influenza vaccines. PMID:28925296

Antigenic and genomic characterization of human influenza A and B viruses circulating in Argentina after the introduction of influenza A(H1N1)pdm09.

PubMed

Russo, Mara L; Pontoriero, Andrea V; Benedetti, Estefania; Czech, Andrea; Avaro, Martin; Periolo, Natalia; Campos, Ana M; Savy, Vilma L; Baumeister, Elsa G

2014-12-01

This study was conducted as part of the Argentinean Influenza and other Respiratory Viruses Surveillance Network, in the context of the Global Influenza Surveillance carried out by the World Health Organization (WHO). The objective was to study the activity and the antigenic and genomic characteristics of circulating viruses for three consecutive seasons (2010, 2011 and 2012) in order to investigate the emergence of influenza viral variants. During the study period, influenza virus circulation was detected from January to December. Influenza A and B, and all current subtypes of human influenza viruses, were present each year. Throughout the 2010 post-pandemic season, influenza A(H1N1)pdm09, unexpectedly, almost disappeared. The haemagglutinin (HA) of the A(H1N1)pdm09 viruses studied were segregated in a different genetic group to those identified during the 2009 pandemic, although they were still antigenically closely related to the vaccine strain A/California/07/2009. Influenza A(H3N2) viruses were the predominant strains circulating during the 2011 season, accounting for nearly 76 % of influenza viruses identified. That year, all HA sequences of the A(H3N2) viruses tested fell into the A/Victoria/208/2009 genetic clade, but remained antigenically related to A/Perth/16/2009 (reference vaccine recommended for this three-year period). A(H3N2) viruses isolated in 2012 were antigenically closely related to A/Victoria/361/2011, recommended by the WHO as the H3 component for the 2013 Southern Hemisphere formulation. B viruses belonging to the B/Victoria lineage circulated in 2010. A mixed circulation of viral variants of both B/Victoria and B/Yamagata lineages was detected in 2012, with the former being predominant. A(H1N1)pdm09 viruses remained antigenically closely related to the vaccine virus A/California/7/2009; A(H3N2) viruses continually evolved into new antigenic clusters and both B lineages, B/Victoria/2/87-like and B/Yamagata/16/88-like viruses, were observed during the study period. The virological surveillance showed that the majority of the circulating strains during the study period were antigenically related to the corresponding Southern Hemisphere vaccine strains except for the 2012 A(H3N2) viruses. © 2014 The Authors.

Genomewide analysis of reassortment and evolution of human influenza A(H3N2) viruses circulating between 1968 and 2011.

PubMed

Westgeest, Kim B; Russell, Colin A; Lin, Xudong; Spronken, Monique I J; Bestebroer, Theo M; Bahl, Justin; van Beek, Ruud; Skepner, Eugene; Halpin, Rebecca A; de Jong, Jan C; Rimmelzwaan, Guus F; Osterhaus, Albert D M E; Smith, Derek J; Wentworth, David E; Fouchier, Ron A M; de Graaf, Miranda

2014-03-01

Influenza A(H3N2) viruses became widespread in humans during the 1968 H3N2 virus pandemic and have been a major cause of influenza epidemics ever since. These viruses evolve continuously by reassortment and genomic evolution. Antigenic drift is the cause for the need to update influenza vaccines frequently. Using two data sets that span the entire period of circulation of human influenza A(H3N2) viruses, it was shown that influenza A(H3N2) virus evolution can be mapped to 13 antigenic clusters. Here we analyzed the full genomes of 286 influenza A(H3N2) viruses from these two data sets to investigate the genomic evolution and reassortment patterns. Numerous reassortment events were found, scattered over the entire period of virus circulation, but most prominently in viruses circulating between 1991 and 1998. Some of these reassortment events persisted over time, and one of these coincided with an antigenic cluster transition. Furthermore, selection pressures and nucleotide and amino acid substitution rates of all proteins were studied, including those of the recently discovered PB1-N40, PA-X, PA-N155, and PA-N182 proteins. Rates of nucleotide and amino acid substitutions were most pronounced for the hemagglutinin, neuraminidase, and PB1-F2 proteins. Selection pressures were highest in hemagglutinin, neuraminidase, matrix 1, and nonstructural protein 1. This study of genotype in relation to antigenic phenotype throughout the period of circulation of human influenza A(H3N2) viruses leads to a better understanding of the evolution of these viruses. Each winter, influenza virus infects approximately 5 to 15% of the world's population, resulting in significant morbidity and mortality. Influenza A(H3N2) viruses evolve continuously by reassortment and genomic evolution. This leads to changes in antigenic recognition (antigenic drift) which make it necessary to update vaccines against influenza A(H3N2) viruses frequently. In this study, the relationship of genetic evolution to antigenic change spanning the entire period of A(H3N2) virus circulation was studied for the first time. The results presented in this study contribute to a better understanding of genetic evolution in correlation with antigenic evolution of influenza A(H3N2) viruses.

Radioimmunoassay of the marjor group specific protein of endogenous baboon type C viruses: relation to the RD-114/CCC group and detection of antigen in normal baboon tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sherr, C.J.; Todaro, G.J.

1974-09-01

The major group specific (gs) protein of the endogenous baboon type C virus M7 was purified to homogeneity by gel filtration and isoelectric focusing. The protein has a molecular weight of approximately 33,000, as determined by electrophoresis in polyacrylamide gels containing sodium dodecyl sulfate, and an isoelectric point (pl) of 7.1, different from the pls of similarly purified gs proteins from six other mammalian type C viruses. Detergent disrupted M7 virus, whether grown in canine thymus or human rhabdomyosarcoma cells, fully displaced radiolabeled M7 gs protein from antigen-antibody complexes in a competitive radioimmunoassay. No antigenic differences were detected among themore » gs proteins of five independent isolates of baboon type C viruses grown in various cultured cell lines. The gs proteins of six independently isolated feline viruses of the RD-114/CCC group were antigenically related to, but could be distinguished from, the gs proteins of baboon type C viruses. No significant cross-reactions were observed in the radioimmunoassay for M7 gs protein using several other type C viruses, including two previously isolated from a woolly monkey and a gibbon ape. Group specific antigen was found in normal baboon testicular and splenic tissues using the M7 radioimmunoassay; no gs antigen could be detected in these same tissues using a radioimmunoassay for the gs protein of the woolly monkey type C virus. No gs antigen was found in baboon liver or in the tissues of several other primates. (auth)« less

Substitutions near the hemagglutinin receptor-binding site determine the antigenic evolution of influenza A H3N2 viruses in U.S. swine.

PubMed

Lewis, Nicola S; Anderson, Tavis K; Kitikoon, Pravina; Skepner, Eugene; Burke, David F; Vincent, Amy L

2014-05-01

Swine influenza A virus is an endemic and economically important pathogen in pigs, with the potential to infect other host species. The hemagglutinin (HA) protein is the primary target of protective immune responses and the major component in swine influenza A vaccines. However, as a result of antigenic drift, vaccine strains must be regularly updated to reflect currently circulating strains. Characterizing the cross-reactivity between strains in pigs and seasonal influenza virus strains in humans is also important in assessing the relative risk of interspecies transmission of viruses from one host population to the other. Hemagglutination inhibition (HI) assay data for swine and human H3N2 viruses were used with antigenic cartography to quantify the antigenic differences among H3N2 viruses isolated from pigs in the United States from 1998 to 2013 and the relative cross-reactivity between these viruses and current human seasonal influenza A virus strains. Two primary antigenic clusters were found circulating in the pig population, but with enough diversity within and between the clusters to suggest updates in vaccine strains are needed. We identified single amino acid substitutions that are likely responsible for antigenic differences between the two primary antigenic clusters and between each antigenic cluster and outliers. The antigenic distance between current seasonal influenza virus H3 strains in humans and those endemic in swine suggests that population immunity may not prevent the introduction of human viruses into pigs, and possibly vice versa, reinforcing the need to monitor and prepare for potential incursions. Influenza A virus (IAV) is an important pathogen in pigs and humans. The hemagglutinin (HA) protein is the primary target of protective immune responses and the major target of vaccines. However, vaccine strains must be updated to reflect current strains. Characterizing the differences between seasonal IAV in humans and swine IAV is important in assessing the relative risk of interspecies transmission of viruses. We found two primary antigenic clusters of H3N2 in the U.S. pig population, with enough diversity to suggest updates in swine vaccine strains are needed. We identified changes in the HA protein that are likely responsible for these differences and that may be useful in predicting when vaccines need to be updated. The difference between human H3N2 viruses and those in swine is enough that population immunity is unlikely to prevent new introductions of human IAV into pigs or vice versa, reinforcing the need to monitor and prepare for potential introductions.

Compensating for cross-reactions using avidity and computation in a suspension multiplex immunoassay for serotyping of Zika versus other flavivirus infections.

PubMed

Rönnberg, Bengt; Gustafsson, Åke; Vapalahti, Olli; Emmerich, Petra; Lundkvist, Åke; Schmidt-Chanasit, Jonas; Blomberg, Jonas

2017-10-01

The recent spread of Zika virus (ZIKV) in the Americas and Asia necessitates an increased preparedness for improved maternal and perinatal health and blood safety. However, serological cross-reactions, especially to Dengue virus (DENV), complicate ZIKV antibody serodiagnosis. A novel "pan-Flavi" suspension multiplex immunoassay (PFSMIA) using 25 antigens, whole virus (WV), non-structural protein 1 (NS1), and envelope (E) proteins, from 7 zoonotic flaviviruses for specific detection of ZIKV and DENV IgM and IgG was developed. Patterns of antibody cross-reactivity, avidity, and kinetics were established in 104 sera from returning travelers with known ZIKV and DENV infections. PFSMIA gave IgM- and IgG-sensitivities for both viruses of 96-100%, compared to an immunofluorescence assay. Main IgM cross-reactions were to NS1, for IgG to the E and WV antigens. Infecting virus yielded reactivity to several antigens of the homologous virus, while cross-reactions tended to occur only to a single antigen from heterologous virus(es). A specificity-enhancing computer procedure took into account antibody isotype, number of antibody-reactive antigens per virus, avidity, average degree of cross-reactivity to heterologous flavivirus antigens, and reactivity changes in serial sera. It classified all 50 cases correctly. Applied to sera from 200 pregnant women and 173 blood donors from Sweden, one blood donor was found ZIKV NS1 IgM positive, and another as ZIKV NS1 IgG positive. These samples did not react with other ZIKV antigens and were thereby judged as false-positives. PFSMIA provided sensitive and specific ZIKV and DENV serology, warranting high-throughput serological surveillance and a minimized need for laborious and expensive virus neutralization assays.

Hepatitis B virus surface antigen (HBsAg)-positive and HBsAg-negative hepatitis B virus infection among mother-teenager pairs 13 years after neonatal hepatitis B virus vaccination.

PubMed

Yao, Qing-Qing; Dong, Xiao-Lian; Wang, Xue-Cai; Ge, Sheng-Xiang; Hu, An-Qun; Liu, Hai-Yan; Wang, Yueping Alex; Yuan, Quan; Zheng, Ying-Jie

2013-02-01

It is unclear whether a mother who is negative for hepatitis B virus surface antigen (HBsAg) but positive for hepatitis B virus (HBV) is at potential risk for mother-to-child transmission of HBV. This study, using a paired mother-teenager population, aimed to assess whether maternal HBsAg-negative HBV infection ((hn)HBI) is a significant source of child HBV infection (HBI). A follow-up study with blood collection has been conducted on the 93 mother-teenager pairs from the initial 135 pregnant woman-newborn pairs 13 years after neonatal HBV vaccination. Serological and viral markers of HBV have been tested, and phylogenetic analysis of HBV isolates has been done. The HBI prevalence was 1.9% (1 (hn)HBI/53) for teenage children of non-HBI mothers, compared with 16.7% (1 (hn)HBI/6) for those of (hn)HBI mothers and 2.9% (1 HBsAg-positive HBV infection [(hp)HBI]/34) for those of (hp)HBI mothers. Similar viral sequences have been found in one pair of whom both the mother and teenager have had (hn)HBI. In comparison with the (hp)HBI cases, those with (hn)HBI had a lower level of HBV load and a higher proportion of genotype-C strains, which were accompanied by differentiated mutations (Q129R, K141E, and Y161N) of the "a" determinant of the HBV surface gene. Our findings suggest that mother-to-teenager transmission of (hn)HBI can occur among those in the neonatal HBV vaccination program.

Hepatitis B Virus Surface Antigen (HBsAg)-Positive and HBsAg-Negative Hepatitis B Virus Infection among Mother-Teenager Pairs 13 Years after Neonatal Hepatitis B Virus Vaccination

PubMed Central

Yao, Qing-Qing; Dong, Xiao-Lian; Wang, Xue-Cai; Ge, Sheng-Xiang; Hu, An-Qun; Liu, Hai-Yan; Wang, Yueping Alex

2013-01-01

It is unclear whether a mother who is negative for hepatitis B virus surface antigen (HBsAg) but positive for hepatitis B virus (HBV) is at potential risk for mother-to-child transmission of HBV. This study, using a paired mother-teenager population, aimed to assess whether maternal HBsAg-negative HBV infection (hnHBI) is a significant source of child HBV infection (HBI). A follow-up study with blood collection has been conducted on the 93 mother-teenager pairs from the initial 135 pregnant woman-newborn pairs 13 years after neonatal HBV vaccination. Serological and viral markers of HBV have been tested, and phylogenetic analysis of HBV isolates has been done. The HBI prevalence was 1.9% (1 hnHBI/53) for teenage children of non-HBI mothers, compared with 16.7% (1 hnHBI/6) for those of hnHBI mothers and 2.9% (1 HBsAg-positive HBV infection [hpHBI]/34) for those of hpHBI mothers. Similar viral sequences have been found in one pair of whom both the mother and teenager have had hnHBI. In comparison with the hpHBI cases, those with hnHBI had a lower level of HBV load and a higher proportion of genotype-C strains, which were accompanied by differentiated mutations (Q129R, K141E, and Y161N) of the “a” determinant of the HBV surface gene. Our findings suggest that mother-to-teenager transmission of hnHBI can occur among those in the neonatal HBV vaccination program. PMID:23254298

Felix Hoppe-Seyler Lecture 1997. Protective antibody responses against viruses.

PubMed

Zinkernagel, R M

1997-08-01

Neutralizing antibody responses against the acute cytopathic vesicular stomatitis virus (VSV) have been studied in mice to evaluate their general characteristics including specificity, self-/non-self discrimination and memory. IgM responses are generated very early, by day 3 to 4, in a T helper cell-independent fashion and without VSV having polyclonal activating capacities. The order of the glycoprotein tips on the virus envelope (multiple, 8-10 nm distance, paracrystalline) exhibiting the neutralizing determinants are key to this prompt response. These paracrystalline identical multimeric antigens are characteristic of infectious agents and are always reacted against by B cells. Self-antigens that are accessible to B cells in the intact host are either monomeric in serum or mobile multimers on cell surfaces; these configurations need contact dependent or contact independent T help, respectively. Because T help is tolerant against self-antigens, no anti-self B cell responses are usually induced against monomeric self-antigens. If collagen or DNA (rigid multimeric self-antigens) become accessible, however, they may become targets of auto-antibody responses. The antibody repertoire against VSV is partially contained in the germline and partially is generated by somatic mutation; they seem not to undergo affinity-maturation. In any case protection against lethal infection is dependent upon strictly T helper cell dependent IgG generated by day 6 to 7 and reaches a protective level of about 1-10 micrograms/ml. Interesting affinity/avidity and onrate above a minimal threshold are of no apparent advantage for protection in vivo. Maintenance of these antibody levels by antigen depots, and not the presence of memory B cells alone, is key to providing protective immunological memory. Collectively these data suggest that studying biologically important protective antibody responses may modify some of the parameters that have been defined by studying hapten specific antibody responses.

Antibody to the rheumatoid arthritis nuclear antigen. Its relationship to in vivo Epstein-Barr virus infection.

PubMed

Catalano, M A; Carson, D A; Niederman, J C; Feorino, P; Vaughan, J H

1980-05-01

Most patients with seropositive rheumatoid arthritis, and a variable but lesser percentage of normal subjects, have precipitating antibodies to a nuclear antigen, rheumatoid arthritis nuclear antigen, present in Epstein-Barr virus-infected human B lymphoblastoid cells. We have used a sensitive indirect immunofluorescence assay for antibody to rheumatoid arthritis nuclear antigen in a study of patients with infectious mononucleosis and healthy control subjects. Of 110 sera from normal, college-age cadets, 58 were from individuals without prior Epstein-Barr virus infection, as indicated by the lack of antibody to viral capsid antigen. All of these also lacked activity to rheumatoid arthritis nuclear antigen. 52 sera were positive for antibody to viral capsid antigen, and antibody to rheumatoid arthritis nuclear antigen was present in 26 (50%) of these. In 67 sequential sera from 11 college-age students with infectious mononucleosis who became positive for antibody to rheumatoid arthritis nuclear antigen, only 2 were positive during the 1 mo. Thereafter the incidence and titers increased progressively through the 1st yr after infection. This time-course resembled that for the development of antibody to Epstein-Barr nuclear antigen, another transformation antigen in Epstein-Barr virus-infected B lymphocytes. The development of positivity for both was much later than that of antibody to the structural viral capsid antigen, which in the current study was always positive by 1 wk. Thus, antibody to rheumatoid arthritis nuclear antigen is present in a large proportion of normal individuals and can now be clearly ascribed, from both in vivo and in vitro studies, to prior infection with Epstein-Barr virus.

Mouse models to study the interaction of risk factors for human liver cancer.

PubMed

Sell, Stewart

2003-11-15

Each of the risk factors for human liver cancer (aflatoxin exposure, hepatitis B virus-associated liver injury, p53 loss, p53ser249 mutation, and male sex) also increases the incidence of hepatocellular carcinoma (HCC) in mouse models of hepatocarcinogenesis. Neonatal mice, partially hepatectomized adult mice, and p53-deficient mice each have a higher hepatocyte proliferation rate, are less able to detoxify AFB1, and form more DNA adducts than do normal wild-type controls. However, transgenic hepatitis B surface antigen mice, expressing hepatitis B surface antigen under control of the albumin promoter (alb/psx), are able to detoxify AFB1 at the same level as do wild-type mice. Thus, AFB1-induced HCC development in neonatal mice and p53+/- mice may be due to "immature" carcinogen metabolism, whereas increased HCC in transgenic hepatitis B virus mice may be due to promotion effects of increased proliferation. Future studies will explore the effects of modifying factors on the development of HCC.

[Application of dhfr gene negative Chinese hamster ovary cell line to express hepatitis B virus surface antigen].

PubMed

Yi, Y; Zhang, M; Liu, C

2001-06-01

To set up an efficient expressing system for recombinant hepatitis B virus surface antigen (HBsAg) in dhfr gene negative CHO cell line. HBsAg gene expressing plasmid pCI-dhfr-S was constructed by integrating HBsAg gene into plasmid pCI which carries dhfr gene. The HBsAg expressing cell line was set up by transfection of plasmid pCI-dhfr-S into dhfr gene negative CHO cell line in the way of lipofectin. Under the selective pressure of MTX, 18 of 28 clonized cell lines expressed HBsAg, 4 of them reached a high titer of 1:32 and protein content 1-3 micrograms/ml. In this study, the high level expression of HBsAg demonstrated that the dhfr negative mammalian cell line when recombined with plasmid harboring the corresponding deleted gene can efficiently express the foreign gene. The further steps toward building optimum conditions of the expressing system and the increase of expressed product are under study.

Changing selective pressure during antigenic changes in human influenza H3.

PubMed

Blackburne, Benjamin P; Hay, Alan J; Goldstein, Richard A

2008-05-02

The rapid evolution of influenza viruses presents difficulties in maintaining the optimal efficiency of vaccines. Amino acid substitutions result in antigenic drift, a process whereby antisera raised in response to one virus have reduced effectiveness against future viruses. Interestingly, while amino acid substitutions occur at a relatively constant rate, the antigenic properties of H3 move in a discontinuous, step-wise manner. It is not clear why this punctuated evolution occurs, whether this represents simply the fact that some substitutions affect these properties more than others, or if this is indicative of a changing relationship between the virus and the host. In addition, the role of changing glycosylation of the haemagglutinin in these shifts in antigenic properties is unknown. We analysed the antigenic drift of HA1 from human influenza H3 using a model of sequence change that allows for variation in selective pressure at different locations in the sequence, as well as at different parts of the phylogenetic tree. We detect significant changes in selective pressure that occur preferentially during major changes in antigenic properties. Despite the large increase in glycosylation during the past 40 years, changes in glycosylation did not correlate either with changes in antigenic properties or with significantly more rapid changes in selective pressure. The locations that undergo changes in selective pressure are largely in places undergoing adaptive evolution, in antigenic locations, and in locations or near locations undergoing substitutions that characterise the change in antigenicity of the virus. Our results suggest that the relationship of the virus to the host changes with time, with the shifts in antigenic properties representing changes in this relationship. This suggests that the virus and host immune system are evolving different methods to counter each other. While we are able to characterise the rapid increase in glycosylation of the haemagglutinin during time in human influenza H3, an increase not present in influenza in birds, this increase seems unrelated to the observed changes in antigenic properties.

Sequential Infection in Ferrets with Antigenically Distinct Seasonal H1N1 Influenza Viruses Boosts Hemagglutinin Stalk-Specific Antibodies

PubMed Central

Kirchenbaum, Greg A.; Carter, Donald M.

2015-01-01

ABSTRACT Broadly reactive antibodies targeting the conserved hemagglutinin (HA) stalk region are elicited following sequential infection or vaccination with influenza viruses belonging to divergent subtypes and/or expressing antigenically distinct HA globular head domains. Here, we demonstrate, through the use of novel chimeric HA proteins and competitive binding assays, that sequential infection of ferrets with antigenically distinct seasonal H1N1 (sH1N1) influenza virus isolates induced an HA stalk-specific antibody response. Additionally, stalk-specific antibody titers were boosted following sequential infection with antigenically distinct sH1N1 isolates in spite of preexisting, cross-reactive, HA-specific antibody titers. Despite a decline in stalk-specific serum antibody titers, sequential sH1N1 influenza virus-infected ferrets were protected from challenge with a novel H1N1 influenza virus (A/California/07/2009), and these ferrets poorly transmitted the virus to naive contacts. Collectively, these findings indicate that HA stalk-specific antibodies are commonly elicited in ferrets following sequential infection with antigenically distinct sH1N1 influenza virus isolates lacking HA receptor-binding site cross-reactivity and can protect ferrets against a pathogenic novel H1N1 virus. IMPORTANCE The influenza virus hemagglutinin (HA) is a major target of the humoral immune response following infection and/or seasonal vaccination. While antibodies targeting the receptor-binding pocket of HA possess strong neutralization capacities, these antibodies are largely strain specific and do not confer protection against antigenic drift variant or novel HA subtype-expressing viruses. In contrast, antibodies targeting the conserved stalk region of HA exhibit broader reactivity among viruses within and among influenza virus subtypes. Here, we show that sequential infection of ferrets with antigenically distinct seasonal H1N1 influenza viruses boosts the antibody responses directed at the HA stalk region. Moreover, ferrets possessing HA stalk-specific antibody were protected against novel H1N1 virus infection and did not transmit the virus to naive contacts. PMID:26559834

Structural and Nonstructural Viral Proteins Are Targets of T-Helper Immune Response against Human Respiratory Syncytial Virus.

PubMed

Lorente, Elena; Barriga, Alejandro; Barnea, Eilon; Mir, Carmen; Gebe, John A; Admon, Arie; López, Daniel

2016-06-01

Proper antiviral humoral and cellular immune responses require previous recognition of viral antigenic peptides that are bound to HLA class II molecules, which are exposed on the surface of antigen-presenting cells. The helper immune response is critical for the control and the clearance of human respiratory syncytial virus (HRSV) infection, a virus with severe health risk in infected pediatric, immunocompromised, and elderly populations. In this study, using a mass spectrometry analysis of complex HLA class II-bound peptide pools that were isolated from large amounts of HRSV-infected cells, 19 naturally processed HLA-DR ligands, most of them included in a complex nested set of peptides, were identified. Both the immunoprevalence and the immunodominance of the HLA class II response to HRSV were focused on one nonstructural (NS1) and two structural (matrix and mainly fusion) proteins of the infective virus. These findings have clear implications for analysis of the helper immune response as well as for antiviral vaccine design. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

A transgenic plant cell-suspension system for expression of epitopes on chimeric Bamboo mosaic virus particles.

PubMed

Muthamilselvan, Thangarasu; Lee, Chin-Wei; Cho, Yu-Hsin; Wu, Feng-Chao; Hu, Chung-Chi; Liang, Yu-Chuan; Lin, Na-Sheng; Hsu, Yau-Heiu

2016-01-01

We describe a novel strategy to produce vaccine antigens using a plant cell-suspension culture system in lieu of the conventional bacterial or animal cell-culture systems. We generated transgenic cell-suspension cultures from Nicotiana benthamiana leaves carrying wild-type or chimeric Bamboo mosaic virus (BaMV) expression constructs encoding the viral protein 1 (VP1) epitope of foot-and-mouth disease virus (FMDV). Antigens accumulated to high levels in BdT38 and BdT19 transgenic cell lines co-expressing silencing suppressor protein P38 or P19. BaMV chimeric virus particles (CVPs) were subsequently purified from the respective cell lines (1.5 and 2.1 mg CVPs/20 g fresh weight of suspended biomass, respectively), and the resulting CVPs displayed VP1 epitope on the surfaces. Guinea pigs vaccinated with purified CVPs produced humoral antibodies. This study represents an important advance in the large-scale production of immunopeptide vaccines in a cost-effective manner using a plant cell-suspension culture system. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Dengue virus-specific human T cell clones. Serotype crossreactive proliferation, interferon gamma production, and cytotoxic activity

PubMed Central

1989-01-01

The severe complications of dengue virus infections, hemorrhagic manifestation and shock, are much more commonly observed during secondary infections caused by a different serotype of dengue virus than that which caused the primary infections. It has been speculated, therefore, that dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) are caused by serotype crossreactive immunopathological mechanisms. We analyzed clones of dengue serotype crossreactive T lymphocytes derived from the PBMC of a donor who had been infected with dengue 3 virus. These PBMC responded best to dengue 3 antigen, but also responded to dengue 1, 2, and 4 antigens, in bulk culture proliferation assays. 12 dengue antigen-specific clones were established using a limiting dilution technique. All of the clones had CD3+ CD4+ CD8 phenotypes. Eight clones responded to dengue 1, 2, 3, and 4 antigens and are crossreactive, while four other clones responded predominantly to dengue 3 antigen. These results indicate that the serotype crossreactive dengue-specific T lymphocyte proliferation observed in bulk cultures reflects the crossreactive responses detected at the clonal level. Serotype crossreactive clones produced high titers of IFN- gamma after stimulation with dengue 3 antigens, and also produced IFN- gamma to lower levels after stimulation with dengue 1, 2, and 4 antigens. The crossreactive clones lysed autologous lymphoblastoid cell line (LCL) pulsed with dengue antigens, and the crossreactivity of CTL lysis by T cell clones was consistent with the crossreactivity observed in proliferation assays. Epidemiological studies have shown that secondary infections with dengue 2 virus cause DHF/DSS at a higher rate than the other serotypes. We hypothesized that the lysis of dengue virus-infected cells by CTL may lead to DHF/DSS; therefore, the clones were examined for cytotoxic activity against dengue 2 virus-infected LCL. All but one of the serotype crossreactive clones lysed dengue 2 virus-infected autologous LCL, and they did not lyse uninfected autologous LCL. The lysis of dengue antigen-pulsed or virus-infected LCL by the crossreactive CTL clones that we have examined is restricted by HLA DP or DQ antigens. These results indicate that primary dengue virus infections induce predominantly crossreactive memory CD4+ T lymphocytes. These crossreactive T lymphocytes proliferate and produce IFN-gamma after stimulation with a virus strain of another serotype, and demonstrate crossreactive cyotoxic activity against autologous cells infected with heterologous dengue viruses.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:2475573

Evaluation of the use of non-pathogenic porcine circovirus type 1 as a vaccine delivery virus vector to express antigenic epitopes of porcine reproductive and respiratory syndrome virus.

PubMed

Piñeyro, Pablo E; Kenney, Scott P; Giménez-Lirola, Luis G; Opriessnig, Tanja; Tian, Debin; Heffron, C Lynn; Meng, Xiang-Jin

2016-02-02

We previously demonstrated that the C-terminus of the capsid gene of porcine circovirus type 2 (PCV2) is an immune reactive epitope displayed on the surface of virions. Insertion of foreign epitope tags in the C-terminus produced infectious virions that elicited humoral immune responses against both PCV2 capsid and the inserted epitope tags, whereas mutation in the N terminus impaired viral replication. Since the non-pathogenic porcine circovirus type 1 (PCV1) shares similar genomic organization and significant sequence identity with pathogenic PCV2, in this study we evaluated whether PCV1 can serve as a vaccine delivery virus vector. Four different antigenic determinants of porcine reproductive and respiratory syndrome virus (PRRSV) were inserted in the C-terminus of the PCV1 capsid gene, the infectivity and immunogenicity of the resulting viruses are determined. We showed that an insertion of 12 (PRRSV-GP2 epitope II, PRRSV-GP3 epitope I, and PRRSV-GP5 epitope I), and 14 (PRRSV-GP5 epitope IV) amino acid residues did not affect PCV1 replication. We successfully rescued and characterized four chimeric PCV1 viruses expressing PRRSV linear antigenic determinants (GP2 epitope II: aa 40-51, ASPSHVGWWSFA; GP3 epitope I: aa 61-72, QAAAEAYEPGRS; GP5 epitope I: aa 35-46, SSSNLQLIYNLT; and GP5 epitope IV: aa 187-200, TPVTRVSAEQWGRP). We demonstrated that all chimeric viruses were stable and infectious in vitro and three chimeric viruses were infectious in vivo. An immunogenicity study in pigs revealed that PCV1-VR2385EPI chimeric viruses elicited neutralizing antibodies against PRRSV-VR2385. The results have important implications for further evaluating PCV1 as a potential vaccine delivery vector. Copyright © 2015 Elsevier B.V. All rights reserved.

Comparative analysis of rabbit hemorrhagic disease virus (RHDV) and new RHDV2 virus antigenicity, using specific virus-like particles.

PubMed

Bárcena, Juan; Guerra, Beatriz; Angulo, Iván; González, Julia; Valcárcel, Félix; Mata, Carlos P; Castón, José R; Blanco, Esther; Alejo, Alí

2015-09-24

In 2010 a new Lagovirus related to rabbit haemorrhagic disease virus (RHDV) emerged in France and has since rapidly spread throughout domestic and wild rabbit populations of several European countries. The new virus, termed RHDV2, exhibits distinctive genetic, antigenic and pathogenic features. Notably, RHDV2 kills rabbits previously vaccinated with RHDV vaccines. Here we report for the first time the generation and characterization of RHDV2-specific virus-like particles (VLPs). Our results further confirmed the differential antigenic properties exhibited by RHDV and RHDV2, highlighting the need of using RHDV2-specific diagnostic assays to monitor the spread of this new virus.

Spontaneous establishment of an Epstein-Barr virus-infected fibroblast line from the synovial tissue of a rheumatoid arthritis patient.

PubMed Central

Koide, J; Takada, K; Sugiura, M; Sekine, H; Ito, T; Saito, K; Mori, S; Takeuchi, T; Uchida, S; Abe, T

1997-01-01

An Epstein-Barr virus (EBV)-infected fibroblast line, designated DSEK, was spontaneously established from synovial tissue of a patient with rheumatoid arthritis (RA). DSEK cells expressed EBV nuclear antigens EBNA-1 and EBNA-2 and latent membrane protein LMP-1. Cell surface markers of DSEK cells were similar to those of EBV-negative fibroblast clones derived from synoviocytes and were negative for lymphocyte and macrophage markers. DSEK cells expressed CD44, CD58, and HLA-DR antigens and spontaneously produced interleukin-10 basic fibroblast growth factor and transforming growth factor beta1. These results indicate that rheumatoid synoviocytes can be a target for EBV infection and suggest that EBV may play a role in the pathogenesis of RA. PMID:9032386

Simian virus 40 large T antigen associates with cyclin A and p33cdk2.

PubMed

Adamczewski, J P; Gannon, J V; Hunt, T

1993-11-01

In this paper we provide evidence that a fraction of large T antigen of simian virus 40 (SV40) interacts with cyclin A and p33cdk2 in both virus-infected and stably transformed cells. Immunoprecipitates of SV40 large T antigen from SV40-infected or SV40 large-T-antigen-transformed cells contain cyclin A, p33cdk2, and histone H1 kinase activity. Conversely, immunoprecipitates of cyclin A from these cells contain SV40 large T antigen. In this respect, SV40 large T antigen has properties similar to those of the E1A oncogene of adenoviruses and the E7 oncogene of human papillomaviruses.

Original antigenic sin responses to influenza viruses.

PubMed

Kim, Jin Hyang; Skountzou, Ioanna; Compans, Richard; Jacob, Joshy

2009-09-01

Most immune responses follow Burnet's rule in that Ag recruits specific lymphocytes from a large repertoire and induces them to proliferate and differentiate into effector cells. However, the phenomenon of "original antigenic sin" stands out as a paradox to Burnet's rule of B cell engagement. Humans, upon infection with a novel influenza strain, produce Abs against older viral strains at the expense of responses to novel, protective antigenic determinants. This exacerbates the severity of the current infection. This blind spot of the immune system and the redirection of responses to the "original Ag" rather than to novel epitopes were described fifty years ago. Recent reports have questioned the existence of this phenomenon. Hence, we revisited this issue to determine the extent to which original antigenic sin is induced by variant influenza viruses. Using two related strains of influenza A virus, we show that original antigenic sin leads to a significant decrease in development of protective immunity and recall responses to the second virus. In addition, we show that sequential infection of mice with two live influenza virus strains leads to almost exclusive Ab responses to the first viral strain, suggesting that original antigenic sin could be a potential strategy by which variant influenza viruses subvert the immune system.

Antigenic variation of H1N1, H1N2 and H3N2 swine influenza viruses in Japan and Vietnam.

PubMed

Takemae, Nobuhiro; Nguyen, Tung; Ngo, Long Thanh; Hiromoto, Yasuaki; Uchida, Yuko; Pham, Vu Phong; Kageyama, Tsutomu; Kasuo, Shizuko; Shimada, Shinichi; Yamashita, Yasutaka; Goto, Kaoru; Kubo, Hideyuki; Le, Vu Tri; Van Vo, Hung; Do, Hoa Thi; Nguyen, Dang Hoang; Hayashi, Tsuyoshi; Matsuu, Aya; Saito, Takehiko

2013-04-01

The antigenicity of the influenza A virus hemagglutinin is responsible for vaccine efficacy in protecting pigs against swine influenza virus (SIV) infection. However, the antigenicity of SIV strains currently circulating in Japan and Vietnam has not been well characterized. We examined the antigenicity of classical H1 SIVs, pandemic A(H1N1)2009 (A(H1N1)pdm09) viruses, and seasonal human-lineage SIVs isolated in Japan and Vietnam. A hemagglutination inhibition (HI) assay was used to determine antigenic differences that differentiate the recent Japanese H1N2 and H3N2 SIVs from the H1N1 and H3N2 domestic vaccine strains. Minor antigenic variation between pig A(H1N1)pdm09 viruses was evident by HI assay using 13 mAbs raised against homologous virus. A Vietnamese H1N2 SIV, whose H1 gene originated from a human strain in the mid-2000s, reacted poorly with post-infection ferret serum against human vaccine strains from 2000-2010. These results provide useful information for selection of optimal strains for SIV vaccine production.

Virus mutations and their impact on vaccination against infectious bursal disease (Gumboro disease).

PubMed

Boudaoud, A; Mamache, B; Tombari, W; Ghram, A

2016-12-01

Infectious bursal disease (also known as Gumboro disease) is an immunosuppressive viral disease specific to chickens. In spite of all the information amassed on the antigenic and immunological characteristics of the virus, the disease has not yet been brought fully under control. It is still prevalent in properly vaccinated flocks carrying specific antibodies at levels normally high enough to prevent the disease. Common causes apart, failure of vaccination against infectious bursal disease is associated mainly with early vaccination in flocks of unknown immune status and with the evolution of viruses circulating in the field, leading to antigenic drift and a sharp rise in pathogenicity. Various highly sensitive molecular techniques have clarified the viral determinants of antigenicity and pathogenicity of the infectious bursal disease virus. However, these markers are not universally recognised and tend to be considered as evolutionary markers. Antigenic variants of the infectious bursal disease virus possess modified neutralising epitopes that allow them to evade the action of maternally-derived or vaccine-induced antibodies. Autogenous or multivalent vaccines are required to control antigenic variants in areas where classical and variant virus strains coexist. Pathotypic variants (very virulent viruses) remain antigenically related to classical viruses. The difficulty in controlling pathotypic variants is linked to the difficulty of eliciting an early immune response, because of the risk of the vaccine virus being neutralised by maternal antibodies. Mathematical calculation of the optimal vaccination time and the use of vaccines resistant to maternally-derived antibodies have improved the control of very virulent viruses. © OIE (World Organisation for Animal Health), 2016.

Humoral markers of active Epstein-Barr virus infection associate with anti-extractable nuclear antigen autoantibodies and plasma galectin-3 binding protein in systemic lupus erythematosus.

PubMed

Rasmussen, N S; Nielsen, C T; Houen, G; Jacobsen, S

2016-12-01

We investigated if signs of active Epstein-Barr virus and cytomegalovirus infections associate with certain autoantibodies and a marker of type I interferon activity in patients with systemic lupus erythematosus. IgM and IgG plasma levels against Epstein-Barr virus early antigen diffuse and cytomegalovirus pp52 were applied as humoral markers of ongoing/recently active Epstein-Barr virus and cytomegalovirus infections, respectively. Plasma galectin-3 binding protein served as a surrogate marker of type I interferon activity. The measurements were conducted in 57 systemic lupus erythematosus patients and 29 healthy controls using ELISAs. Regression analyses and univariate comparisons were performed for associative evaluation between virus serology, plasma galectin-3 binding protein and autoantibodies, along with other clinical and demographic parameters. Plasma galectin-3 binding protein concentrations were significantly higher in systemic lupus erythematosus patients (P = 0.009) and associated positively with Epstein-Barr virus early antigen diffuse-directed antibodies and the presence of autoantibodies against extractable nuclear antigens in adjusted linear regressions (B = 2.02 and 2.02, P = 0.02 and P = 0.002, respectively). Furthermore, systemic lupus erythematosus patients with anti-extractable nuclear antigens had significantly higher antibody levels against Epstein-Barr virus early antigen diffuse (P = 0.02). Our study supports a link between active Epstein-Barr virus infections, positivity for anti-extractable nuclear antigens and increased plasma galectin-3 binding protein concentrations/type I interferon activity in systemic lupus erythematosus patients. © The Author(s) 2016.

Conjugating influenza a (H1N1) antigen to n-trimethylaminoethylmethacrylate chitosan nanoparticles improves the immunogenicity of the antigen after nasal administration.

PubMed

Liu, Qingfeng; Zheng, Xiaoyao; Zhang, Chi; Shao, Xiayan; Zhang, Xi; Zhang, Qizhi; Jiang, Xinguo

2015-11-01

As one of the most serious infectious respiratory diseases, influenza A (H1N1) is a great threat to human health, and it has created an urgent demand for effective vaccines. Nasal immunization can induce both systemic and mucosal immune responses against viruses, and it can serve as an ideal route for vaccination. However, the low immunogenicity of antigens on nasal mucosa is a high barrier for the development of nasal vaccines. In this study, we covalently conjugated an influenza A (H1N1) antigen to the surface of N-trimethylaminoethylmethacrylate chitosan (TMC) nanoparticles (H1N1-TMC/NP) through thioester bonds to increase the immunogenicity of the antigen after nasal administration. SDS-PAGE revealed that most of the antigen was conjugated on TMC nanoparticles, and an in vitro biological activity assay confirmed the stability of the antigen after conjugation. After three nasal immunizations, the H1N1-TMC/NP induced significantly higher levels of serum IgG and mucosal sIgA compared with free antigen. A hemagglutination inhibition assay showed that H1N1-TMC/NP induced much more protective antibodies than antigen-encapsulated nanoparticles or alum-precipitated antigen (I.M.). In the mechanistic study, H1N1-TMC/NP was shown to stimulate macrophages to produce IL-1β and IL-6 and to stimulate spleen lymphocytes to produce IL-2 and IFN-γ. These results indicated that H1N1-TMC/NP may be an effective vaccine against influenza A (H1N1) viruses for use in nasal immunization. © 2015 Wiley Periodicals, Inc.

Vaccination with Recombinant Parainfluenza Virus 5 Expressing Neuraminidase Protects against Homologous and Heterologous Influenza Virus Challenge

PubMed Central

Mooney, Alaina J.; Gabbard, Jon D.; Li, Zhuo; Dlugolenski, Daniel A.; Johnson, Scott K.

2017-01-01

ABSTRACT Seasonal human influenza virus continues to cause morbidity and mortality annually, and highly pathogenic avian influenza (HPAI) viruses along with other emerging influenza viruses continue to pose pandemic threats. Vaccination is considered the most effective measure for controlling influenza; however, current strategies rely on a precise vaccine match with currently circulating virus strains for efficacy, requiring constant surveillance and regular development of matched vaccines. Current vaccines focus on eliciting specific antibody responses against the hemagglutinin (HA) surface glycoprotein; however, the diversity of HAs across species and antigenic drift of circulating strains enable the evasion of virus-inhibiting antibody responses, resulting in vaccine failure. The neuraminidase (NA) surface glycoprotein, while diverse, has a conserved enzymatic site and presents an appealing target for priming broadly effective antibody responses. Here we show that vaccination with parainfluenza virus 5 (PIV5), a promising live viral vector expressing NA from avian (H5N1) or pandemic (H1N1) influenza virus, elicited NA-specific antibody and T cell responses, which conferred protection against homologous and heterologous influenza virus challenges. Vaccination with PIV5-N1 NA provided cross-protection against challenge with a heterosubtypic (H3N2) virus. Experiments using antibody transfer indicate that antibodies to NA have an important role in protection. These findings indicate that PIV5 expressing NA may be effective as a broadly protective vaccine against seasonal influenza and emerging pandemic threats. IMPORTANCE Seasonal influenza viruses cause considerable morbidity and mortality annually, while emerging viruses pose potential pandemic threats. Currently licensed influenza virus vaccines rely on the antigenic match of hemagglutinin (HA) for vaccine strain selection, and most vaccines rely on HA inhibition titers to determine efficacy, despite the growing awareness of the contribution of neuraminidase (NA) to influenza virus vaccine efficacy. Although NA is immunologically subdominant to HA, and clinical studies have shown variable NA responses to vaccination, in this study, we show that vaccination with a parainfluenza virus 5 recombinant vaccine candidate expressing NA (PIV5-NA) from a pandemic influenza (pdmH1N1) virus or highly pathogenic avian influenza (H5N1) virus elicits robust, cross-reactive protection from influenza virus infection in two animal models. New vaccination strategies incorporating NA, including PIV5-NA, could improve seasonal influenza virus vaccine efficacy and provide protection against emerging influenza viruses. PMID:28931689

Electron microscopy and antigenic studies of uncharacterized viruses. I. Evidence suggesting the placement of viruses in families Arenaviridae, Paramyxoviridae, or Poxviridae.

PubMed

Zeller, H G; Karabatsos, N; Calisher, C H; Digoutte, J P; Murphy, F A; Shope, R E

1989-01-01

During approximately 35 years, investigators in various laboratories studying arbovirus ecology and epidemiology accumulated many virus isolates, more than 60 of which were not characterized or placed in taxa. By a combination of electron microscopic and antigenic studies we collected information sufficient to provisionally classify 60 isolates. Electron microscopic observations suggest that 20 are members of the virus family Bunyaviridae, 20 Rhabdoviridae, 14 Reoviridae, one Togaviridae, one Paramyxoviridae (Mapuera virus, from a bat), and one Poxviridae (Yoka virus, from mosquitoes). Serologic studies provided evidence sufficient to place some of these viruses in recognized antigenic groups, within families and genera, and to establish new antigenic groups and taxa for others. Three viruses were found to have morphologic and morphogenetic characteristics consistent with those of members of the family Arenaviridae: Quaranfil virus, a human pathogen, Johnston Atoll virus, isolated from birds and ticks, and Araguari virus, isolated from an opossum. This, the first in a series of three papers, described methods used for these investigations and also presents descriptions of viruses provisionally placed in the families Arenaviridae, Paramyxoviridae, or Poxviridae. Descriptions of viruses provisionally placed in families Bunyaviridae and Reoviridae are published in the second and third papers, respectively. Viruses of the family Rhabdoviridae have been described separately.

Steric Shielding of Surface Epitopes and Impaired Immune Recognition Induced by the Ebola Virus Glycoprotein

PubMed Central

Francica, Joseph R.; Varela-Rohena, Angel; Medvec, Andrew; Plesa, Gabriela; Riley, James L.; Bates, Paul

2010-01-01

Many viruses alter expression of proteins on the surface of infected cells including molecules important for immune recognition, such as the major histocompatibility complex (MHC) class I and II molecules. Virus-induced downregulation of surface proteins has been observed to occur by a variety of mechanisms including impaired transcription, blocks to synthesis, and increased turnover. Viral infection or transient expression of the Ebola virus (EBOV) glycoprotein (GP) was previously shown to result in loss of staining of various host cell surface proteins including MHC1 and β1 integrin; however, the mechanism responsible for this effect has not been delineated. In the present study we demonstrate that EBOV GP does not decrease surface levels of β1 integrin or MHC1, but rather impedes recognition by steric occlusion of these proteins on the cell surface. Furthermore, steric occlusion also occurs for epitopes on the EBOV glycoprotein itself. The occluded epitopes in host proteins and EBOV GP can be revealed by removal of the surface subunit of GP or by removal of surface N- and O- linked glycans, resulting in increased surface staining by flow cytometry. Importantly, expression of EBOV GP impairs CD8 T-cell recognition of MHC1 on antigen presenting cells. Glycan-mediated steric shielding of host cell surface proteins by EBOV GP represents a novel mechanism for a virus to affect host cell function, thereby escaping immune detection. PMID:20844579

Antigenic Relationships Among Four Herpesviruses

PubMed Central

Blue, W. T.; Plummer, G.

1973-01-01

Common viral antigens were detected, by fluorescent-antibody studies, in cells infected with herpes simplex virus 1, squirrel monkey herpesvirus 1, bovine rhinotracheitis, and equine abortion viruses. The two primate viruses showed slight cross-neutralization. PMID:4351969

Oral immunization with hepatitis B surface antigen expressed in transgenic plants

PubMed Central

Kong, Qingxian; Richter, Liz; Yang, Yu Fang; Arntzen, Charles J.; Mason, Hugh S.; Thanavala, Yasmin

2001-01-01

Oral immunogenicity of recombinant hepatitis B surface antigen (HBsAg) derived from yeast (purified product) or in transgenic potatoes (uncooked unprocessed sample) was compared. An oral adjuvant, cholera toxin, was used to increase immune responses. Transgenic plant material containing HBsAg was the superior means of both inducing a primary immune response and priming the mice to respond to a subsequent parenteral injection of HBsAg. Electron microscopy of transgenic plant samples revealed evidence that the HBsAg accumulated intracellularly; we conclude that natural bioencapsulation of the antigen may provide protection from degradation in the digestive tract until plant cell degradation occurs near an immune effector site in the gut. The correlate of protection from hepatitis B virus infection is serum antibody titers induced by vaccination; the protective level in humans is 10 milliunits/ml or greater. Mice fed HBsAg-transgenic potatoes produced HBsAg-specific serum antibodies that exceeded the protective level and, on parenteral boosting, generated a strong long-lasting secondary antibody response. We have also shown the effectiveness of oral delivery by using a parenteral prime-oral boost immunization schedule. The demonstrated success of oral immunization for hepatitis B virus with an “edible vaccine” provides a strategy for contributing a means to achieve global immunization for hepatitis B prevention and eradication. PMID:11553782

Substitutions near the Hemagglutinin Receptor-Binding Site Determine the Antigenic Evolution of Influenza A H3N2 Viruses in U.S. Swine

PubMed Central

Lewis, Nicola S.; Anderson, Tavis K.; Kitikoon, Pravina; Skepner, Eugene; Burke, David F.

2014-01-01

ABSTRACT Swine influenza A virus is an endemic and economically important pathogen in pigs, with the potential to infect other host species. The hemagglutinin (HA) protein is the primary target of protective immune responses and the major component in swine influenza A vaccines. However, as a result of antigenic drift, vaccine strains must be regularly updated to reflect currently circulating strains. Characterizing the cross-reactivity between strains in pigs and seasonal influenza virus strains in humans is also important in assessing the relative risk of interspecies transmission of viruses from one host population to the other. Hemagglutination inhibition (HI) assay data for swine and human H3N2 viruses were used with antigenic cartography to quantify the antigenic differences among H3N2 viruses isolated from pigs in the United States from 1998 to 2013 and the relative cross-reactivity between these viruses and current human seasonal influenza A virus strains. Two primary antigenic clusters were found circulating in the pig population, but with enough diversity within and between the clusters to suggest updates in vaccine strains are needed. We identified single amino acid substitutions that are likely responsible for antigenic differences between the two primary antigenic clusters and between each antigenic cluster and outliers. The antigenic distance between current seasonal influenza virus H3 strains in humans and those endemic in swine suggests that population immunity may not prevent the introduction of human viruses into pigs, and possibly vice versa, reinforcing the need to monitor and prepare for potential incursions. IMPORTANCE Influenza A virus (IAV) is an important pathogen in pigs and humans. The hemagglutinin (HA) protein is the primary target of protective immune responses and the major target of vaccines. However, vaccine strains must be updated to reflect current strains. Characterizing the differences between seasonal IAV in humans and swine IAV is important in assessing the relative risk of interspecies transmission of viruses. We found two primary antigenic clusters of H3N2 in the U.S. pig population, with enough diversity to suggest updates in swine vaccine strains are needed. We identified changes in the HA protein that are likely responsible for these differences and that may be useful in predicting when vaccines need to be updated. The difference between human H3N2 viruses and those in swine is enough that population immunity is unlikely to prevent new introductions of human IAV into pigs or vice versa, reinforcing the need to monitor and prepare for potential introductions. PMID:24522915

Antiviral efficacy against hepatitis B virus replication of oleuropein isolated from Jasminum officinale L. var. grandiflorum.

PubMed

Zhao, Guiqin; Yin, Zhifeng; Dong, Junxing

2009-09-07

Jasminum officinale L. var. grandiflorum (JOG) is a folk medicine used for the treatment of hepatitis in south of China. Phytochemical studies showed that secoiridoid glycosides are the typical constituents of this plant. The present study was undertaken to evaluate the effect of oleuropein (Ole) derived from the flowers of JOG on hepatitis B virus (HBV) replication in HepG2 2.2.15 cell line in vitro and duck hepatitis B virus (DHBV) replication in ducklings in vivo. The extracellular hepatitis B e antigen (HBeAg) and hepatitis B surface antigen (HBsAg) concentrations in cell culture medium were determined by ELISA. DHBV in duck serum was analyzed by dot blot. Ole blocks effectively HBsAg secretion in HepG2 2.2.15 cells in a dose-dependent manner (IC(50)=23.2 microg/ml). Ole (80 mg/kg, intraperitoneally, twice daily) also reduced viremia in DHBV-infected ducks. Ole therefore warrants further investigation as a potential therapeutic agent for HBV infection.

Heteroassembled gold nanoparticles with sandwich-immunoassay LSPR chip format for rapid and sensitive detection of hepatitis B virus surface antigen (HBsAg).

PubMed

Kim, Jinwoon; Oh, Seo Yeong; Shukla, Shruti; Hong, Seok Bok; Heo, Nam Su; Bajpai, Vivek K; Chun, Hyang Sook; Jo, Cheon-Ho; Choi, Bong Gill; Huh, Yun Suk; Han, Young-Kyu

2018-06-01

This study aimed to develop a more sensitive method for the detection of hepatitis B surface antigen (HBsAg) using heteroassembled gold nanoparticles (AuNPs). A single layered localized surface plasmon resonance (LSPR) chip format was developed with antigen-antibody reaction-based detection symmetry using AuNPs, which detected HBsAg at 10 pg/mL. To further improve the detection limit, a modified detection format was fabricated by fixing a secondary antibody (to form a heteroassembled sandwich format) to the AuNP monolayer, which enhanced the detection sensitivity by about 100 times. The developed heteroassembled AuNPs sandwich-immunoassay LSPR chip format was able to detect as little as 100 fg/mL of HBsAg within 10-15 min. In addition, the heteroassembled AuNPs sandwich-immunoassay LSPR chip format did not show any non-specific binding to other tested antigens, including alpha fetoprotein (AFP), C-reactive protein (CRP), and prostate-specific antigen (PSA). These findings confirm that the proposed detection strategy of heteroassembled AuNPs sandwich-immunoassay LSPR chip format may provide a new platform for early diagnosis of various human diseases. Copyright © 2018 Elsevier B.V. All rights reserved.

In vitro and in vivo evidence of a potential A(H1N1)pdm09 antigenic drift mediated by escape mutations in the haemagglutinin Sa antigenic site.

PubMed

Retamal, Miguel; Abed, Yacine; Rhéaume, Chantal; Baz, Mariana; Boivin, Guy

2017-06-01

Influenza A(H1N1)pdm09 virus continues to circulate worldwide without evidence of significant antigenic drift between 2009 and 2016. By using escape mutants, we previously identified six haemagglutinin (HA) changes (T80R, G143E, G158E, N159D, K166E and A198E) that were located within antigenic sites. Combinations of these mutations were introduced into the A(H1N1)pdm09 HA plasmid by mutagenesis. Reassortant 6 : 2 viruses containing both the HA and NA genes of the A(H1N1)pdm09 and the six internal gene segments of A/PR/8/34 were rescued by reverse genetics. In vitro, HA inhibition and microneutralization assays showed that the HA hexa-mutant reassortant virus (RG1) escaped A(H1N1)pdm09 hyper-immune ferret antiserum recognition. C57Black/6 mice that received the vaccine formulated with A/California/07/09 were challenged with 2×104 p.f.u. of either the 6 : 2 wild-type (WT) or RG1 viruses. Reductions in body weight loss, mortality rate and lung viral titre were observed in immunized animals challenged with the 6 : 2 WT virus compared to non-immunized mice. However, immunization did not protect mice challenged with RG1 virus. To further characterize the mutations causing this antigenic change, 11 additional RG viruses whose HA gene contained single or combinations of mutations were evaluated in vitro. Although the RG1 virus was still the least reactive against hyper-immune serum by HAI testing, mutations G158E and N159D within the Sa antigenic site appeared to play the major role in the altered antigenicity of the A(H1N1)pdm09 virus. These results show that the Sa antigenic site contains the most prominent epitopes susceptible to cause an antigenic drift, escaping actual vaccine protection.

Need for immunization against hepatotropic viruses in children with chronic liver disease.

PubMed

Srivastava, Anshu; Mathias, Amrita; Yachha, Surender Kumar; Agarwal, Jaya; Aggarwal, Rakesh

2014-09-01

Infection with hepatotropic viruses is a common cause of acute deterioration and adverse outcome in children with chronic liver disease (CLD). Such superimposed infections may be preventable through vaccination. The present study aimed to evaluate the exposure rates of hepatitis A, B, and E viruses in children with CLD and suggest an optimal vaccination strategy. Children with CLD were prospectively evaluated with a demographic, clinical, and investigative proforma. Hepatitis B surface antigen positive cases were labeled as hepatitis B virus-CLD, and all other etiologies as non-HBV-related CLD. Patients were tested for exposure to hepatitis A (total anti-hepatitis A virus [HAV], immunoglobulin M anti-HAV), hepatitis B (hepatitis B surface antigen, total anti-hepatitis B core, anti-hepatitis B surface), and hepatitis E (IgG anti-hepatitis E virus). A total of 142 children with CLD (age 9.1 ± 3.7 years, 83 [58.5%] boys) were enrolled. A total of 3.5% (5/142) and 38.7% (55/142) had received HAV and HBV vaccines, respectively. A total of 134 (94.4%) were total anti-HAV positive including 5 postimmunization patients, with higher positivity in those older than 5 years (19/25 vs 115/117; P = 0.001). Of the 115 patients with non-HBV-related CLD, 45 (39.1%) had exposure to HBV (40 total anti-hepatitis B core positive and 5 anti-HBs positive without immunization). Only 28 of 142 (19.7%) patients were IgG anti-HEV positive, with no difference across age. A total of 90.8%, 39.1%, and 19.7% of children with CLD from the developing world are exposed to hepatitis A, B, and E infections, respectively. Selective hepatitis A vaccination (patients younger than 5 years of age) and universal hepatitis B vaccination are required to protect children with CLD. Sanitation improvement and HEV vaccine trial are needed for prevention against HEV.

Hepatitis B virus DNA-positive, hepatitis B surface antigen-negative blood donations intercepted by anti-hepatitis B core antigen testing: the Canadian Blood Services experience.

PubMed

O'Brien, Sheila F; Fearon, Margaret A; Yi, Qi-Long; Fan, Wenli; Scalia, Vito; Muntz, Irene R; Vamvakas, Eleftherios C

2007-10-01

The benefit of introducing anti-hepatitis B core antigen (HBc) screening for intercepting potentially infectious donations missed by hepatitis B surface antigen (HBsAg) screening in Canada was studied. Anti-HBc testing of all donations was implemented in April 2005, along with antibody to hepatitis B surface antigen (anti-HBs) and hepatitis B virus (HBV) DNA supplemental testing of anti-HBc repeat-reactive, HBsAg-negative donations. The proportion of potentially infectious donations intercepted by anti-HBc over the initial 18 months of testing was calculated based on three assumptions relating infectivity of HBV DNA-positive units to anti-HBs levels. Lookback was conducted for all DNA-positive donations. Of 493,344 donors, 5,585 (1.13%) were repeat-reactive for the presence of anti-HBc, with 29 (0.52%) being HBV DNA-positive and HBsAg-negative. The proportion of potentially infectious donations intercepted by anti-HBc screening was 1 in 17,800 if all HBV DNA-positive donations were infectious, 1 in 26,900 if infectivity was limited to donations with an anti-HBs level of not more than 100 mIU per mL, and 1 in 69,300 if only donations with undetectable anti-HBs were infectious. For 279 components in the lookback study, no traced recipients were HBsAg-positive and 7 recipients were anti-HBc-reactive in association with 4 donors, 3 of whom had an anti-HBs level of more than 100 mIU per mL and 1 of whom had a level of 61 mIU per mL. Implementation of anti-HBc screening reduced the risk of transfusing potentially infectious units by at least as much as had been expected based on the literature. The lookback did not provide proof of transfusion transmission of HBV from HBV DNA-positive, anti-HBc-reactive, HBsAg-negative donors but it did not establish lack of transmission either.

Non-invasive optical detection of HBV based on serum surface-enhanced Raman spectroscopy

NASA Astrophysics Data System (ADS)

Zheng, Zuci; Wang, Qiwen; Weng, Cuncheng; Lin, Xueliang; Lin, Yao; Feng, Shangyuan

2016-10-01

An optical method of surface-enhanced Raman spectroscopy (SERS) was developed for non-invasive detection of hepatitis B surface virus (HBV). Hepatitis B virus surface antigen (HBsAg) is an established serological marker that is routinely used for the diagnosis of acute or chronic hepatitis B virus(HBV) infection. Utilizing SERS to analyze blood serum for detecting HBV has not been reported in previous literature. SERS measurements were performed on two groups of serum samples: one group for 50 HBV patients and the other group for 50 healthy volunteers. Blood serum samples are collected from healthy control subjects and patients diagnosed with HBV. Furthermore, principal components analysis (PCA) combined with linear discriminant analysis (LDA) were employed to differentiate HBV patients from healthy volunteer and achieved sensitivity of 80.0% and specificity of 74.0%. This exploratory work demonstrates that SERS serum analysis combined with PCA-LDA has tremendous potential for the non-invasive detection of HBV.

Genome-wide evolutionary dynamics of influenza B viruses on a global scale

PubMed Central

Langat, Pinky; Bowden, Thomas A.; Edwards, Stephanie; Gall, Astrid; Rambaut, Andrew; Daniels, Rodney S.; Russell, Colin A.; Pybus, Oliver G.; McCauley, John

2017-01-01

The global-scale epidemiology and genome-wide evolutionary dynamics of influenza B remain poorly understood compared with influenza A viruses. We compiled a spatio-temporally comprehensive dataset of influenza B viruses, comprising over 2,500 genomes sampled worldwide between 1987 and 2015, including 382 newly-sequenced genomes that fill substantial gaps in previous molecular surveillance studies. Our contributed data increase the number of available influenza B virus genomes in Europe, Africa and Central Asia, improving the global context to study influenza B viruses. We reveal Yamagata-lineage diversity results from co-circulation of two antigenically-distinct groups that also segregate genetically across the entire genome, without evidence of intra-lineage reassortment. In contrast, Victoria-lineage diversity stems from geographic segregation of different genetic clades, with variability in the degree of geographic spread among clades. Differences between the lineages are reflected in their antigenic dynamics, as Yamagata-lineage viruses show alternating dominance between antigenic groups, while Victoria-lineage viruses show antigenic drift of a single lineage. Structural mapping of amino acid substitutions on trunk branches of influenza B gene phylogenies further supports these antigenic differences and highlights two potential mechanisms of adaptation for polymerase activity. Our study provides new insights into the epidemiological and molecular processes shaping influenza B virus evolution globally. PMID:29284042

Simian virus 40 large T antigen associates with cyclin A and p33cdk2.

PubMed Central

Adamczewski, J P; Gannon, J V; Hunt, T

1993-01-01

In this paper we provide evidence that a fraction of large T antigen of simian virus 40 (SV40) interacts with cyclin A and p33cdk2 in both virus-infected and stably transformed cells. Immunoprecipitates of SV40 large T antigen from SV40-infected or SV40 large-T-antigen-transformed cells contain cyclin A, p33cdk2, and histone H1 kinase activity. Conversely, immunoprecipitates of cyclin A from these cells contain SV40 large T antigen. In this respect, SV40 large T antigen has properties similar to those of the E1A oncogene of adenoviruses and the E7 oncogene of human papillomaviruses. Images PMID:8411358

[Behavior of Orf virus in permissive and nonpermissive systems].

PubMed

Büttner, M; Czerny, C P; Schumm, M

1995-04-01

Dogs were immunized i.m. with attenuated poxvirus vaccines (vaccinia virus, Orf-virus) and a bovine herpesvirus-1 (BHV-1) vaccine. After intradermal (i.d.) application of the vaccine viruses a specific delayed type hypersensitivity (DTH) reaction of the skin occurred only with vaccinia virus. The i.d. application of Orf-virus caused a short-term, non-specific inflammatory reaction of the skin, even in dogs not immunized with Orf-virus. Out of 30 sera from Orf-virus immunized beagles (n = 4) only eight were found reactive to Orf-virus in a competition ELISA. Three sera from dogs not Orf-virus immunized but skin-tested with the virus contained low antibody titers. Using indirect immunofluorescence (IIF) in flow cytometry, the existence of Orf-virus antigens was examined on the surface and in the cytoplasm of permissive (BFK and Vero)- and questionable permissive MDCK cells. The canine kidney MDCK cell line was found to be non-permissive for Orf-virus replication; the occurrence of an Orf-(ecthyma contagiosum) like disease in dogs is unlikely.

Virus-like particles as universal influenza vaccines

PubMed Central

Kang, Sang-Moo; Kim, Min-Chul; Compans, Richard W

2012-01-01

Current influenza vaccines are primarily targeted to induce immunity to the influenza virus strain-specific hemagglutinin antigen and are not effective in controlling outbreaks of new pandemic viruses. An approach for developing universal vaccines is to present highly conserved antigenic epitopes in an immunogenic conformation such as virus-like particles (VLPs) together with an adjuvant to enhance the vaccine immunogenicity. In this review, the authors focus on conserved antigenic targets and molecular adjuvants that were presented in VLPs. Conserved antigenic targets that include the hemagglutinin stalk domain, the external domain of influenza M2 and neuraminidase are discussed in addition to molecular adjuvants that are engineered to be incorporated into VLPs in a membrane-anchored form. PMID:23002980

Vaccine efficacy of live-attenuated virus, whole inactivated virus and alphavirus vectored subunit vaccines against antigenically distinct H3N2 swine influenza A viruses

USDA-ARS?s Scientific Manuscript database

Introduction Influenza A virus (IAV) is an important pathogen in swine, and the main intervention strategy is vaccination to induce neutralizing antibodies against the hemagglutinin (HA). Three major antigenic clusters, cyan, red, and green, were identified among H3N2 viruses circulating in pigs in ...

Reemergence of enterovirus 71 epidemic in northern Taiwan, 2012.

PubMed

Luo, Shu-Ting; Chiang, Pai-Shan; Chung, Wan-Yu; Chia, Min-Yuan; Tsao, Kuo-Chien; Wang, Ying-Hsiang; Lin, Tzou-Yien; Lee, Min-Shi

2015-01-01

Enterovirus 71 (EV71) belongs to picornavirus family and could be classified phylogenetically into three major genogroups (A, B and C) including 11 genotypes (A, B1-B5 and C1-C5). Since 1997, EV71 has caused large-scale of epidemics with neurological complications in Asian children. In Taiwan, nationwide EV71 epidemics with different predominant genotypes have occurred cyclically since 1998. A nationwide EV71 epidemic occurred again in 2012. We conducted genetic and antigenic characterizations of the 2012 epidemic. Chang Gung Memorial Hospital (CGMH) is a medical center in northern Taiwan. In CGMH, specimens were collected from pediatric inpatients with suspected enterovirus infections for virus isolation. Enterovirus isolates were serotyped and genotyped and sera from EV71 inpatients were collected for measuring neutralizing antibody titers. There were 10, 16 and 99 EV71 inpatients identified in 2010, 2011 and 2012, respectively. There were 82 EV71 isolates genotyped, which identified 17 genotype C4a viruses and 65 genotype B5 viruses. The genotype B5 viruses were not detected until November 2011 and caused epidemics in 2012. Interestingly, the B5-2011 viruses were genetically distinguishable from the B5 viruses causing the 2008 epidemic and are likely introduced from China or Southeastern Asia. Based on antigenic analysis, minor antigenic variations were detected among the B5-2008, B5-2011, C4a-2008 and C4a-2012 viruses but these viruses antigenically differed from genotype A. Genotype B5 and C4a viruses antigenically differ from genotype A viruses which have disappeared globally for 30 years but have been detected in China since 2008. Enterovirus surveillance should monitor genetic and antigenic variations of EV71.

Impact of Human Immunodeficiency Virus Infection in Pregnant Women on Variant-Specific Immunity to Malaria▿

PubMed Central

Dembo, Edson G.; Mwapasa, Victor; Montgomery, Jacqui; Craig, Alister G.; Porter, Kimberly A.; Meshnick, Steven R.; Molyneux, Malcolm E.; Rogerson, Stephen J.

2008-01-01

Human immunodeficiency virus (HIV) increases susceptibility to Plasmodium falciparum infection, and this has most clearly been demonstrated in pregnant women. Variant surface antigens on the surfaces of erythrocytes infected with P. falciparum are major targets of protective immunity. We studied the impact of HIV infection on pregnant women's humoral immunity to variant surface antigens expressed by placental and pediatric isolates of P. falciparum. By flow cytometry, sera from HIV-infected women more frequently lacked antibodies to these antigens than sera from HIV-uninfected women. This difference was similar in magnitude for pediatric isolates (unadjusted odds ratio [OR] = 6.36; 95% confidence interval [CI] = 1.14, 35.32; P < 0.05) and placental isolates (unadjusted OR = 6.47; 95% CI = 0.75, 55.64; P < 0.10). We divided women into high and low responders on the basis of their antibody levels. After adjustment for CD4 count, maternal age, and gravidity, we found that HIV-infected women more frequently had low responses to both pediatric isolates (OR = 5.34; 95% CI = 1.23, 23.16; P = 0.025) and placental isolates (OR = 4.14; 95% CI = 1.71, 10.02; P = 0.002). The relative quantity of antibodies to both pediatric isolates (P = 0.035) and placental isolates (P = 0.005) was lower in HIV-infected women than in HIV-uninfected women. HIV infection has a broad impact on variant-specific immunity, which may explain the susceptibility of infected individuals to clinical malaria episodes. PMID:18199738

Optimization of peptide arrays for studying antibodies to hepatitis C virus continuous epitopes

PubMed Central

Ruwona, Tinashe B; Mcbride, Ryan; Chappel, Rebecca; Head, Steven R; Ordoukhanian, Phillip; Burton, Dennis R.; Law, Mansun

2014-01-01

Accurate and in-depth mapping of antibody responses is of great value in vaccine and antibody research. Using hepatitis C virus (HCV) as a model, we developed an affordable and high-throughput microarray-based assay for mapping antibody specificities to continuous antibody epitopes of HCV at high resolution. Important parameters in the chemistry for conjugating peptides/antigens to the array surface, the array layout, fluorophore choice and the methods for data analysis were investigated. Microscopic glass slide pre-coated with N-Hydroxysuccinimide (NHS)-ester (Slide H) was the preferred surface for conjugation of aminooxy-tagged peptides. This combination provides a simple chemical means to orient the peptides to the conjugation surface via an orthogonal covalent linkage at the N- or C-terminus of each peptide. The addition of polyvinyl alcohol to printing buffer gave uniform spot morphology, improved sensitivity and specificity of binding signals. Libraries of overlapping peptides covering the HCV E1 and E2 glycoprotein polypeptides (15-mer, 10 amino acids overlap) of 6 major HCV genotypes and the entire polypeptide sequence of the prototypic strain H77 were synthesized and printed in quadruplets in the assays. The utility of the peptide arrays were confirmed using HCV monoclonal antibodies (mAbs) specific to known continuous epitopes and immune sera of rabbits immunized with HCV antigens. The methods developed here can be easily adapted to studying antibody responses to antigens relevant in vaccine and autoimmune research. PMID:24269751

A Longitudinal Hepatitis B Vaccine Cohort Demonstrates Long-lasting Hepatitis B Virus (HBV) Cellular Immunity Despite Loss of Antibody Against HBV Surface Antigen

PubMed Central

Simons, Brenna C.; Spradling, Philip R.; Bruden, Dana J. T.; Zanis, Carolyn; Case, Samantha; Choromanski, Tammy L.; Apodaca, Minjun; Brogdon, Hazel D.; Dwyer, Gaelen; Snowball, Mary; Negus, Susan; Bruce, Michael G.; Morishima, Chihiro; Knall, Cindy; McMahon, Brian J.

2016-01-01

Background. Long-lasting protection resulting from hepatitis B vaccine, despite loss of antibody against hepatitis B virus (HBV) surface antigen (anti-HBs), is undetermined. Methods. We recruited persons from a cohort vaccinated with plasma-derived hepatitis B vaccine in 1981 who have been followed periodically since. We performed serological testing for anti-HBs and microRNA-155 and assessed HBV-specific T-cell responses by enzyme-linked immunospot and cytometric bead array. Study subgroups were defined 32 years after vaccination as having an anti-HBs level of either ≥10 mIU/mL (group 1; n = 13) or <10 mIU/mL (group 2; n = 31). Results. All 44 participants, regardless of anti-HBs level, tested positive for tumor necrosis factor α, interleukin 10, or interleukin 6 production by HBV surface antigen–specific T cells. The frequency of natural killer T cells correlated with the level of anti-HBs (P = .008). The proportion of participants who demonstrated T-cell responses to HBV core antigen varied among the cytokines measured, suggesting some natural exposure to HBV in the study group. No participant had evidence of breakthrough HBV infection. Conclusions. Evidence of long-lasting cellular immunity, regardless of anti-HBs level, suggests that protection afforded by primary immunization with plasma-derived hepatitis B vaccine during childhood and adulthood lasts at least 32 years. PMID:27056956

Characterization of an antigenic site that contains a dominant, type-specific neutralization determinant on the envelope protein domain III (ED3) of dengue 2 virus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gromowski, Gregory D.; Barrett, Alan D.T.

2007-09-30

The surface of the mature dengue virus (DENV) particle consists of 90 envelope (E) protein dimers that mediate both receptor binding and fusion. The E protein ectodomain can be divided into three structural domains designated ED1, ED2, and ED3, of which ED3 contains the critical and dominant virus-specific neutralization sites. In this study the ED3 epitopes recognized by seven, murine, IgG1 DENV-2 type-specific, monoclonal antibodies (MAbs) were determined using site-directed mutagenesis of a recombinant DENV-2 ED3 (rED3) protein. A total of 41 single amino acid substitutions were introduced into the rED3 at 30 different surface accessible residues. The affinity ofmore » each MAb with the mutant rED3s was assessed by indirect ELISA and the results indicate that all seven MAbs recognize overlapping epitopes with residues K305 and P384 critical for binding. These residues are conserved among DENV-2 strains and cluster together on the upper lateral face of ED3. A linear relationship was observed between relative occupancy of ED3 on the virion by MAb and neutralization of the majority of virus infectivity ({approx} 90%) for all seven MAbs. Depending on the MAb, it is predicted that between 10% and 50% relative occupancy of ED3 on the virion is necessary for virus neutralization and for all seven MAbs occupancy levels approaching saturation were required for 100% neutralization of virus infectivity. Overall, the conserved antigenic site recognized by all seven MAbs is likely to be a dominant DENV-2 type-specific, neutralization determinant.« less

Antigenic variants of influenza A virus, PR8 strain. I. Their development during serial passage in the lungs of partially immune mice.

PubMed

GERBER, P; LOOSLI, C G; HAMBRE, D

1955-06-01

Antigenically different strains of mouse-adapted PR8 influenza A virus have been produced by 17 serial passages of the virus in the lungs of mice immunized with the homologous agent. Comparative serological tests show that the variant strains share antigenic components with the parent strain but the dominant antigen is different. By means of antibody absorption it was shown that the "new" antigenic component of the variant was already present in minor amounts up to the eighth passage and thereafter gained prominence with continued passage in vaccinated mice. Groups of mice vaccinated with either the PR8-S or T(21) virus and having comparable antibody titers showed no growth of virus in the lungs following aid-borne challenge with homologous strains. On the other hand, following heterologous air-borne challenge no deaths occurred, but virus grew in the lungs of both groups of vaccinated mice. Almost unrestricted virus multiplication took place in the lungs of mice vaccinated with the parent strain and challenged with the PR8-T(21) virus which resulted in extensive consolidation. Less virus grew in the lungs of the mice vaccinated with the variant strains and challenged with the PR8-S virus. In these animals only microscopic evidence of changes due to virus growth in the lungs was observed. The successful serial passage of PR8 influenza A virus in immunized animals was dependent on the initial selection of mice with uniformly low H.I. antibody titers as determined on tail blood, and the intranasal instillation of sufficient virus to favor the survival of those virus particles least related to the antibodies present. The epidemiological implications of these observations are discussed briefly.

ANTIGENIC VARIANTS OF INFLUENZA A VIRUS (PR8 STRAIN)

PubMed Central

Gerber, Paul; Loosli, Clayton G.; Hamre, Dorothy

1955-01-01

Antigenically different strains of mouse-adapted PR8 influenza A virus have been produced by 17 serial passages of the virus in the lungs of mice immunized with the homologous agent. Comparative serological tests show that the variant strains share antigenic components with the parent strain but the dominant antigen is different. By means of antibody absorption it was shown that the "new" antigenic component of the variant was already present in minor amounts up to the eighth passage and thereafter gained prominence with continued passage in vaccinated mice. Groups of mice vaccinated with either the PR8-S or T21 virus and having comparable antibody titers showed no growth of virus in the lungs following aid-borne challenge with homologous strains. On the other hand, following heterologous air-borne challenge no deaths occurred, but virus grew in the lungs of both groups of vaccinated mice. Almost unrestricted virus multiplication took place in the lungs of mice vaccinated with the parent strain and challenged with the PR8-T21 virus which resulted in extensive consolidation. Less virus grew in the lungs of the mice vaccinated with the variant strains and challenged with the PR8-S virus. In these animals only microscopic evidence of changes due to virus growth in the lungs was observed. The successful serial passage of PR8 influenza A virus in immunized animals was dependent on the initial selection of mice with uniformly low H.I. antibody titers as determined on tail blood, and the intranasal instillation of sufficient virus to favor the survival of those virus particles least related to the antibodies present. The epidemiological implications of these observations are discussed briefly. PMID:14367684

Detection of avian influenza antigens in proximity fiber, droplet, and optical waveguide microfluidics

NASA Astrophysics Data System (ADS)

Yoon, Jeong-Yeol; Heinze, Brian C.; Gamboa, Jessica; You, David J.

2009-05-01

Virus antigens of avian influenza subtype H3N2 were detected on two different microfluidic platforms: microchannel and droplet. Latex immunoagglutination assays were performed using 920-nm highly carboxylated polystyrene beads that are conjugated with antibody to avian influenza virus. The bead suspension was merged with the solutions of avian influenza virus antigens in a Y-junction of a microchannel made by polydimethylsiloxane soft lithography. The resulting latex immunoagglutinations were measured with two optical fibers in proximity setup to detect 45° forward light scattering. Alternatively, 10 μL droplets of a bead suspension and an antigen solution were merged on a superhydrophobic surface (water contact angle = 155°), whose movement was guided by a metal wire, and 180° back light scattering is measured with a backscattering optical probe. Detection limits were 0.1 pg mL-1 for both microchannel with proximity fibers and droplet microfluidics, thanks to the use of micro-positioning stages to help generate reproducible optical signals. Additionally, optical waveguide was tested by constructing optical waveguide channels (filled with mineral oil) within a microfluidic device to detect the same light scattering. Detection limit was 0.1 ng mL-1 for an optical waveguide device, with a strong potential of improvement in the near future. The use of optical waveguide enabled smaller device setup, easier operation, smaller standard deviations and broader linear range of assay than proximity fiber microchannel and droplet microfluidics. Total assay time was less than 10 min.

Immunization of Pigs by DNA Prime and Recombinant Vaccinia Virus Boost To Identify and Rank African Swine Fever Virus Immunogenic and Protective Proteins

PubMed Central

2018-01-01

ABSTRACT African swine fever virus (ASFV) causes an acute hemorrhagic fever in domestic pigs, with high socioeconomic impact. No vaccine is available, limiting options for control. Although live attenuated ASFV can induce up to 100% protection against lethal challenge, little is known of the antigens which induce this protective response. To identify additional ASFV immunogenic and potentially protective antigens, we cloned 47 viral genes in individual plasmids for gene vaccination and in recombinant vaccinia viruses. These antigens were selected to include proteins with different functions and timing of expression. Pools of up to 22 antigens were delivered by DNA prime and recombinant vaccinia virus boost to groups of pigs. Responses of immune lymphocytes from pigs to individual recombinant proteins and to ASFV were measured by interferon gamma enzyme-linked immunosorbent spot (ELISpot) assays to identify a subset of the antigens that consistently induced the highest responses. All 47 antigens were then delivered to pigs by DNA prime and recombinant vaccinia virus boost, and pigs were challenged with a lethal dose of ASFV isolate Georgia 2007/1. Although pigs developed clinical and pathological signs consistent with acute ASFV, viral genome levels were significantly reduced in blood and several lymph tissues in those pigs immunized with vectors expressing ASFV antigens compared with the levels in control pigs. IMPORTANCE The lack of a vaccine limits the options to control African swine fever. Advances have been made in the development of genetically modified live attenuated ASFV that can induce protection against challenge. However, there may be safety issues relating to the use of these in the field. There is little information about ASFV antigens that can induce a protective immune response against challenge. We carried out a large screen of 30% of ASFV antigens by delivering individual genes in different pools to pigs by DNA immunization prime and recombinant vaccinia virus boost. The responses in immunized pigs to these individual antigens were compared to identify the most immunogenic. Lethal challenge of pigs immunized with a pool of antigens resulted in reduced levels of virus in blood and lymph tissues compared to those in pigs immunized with control vectors. Novel immunogenic ASFV proteins have been identified for further testing as vaccine candidates. PMID:29386289

Immunization of Pigs by DNA Prime and Recombinant Vaccinia Virus Boost To Identify and Rank African Swine Fever Virus Immunogenic and Protective Proteins.

PubMed

Jancovich, James K; Chapman, Dave; Hansen, Debra T; Robida, Mark D; Loskutov, Andrey; Craciunescu, Felicia; Borovkov, Alex; Kibler, Karen; Goatley, Lynnette; King, Katherine; Netherton, Christopher L; Taylor, Geraldine; Jacobs, Bertram; Sykes, Kathryn; Dixon, Linda K

2018-04-15

African swine fever virus (ASFV) causes an acute hemorrhagic fever in domestic pigs, with high socioeconomic impact. No vaccine is available, limiting options for control. Although live attenuated ASFV can induce up to 100% protection against lethal challenge, little is known of the antigens which induce this protective response. To identify additional ASFV immunogenic and potentially protective antigens, we cloned 47 viral genes in individual plasmids for gene vaccination and in recombinant vaccinia viruses. These antigens were selected to include proteins with different functions and timing of expression. Pools of up to 22 antigens were delivered by DNA prime and recombinant vaccinia virus boost to groups of pigs. Responses of immune lymphocytes from pigs to individual recombinant proteins and to ASFV were measured by interferon gamma enzyme-linked immunosorbent spot (ELISpot) assays to identify a subset of the antigens that consistently induced the highest responses. All 47 antigens were then delivered to pigs by DNA prime and recombinant vaccinia virus boost, and pigs were challenged with a lethal dose of ASFV isolate Georgia 2007/1. Although pigs developed clinical and pathological signs consistent with acute ASFV, viral genome levels were significantly reduced in blood and several lymph tissues in those pigs immunized with vectors expressing ASFV antigens compared with the levels in control pigs. IMPORTANCE The lack of a vaccine limits the options to control African swine fever. Advances have been made in the development of genetically modified live attenuated ASFV that can induce protection against challenge. However, there may be safety issues relating to the use of these in the field. There is little information about ASFV antigens that can induce a protective immune response against challenge. We carried out a large screen of 30% of ASFV antigens by delivering individual genes in different pools to pigs by DNA immunization prime and recombinant vaccinia virus boost. The responses in immunized pigs to these individual antigens were compared to identify the most immunogenic. Lethal challenge of pigs immunized with a pool of antigens resulted in reduced levels of virus in blood and lymph tissues compared to those in pigs immunized with control vectors. Novel immunogenic ASFV proteins have been identified for further testing as vaccine candidates. Copyright © 2018 Jancovich et al.

Effect of respiratory syncytial virus (RSV) infection on the adherence of pathogenic bacteria to human epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Faden, H.; Hong, J.J.; Ogra, P.L.

1986-03-01

The effect of RSV infection on the adherence of Streptococcus pneumoniae (SP), Haemophilus influenzae (HI) and Staphylococcus aureus (SA) to human epithelial cells was determined. RSV-infected Hep-2 cell cultures at different stages of expression of surface viral antigens and bacteria labeled with /sup 3/H-thymidine were employed to examine the kinetics of bacterial adherence to virus-infected cells. RSV infection did not alter the magnitude of adherence of HI or SA to HEp-2 cells. However, adherence of SP to HEp-2 cells was significantly (P < 0.01) enhanced by prior RSV infection. The degree of adherence was directly related to the amount ofmore » viral antigen expressed on the cell surface. The adherence was temperature dependent, with maximal adherence observed at 37/sup 0/C. Heat-inactivation of SP did not alter adherence characteristics. These data suggest that RSV infection increases adherence of SP to the surface of epithelial cells in vitro. Since attachment of bacteria to mucosal surfaces is the first step in many infections, it is suggested that viral infections of epithelial cells render them more susceptible to bacterial adherence. Thus, RSV infection in vivo may predispose children to SP infections, such as in otitis media, by increasing colonization with SP.« less

Characterization of the interface of the bone marrow stromal cell antigen 2-Vpu protein complex via computational chemistry.

PubMed

Zhou, Jinming; Zhang, Zhixin; Mi, Zeyun; Wang, Xin; Zhang, Quan; Li, Xiaoyu; Liang, Chen; Cen, Shan

2012-02-14

Bone marrow stromal cell antigen 2 (BST-2) inhibits the release of enveloped viruses from the cell surface. Various viral counter measures have been discovered, which allow viruses to escape BST-2 restriction. Human immunodeficiency virus type 1 (HIV-1) encodes viral protein U (Vpu) that interacts with BST-2 through their transmembrane domains and causes the downregulation of cell surface BST-2. In this study, we used a computer modeling method to establish a molecular model to investigate the binding interface of the transmembrane domains of BST-2 and Vpu. The model predicts that the interface is composed of Vpu residues I6, A10, A14, A18, V25, and W22 and BST-2 residues L23, I26, V30, I34, V35, L41, I42, and T45. Introduction of mutations that have been previously reported to disrupt the Vpu-BST-2 interaction led to a calculated higher binding free energy (MMGBSA), which supports our molecular model. A pharmacophore was also generated on the basis of this model. Our results provide a precise model that predicts the detailed interaction occurring between the transmembrane domains of Vpu and BST-2 and should facilitate the design of anti-HIV agents that are able to disrupt this interaction.

Benchtop Antigen Detection Technique using Nanofiltration and Fluorescent Dyes

NASA Technical Reports Server (NTRS)

Scardelletti, Maximilian C.; Varaljay, Vanessa

2009-01-01

The designed benchtop technique is primed to detect bacteria and viruses from antigenic surface marker proteins in solutions, initially water. This inclusive bio-immunoassay uniquely combines nanofiltration and near infrared (NIR) dyes conjugated to antibodies to isolate and distinguish microbial antigens, using laser excitation and spectrometric analysis. The project goals include detecting microorganisms aboard the International Space Station, space shuttle, Crew Exploration Vehicle (CEV), and human habitats on future Moon and Mars missions, ensuring astronaut safety. The technique is intended to improve and advance water contamination testing both commercially and environmentally as well. Lastly, this streamlined technique poses to greatly simplify and expedite testing of pathogens in complex matrices, such as blood, in hospital and laboratory clinics.

Development of an algorithm for production of inactivated arbovirus antigens in cell culture

PubMed Central

Goodman, C.H.; Russell, B.J.; Velez, J.O.; Laven, J.J.; Nicholson, W.L; Bagarozzi, D.A.; Moon, J.L.; Bedi, K.; Johnson, B.W.

2015-01-01

Arboviruses are medically important pathogens that cause human disease ranging from a mild fever to encephalitis. Laboratory diagnosis is essential to differentiate arbovirus infections from other pathogens with similar clinical manifestations. The Arboviral Diseases Branch (ADB) reference laboratory at the CDC Division of Vector-Borne Diseases (DVBD) produces reference antigens used in serological assays such as the virus-specific immunoglobulin M antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA). Antigen production in cell culture has largely replaced the use of suckling mice; however, the methods are not directly transferable. The development of a cell culture antigen production algorithm for nine arboviruses from the three main arbovirus families, Flaviviridae, Togaviridae, and Bunyaviridae, is described here. Virus cell culture growth and harvest conditions were optimized, inactivation methods were evaluated, and concentration procedures were compared for each virus. Antigen performance was evaluated by the MAC-ELISA at each step of the procedure. The antigen production algorithm is a framework for standardization of methodology and quality control; however, a single antigen production protocol was not applicable to all arboviruses and needed to be optimized for each virus. PMID:25102428

A small antigenic determinant of the Chikungunya virus E2 protein is sufficient to induce neutralizing antibodies which are partially protective in mice.

PubMed

Weber, Christopher; Büchner, Sarah M; Schnierle, Barbara S

2015-04-01

The mosquito-borne Chikungunya virus (CHIKV) causes high fever and severe joint pain in humans. It is expected to spread in the future to Europe and has recently reached the USA due to globalization, climate change and vector switch. Despite this, little is known about the virus life cycle and, so far, there is no specific treatment or vaccination against Chikungunya infections. We aimed here to identify small antigenic determinants of the CHIKV E2 protein able to induce neutralizing immune responses. E2 enables attachment of the virus to target cells and a humoral immune response against E2 should protect from CHIKV infections. Seven recombinant proteins derived from E2 and consisting of linear and/or structural antigens were created, and were expressed in and purified from E. coli. BALB/c mice were vaccinated with these recombinant proteins and the mouse sera were screened for neutralizing antibodies. Whereas a linear N-terminally exposed peptide (L) and surface-exposed parts of the E2 domain A (sA) alone did not induce neutralizing antibodies, a construct containing domain B and a part of the β-ribbon (called B+) was sufficient to induce neutralizing antibodies. Furthermore, domain sA fused to B+ (sAB+) induced the highest amount of neutralizing antibodies. Therefore, the construct sAB+ was used to generate a recombinant modified vaccinia virus Ankara (MVA), MVA-CHIKV-sAB+. Mice were vaccinated with MVA-CHIKV-sAB+ and/or the recombinant protein sAB+ and were subsequently challenged with wild-type CHIKV. Whereas four vaccinations with MVA-CHIKV-sAB+ were not sufficient to protect mice from a CHIKV infection, protein vaccination with sAB+ markedly reduced the viral titers of vaccinated mice. The recombinant protein sAB+ contains important structural antigens for a neutralizing antibody response in mice and its formulation with appropriate adjuvants might lead to a future CHIKV vaccine.

Alanine scanning of the rabies virus glycoprotein antigenic site III using recombinant rabies virus: implication for post-exposure treatment.

PubMed

Papaneri, Amy B; Wirblich, Christoph; Marissen, Wilfred E; Schnell, Matthias J

2013-12-02

The safety and availability of the human polyclonal sera that is currently utilized for post-exposure treatment (PET) of rabies virus (RABV) infection remain a concern. Recombinant monoclonal antibodies have been postulated as suitable alternatives by WHO. To this extent, CL184, the RABV human antibody combination comprising monoclonal antibodies (mAbs) CR57 and CR4098, has been developed and has delivered promising clinical data to support its use for RABV PET. For this fully human IgG1 cocktail, mAbs CR57 and CR4098 are produced in the PER.C6 human cell line and combined in equal amounts in the final product. During preclinical evaluation, CR57 was shown to bind to antigenic site I whereas CR4098 neutralization was influenced by a mutation of position 336 (N336) located within antigenic site III. Here, alanine scanning was used to analyze the influence of mutations within the potential binding site for CR4098, antigenic site III, in order to evaluate the possibility of mutated rabies viruses escaping neutralization. For this approach, twenty flanking amino acids (10 upstream and 10 downstream) of the RABV glycoprotein (G) asparagine (N336) were exchanged to alanine (or serine, if already alanine) by site-directed mutagenesis. Analysis of G expression revealed four of the twenty mutant Gs to be non-functional, as shown by their lack of cell surface expression, which is a requirement for the production of infectious RABV. Therefore, these mutants were excluded from further study. The remaining sixteen mutants were introduced in an infectious clone of RABV, and recombinant RABVs (rRABVs) were recovered and utilized for in vitro neutralization assays. All of the viruses were effectively neutralized by CR4098 as well as by CR57, indicating that single amino acid exchanges in this region does not affect the broad neutralizing capability of the CL184 mAb combination. Copyright © 2013 Elsevier Ltd. All rights reserved.

Detection of Antigenic Variants of Subtype H3 Swine Influenza A Viruses from Clinical Samples

PubMed Central

Martin, Brigitte E.; Li, Lei; Nolting, Jacqueline M.; Smith, David R.; Hanson, Larry A.

2017-01-01

ABSTRACT A large population of genetically and antigenically diverse influenza A viruses (IAVs) are circulating among the swine population, playing an important role in influenza ecology. Swine IAVs not only cause outbreaks among swine but also can be transmitted to humans, causing sporadic infections and even pandemic outbreaks. Antigenic characterizations of swine IAVs are key to understanding the natural history of these viruses in swine and to selecting strains for effective vaccines. However, influenza outbreaks generally spread rapidly among swine, and the conventional methods for antigenic characterization require virus propagation, a time-consuming process that can significantly reduce the effectiveness of vaccination programs. We developed and validated a rapid, sensitive, and robust method, the polyclonal serum-based proximity ligation assay (polyPLA), to identify antigenic variants of subtype H3N2 swine IAVs. This method utilizes oligonucleotide-conjugated polyclonal antibodies and quantifies antibody-antigen binding affinities by quantitative reverse transcription-PCR (RT-PCR). Results showed the assay can rapidly detect H3N2 IAVs directly from nasal wash or nasal swab samples collected from laboratory-challenged animals or during influenza surveillance at county fairs. In addition, polyPLA can accurately separate the viruses at two contemporary swine IAV antigenic clusters (H3N2 swine IAV-α and H3N2 swine IAV-ß) with a sensitivity of 84.9% and a specificity of 100.0%. The polyPLA can be routinely used in surveillance programs to detect antigenic variants of influenza viruses and to select vaccine strains for use in controlling and preventing disease in swine. PMID:28077698

Predominance of influenza A(H3N2) viruses during the 2016/2017 season in Bulgaria.

PubMed

Korsun, Neli; Angelova, Svetla; Trifonova, Ivelina; Tzotcheva, Iren; Mileva, Sirma; Voleva, Silvia; Georgieva, Irina; Perenovska, Penka

2018-02-01

Influenza viruses are characterised by high variability, which makes them able to cause annual epidemics. The aim of this study is to determine the antigenic and genetic characteristics of influenza viruses circulating in Bulgaria during the 2016/2017 season. The detection and typing/subtyping of influenza viruses were performed using real time RT-PCR. Results of antigenic characterisation, phylogenetic and amino acid sequence analyses of representative influenza strains are presented herein. The 2016/2017 season was characterised by an early start, an exclusive dominance of A(H3N2) viruses accounting for 93 % of total influenza virus detections, and a low circulation of A(H1N1)pdm09 (4.2 %) and type B (2.5 %) viruses. The analysed A(H3N2) viruses belonged to subclades 3C.2a (52 %) and 3C.2a1 (48 %); all studied A(H1N1)pdm09 and B/Victoria-lineage viruses belonged to subclades 6B.1 and 1A, respectively. The amino acid sequence analysis of 56 A(H3N2) isolates revealed the presence of substitutions in 18 positions in haemagglutinin (HA) as compared to the A/Hong Kong/4801/2014 vaccine virus, seven of which occurred in four antigenic sites, together with changes in 23 positions in neuraminidase (NA), and a number of substitutions in internal proteins PB2, PB1, PB1-F2, PA, NP and NS1. Despite the many amino acid substitutions, A(H3N2) viruses remained antigenically similar to the vaccine strain. Substitutions in HA and NA sequences of A(H1N1)pdm09 and B/Victoria-lineage strains were also identified, including in antigenic sites. The results of this study confirm the genetic variability of circulating influenza viruses, particularly A(H3N2), and the need for continued antigenic and molecular surveillance.

Tandem Fusion of Hepatitis B Core Antigen Allows Assembly of Virus-Like Particles in Bacteria and Plants with Enhanced Capacity to Accommodate Foreign Proteins

PubMed Central

Peyret, Hadrien; Gehin, Annick; Thuenemann, Eva C.; Blond, Donatienne; El Turabi, Aadil; Beales, Lucy; Clarke, Dean; Gilbert, Robert J. C.; Fry, Elizabeth E.; Stuart, David I.; Holmes, Kris; Stonehouse, Nicola J.; Whelan, Mike; Rosenberg, William; Lomonossoff, George P.; Rowlands, David J.

2015-01-01

The core protein of the hepatitis B virus, HBcAg, assembles into highly immunogenic virus-like particles (HBc VLPs) when expressed in a variety of heterologous systems. Specifically, the major insertion region (MIR) on the HBcAg protein allows the insertion of foreign sequences, which are then exposed on the tips of surface spike structures on the outside of the assembled particle. Here, we present a novel strategy which aids the display of whole proteins on the surface of HBc particles. This strategy, named tandem core, is based on the production of the HBcAg dimer as a single polypeptide chain by tandem fusion of two HBcAg open reading frames. This allows the insertion of large heterologous sequences in only one of the two MIRs in each spike, without compromising VLP formation. We present the use of tandem core technology in both plant and bacterial expression systems. The results show that tandem core particles can be produced with unmodified MIRs, or with one MIR in each tandem dimer modified to contain the entire sequence of GFP or of a camelid nanobody. Both inserted proteins are correctly folded and the nanobody fused to the surface of the tandem core particle (which we name tandibody) retains the ability to bind to its cognate antigen. This technology paves the way for the display of natively folded proteins on the surface of HBc particles either through direct fusion or through non-covalent attachment via a nanobody. PMID:25830365

The Antigenic Structure of Zika Virus and Its Relation to Other Flaviviruses: Implications for Infection and Immunoprophylaxis.

PubMed

Heinz, Franz X; Stiasny, Karin

2017-03-01

Zika virus was discovered ∼70 years ago in Uganda and maintained a low profile as a human disease agent in Africa and Asia. Only recently has it caused explosive outbreaks in previously unaffected regions, first in Oceania and then in the Americas since 2015. Of special concern is the newly identified link between congenital malformations (especially microcephaly) and Zika virus infections during pregnancy. At present, it is unclear whether Zika virus changed its pathogenicity or whether the huge number of infections allowed the recognition of a previously cryptic pathogenic property. The purpose of this review is to discuss recent data on the molecular antigenic structure of Zika virus in the context of antibody-mediated neutralization and antibody-dependent enhancement (ADE) of infection, a phenomenon that has been implicated in the development of severe disease caused by the related dengue viruses. Emphasis is given to epitopes of antibodies that potently neutralize Zika virus and also to epitopes that provide antigenic links to other important human-pathogenic flaviviruses such as dengue, yellow fever, West Nile, Japanese encephalitis, and tick-borne encephalitis viruses. The antigenic cross talk between Zika and dengue viruses appears to be of special importance, since they cocirculate in many regions of endemicity and sequential infections are likely to occur frequently. New insights into the molecular antigenic structure of Zika virus and flaviviruses in general have provided the foundation for great progress made in developing Zika virus vaccines and antibodies for passive immunization. Copyright © 2017 American Society for Microbiology.

The Antigenic Structure of Zika Virus and Its Relation to Other Flaviviruses: Implications for Infection and Immunoprophylaxis

PubMed Central

Stiasny, Karin

2017-01-01

SUMMARY Zika virus was discovered ∼70 years ago in Uganda and maintained a low profile as a human disease agent in Africa and Asia. Only recently has it caused explosive outbreaks in previously unaffected regions, first in Oceania and then in the Americas since 2015. Of special concern is the newly identified link between congenital malformations (especially microcephaly) and Zika virus infections during pregnancy. At present, it is unclear whether Zika virus changed its pathogenicity or whether the huge number of infections allowed the recognition of a previously cryptic pathogenic property. The purpose of this review is to discuss recent data on the molecular antigenic structure of Zika virus in the context of antibody-mediated neutralization and antibody-dependent enhancement (ADE) of infection, a phenomenon that has been implicated in the development of severe disease caused by the related dengue viruses. Emphasis is given to epitopes of antibodies that potently neutralize Zika virus and also to epitopes that provide antigenic links to other important human-pathogenic flaviviruses such as dengue, yellow fever, West Nile, Japanese encephalitis, and tick-borne encephalitis viruses. The antigenic cross talk between Zika and dengue viruses appears to be of special importance, since they cocirculate in many regions of endemicity and sequential infections are likely to occur frequently. New insights into the molecular antigenic structure of Zika virus and flaviviruses in general have provided the foundation for great progress made in developing Zika virus vaccines and antibodies for passive immunization. PMID:28179396

Novel 6xHis tagged foot-and-mouth disease virus vaccine bound to nanolipoprotein adjuvant via metal ions provides antigenic distinction and effective protective immunity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rai, Devendra K.; Segundo, Fayna Diaz-San; Department of Pathobiology and Veterinary Science, CANR, University of Connecticut, Storrs, CT 06269

Here, we engineered two FMD viruses with histidine residues inserted into or fused to the FMDV capsid. Both 6xHis viruses exhibited growth kinetics, plaque morphologies and antigenic characteristics similar to wild-type virus. The 6xHis tag allowed one-step purification of the mutant virions by Co{sup 2+} affinity columns. Electron microscopy and biochemical assays showed that the 6xHis FMDVs readily assembled into antigen: adjuvant complexes in solution, by conjugating with Ni{sup 2+}-chelated nanolipoprotein and monophosphoryl lipid A adjuvant (MPLA:NiNLP). Animals Immunized with the inactivated 6xHis-FMDV:MPLA:NiNLP vaccine acquired enhanced protective immunity against FMDV challenge compared to virions alone. Induction of anti-6xHis and anti-FMDVmore » neutralizing antibodies in the immunized animals could be exploited in the differentiation of vaccinated from infected animals needed for the improvement of FMD control measures. The novel marker vaccine/nanolipid technology described here has broad applications for the development of distinctive and effective immune responses to other pathogens of importance. - Highlights: • 6xHis-tags in A{sub 24} FMDV enable purification and biding to adjuvants via metal ions. • 6xHis A{sub 24} FMDV:MPLA:NiNLP vaccine enhanced protective immunity against FMDV. • Surface exposed capsid tags allow distinction of infected from vaccinated animals.« less

Production of Antigens and Antibodies for Diagnosis of Arbovirus Diseases.

DTIC Science & Technology

1994-05-20

for Germiston, Qalyub, Sicilian, vesicular stomatitis Indiana, and Ganjam viruses. The antigens were inactivated with beta-propiolactone. Rabbits were...vesicular stomatitis Indiana, and Ganjam viruses. The antigens were inactivated with beta-propiolactone. Rabbits were immunized successfully intravenously...370 sm4 6 229 Sicilian Sabin sm37,Vero2 1 23 VS-Indiana Ind. Lab sm7 1 45 Ganjam IG 619 sm5 1 67 Additionally, 22 viruses were passaged in baby mice

Adult T-cell leukemia-associated antigen (ATLA): detection of a glycoprotein in cell- and virus-free supernatant.

PubMed

Yamamoto, N; Schneider, J; Hinuma, Y; Hunsmann, G

1982-01-01

A glycoprotein of an apparent molecular mass of 46,000, gp 46, was enriched by affinity chromatography from the virus- and cell-free culture medium of adult T-cell leukemia virus (ATLV) infected cells. gp 46 was specifically precipitated with sera from patients with adult T-cell leukemia associated antigen (ATLA). Thus, gp 46 is a novel component of the ATLA antigen complex.

Antigenic Properties of the HIV Envelope on Virions in Solution

PubMed Central

Mengistu, Meron; Lewis, George K.; Lakowicz, Joseph R.

2014-01-01

The structural flexibility found in human immunodeficiency virus (HIV) envelope glycoproteins creates a complex relationship between antigenicity and sensitivity to antiviral antibodies. The study of this issue in the context of viral particles is particularly problematic as conventional virus capture approaches can perturb antigenicity profiles. Here, we employed a unique analytical system based on fluorescence correlation spectroscopy (FCS), which measures antibody-virion binding with all reactants continuously in solution. Panels of nine anti-envelope monoclonal antibodies (MAbs) and five virus types were used to connect antibody binding profiles with neutralizing activities. Anti-gp120 MAbs against the 2G12 or b12 epitope, which marks functional envelope structures, neutralized viruses expressing CCR5-tropic envelopes and exhibited efficient virion binding in solution. MAbs against CD4-induced (CD4i) epitopes considered hidden on functional envelope structures poorly bound these viruses and were not neutralizing. Anti-gp41 MAb 2F5 was neutralizing despite limited virion binding. Similar antigenicity patterns occurred on CXCR4-tropic viruses, except that anti-CD4i MAbs 17b and 19e were neutralizing despite little or no virion binding. Notably, anti-gp120 MAb PG9 and anti-gp41 MAb F240 bound to both CCR5-tropic and CXCR4-tropic viruses without exerting neutralizing activity. Differences in the virus production system altered the binding efficiencies of some antibodies but did not enhance antigenicity of aberrant gp120 structures. Of all viruses tested, only JRFL pseudoviruses showed a direct relationship between MAb binding efficiency and neutralizing potency. Collectively, these data indicate that the antigenic profiles of free HIV particles generally favor the exposure of functional over aberrant gp120 structures. However, the efficiency of virion-antibody interactions in solution inconsistently predicts neutralizing activity in vitro. PMID:24284318

ɣδ T cell receptor ligands and modes of antigen recognition

PubMed Central

Champagne, Eric

2011-01-01

T lymphocytes expressing the γδ-type of T cell receptors for antigens contribute to all aspects of immune responses, including defenses against viruses, bacteria, parasites and tumors, allergy and autoimmunity. Multiple subsets have been individualized in humans as well as in mice and they appear to recognize in a TCR-dependent manner antigens as diverse as small non-peptidic molecules, soluble or membrane-anchored polypeptides and molecules related to MHC antigens on cell surfaces, implying diverse modes of antigen recognition. We review here the γδ TCR ligands which have been identified along the years and their characteristics, with emphasis on a few systems which have been extensively studied such as human γδ T cells responding to phosphoantigens or murine γδ T cells activated by allogeneic MHC antigens. We discuss a speculative model of antigen recognition involving simultaneous TCR recognition of MHC-like and non-MHC ligands which could fit with most available data and shares many similarities with the classical model of MHC-restricted antigen recognition for peptides or lipids by T cells subsets with αβ-type TCRs. PMID:21298486

γδ T cell receptor ligands and modes of antigen recognition.

PubMed

Champagne, Eric

2011-04-01

T lymphocytes expressing the γδ-type of T cell receptors (TCRs) for antigens contribute to all aspects of immune responses, including defenses against viruses, bacteria, parasites and tumors, allergy and autoimmunity. Multiple subsets have been individualized in humans as well as in mice and they appear to recognize in a TCR-dependent manner antigens as diverse as small non-peptidic molecules, soluble or membrane-anchored polypeptides and molecules related to MHC antigens on cell surfaces, implying diverse modes of antigen recognition. We review here the γδ TCR ligands which have been identified along the years and their characteristics, with emphasis on a few systems which have been extensively studied such as human γδ T cells responding to phosphoantigens or murine γδ T cells activated by allogeneic MHC antigens. We discuss a speculative model of antigen recognition involving simultaneous TCR recognition of MHC-like and non-MHC ligands which could fit with most available data and shares many similarities with the classical model of MHC-restricted antigen recognition for peptides or lipids by T cells subsets with αβ-type TCRs.

Antigenic Drift of the Pandemic 2009 A(H1N1) Influenza Virus in a Ferret Model

PubMed Central

Guarnaccia, Teagan; Carolan, Louise A.; Maurer-Stroh, Sebastian; Lee, Raphael T. C.; Job, Emma; Reading, Patrick C.; Petrie, Stephen; McCaw, James M.; McVernon, Jodie; Hurt, Aeron C.; Kelso, Anne; Mosse, Jennifer; Barr, Ian G.; Laurie, Karen L.

2013-01-01

Surveillance data indicate that most circulating A(H1N1)pdm09 influenza viruses have remained antigenically similar since they emerged in humans in 2009. However, antigenic drift is likely to occur in the future in response to increasing population immunity induced by infection or vaccination. In this study, sequential passaging of A(H1N1)pdm09 virus by contact transmission through two independent series of suboptimally vaccinated ferrets resulted in selection of variant viruses with an amino acid substitution (N156K, H1 numbering without signal peptide; N159K, H3 numbering without signal peptide; N173K, H1 numbering from first methionine) in a known antigenic site of the viral HA. The N156K HA variant replicated and transmitted efficiently between naïve ferrets and outgrew wildtype virus in vivo in ferrets in the presence and absence of immune pressure. In vitro, in a range of cell culture systems, the N156K variant rapidly adapted, acquiring additional mutations in the viral HA that also potentially affected antigenic properties. The N156K escape mutant was antigenically distinct from wildtype virus as shown by binding of HA-specific antibodies. Glycan binding assays demonstrated the N156K escape mutant had altered receptor binding preferences compared to wildtype virus, which was supported by computational modeling predictions. The N156K substitution, and culture adaptations, have been detected in human A(H1N1)pdm09 viruses with N156K preferentially reported in sequences from original clinical samples rather than cultured isolates. This study demonstrates the ability of the A(H1N1)pdm09 virus to undergo rapid antigenic change to evade a low level vaccine response, while remaining fit in a ferret transmission model of immunization and infection. Furthermore, the potential changes in receptor binding properties that accompany antigenic changes highlight the importance of routine characterization of clinical samples in human A(H1N1)pdm09 influenza surveillance. PMID:23671418

Seroprevalence of hepatitis B surface antigenemia and its effects on hematological parameters in pregnant women in Osogbo, Nigeria.

PubMed

Kolawole, Olatunji M; Wahab, Abideen A; Adekanle, Daniel A; Sibanda, Timothy; Okoh, Anthony I

2012-12-27

The transmission of the hepatitis B virus (HBV) is parenteral, sexual and perinatal. Prevention of vertical transmission of HBV is extremely important because HBV infection in early life usually results in a chronic carrier State. A descriptive seroepidemiological study of hepatitis B virus and its effects on hematological parameters was investigated in pregnant women attending antenatal clinic of LAUTECH Teaching Hospital, Osogbo, Nigeria. 200 venous samples were subjected to full blood count and its sera were subjected to enzyme-linked immunosorbent assay for the detection of surface antigen of hepatitis B virus. Prevalence rate of 16.5% was obtained for hepatitis B surface antigen in pregnant women. The highest HBsAg prevalence rate recorded was 23.3% for pregnant women between aged 30-34 years while the lowest recorded was zero percent for those aged greater than 40 years. RBC, WBC, neutrophil, hemoglobin lymphocyte and platelet counts have no significant effects on HBsAg positivity of pregnant women (p=0.801). There was no significant difference in HBsAg positivity in relation to maternal age, gravidity, gestational age, family type, level of education and occupation (p=0.073). Among the potential risk factors, there was significant difference in HBsAg positivity in the pregnant women in relation to their history of HBV vaccination (p=0.039). We advocate universal free screening of pregnant women as the endemicity of HBV infections is thus being propagated.

Seroprevalence of hepatitis B surface antigenemia and its effects on hematological parameters in pregnant women in Osogbo, Nigeria

PubMed Central

2012-01-01

Background The transmission of the hepatitis B virus (HBV) is parenteral, sexual and perinatal. Prevention of vertical transmission of HBV is extremely important because HBV infection in early life usually results in a chronic carrier State. Methods A descriptive seroepidemiological study of hepatitis B virus and its effects on hematological parameters was investigated in pregnant women attending antenatal clinic of LAUTECH Teaching Hospital, Osogbo, Nigeria. 200 venous samples were subjected to full blood count and its sera were subjected to enzyme–linked immunosorbent assay for the detection of surface antigen of hepatitis B virus. Results Prevalence rate of 16.5% was obtained for hepatitis B surface antigen in pregnant women. The highest HBsAg prevalence rate recorded was 23.3% for pregnant women between aged 30–34 years while the lowest recorded was zero percent for those aged greater than 40 years. RBC, WBC, neutrophil, hemoglobin lymphocyte and platelet counts have no significant effects on HBsAg positivity of pregnant women (p = 0.801). There was no significant difference in HBsAg positivity in relation to maternal age, gravidity, gestational age, family type, level of education and occupation (p = 0.073). Among the potential risk factors, there was significant difference in HBsAg positivity in the pregnant women in relation to their history of HBV vaccination (p = 0.039). Conclusions We advocate universal free screening of pregnant women as the endemicity of HBV infections is thus being propagated. PMID:23268985

Detection of hepatitis B surface antigen in pregnant women attending a public hospital for delivery: implication for vaccination strategy in Bangladesh.

PubMed

Rumi, M A; Begum, K; Hassan, M S; Hasan, S M; Azam, M G; Hasan, K N; Shirin, M; Khan, A K

1998-08-01

Routine antenatal hepatitis B surface antigen (HBsAg) screening and immunization of risk babies is very effective in preventing perinatal transmission of hepatitis B virus (HBV). We studied 1,800 parturients attending a public hospital to assess the rationale for such vaccination in Bangladesh. In one in every 29 deliveries (63 of 1,800 or 3.5%), the mother was found to be HBsAg positive. All were asymptomatic and many (41 of 63 or 65%) without risk factors would remain undetected if HBsAg screening were performed on selected groups. Most of the HBsAg-positive mothers (54 of 63 or 85.7%) were found to be chronic carriers and 30.2% (19 of 63) were also hepatitis B e antigen (HBeAg) positive, indicating high infectivity. Although 23 cord blood were positive for HBsAg or HBeAg, none were positive for IgM antibody to hepatitis B core antigen (IgM anti-HBc), suggesting transplacental transmission of the antigens rather than intrauterine infection. These findings are discussed in relation to the cost-effectiveness of routine prenatal screening and immunization of risk babies compared with universal infant immunization.

Evolution of canine parvovirus--a need for new vaccines?

PubMed

Truyen, Uwe

2006-10-05

Canine parvovirus (CPV) is a new virus, which is continuing to evolve, giving rise to new antigenic types and virus mutants that spread through the dog population. The most successful mutants, from an evolutionary perspective, appear to be selected by improved binding to the CPV receptor, the canine transferrin receptor, and by an extended host range, which for the newer antigenic types now includes both the dog and the cat. The new viruses also show antigenic differences that can be defined by binding of certain monoclonal antibodies; they also differ in their reactivity in virus neutralisation tests, using immune sera raised against the various antigenic types. These differences may influence the susceptibility of young animals to infection at the time when the level of maternally derived antibody decreases to the minimum protective titre. This minimum protective titre may vary depending on the infecting virus type. There is, however, a high degree of cross-protection between the virus types and the true relevance of the differences in neutralisation titer is currently not known.

Molecular and antigenic traits on hemagglutinin gene of avian influenza H9N2 viruses: Evidence of a new escape mutant in Egypt adapted in quails.

PubMed

Adel, Amany; Arafa, Abdelsatar; Hussein, Hussein A; El-Sanousi, Ahmed A

2017-06-01

The LPAI viruses of H9N2 subtype became widely distributed in Middle Eastern countries, causing great economic losses in poultry industry especially when complicated with other pathogens. The H9N2 viruses in Egypt have a wide spread nature since its first occurrence in 2011. In this study, we collected cloacal and tracheal samples from 19 flocks for detection and propagation of H9N2 virus using real-time RT-PCR and egg inoculation. We studied the molecular evolution of the Hemagglutinin gene of H9N2 viruses by full HA gene sequencing, then the antigenic characterization was implemented using the cross HI assay and analyzed using 3D Bioinformatics cartography software. The phylogenetic analysis of the HA gene of Egyptian H9N2 viruses clearly points out the presence of only one group (Egy/G1) of originally introduced viruses in 2011 related to the G1 lineage within group B, with the presence of multiple minor clusters includes viruses from 2011 to 2015. However, a new variant (Egy/G1var) cluster was detected in quails since 2012. Genetically, Egy/G1var viruses characterized by presence of 20 amino acid substitutions within and adjacent to the antigenic sites in comparison to other Egyptian viruses. In addition, two glycosylation sites at amino acid residues 127 and 189 were determined in close to the receptor binding and antigenic sites. The antigenic analysis based on 3D antigenic mapping showed that the Egy/G1var cluster was clearly distinct from the original Egy/G1 viruses. In conclusion, Egy/G1var is shown to be a new escape mutant variant cluster with an adaptive evolution in quails. Copyright © 2017 Elsevier Ltd. All rights reserved.

Study design for a hepatitis B vaccine trial.

PubMed Central

Lustbader, E D; London, W T; Blumberg, B S

1976-01-01

A short-time trial of small sample size for an evaluation of the hepatitis B vaccine is proposed and designed. The vaccine is based on the premise that antibody to the surface antigen of the hepatitis B virus is protective against viral infection. This premise is verified by using the presence of the surface antigen as the marker of infection and comparing infection rates in renal dialysis patients who had naturally acquired antibody to patients without antibody. Patients with antibody have an extremely low risk of infection. The probability of remaining uninfected decreases at an exponential rate for patients without antibody, implying a constant risk of infection at any point in time. The study design described makes use of this time independence and the observed infection rates to formulate a clinical trial which can be accomplished with a relatively small number of patients. This design might be useful if, in preliminary studies, it is shown that the vaccine produces antibody in the patients and that protection against hepatitis B virus would be beneficial to the patients. PMID:1062809

Stable expression of the hepatitis B virus surface antigen containing pre-S2 protein in mouse cells using a bovine papillomavirus vector.

PubMed

Yoneyama, T; Akatsuka, T; Miyamura, T

1988-08-01

The large BglII fragment (2.8 kilobases) of hepatitis B virus DNA including the transcription unit for the hepatitis B surface antigen (HBsAg) was inserted into a bovine papillomavirus vector containing the neomycin resistance gene. The recombinant DNA was transfected into mouse C127 cells. A stable transformed cell line (MS128) secreting a large amount of 22 nm HBsAg particles containing pre-S2 protein was established. The secreted HBsAg particles had the receptor for polymerized human serum albumin. Immunoprecipitation and Western blot analyses showed that HBsAg particles consisted of two major proteins of 22K and 26K encoded by the S gene and a minor protein of 35K encoded by the pre-S2 and S genes. Southern blot analysis revealed that the transfected plasmid was integrated into the host chromosomal DNA and that most of the plasmid sequences were present. These results suggest that the stable expression of the HBsAg in MS128 cells is related to the integrated state of the recombinant DNA.

Hemagglutinin-specific neutralization of subacute sclerosing panencephalitis viruses

PubMed Central

Muller, Claude P.; Russell, Stephen J.

2018-01-01

Subacute sclerosing panencephalitis (SSPE) is a progressive, lethal complication of measles caused by particular mutants of measles virus (MeV) that persist in the brain despite high levels of neutralizing antibodies. We addressed the hypothesis that antigenic drift is involved in the pathogenetic mechanism of SSPE by analyzing antigenic alterations in the MeV envelope hemagglutinin protein (MeV-H) found in patients with SSPE in relation to major circulating MeV genotypes. To this aim, we obtained cDNA for the MeV-H gene from tissue taken at brain autopsy from 3 deceased persons with SSPE who had short (3–4 months, SMa79), average (3.5 years, SMa84), and long (18 years, SMa94) disease courses. Recombinant MeVs with a substituted MeV-H gene were generated by a reverse genetic system. Virus neutralization assays with a panel of anti-MeV-H murine monoclonal antibodies (mAbs) or vaccine-immunized mouse anti-MeV-H polyclonal sera were performed to determine the antigenic relatedness. Functional and receptor-binding analysis of the SSPE MeV-H showed activity in a SLAM/nectin-4–dependent manner. Similar to our panel of wild-type viruses, our SSPE viruses showed an altered antigenic profile. Genotypes A, G3, and F (SSPE case SMa79) were the exception, with an intact antigenic structure. Genotypes D7 and F (SSPE SMa79) showed enhanced neutralization by mAbs targeting antigenic site IIa. Genotypes H1 and the recently reported D4.2 were the most antigenically altered genotypes. Epitope mapping of neutralizing mAbs BH015 and BH130 reveal a new antigenic site on MeV-H, which we designated Φ for its intermediate position between previously defined antigenic sites Ia and Ib. We conclude that SSPE-causing viruses show similar antigenic properties to currently circulating MeV genotypes. The absence of a direct correlation between antigenic changes and predisposition of a certain genotype to cause SSPE does not lend support to the proposed antigenic drift as a pathogenetic mechanism in SSPE. PMID:29466428

Hemagglutinin-specific neutralization of subacute sclerosing panencephalitis viruses.

PubMed

Muñoz-Alía, Miguel Ángel; Muller, Claude P; Russell, Stephen J

2018-01-01

Subacute sclerosing panencephalitis (SSPE) is a progressive, lethal complication of measles caused by particular mutants of measles virus (MeV) that persist in the brain despite high levels of neutralizing antibodies. We addressed the hypothesis that antigenic drift is involved in the pathogenetic mechanism of SSPE by analyzing antigenic alterations in the MeV envelope hemagglutinin protein (MeV-H) found in patients with SSPE in relation to major circulating MeV genotypes. To this aim, we obtained cDNA for the MeV-H gene from tissue taken at brain autopsy from 3 deceased persons with SSPE who had short (3-4 months, SMa79), average (3.5 years, SMa84), and long (18 years, SMa94) disease courses. Recombinant MeVs with a substituted MeV-H gene were generated by a reverse genetic system. Virus neutralization assays with a panel of anti-MeV-H murine monoclonal antibodies (mAbs) or vaccine-immunized mouse anti-MeV-H polyclonal sera were performed to determine the antigenic relatedness. Functional and receptor-binding analysis of the SSPE MeV-H showed activity in a SLAM/nectin-4-dependent manner. Similar to our panel of wild-type viruses, our SSPE viruses showed an altered antigenic profile. Genotypes A, G3, and F (SSPE case SMa79) were the exception, with an intact antigenic structure. Genotypes D7 and F (SSPE SMa79) showed enhanced neutralization by mAbs targeting antigenic site IIa. Genotypes H1 and the recently reported D4.2 were the most antigenically altered genotypes. Epitope mapping of neutralizing mAbs BH015 and BH130 reveal a new antigenic site on MeV-H, which we designated Φ for its intermediate position between previously defined antigenic sites Ia and Ib. We conclude that SSPE-causing viruses show similar antigenic properties to currently circulating MeV genotypes. The absence of a direct correlation between antigenic changes and predisposition of a certain genotype to cause SSPE does not lend support to the proposed antigenic drift as a pathogenetic mechanism in SSPE.

Preparation of protein loaded poly(D,L-lactide-co-glycolide) microparticles for the antigen delivery to dendritic cells using a static micromixer.

PubMed

Wischke, Christian; Lorenzen, Dirk; Zimmermann, Julian; Borchert, Hans-Hubert

2006-04-01

The cellular immune response against tumors, viruses, or intracellular bacteria requires adequate antigen delivery to professional phagocytes, their processing and the presentation of antigenic peptides to T-cells. Biodegradable microparticles to enhance antigen phagocytosis and the response of cytotoxic lymphocytes have been proposed. The aim of the present study was to formulate poly(lactide-co-glycolide) (PLGA) microparticles using a w/o/w solvent evaporation procedure in order to obtain suitable vehicles for vaccination. Bovine serum albumin bearing fluorescein isothiocyanate (FITC-BSA) was used as a model antigen. For microparticle preparation a static micromixer was employed. Microparticles of 2-3 microm can be produced with good reproducibility by applying high flow rates at the micromixer. Microparticles with a smooth surface and only one pore were observed using scanning electron microscopy (SEM). Confocal laser scanning microscopy (CLSM) allowed localisation of the FITC-BSA near the surface of the microparticle. Microencapsulation of FITC-BSA did not altered the polymer characteristics, as determined by measuring the glass transition temperature. Additionally we could determine residual methylene chloride, employed as solvent in microparticle preparation, to be less than 1/1000 of the USP and Ph. Eur. limit. The microparticles described herein were able to deliver the model antigen to human dendritic cells (DC).

Antigenically Diverse Swine Origin H1N1 Variant Influenza Viruses Exhibit Differential Ferret Pathogenesis and Transmission Phenotypes.

PubMed

Pulit-Penaloza, Joanna A; Jones, Joyce; Sun, Xiangjie; Jang, Yunho; Thor, Sharmi; Belser, Jessica A; Zanders, Natosha; Creager, Hannah M; Ridenour, Callie; Wang, Li; Stark, Thomas J; Garten, Rebecca; Chen, Li-Mei; Barnes, John; Tumpey, Terrence M; Wentworth, David E; Maines, Taronna R; Davis, C Todd

2018-06-01

Influenza A(H1) viruses circulating in swine represent an emerging virus threat, as zoonotic infections occur sporadically following exposure to swine. A fatal infection caused by an H1N1 variant (H1N1v) virus was detected in a patient with reported exposure to swine and who presented with pneumonia, respiratory failure, and cardiac arrest. To understand the genetic and phenotypic characteristics of the virus, genome sequence analysis, antigenic characterization, and ferret pathogenesis and transmissibility experiments were performed. Antigenic analysis of the virus isolated from the fatal case, A/Ohio/09/2015, demonstrated significant antigenic drift away from the classical swine H1N1 variant viruses and H1N1 pandemic 2009 viruses. A substitution in the H1 hemagglutinin (G155E) was identified that likely impacted antigenicity, and reverse genetics was employed to understand the molecular mechanism of antibody escape. Reversion of the substitution to 155G, in a reverse genetics A/Ohio/09/2015 virus, showed that this residue was central to the loss of hemagglutination inhibition by ferret antisera raised against a prototypical H1N1 pandemic 2009 virus (A/California/07/2009), as well as gamma lineage classical swine H1N1 viruses, demonstrating the importance of this residue for antibody recognition of this H1 lineage. When analyzed in the ferret model, A/Ohio/09/2015 and another H1N1v virus, A/Iowa/39/2015, as well as A/California/07/2009, replicated efficiently in the respiratory tract of ferrets. The two H1N1v viruses transmitted efficiently among cohoused ferrets, but respiratory droplet transmission studies showed that A/California/07/2009 transmitted through the air more efficiently. Preexisting immunity to A/California/07/2009 did not fully protect ferrets from challenge with A/Ohio/09/2015. IMPORTANCE Human infections with classical swine influenza A(H1N1) viruses that circulate in pigs continue to occur in the United States following exposure to swine. To understand the genetic and virologic characteristics of a virus (A/Ohio/09/2015) associated with a fatal infection and a virus associated with a nonfatal infection (A/Iowa/39/2015), we performed genome sequence analysis, antigenic testing, and pathogenicity and transmission studies in a ferret model. Reverse genetics was employed to identify a single antigenic site substitution (HA G155E) responsible for antigenic variation of A/Ohio/09/2015 compared to related classical swine influenza A(H1N1) viruses. Ferrets with preexisting immunity to the pandemic A(H1N1) virus were challenged with A/Ohio/09/2015, demonstrating decreased protection. These data illustrate the potential for currently circulating swine influenza viruses to infect and cause illness in humans with preexisting immunity to H1N1 pandemic 2009 viruses and a need for ongoing risk assessment and development of candidate vaccine viruses for improved pandemic preparedness. This is a work of the U.S. Government and is not subject to copyright protection in the United States. Foreign copyrights may apply.

A STING-activating nanovaccine for cancer immunotherapy

NASA Astrophysics Data System (ADS)

Luo, Min; Wang, Hua; Wang, Zhaohui; Cai, Haocheng; Lu, Zhigang; Li, Yang; Du, Mingjian; Huang, Gang; Wang, Chensu; Chen, Xiang; Porembka, Matthew R.; Lea, Jayanthi; Frankel, Arthur E.; Fu, Yang-Xin; Chen, Zhijian J.; Gao, Jinming

2017-07-01

The generation of tumour-specific T cells is critically important for cancer immunotherapy. A major challenge in achieving a robust T-cell response is the spatiotemporal orchestration of antigen cross-presentation in antigen-presenting cells with innate stimulation. Here, we report a minimalist nanovaccine, comprising a simple physical mixture of an antigen and a synthetic polymeric nanoparticle, PC7A NP, which generates a strong cytotoxic T-cell response with low systemic cytokine expression. Mechanistically, the PC7A NP achieves efficient cytosolic delivery of tumour antigens to antigen-presenting cells in draining lymph nodes, leading to increased surface presentation while simultaneously activating type I interferon-stimulated genes. This effect is dependent on stimulator of interferon genes (STING), but not the Toll-like receptor or the mitochondrial antiviral-signalling protein (MAVS) pathway. The nanovaccine led to potent tumour growth inhibition in melanoma, colon cancer and human papilloma virus-E6/E7 tumour models. The combination of the PC7A nanovaccine and an anti-PD-1 antibody showed great synergy, with 100% survival over 60 days in a TC-1 tumour model. Rechallenging of these tumour-free animals with TC-1 cells led to complete inhibition of tumour growth, suggesting the generation of long-term antitumour memory. The STING-activating nanovaccine offers a simple, safe and robust strategy in boosting anti-tumour immunity for cancer immunotherapy.

Virus like particles as a platform for cancer vaccine development.

PubMed

Ong, Hui Kian; Tan, Wen Siang; Ho, Kok Lian

2017-01-01

Cancers have killed millions of people in human history and are still posing a serious health problem worldwide. Therefore, there is an urgent need for developing preventive and therapeutic cancer vaccines. Among various cancer vaccine development platforms, virus-like particles (VLPs) offer several advantages. VLPs are multimeric nanostructures with morphology resembling that of native viruses and are mainly composed of surface structural proteins of viruses but are devoid of viral genetic materials rendering them neither infective nor replicative. In addition, they can be engineered to display multiple, highly ordered heterologous epitopes or peptides in order to optimize the antigenicity and immunogenicity of the displayed entities. Like native viruses, specific epitopes displayed on VLPs can be taken up, processed, and presented by antigen-presenting cells to elicit potent specific humoral and cell-mediated immune responses. Several studies also indicated that VLPs could overcome the immunosuppressive state of the tumor microenvironment and break self-tolerance to elicit strong cytotoxic lymphocyte activity, which is crucial for both virus clearance and destruction of cancerous cells. Collectively, these unique characteristics of VLPs make them optimal cancer vaccine candidates. This review discusses current progress in the development of VLP-based cancer vaccines and some potential drawbacks of VLPs in cancer vaccine development. Extracellular vesicles with close resembling to viral particles are also discussed and compared with VLPs as a platform in cancer vaccine developments.

Antibody responses to Herpesvirus papio antigens in baboons with lymphoma.

PubMed

Neubauer, R H; Rabin, H; Strnad, B C; Lapin, B A; Yakovleva, L A; Indzie, E

1979-02-01

An Epstein-Barr virus-related herpesvirus, termed Herpesvirus papio (HVP), was isolated from baboons (Papio hamadryas) at the Institute of Experimental Pathology and Therapy, Sukhumi, USSR, where there is a continuing outbreak of lymphoma. In the present study sera from diseased baboons and from age- and sex-matched control animals were examined for antibodies to HVP antigens. Results showed that animals with lymphoid disease had antibodies to HVP virus capsid, early, soluble, and nuclear antigens at higher frequencies and at higher titers than did control animals. Antibody titers were not age- or sex-related. No concordancy was detected for antibodies to soluble and nuclear antigens. The sera were also examined for antibodies to two other widely distributed viruses of hamadryas baboons, cytomegalovirus and foamy virus. The results of these studies did not indicate a disease-related role for either of these viruses.

Increasing Type 1 Poliovirus Capsid Stability by Thermal Selection

PubMed Central

Adeyemi, Oluwapelumi O.; Nicol, Clare

2016-01-01

ABSTRACT Poliomyelitis is a highly infectious disease caused by poliovirus (PV). It can result in paralysis and may be fatal. Integrated global immunization programs using live-attenuated oral (OPV) and/or inactivated (IPV) PV vaccines have systematically reduced its spread and paved the way for eradication. Immunization will continue posteradication to ensure against reintroduction of the disease, but there are biosafety concerns for both OPV and IPV. They could be addressed by the production and use of virus-free virus-like particle (VLP) vaccines that mimic the “empty” capsids (ECs) normally produced in viral infection. Although ECs are antigenically indistinguishable from mature virus particles, they are less stable and readily convert into an alternative conformation unsuitable for vaccine purposes. Stabilized ECs, expressed recombinantly as VLPs, could be ideal candidate vaccines for a polio-free world. However, although genome-free PV ECs have been expressed as VLPs in a variety of systems, their inherent antigenic instability has proved a barrier to further development. In this study, we selected thermally stable ECs of type 1 PV (PV-1). The ECs are antigenically stable at temperatures above the conversion temperature of wild-type (wt) virions. We have identified mutations on the capsid surface and in internal networks that are responsible for EC stability. With reference to the capsid structure, we speculate on the roles of these residues in capsid stability and postulate that such stabilized VLPs could be used as novel vaccines. IMPORTANCE Poliomyelitis is a highly infectious disease caused by PV and is on the verge of eradication. There are biosafety concerns about reintroduction of the disease from current vaccines that require live virus for production. Recombinantly expressed virus-like particles (VLPs) could address these inherent problems. However, the genome-free capsids (ECs) of wt PV are unstable and readily change antigenicity to a form not suitable as a vaccine. Here, we demonstrate that the ECs of type 1 PV can be stabilized by selecting heat-resistant viruses. Our data show that some capsid mutations stabilize the ECs and could be applied as candidates to synthesize stable VLPs as future genome-free poliovirus vaccines. PMID:27928008

Immunization of dogs with a canine herpesvirus vector expressing Neospora caninum surface protein, NcSRS2.

PubMed

Nishikawa, Y; Ikeda, H; Fukumoto, S; Xuan, X; Nagasawa, H; Otsuka, H; Mikami, T

2000-10-01

In order to develop a vaccine against Neospora caninum in dogs, we constructed recombinant canine herpesvirus (CHV) expressing N. caninum surface protein, NcSRS2. Indirect immunofluorescence indicated that the antigenic structure of the recombinant NcSRS2 was similar to the authentic parasite protein. The dogs immunised with recombinant virus produced IgG antibody to N. caninum, and their sera recognised the parasite protein on Western blot. The dogs inoculated with recombinant virus showed no clinical symptoms and infectious CHV was not recovered from the dogs, suggesting that recombinant CHV expressing N. caninum proteins may lead to a vaccine against neosporosis in dogs.

Antibody-Induced Internalization of the Human Respiratory Syncytial Virus Fusion Protein.

PubMed

Leemans, A; De Schryver, M; Van der Gucht, W; Heykers, A; Pintelon, I; Hotard, A L; Moore, M L; Melero, J A; McLellan, J S; Graham, B S; Broadbent, L; Power, U F; Caljon, G; Cos, P; Maes, L; Delputte, P

2017-07-15

Respiratory syncytial virus (RSV) infections remain a major cause of respiratory disease and hospitalizations among infants. Infection recurs frequently and establishes a weak and short-lived immunity. To date, RSV immunoprophylaxis and vaccine research is mainly focused on the RSV fusion (F) protein, but a vaccine remains elusive. The RSV F protein is a highly conserved surface glycoprotein and is the main target of neutralizing antibodies induced by natural infection. Here, we analyzed an internalization process of antigen-antibody complexes after binding of RSV-specific antibodies to RSV antigens expressed on the surface of infected cells. The RSV F protein and attachment (G) protein were found to be internalized in both infected and transfected cells after the addition of either RSV-specific polyclonal antibodies (PAbs) or RSV glycoprotein-specific monoclonal antibodies (MAbs), as determined by indirect immunofluorescence staining and flow-cytometric analysis. Internalization experiments with different cell lines, well-differentiated primary bronchial epithelial cells (WD-PBECs), and RSV isolates suggest that antibody internalization can be considered a general feature of RSV. More specifically for RSV F, the mechanism of internalization was shown to be clathrin dependent. All RSV F-targeted MAbs tested, regardless of their epitopes, induced internalization of RSV F. No differences could be observed between the different MAbs, indicating that RSV F internalization was epitope independent. Since this process can be either antiviral, by affecting virus assembly and production, or beneficial for the virus, by limiting the efficacy of antibodies and effector mechanism, further research is required to determine the extent to which this occurs in vivo and how this might impact RSV replication. IMPORTANCE Current research into the development of new immunoprophylaxis and vaccines is mainly focused on the RSV F protein since, among others, RSV F-specific antibodies are able to protect infants from severe disease, if administered prophylactically. However, antibody responses established after natural RSV infections are poorly protective against reinfection, and high levels of antibodies do not always correlate with protection. Therefore, RSV might be capable of interfering, at least partially, with antibody-induced neutralization. In this study, a process through which surface-expressed RSV F proteins are internalized after interaction with RSV-specific antibodies is described. One the one hand, this antigen-antibody complex internalization could result in an antiviral effect, since it may interfere with virus particle formation and virus production. On the other hand, this mechanism may also reduce the efficacy of antibody-mediated effector mechanisms toward infected cells. Copyright © 2017 American Society for Microbiology.

Antibody-Induced Internalization of the Human Respiratory Syncytial Virus Fusion Protein

PubMed Central

Leemans, A.; De Schryver, M.; Van der Gucht, W.; Heykers, A.; Pintelon, I.; Hotard, A. L.; Moore, M. L.; Melero, J. A.; McLellan, J. S.; Graham, B. S.; Broadbent, L.; Power, U. F.; Caljon, G.; Cos, P.; Maes, L.

2017-01-01

ABSTRACT Respiratory syncytial virus (RSV) infections remain a major cause of respiratory disease and hospitalizations among infants. Infection recurs frequently and establishes a weak and short-lived immunity. To date, RSV immunoprophylaxis and vaccine research is mainly focused on the RSV fusion (F) protein, but a vaccine remains elusive. The RSV F protein is a highly conserved surface glycoprotein and is the main target of neutralizing antibodies induced by natural infection. Here, we analyzed an internalization process of antigen-antibody complexes after binding of RSV-specific antibodies to RSV antigens expressed on the surface of infected cells. The RSV F protein and attachment (G) protein were found to be internalized in both infected and transfected cells after the addition of either RSV-specific polyclonal antibodies (PAbs) or RSV glycoprotein-specific monoclonal antibodies (MAbs), as determined by indirect immunofluorescence staining and flow-cytometric analysis. Internalization experiments with different cell lines, well-differentiated primary bronchial epithelial cells (WD-PBECs), and RSV isolates suggest that antibody internalization can be considered a general feature of RSV. More specifically for RSV F, the mechanism of internalization was shown to be clathrin dependent. All RSV F-targeted MAbs tested, regardless of their epitopes, induced internalization of RSV F. No differences could be observed between the different MAbs, indicating that RSV F internalization was epitope independent. Since this process can be either antiviral, by affecting virus assembly and production, or beneficial for the virus, by limiting the efficacy of antibodies and effector mechanism, further research is required to determine the extent to which this occurs in vivo and how this might impact RSV replication. IMPORTANCE Current research into the development of new immunoprophylaxis and vaccines is mainly focused on the RSV F protein since, among others, RSV F-specific antibodies are able to protect infants from severe disease, if administered prophylactically. However, antibody responses established after natural RSV infections are poorly protective against reinfection, and high levels of antibodies do not always correlate with protection. Therefore, RSV might be capable of interfering, at least partially, with antibody-induced neutralization. In this study, a process through which surface-expressed RSV F proteins are internalized after interaction with RSV-specific antibodies is described. One the one hand, this antigen-antibody complex internalization could result in an antiviral effect, since it may interfere with virus particle formation and virus production. On the other hand, this mechanism may also reduce the efficacy of antibody-mediated effector mechanisms toward infected cells. PMID:28468888

Poxvirus Antigen Staining of Immune Cells as a Biomarker to Predict Disease Outcome in Monkeypox and Cowpox Virus Infection in Non-Human Primates

PubMed Central

Song, Haifeng; Janosko, Krisztina; Johnson, Reed F.; Qin, Jing; Josleyn, Nicole; Jett, Catherine; Byrum, Russell; Claire, Marisa St.; Dyall, Julie; Blaney, Joseph E.; Jennings, Gerald; Jahrling, Peter B.

2013-01-01

Infection of non-human primates (NHPs) such as rhesus and cynomolgus macaques with monkeypox virus (MPXV) or cowpox virus (CPXV) serve as models to study poxvirus pathogenesis and to evaluate vaccines and anti-orthopox therapeutics. Intravenous inoculation of macaques with high dose of MPXV (>1–2×107 PFU) or CPXV (>102 PFU) results in 80% to 100% mortality and 66 to 100% mortality respectively. Here we report that NHPs with positive detection of poxvirus antigens in immune cells by flow cytometric staining, especially in monocytes and granulocytes succumbed to virus infection and that early positive pox staining is a strong predictor for lethality. Samples from four independent studies were analyzed. Eighteen NHPs from three different experiments were inoculated with two different MPXV strains at lethal doses. Ten NHPs displayed positive pox-staining and all 10 NHPs reached moribund endpoint. In contrast, none of the three NHPs that survived anticipated lethal virus dose showed apparent virus staining in the monocytes and granulocytes. In addition, three NHPs that were challenged with a lethal dose of MPXV and received cidofovir treatment were pox-antigen negative and all three NHPs survived. Furthermore, data from a CPXV study also demonstrated that 6/9 NHPs were pox-antigen staining positive and all 6 NHPs reached euthanasia endpoint, while the three survivors were pox-antigen staining negative. Thus, we conclude that monitoring pox-antigen staining in immune cells can be used as a biomarker to predict the prognosis of virus infection. Future studies should focus on the mechanisms and implications of the pox-infection of immune cells and the correlation between pox-antigen detection in immune cells and disease progression in human poxviral infection. PMID:23577120

Unusual Features of Sodium Taurocholate Cotransporting Polypeptide as a Hepatitis B Virus Receptor

PubMed Central

Zong, Li; Sureau, Camille; Barker, Luke; Wands, Jack R.; Tong, Shuping

2016-01-01

ABSTRACT Cell culture (cc)-derived hepatitis B virus (HBV) can infect differentiated HepaRG cells, but efficient infection requires addition of polyethylene glycol (PEG) during inoculation. Identification of sodium taurocholate cotransporting polypeptide (NTCP) as an HBV receptor enabled ccHBV infection of NTCP reconstituted HepG2 cells, although very little hepatitis B surface antigen (HBsAg) is produced. We found infection by patient serum-derived HBV (sHBV), which required purification of viral particles through ultracentrifugation or PEG precipitation, was PEG independent and much more efficient in HepaRG cells than in HepG2/NTCP cells. In contrast to hepatitis B e antigen (HBeAg), HBsAg was not a reliable marker of productive sHBV infection at early time points. A low HBsAg/HBeAg ratio by ccHBV-infected HepG2/NTCP cells was attributable to dimethyl sulfoxide (DMSO) in culture medium, NTCP overexpression, and HBV genotype D. HepG2/NTCP cells released more viral antigens than HepG2 cells after HBV genome delivery by adeno-associated virus, and stable expression of NTCP in a ccHBV producing cell line increased viral mRNAs, proteins, replicative DNA, and covalently closed circular DNA. NTCP protein expression in HepG2/NTCP cells, despite being driven by the cytomegalovirus promoter, was markedly increased by DMSO treatment. This at least partly explains ability of DMSO to promote ccHBV infection in such cell lines. In conclusion, NTCP appeared inefficient to mediate infection by serum-derived HBV. It could promote HBV RNA transcription while inhibiting HBsAg secretion. Efficient PEG-independent sHBV infection of HepaRG cells permits comparative studies of diverse clinical HBV isolates and will help identify additional factors on virion surface promoting attachment to hepatocytes. IMPORTANCE Currently in vitro infection with hepatitis B virus (HBV) depends on cell culture-derived HBV inoculated in the presence of polyethylene glycol. We found patient serum-derived HBV could efficiently infect differentiated HepaRG cells independent of polyethylene glycol, which represents a more physiological infection system. Serum-derived HBV has poor infectivity in HepG2 cells reconstituted with sodium taurocholate cotransporting polypeptide (NTCP), the currently accepted HBV receptor. Moreover, HepG2/NTCP cells secreted very little hepatitis B surface antigen after infection with cell culture-derived HBV, which was attributed to NTCP overexpression, genotype D virus, and dimethyl sulfoxide added to culture medium. NTCP could promote HBV RNA transcription, protein expression, and DNA replication in HepG2 cells stably transfected with HBV DNA, while dimethyl sulfoxide could increase NTCP protein level despite transcriptional control by a cytomegalovirus promoter. Therefore, this study revealed several unusual features of NTCP as an HBV receptor and established conditions for efficient serum virus infection in vitro. PMID:27384660

Long-Term Serologic Follow-Up of Isolated Hepatitis B Core Antibody in HIV-Infected and HIV-Uninfected Women

PubMed Central

French, Audrey L.; Lin, Michael Y.; Evans, Charlesnika T.; Benning, Lorie; Glesby, Marshall J.; Young, Mary A.; Operskalski, Eva A.; Augenbraun, Michael; Peters, Marion

2009-01-01

Background Isolated antibody to hepatitis B core antigen (anti-HBc) is a common serologic finding in persons infected with human immunodeficiency virus (HIV), but the outcome and clinical significance are uncertain. Methods We performed repeated hepatitis B virus (HBV) serologic tests on women who participated in the Women’s Interagency HIV Study and who had isolated anti-HBc at study entry. Results Repeated serologic tests were performed for 322 women (282 HIV-infected and 40 HIV-uninfected) at a median of 7.5 years after study entry. Seventy-one percent of women retained isolated anti-HBc serologic status, 20% acquired antibody to hepatitis B surface antigen (anti-HBs), and 2% acquired hepatitis B surface antigen (HBsAg). In unadjusted analysis, increasing age, injection drug use, and hepatitis C viremia were negatively associated with acquisition of anti-HBs. For HIV-infected women, predictors of acquisition of anti-HBs were an increase in CD4 cell count and the use of highly active antiretroviral therapy (HAART). Receipt of drugs with activity against HBV and self-reported HBV vaccination did not predict anti-HBs acquisition. In the multivariable regression model, HAART use remained a significant predictor of anti-HBs acquisition, whereas women with hepatitis C viremia were more likely to retain isolated anti-HBc serologic status. Conclusions Isolated anti-HBc status remained stable over time for the majority of women, especially women with chronic hepatitis C virus infection. Development of anti-HBs was predicted by HAART use and an increase in CD4 cell count. We conclude that a proportion of HIV-infected women with isolated anti-HBc have prior natural HBV infection with anti-HBs that is at an undetectable level because of immune dysfunction. Isolated anti-HBc in the presence of chronic hepatitis C virus infection may be attributable to a different phenomenon, such as dysfunctional antibody production. PMID:19480573

Biosynthesis and processing of a human T lymphocyte antigen.

PubMed

Bergman, Y; Levy, R

1982-03-01

The biosynthesis and processing of Leu-1, a human T lymphocyte antigen, has been studied with the use of a monoclonal antibody. This molecule exists on the cell surface as a 67,000 m.w. glycoprotein. Through a series of pulse-labeling studies, in conjunction with the use of the antibiotic tunicamycin and the enzyme Endo-H, the details of glycosylation, processing, and deposition at the cell membrane were examined. The protein backbone of the molecule is 58,000 m.w. High-mannose sugars are added to asparagine residues during synthesis. Within 20 min, these high mannose sugars are converted to complex type carbohydrates, including fucose. The fully processed glycoprotein appears at the cell surface within 30 min after synthesis. This sequence of events is similar to that for other cell surface glycoproteins, including HLA and vesicular stomatitus virus glycoprotein.

Viral Inhibition of the Transporter Associated with Antigen Processing (TAP): A Striking Example of Functional Convergent Evolution

PubMed Central

Verweij, Marieke C.; Horst, Daniëlle; Griffin, Bryan D.; Luteijn, Rutger D.; Davison, Andrew J.; Ressing, Maaike E.; Wiertz, Emmanuel J. H. J.

2015-01-01

Herpesviruses are large DNA viruses that are highly abundant within their host populations. Even in the presence of a healthy immune system, these viruses manage to cause lifelong infections. This persistence is partially mediated by the virus entering latency, a phase of infection characterized by limited viral protein expression. Moreover, herpesviruses have devoted a significant part of their coding capacity to immune evasion strategies. It is believed that the close coexistence of herpesviruses and their hosts has resulted in the evolution of viral proteins that specifically attack multiple arms of the host immune system. Cytotoxic T lymphocytes (CTLs) play an important role in antiviral immunity. CTLs recognize their target through viral peptides presented in the context of MHC molecules at the cell surface. Every herpesvirus studied to date encodes multiple immune evasion molecules that effectively interfere with specific steps of the MHC class I antigen presentation pathway. The transporter associated with antigen processing (TAP) plays a key role in the loading of viral peptides onto MHC class I molecules. This is reflected by the numerous ways herpesviruses have developed to block TAP function. In this review, we describe the characteristics and mechanisms of action of all known virus-encoded TAP inhibitors. Orthologs of these proteins encoded by related viruses are identified, and the conservation of TAP inhibition is discussed. A phylogenetic analysis of members of the family Herpesviridae is included to study the origin of these molecules. In addition, we discuss the characteristics of the first TAP inhibitor identified outside the herpesvirus family, namely, in cowpox virus. The strategies of TAP inhibition employed by viruses are very distinct and are likely to have been acquired independently during evolution. These findings and the recent discovery of a non-herpesvirus TAP inhibitor represent a striking example of functional convergent evolution. PMID:25880312

Directional budding of human immunodeficiency virus from monocytes.

PubMed Central

Perotti, M E; Tan, X; Phillips, D M

1996-01-01

Time-lapse cinematography revealed that activated human immunodeficiency virus (HIV)-infected monocytes crawl along surfaces, putting forward a leading pseudopod. Scanning electron micrographs showed monocyte pseudopods associated with spherical structures the size of HIV virions, and transmission electron micrographs revealed HIV virions budding from pseudopods. Filamentous actin (F-actin) was localized by electron microscopy in the pseudopod by heavy meromyosin decoration. Colocalization of F-actin and p24 viral antigen by light microscopy immunofluorescence indicated that F-actin and virus were present on the same pseudopod. These observations indicate that monocytes produce virus from a leading pseudopod. We suggest that HIV secretion at the leading edges of donor monocytes/macrophages may be an efficient way for HIV to infect target cells. PMID:8709212

H13 influenza viruses in wild birds have undergone genetic and antigenic diversification in nature.

PubMed

Wang, Zu-Jyun; Kikutani, Yuto; Nguyen, Lam Thanh; Hiono, Takahiro; Matsuno, Keita; Okamatsu, Masatoshi; Krauss, Scott; Webby, Richard; Lee, Youn-Jeong; Kida, Hiroshi; Sakoda, Yoshihiro

2018-05-23

Among 16 haemagglutinin (HA) subtypes of avian influenza viruses (AIVs), H13 AIVs have rarely been isolated in wild waterfowl. H13 AIVs cause asymptomatic infection and are maintained mainly in gull and tern populations; however, the recorded antigenic information relating to the viruses has been limited. In this study, 2 H13 AIVs, A/duck/Hokkaido/W345/2012 (H13N2) and A/duck/Hokkaido/WZ68/2012 (H13N2), isolated from the same area in the same year in our surveillance, were genetically and antigenically analyzed with 10 representative H13 strains including a prototype strain, A/gull/Maryland/704/1977 (H13N6). The HA genes of H13 AIVs were phylogenetically divided into 3 groups (I, II, and III). A/duck/Hokkaido/W345/2012 (H13N2) was genetically classified into Group III. This virus was distinct from a prototype strain, A/gull/Maryland/704/1977 (H13N6), and the virus, A/duck/Hokkaido/WZ68/2012 (H13N2), both belonging to Group I. Antigenic analysis indicated that the viruses of Group I were antigenically closely related to those of Group II, but distinct from those of Group III, including A/duck/Hokkaido/W345/2012 (H13N2). In summary, our study indicates that H13 AIVs have undergone antigenic diversification in nature.

Human monoclonal antibodies reactive with human myelomonocytic leukemia cells.

PubMed

Posner, M R; Santos, D J; Elboim, H S; Tumber, M B; Frackelton, A R

1989-04-01

Peripheral blood mononuclear cells from a patient with chronic myelogenous leukemia (CML), in remission, were depleted of CD8-positive T-cells and cultured with Epstein-Barr virus. Four of 20 cultures (20%) secreted human IgG antibodies selectively reactive with the cell surfaces of certain human leukemia cell lines. Three polyclonal, Epstein-Barr virus-transformed, B-cell lines were expanded and fused with the human-mouse myeloma analogue HMMA2.11TG/O. Antibody from secreting clones HL 1.2 (IgG1), HL 2.1 (IgG3), and HL 3.1 (IgG1) have been characterized. All three react with HL-60 (promyelocytic), RWLeu4 (CML promyelocytic), and U937 (monocytic), but not with KG-1 (myeloblastic) or K562 (CML erythroid). There is no reactivity with T-cell lines, Burkitt's cell lines, pre-B-leukemia cell lines, or an undifferentiated CML cell line, BV173. Leukemic cells from two of seven patients with acute myelogenous leukemia and one of five with acute lymphocytic leukemia react with all three antibodies. Normal lymphocytes, monocytes, polymorphonuclear cells, red blood cells, bone marrow cells, and platelets do not react. Samples from patients with other diverse hematopoietic malignancies showed no reactivity. Immunoprecipitations suggest that the reactive antigen(s) is a lactoperoxidase iodinatable series of cell surface proteins with molecular weights of 42,000-54,000 and a noniodinatable protein with a molecular weight of 82,000. Based on these data these human monoclonal antibodies appear to react with myelomonocytic leukemic cells and may detect a leukemia-specific antigen or a highly restricted differentiation antigen.

An improved Abbott ARCHITECT assay for the detection of hepatitis B virus surface antigen (HBsAg).

PubMed

Lou, Sheng C; Pearce, Sandra K; Lukaszewska, Teresa X; Taylor, Russell E; Williams, Gregg T; Leary, Thomas P

2011-05-01

The sensitive and accurate detection of hepatitis B virus surface antigen (HBsAg) is critical to the identification of infection and the prevention of transfusion transmitted disease. Improvement in HBsAg assay sensitivity is essential to reduce the window to detect an acute HBV infection. Additionally, the sensitive detection of HBsAg mutants that continue to evolve due to vaccine escape, immune selection and an error prone reverse transcriptase is a necessity. A fully automated HBsAg prototype assay on the Abbott ARCHITECT instrument was developed to improve sensitivity and mutant detection. This magnetic microparticle-based assay utilizes anti-HBsAg monoclonal antibodies to capture antigen present in serum or plasma. Captured antigen is then detected using anti-HBsAg antibody conjugated with the chemiluminescent compound, acridinium. The sensitivity of the ARCHITECT HBsAg prototype assay was improved as compared to the current ARCHITECT, PRISM, and competitor HBsAg assays. The enhancement in assay sensitivity was demonstrated by the use of commercially available HBV seroconversion panels. The prototype assay detected more panel members (185 of 383) vs. the current ARCHITECT (171), PRISM (181), or competitor HBsAg assays (73/140 vs. 62/140, respectively). The ARCHITECT prototype assay also efficiently detected all mutants evaluated. Finally, the sensitivity improvement did not compromise the specificity of the assay (99.94%). An improved Abbott ARCHITECT HBsAg prototype assay with enhanced detection of HBsAg and HBsAg mutants, as well as equivalent specificity was developed for the detection, diagnosis, and management of HBV infection. Copyright © 2011 Elsevier B.V. All rights reserved.

Detection of Antigenic Variants of Subtype H3 Swine Influenza A Viruses from Clinical Samples.

PubMed

Martin, Brigitte E; Bowman, Andrew S; Li, Lei; Nolting, Jacqueline M; Smith, David R; Hanson, Larry A; Wan, Xiu-Feng

2017-04-01

A large population of genetically and antigenically diverse influenza A viruses (IAVs) are circulating among the swine population, playing an important role in influenza ecology. Swine IAVs not only cause outbreaks among swine but also can be transmitted to humans, causing sporadic infections and even pandemic outbreaks. Antigenic characterizations of swine IAVs are key to understanding the natural history of these viruses in swine and to selecting strains for effective vaccines. However, influenza outbreaks generally spread rapidly among swine, and the conventional methods for antigenic characterization require virus propagation, a time-consuming process that can significantly reduce the effectiveness of vaccination programs. We developed and validated a rapid, sensitive, and robust method, the polyclonal serum-based proximity ligation assay (polyPLA), to identify antigenic variants of subtype H3N2 swine IAVs. This method utilizes oligonucleotide-conjugated polyclonal antibodies and quantifies antibody-antigen binding affinities by quantitative reverse transcription-PCR (RT-PCR). Results showed the assay can rapidly detect H3N2 IAVs directly from nasal wash or nasal swab samples collected from laboratory-challenged animals or during influenza surveillance at county fairs. In addition, polyPLA can accurately separate the viruses at two contemporary swine IAV antigenic clusters (H3N2 swine IAV-α and H3N2 swine IAV-ß) with a sensitivity of 84.9% and a specificity of 100.0%. The polyPLA can be routinely used in surveillance programs to detect antigenic variants of influenza viruses and to select vaccine strains for use in controlling and preventing disease in swine. Copyright © 2017 American Society for Microbiology.

Advances in alfalfa mosaic virus-mediated expression of anthrax antigen in planta

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brodzik, R.; Bandurska, K.; Deka, D.

2005-12-16

Plant viruses show great potential for production of pharmaceuticals in plants. Such viruses can harbor a small antigenic peptide(s) as a part of their coat proteins (CP) and elicit an antigen-specific immune response. Here, we report the high yield and consistency in production of recombinant alfalfa mosaic virus (AlMV) particles for specific presentation of the small loop 15 amino acid epitope from domain-4 of the Bacillus anthracis protective antigen (PA-D4s). The epitope was inserted immediately after the first 25 N-terminal amino acids of AlMV CP to retain genome activation and binding of CP to viral RNAs. Recombinant AlMV particles weremore » efficiently produced in tobacco, easily purified for immunological analysis, and exhibited extended stability and systemic proliferation in planta. Intraperitional injections of mice with recombinant plant virus particles harboring the PA-D4s epitope elicited a distinct immune response. Western blotting and ELISA analysis showed that sera from immunized mice recognized both native PA antigen and the AlMV CP.« less

Production and Evaluation of a Purified Adenovirus Group-Specific (Hexon) Antigen for Use in the Diagnostic Complement Fixation Test

PubMed Central

Dowdle, W. R.; Lambriex, M.; Hierholzer, J. C.

1971-01-01

A simple procedure for the production of large volumes of purified adenovirus group-specific complement-fixing (CF) (hexon) antigen by selective adsorption to and elution from CaHPO4 is described. Results of immunodiffusion tests, electrophoresis, electron microscopy, and tests for hemagglutination and infectivity indicate that the purified antigen consisted of a single virus component (hexon). The purified product contained little host materials. Unlike the crude virus harvest usually employed for serodiagnostic CF tests, the purified antigen demonstrated no anticomplementary activity and did not develop such activity during storage. The purified antigen was equal to or slightly more sensitive than crude virus harvests for serodiagnosis of adenovirus infections. Images PMID:4325021

Analysis of hepatitis B surface antigen (HBsAg) using high-sensitivity HBsAg assays in hepatitis B virus carriers in whom HBsAg seroclearance was confirmed by conventional assays.

PubMed

Ozeki, Itaru; Nakajima, Tomoaki; Suii, Hirokazu; Tatsumi, Ryoji; Yamaguchi, Masakatsu; Kimura, Mutsuumi; Arakawa, Tomohiro; Kuwata, Yasuaki; Ohmura, Takumi; Hige, Shuhei; Karino, Yoshiyasu; Toyota, Joji

2018-02-01

We investigated the utility of high-sensitivity hepatitis B surface antigen (HBsAg) assays compared with conventional HBsAg assays. Using serum samples from 114 hepatitis B virus (HBV) carriers in whom HBsAg seroclearance was confirmed by conventional HBsAg assays (cut-off value, 0.05 IU/mL), the amount of HBsAg was re-examined by high-sensitivity HBsAg assays (cut-off value, 0.005 IU/mL). Cases negative for HBsAg in both assays were defined as consistent cases, and cases positive for HBsAg in the high-sensitivity HBsAg assay only were defined as discrepant cases. There were 55 (48.2%) discrepant cases, and the range of HBsAg titers determined by high-sensitivity HBsAg assays was 0.005-0.056 IU/mL. Multivariate analysis showed that the presence of nucleos(t)ide analog therapy, liver cirrhosis, and negative anti-HBs contributed to the discrepancies between the two assays. Cumulative anti-HBs positivity rates among discrepant cases were 12.7%, 17.2%, 38.8%, and 43.9% at baseline, 1 year, 3 years, and 5 years, respectively, whereas the corresponding rates among consistent cases were 50.8%, 56.0%, 61.7%, and 68.0%, respectively. Hepatitis B virus DNA negativity rates were 56.4% and 81.4% at baseline, 51.3% and 83.3% at 1 year, and 36.8% and 95.7% at 3 years, among discrepant and consistent cases, respectively. Hepatitis B surface antigen reversion was observed only in discrepant cases. Re-examination by high-sensitivity HBsAg assays revealed that HBsAg was positive in approximately 50% of cases. Cumulative anti-HBs seroconversion rates and HBV-DNA seroclearance rates were lower in these cases, suggesting a population at risk for HBsAg reversion. © 2017 The Japan Society of Hepatology.

Vaccination with Recombinant Parainfluenza Virus 5 Expressing Neuraminidase Protects against Homologous and Heterologous Influenza Virus Challenge.

PubMed

Mooney, Alaina J; Gabbard, Jon D; Li, Zhuo; Dlugolenski, Daniel A; Johnson, Scott K; Tripp, Ralph A; He, Biao; Tompkins, S Mark

2017-12-01

Seasonal human influenza virus continues to cause morbidity and mortality annually, and highly pathogenic avian influenza (HPAI) viruses along with other emerging influenza viruses continue to pose pandemic threats. Vaccination is considered the most effective measure for controlling influenza; however, current strategies rely on a precise vaccine match with currently circulating virus strains for efficacy, requiring constant surveillance and regular development of matched vaccines. Current vaccines focus on eliciting specific antibody responses against the hemagglutinin (HA) surface glycoprotein; however, the diversity of HAs across species and antigenic drift of circulating strains enable the evasion of virus-inhibiting antibody responses, resulting in vaccine failure. The neuraminidase (NA) surface glycoprotein, while diverse, has a conserved enzymatic site and presents an appealing target for priming broadly effective antibody responses. Here we show that vaccination with parainfluenza virus 5 (PIV5), a promising live viral vector expressing NA from avian (H5N1) or pandemic (H1N1) influenza virus, elicited NA-specific antibody and T cell responses, which conferred protection against homologous and heterologous influenza virus challenges. Vaccination with PIV5-N1 NA provided cross-protection against challenge with a heterosubtypic (H3N2) virus. Experiments using antibody transfer indicate that antibodies to NA have an important role in protection. These findings indicate that PIV5 expressing NA may be effective as a broadly protective vaccine against seasonal influenza and emerging pandemic threats. IMPORTANCE Seasonal influenza viruses cause considerable morbidity and mortality annually, while emerging viruses pose potential pandemic threats. Currently licensed influenza virus vaccines rely on the antigenic match of hemagglutinin (HA) for vaccine strain selection, and most vaccines rely on HA inhibition titers to determine efficacy, despite the growing awareness of the contribution of neuraminidase (NA) to influenza virus vaccine efficacy. Although NA is immunologically subdominant to HA, and clinical studies have shown variable NA responses to vaccination, in this study, we show that vaccination with a parainfluenza virus 5 recombinant vaccine candidate expressing NA (PIV5-NA) from a pandemic influenza (pdmH1N1) virus or highly pathogenic avian influenza (H5N1) virus elicits robust, cross-reactive protection from influenza virus infection in two animal models. New vaccination strategies incorporating NA, including PIV5-NA, could improve seasonal influenza virus vaccine efficacy and provide protection against emerging influenza viruses. Copyright © 2017 American Society for Microbiology.

Hepatitis B core-related antigen (HBcrAg) levels in the natural history of hepatitis B virus infection in a large European cohort predominantly infected with genotypes A and D.

PubMed

Maasoumy, B; Wiegand, S B; Jaroszewicz, J; Bremer, B; Lehmann, P; Deterding, K; Taranta, A; Manns, M P; Wedemeyer, H; Glebe, D; Cornberg, M

2015-06-01

Hepatitis B core-related antigen (HBcrAg) has been suggested as an additional marker of hepatitis B virus (HBV) infection. HBcrAg combines the antigenic reactivity resulting from denatured hepatitis B e antigen (HBeAg), HBV core antigen and an artificial core-related protein (p22cr). In Asian patients, high levels of HBcrAg have been suggested to be an independent risk factor for hepatocellular carcinoma, while low levels could guide safe cessation of treatment with nucleos(t)ide analogues. We here studied HBcrAg levels in different phases of HBV infection in a large European cohort predominantly infected with genotypes A and D: HBeAg-positive immune tolerance (n = 30), HBeAg-positive immune clearance (IC) (n = 60), HBeAg-negative hepatitis (ENH) (n = 50), HBeAg-negative inactive/quiescent carrier phase (c) (n = 109) and acute hepatitis B (n = 8). Median HBcrAg levels were high in the immune tolerance and immune clearance phases (8.41 and 8.11 log U/mL, respectively), lower in ENH subjects (4.82 log U/mL) but only 2.00 log U/mL in ENQ subjects. Correlation between HBcrAg and HBV DNA varied among the different phases of HBV infection, while HBcrAg moderately correlated with hepatitis B surface antigen in all phases. ENQ patients had HBcrAg levels <3 log U/mL in 79%, in contrast to only 12% in the ENH group. HBcrAg levels vary significantly during the different phases of HBV infection. HBcrAg may serve as valuable marker for virus replication and reflect the transcriptional activity of intrahepatic cccDNA. In HBeAg-negative patients, HBcrAg may help to distinguish between inactive carriers (ENQ) and those with active disease (ENH). Copyright © 2015 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Characterization of recombinant yellow fever-dengue vaccine viruses with human monoclonal antibodies targeting key conformational epitopes.

PubMed

Lecouturier, Valerie; Berry, Catherine; Saulnier, Aure; Naville, Sophie; Manin, Catherine; Girerd-Chambaz, Yves; Crowe, James E; Jackson, Nicholas; Guy, Bruno

2018-04-26

The recombinant yellow fever-17D-dengue virus, live, attenuated, tetravalent dengue vaccine (CYD-TDV) is licensed in several dengue-endemic countries. Although the vaccine provides protection against dengue, the level of protection differs by serotype and warrants further investigation. We characterized the antigenic properties of each vaccine virus serotype using highly neutralizing human monoclonal antibodies (hmAbs) that bind quaternary structure-dependent epitopes. Specifically, we monitored the binding of dengue virus-1 (DENV-1; 1F4), DENV-2 (2D22) or DENV-3 (5J7) serotype-specific or DENV-1-4 cross-reactive (1C19) hmAbs to the four chimeric yellow fever-dengue vaccine viruses (CYD-1-4) included in phase III vaccine formulations using a range of biochemical and functional assays (dot blot, ELISA, surface plasmon resonance and plaque reduction neutralization assays). In addition, we used the "classic" live, attenuated DENV-2 vaccine serotype, immature CYD-2 viruses and DENV-2 virus-like particles as control antigens for anti-serotype-2 reactivity. The CYD vaccine serotypes were recognized by each hmAbs with the expected specificity, moreover, surface plasmon resonance indicated a high functional affinity interaction with the CYD serotypes. In addition, the hmAbs provided similar protection against CYD and wild-type dengue viruses in the in vitro neutralization assay. Overall, these findings demonstrate that the four CYD viruses used in clinical trials display key conformational and functional epitopes targeted by serotype-specific and/or cross-reactive neutralizing human antibodies. More specifically, we showed that CYD-2 displays serotype- specific epitopes present only on the mature virus. This indicates that the CYD-TDV has the ability to elicit antibody specificities which are similar to those induced by the wild type DENV. Future investigations will be needed to address the nature of CYD-TDV-induced responses after vaccine administration, and how these laboratory markers relate to vaccine efficacy and safety. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Evaluation of the Potential Risk of Hepatitis B Virus Transmission in Skin Allografting.

PubMed

Wang, D; Xie, W; Chen, T; Dong, C; Zhao, C; Tan, H; Tian, H; Xie, Q

2015-01-01

Skin transplantation is associated with potential risk of infectious disease transmission; however, the exclusion of donors owing to hepatitis B virus (HBV) infection will worsen the shortage of allograft skin supply. We report a paired study to evaluate the potential risk of HBV transmission in skin allografting. The presence of HBV DNA in the serum and skin from 37 burn patients with chronic HBV infection (CHB) was monitored by a HBV polymerase chain reaction (PCR) and the positive rates were compared by Fisher's exact probability test. There was a high consistency in the HBV serology profile between HBV DNA PCR (83.78%) and the clinical HBV test. Only 2 patients who were positive for hepatitis B surface antigen, hepatitis B e antigen, and hepatitis B core antibody had detectable HBV DNA in the skin tissue; however, no hepatitis B surface antigen was detected as examined by immunohistochemistry staining. There was a significant difference between the positive rates of HBV DNA in the serum and skin (χc(2) = 27.03; P < .001). The potential risk for HBV transmission by skin allografting is very low. Given that China has a large population of patients with HBV, the acceptance of skin from donors with CHB to the skin bank would increase the number of tissue donations to meet the urgent medical need for skin transplantation. Copyright © 2015 Elsevier Inc. All rights reserved.

Antigenic variation of the human influenza A (H3N2) virus during the 2014-2015 winter season.

PubMed

Hua, Sha; Li, XiYan; Liu, Mi; Cheng, YanHui; Peng, YouSong; Huang, WeiJuan; Tan, MinJu; Wei, HeJiang; Guo, JunFeng; Wang, DaYan; Wu, AiPing; Shu, YueLong; Jiang, TaiJiao

2015-09-01

The human influenza A (H3N2) virus dominated the 2014-2015 winter season in many countries and caused massive morbidity and mortality because of its antigenic variation. So far, very little is known about the antigenic patterns of the recent H3N2 virus. By systematically mapping the antigenic relationships of H3N2 strains isolated since 2010, we discovered that two groups with obvious antigenic divergence, named SW13 (A/Switzerland/9715293/2013-like strains) and HK14 (A/Hong Kong/5738/2014-like strains), co-circulated during the 2014-2015 winter season. HK14 group co-circulated with SW13 in Europe and the United States during this season, while there were few strains of HK14 in mainland China, where SW13 has dominated since 2012. Furthermore, we found that substitutions near the receptor-binding site on hemagglutinin played an important role in the antigenic variation of both the groups. These findings provide a comprehensive understanding of the recent antigenic evolution of H3N2 virus and will aid in the selection of vaccine strains.

Matrix-M™ adjuvant enhances immunogenicity of both protein- and modified vaccinia virus Ankara-based influenza vaccines in mice.

PubMed

Magnusson, Sofia E; Altenburg, Arwen F; Bengtsson, Karin Lövgren; Bosman, Fons; de Vries, Rory D; Rimmelzwaan, Guus F; Stertman, Linda

2018-04-01

Influenza viruses continuously circulate in the human population and escape recognition by virus neutralizing antibodies induced by prior infection or vaccination through accumulation of mutations in the surface proteins hemagglutinin (HA) and neuraminidase (NA). Various strategies to develop a vaccine that provides broad protection against different influenza A viruses are under investigation, including use of recombinant (r) viral vectors and adjuvants. The replication-deficient modified vaccinia virus Ankara (MVA) is a promising vaccine vector that efficiently induces B and T cell responses specific for the antigen of interest. It is assumed that live vaccine vectors do not require an adjuvant to be immunogenic as the vector already mediates recruitment and activation of immune cells. To address this topic, BALB/c mice were vaccinated with either protein- or rMVA-based HA influenza vaccines, formulated with or without the saponin-based Matrix-M™ adjuvant. Co-formulation with Matrix-M significantly increased HA vaccine immunogenicity, resulting in antigen-specific humoral and cellular immune responses comparable to those induced by unadjuvanted rMVA-HA. Of special interest, rMVA-HA immunogenicity was also enhanced by addition of Matrix-M, demonstrated by enhanced HA inhibition antibody titres and cellular immune responses. Matrix-M added to either protein- or rMVA-based HA vaccines mediated recruitment and activation of antigen-presenting cells and lymphocytes to the draining lymph node 24 and 48 h post-vaccination. Taken together, these results suggest that adjuvants can be used not only with protein-based vaccines but also in combination with rMVA to increase vaccine immunogenicity, which may be a step forward to generate new and more effective influenza vaccines.

Issues encountered in development of enzyme-linked immunosorbent assay for use in detecting Influenza A virus subtype H5N1 exposure in swine.

PubMed

Buehler, Jason; Lager, Kelly; Vincent, Amy; Miller, Cathy; Thacker, Eileen; Janke, Bruce

2014-03-01

A potential mechanism by which highly pathogenic avian Influenza A virus subtype H5N1 could more readily infect human beings is through the infection of and adaptation in pigs. To detect the occurrence of such infection, monitoring of pig populations through serological screening would be highly desirable. In the current study, hemagglutination inhibition assays were able to detect antibodies against H5N1 developed in pigs, but because of antigenic variation between clades, the use of multiple virus strains were required. Whole recombinant virus and recombinant hemagglutinin antigen enzyme-linked immunosorbent assays (ELISAs) were generated that could detect antibody against multiple H5N1 strains, but which also detected antibody against endemic swine influenza viruses. A recombinant hemagglutinin antigen-based ELISA was as effective as the whole virus antigen ELISAs in detecting antibody against the H5N1 virus strains used and eliminated nearly all of the cross-reactivity with non-H5N1 virus antibody. The current study also highlighted the difficulty in establishing a decision (cutoff) value that would effectively counterbalance nonspecific reactivity against sensitivity. The results provide important information and considerations for the development of serological screening assays for highly pathogenic avian H5N1 viruses.

Genetic and antigenic characterisation of serotype A FMD viruses from East Africa to select new vaccine strains

PubMed Central

Bari, Fufa D.; Parida, Satya; Tekleghiorghis, Tesfaalem; Dekker, Aldo; Sangula, Abraham; Reeve, Richard; Haydon, Daniel T.; Paton, David J.; Mahapatra, Mana

2014-01-01

Vaccine strain selection for emerging foot-and-mouth disease virus (FMDV) outbreaks in enzootic countries can be addressed through antigenic and genetic characterisation of recently circulating viruses. A total of 56 serotype A FMDVs isolated between 1998 and 2012, from Central, East and North African countries were characterised antigenically by virus neutralisation test using antisera to three existing and four candidate vaccine strains and, genetically by characterising the full capsid sequence data. A Bayesian analysis of the capsid sequence data revealed the viruses to be of either African or Asian topotypes with subdivision of the African topotype viruses into four genotypes (Genotypes I, II, IV and VII). The existing vaccine strains were found to be least cross-reactive (good matches observed for only 5.4–46.4% of the sampled viruses). Three bovine antisera, raised against A-EA-2007, A-EA-1981 and A-EA-1984 viruses, exhibited broad cross-neutralisation, towards more than 85% of the circulating viruses. Of the three vaccines, A-EA-2007 was the best showing more than 90% in-vitro cross-protection, as well as being the most recent amongst the vaccine strains used in this study. It therefore appears antigenically suitable as a vaccine strain to be used in the region in FMD control programmes. PMID:25171846

Molecular Characterizations of Surface Proteins Hemagglutinin and Neuraminidase from Recent H5Nx Avian Influenza Viruses.

PubMed

Yang, Hua; Carney, Paul J; Mishin, Vasiliy P; Guo, Zhu; Chang, Jessie C; Wentworth, David E; Gubareva, Larisa V; Stevens, James

2016-06-15

During 2014, a subclade 2.3.4.4 highly pathogenic avian influenza (HPAI) A(H5N8) virus caused poultry outbreaks around the world. In late 2014/early 2015, the virus was detected in wild birds in Canada and the United States, and these viruses also gave rise to reassortant progeny, composed of viral RNA segments (vRNAs) from both Eurasian and North American lineages. In particular, viruses were found with N1, N2, and N8 neuraminidase vRNAs, and these are collectively referred to as H5Nx viruses. In the United States, more than 48 million domestic birds have been affected. Here we present a detailed structural and biochemical analysis of the surface antigens of H5N1, H5N2, and H5N8 viruses in addition to those of a recent human H5N6 virus. Our results with recombinant hemagglutinin reveal that these viruses have a strict avian receptor binding preference, while recombinantly expressed neuraminidases are sensitive to FDA-approved and investigational antivirals. Although H5Nx viruses currently pose a low risk to humans, it is important to maintain surveillance of these circulating viruses and to continually assess future changes that may increase their pandemic potential. The H5Nx viruses emerging in North America, Europe, and Asia pose a great public health concern. Here we report a molecular and structural study of the major surface proteins of several H5Nx influenza viruses. Our results improve the understanding of these new viruses and provide important information on their receptor preferences and susceptibilities to antivirals, which are central to pandemic risk assessment. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Molecular Characterizations of Surface Proteins Hemagglutinin and Neuraminidase from Recent H5Nx Avian Influenza Viruses

PubMed Central

Yang, Hua; Carney, Paul J.; Mishin, Vasiliy P.; Guo, Zhu; Chang, Jessie C.; Wentworth, David E.; Gubareva, Larisa V.

2016-01-01

ABSTRACT During 2014, a subclade 2.3.4.4 highly pathogenic avian influenza (HPAI) A(H5N8) virus caused poultry outbreaks around the world. In late 2014/early 2015, the virus was detected in wild birds in Canada and the United States, and these viruses also gave rise to reassortant progeny, composed of viral RNA segments (vRNAs) from both Eurasian and North American lineages. In particular, viruses were found with N1, N2, and N8 neuraminidase vRNAs, and these are collectively referred to as H5Nx viruses. In the United States, more than 48 million domestic birds have been affected. Here we present a detailed structural and biochemical analysis of the surface antigens of H5N1, H5N2, and H5N8 viruses in addition to those of a recent human H5N6 virus. Our results with recombinant hemagglutinin reveal that these viruses have a strict avian receptor binding preference, while recombinantly expressed neuraminidases are sensitive to FDA-approved and investigational antivirals. Although H5Nx viruses currently pose a low risk to humans, it is important to maintain surveillance of these circulating viruses and to continually assess future changes that may increase their pandemic potential. IMPORTANCE The H5Nx viruses emerging in North America, Europe, and Asia pose a great public health concern. Here we report a molecular and structural study of the major surface proteins of several H5Nx influenza viruses. Our results improve the understanding of these new viruses and provide important information on their receptor preferences and susceptibilities to antivirals, which are central to pandemic risk assessment. PMID:27053557

Molecular Characterizations of Surface Proteins Hemagglutinin and Neuraminidase from Recent H5Nx Avian Influenza Viruses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Hua; Carney, Paul J.; Mishin, Vasiliy P.

ABSTRACT During 2014, a subclade 2.3.4.4 highly pathogenic avian influenza (HPAI) A(H5N8) virus caused poultry outbreaks around the world. In late 2014/early 2015, the virus was detected in wild birds in Canada and the United States, and these viruses also gave rise to reassortant progeny, composed of viral RNA segments (vRNAs) from both Eurasian and North American lineages. In particular, viruses were found with N1, N2, and N8 neuraminidase vRNAs, and these are collectively referred to as H5Nx viruses. In the United States, more than 48 million domestic birds have been affected. Here we present a detailed structural and biochemicalmore » analysis of the surface antigens of H5N1, H5N2, and H5N8 viruses in addition to those of a recent human H5N6 virus. Our results with recombinant hemagglutinin reveal that these viruses have a strict avian receptor binding preference, while recombinantly expressed neuraminidases are sensitive to FDA-approved and investigational antivirals. Although H5Nx viruses currently pose a low risk to humans, it is important to maintain surveillance of these circulating viruses and to continually assess future changes that may increase their pandemic potential. IMPORTANCEThe H5Nx viruses emerging in North America, Europe, and Asia pose a great public health concern. Here we report a molecular and structural study of the major surface proteins of several H5Nx influenza viruses. Our results improve the understanding of these new viruses and provide important information on their receptor preferences and susceptibilities to antivirals, which are central to pandemic risk assessment.« less

Cloning and Expression of Genes for Dengue Virus Type-2 Encoded-Antigens for Rapid Diagnosis and Vaccine Development

DTIC Science & Technology

1988-10-31

00 0 Cloning and Expression of Genes for Dengue Virus (Type-2 Encoded-Antigens for Rapid ODiagnosis and Vaccine DevelopmentN| ANNUAL PROGRESS REPORT...11. TITLE (include Security Classification) Cloning and Expression of Genes f or Dengue Virus Type 2 Fncoded Antigens for Rapid Diagnosis and Vaccine ...epidemics in Central and South Americas and the Caribbean is a cause of major concern. An effective vaccine is not available to protect individuals

[One example of false negative hepatitis B surface antigen (EIA) result due to variant S area strain and reagment reactiveness related to hepatitis B surface antigen].

PubMed

Matsuda, Chikashi; Moriyama, Hidehiko; Taketani, Takeshi; Shibata, Hiroshi; Nagai, Atsushi

2011-01-01

The presence in serum of the Hepatitis B surface antigen (HBsAg), the outer envelope of the hepatitis B virus (HBV), indicates viral infection, used in laboratory tests to confirm this. We report a case of discrepancy among HBsAg test results detected between measurements in a subject with HB infection. Gene analysis demonstrated several S region gene mutations, not detected previously. We tested 12 measurements e.g., EIA, CLIA, CLEIA, F-EIA, MAT, and IC for whether they could detect our subject's HBsAg and found that it was not recognized by a method using only a single monoclonal antibody to detect HBsAg in two detection processes, in contrast to the 11 other measurements, which used two different antibodies. This case shows that amino acid substitution may cause a false negative result for HBsAg. Gene mutations known to occur in HBV, should thus trigger an awareness of the need to keep in mind that false negative results can happen in case such as ours.

Antigenic change in feline calicivirus during persistent infection.

PubMed Central

Johnson, R P

1992-01-01

To determine if antigenic variation occurred during persistent infection of cats with feline caliciviruses (FCV), nine persistent (progeny) isolates from nine different carrier cats were compared antigenically to the original infecting parent strain, FCV 255, by two-way cross-neutralization tests with rabbit antisera. Five of the nine progeny viruses isolated 35 to 169 days after initial infection were antigenically different from the parent strain. These five isolates represented four distinct antigenic phenotypes. The emergence of four distinctly different antigenic variants from a single parent strain indicates that FCV, like many other RNA viruses, exhibits considerable antigenic heterogeneity during replication in its natural host, and supports the hypothesis that antigenic variation contributes to chronic FCV infection. PMID:1335833

Antigenic Maps of Influenza A(H3N2) Produced With Human Antisera Obtained After Primary Infection.

PubMed

Fonville, Judith M; Fraaij, Pieter L A; de Mutsert, Gerrie; Wilks, Samuel H; van Beek, Ruud; Fouchier, Ron A M; Rimmelzwaan, Guus F

2016-01-01

Antigenic characterization of influenza viruses is typically based on hemagglutination inhibition (HI) assay data for viral isolates tested against strain-specific postinfection ferret antisera. Here, similar virus characterizations were performed using serological data from humans with primary influenza A(H3N2) infection. We screened sera collected between 1995 and 2011 from children between 9 and 24 months of age for influenza virus antibodies, performed HI tests for the positive sera against 23 influenza viruses isolated between 1989 and 2011, and measured HI titers of antisera against influenza A(H3N2) from 24 ferrets against the same panel of viruses. Of the 17 positive human sera, 6 had a high response, showing HI patterns that would be expected from primary infection antisera, while 11 sera had lower, more dispersed patterns of reactivity that are not easily explained. The antigenic map based on the high-response human HI data was similar to the map created using ferret data. Although the overall structure of the ferret and human antigenic maps is similar, local differences in virus positions indicate that the human and ferret immune system might see antigenic properties of viruses differently. Further studies are needed to establish the degree of similarity between serological patterns in ferret and human data. © The Author 2015. Published by Oxford University Press for the Infectious Diseases Society of America.

Hepatitis B virus surface antigen impairs myeloid dendritic cell function: a possible immune escape mechanism of hepatitis B virus

PubMed Central

Op den Brouw, Marjoleine L; Binda, Rekha S; van Roosmalen, Mark H; Protzer, Ulrike; Janssen, Harry L A; van der Molen, Renate G; Woltman, Andrea M

2009-01-01

Chronic hepatitis B virus (HBV) infection is the result of an inadequate immune response towards the virus. Myeloid dendritic cells (mDC) of patients with chronic HBV are impaired in their maturation and function, resulting in more tolerogenic rather than immunogenic responses, which may contribute to viral persistence. The mechanism responsible for altered mDC function remains unclear. The HBV-infected patients display large amounts of HBV particles and viral proteins in their circulation, especially the surface antigen HBsAg, which allows multiple interactions between the virus, its viral proteins and DC. To assess whether HBV directly influences mDC function, the effects of HBV and HBsAg on human mDC maturation and function were investigated in vitro. As already described for internalization of HBV by DC, the present study shows that peripheral blood-derived mDC of healthy controls also actively take up HBsAg in a time-dependent manner. Cytokine-induced maturation in the presence of HBV or HBsAg resulted in a significantly more tolerogenic mDC phenotype as demonstrated by a diminished up-regulation of costimulatory molecules and a decreased T-cell stimulatory capacity, as assessed by T-cell proliferation and interferon-γ production. In addition, the presence of HBV significantly reduced interleukin-12 production by mDC. These results show that both HBV particles and purified HBsAg have an immune modulatory capacity and may directly contribute to the dysfunction of mDC in patients with chronic HBV. The direct immune regulatory effect of HBV and circulating HBsAg particles on the function of DC can be considered as part of the mechanism by which HBV escapes immunity. PMID:18624732

Antigenicity of the 2015-2016 seasonal H1N1 human influenza virus HA and NA proteins.

PubMed

Clark, Amelia M; DeDiego, Marta L; Anderson, Christopher S; Wang, Jiong; Yang, Hongmei; Nogales, Aitor; Martinez-Sobrido, Luis; Zand, Martin S; Sangster, Mark Y; Topham, David J

2017-01-01

Antigenic drift of the hemagglutinin (HA) and neuraminidase (NA) influenza virus proteins contributes to reduced vaccine efficacy. To analyze antigenic drift in human seasonal H1N1 viruses derived from the 2009 pandemic H1N1 virus (pH1N1-like viruses) accounts for the limited effectiveness (around 40%) of vaccination against pH1N1-like viruses during the 2015-2016 season, nasal washes/swabs collected from adult subjects in the Rochester, NY area, were used to sequence and isolate the circulating viruses. The HA and NA proteins from viruses circulating during the 2015-2016 season encoded eighteen and fourteen amino acid differences, respectively, when compared to A/California/04/2009, a strain circulating at the origin of the 2009 pandemic. The circulating strains belonged to subclade 6B.1, defined by HA amino acid substitutions S101N, S179N, and I233T. Hemagglutination-inhibiting (HAI) and HA-specific neutralizing serum antibody (Ab) titers from around 50% of pH1N1-like virus-infected subjects and immune ferrets were 2-4 fold lower for the 2015-2016 circulating strains compared to the vaccine strain. In addition, using a luminex-based mPlex HA assay, the binding of human sera from subjects infected with pH1N1-like viruses to the HA proteins from circulating and vaccine strains was not identical, strongly suggesting antigenic differences in the HA protein. Additionally, NA inhibition (NAI) Ab titers in human sera from pH1N1-like virus-infected subjects increased after the infection and there were measurable antigenic differences between the NA protein of circulating strains and the vaccine strain using both ferret and human antisera. Despite having been vaccinated, infected subjects exhibited low HAI Ab titers against the vaccine and circulating strains. This suggests that poor responses to the H1N1 component of the vaccine as well as antigenic differences in the HA and NA proteins of currently circulating pH1N1-like viruses could be contributing to risk of infection even after vaccination.

Antigenicity of the 2015–2016 seasonal H1N1 human influenza virus HA and NA proteins

PubMed Central

Anderson, Christopher S.; Wang, Jiong; Yang, Hongmei; Nogales, Aitor; Martinez-Sobrido, Luis; Zand, Martin S.; Sangster, Mark Y.; Topham, David J.

2017-01-01

Antigenic drift of the hemagglutinin (HA) and neuraminidase (NA) influenza virus proteins contributes to reduced vaccine efficacy. To analyze antigenic drift in human seasonal H1N1 viruses derived from the 2009 pandemic H1N1 virus (pH1N1-like viruses) accounts for the limited effectiveness (around 40%) of vaccination against pH1N1-like viruses during the 2015–2016 season, nasal washes/swabs collected from adult subjects in the Rochester, NY area, were used to sequence and isolate the circulating viruses. The HA and NA proteins from viruses circulating during the 2015–2016 season encoded eighteen and fourteen amino acid differences, respectively, when compared to A/California/04/2009, a strain circulating at the origin of the 2009 pandemic. The circulating strains belonged to subclade 6B.1, defined by HA amino acid substitutions S101N, S179N, and I233T. Hemagglutination-inhibiting (HAI) and HA-specific neutralizing serum antibody (Ab) titers from around 50% of pH1N1-like virus-infected subjects and immune ferrets were 2–4 fold lower for the 2015–2016 circulating strains compared to the vaccine strain. In addition, using a luminex-based mPlex HA assay, the binding of human sera from subjects infected with pH1N1-like viruses to the HA proteins from circulating and vaccine strains was not identical, strongly suggesting antigenic differences in the HA protein. Additionally, NA inhibition (NAI) Ab titers in human sera from pH1N1-like virus-infected subjects increased after the infection and there were measurable antigenic differences between the NA protein of circulating strains and the vaccine strain using both ferret and human antisera. Despite having been vaccinated, infected subjects exhibited low HAI Ab titers against the vaccine and circulating strains. This suggests that poor responses to the H1N1 component of the vaccine as well as antigenic differences in the HA and NA proteins of currently circulating pH1N1-like viruses could be contributing to risk of infection even after vaccination. PMID:29145498

Monoclonal antibodies of African swine fever virus: antigenic differences among field virus isolates and viruses passaged in cell culture.

PubMed Central

García-Barreno, B; Sanz, A; Nogal, M L; Viñuela, E; Enjuanes, L

1986-01-01

An analysis of the binding properties of a collection of monoclonal antibodies to African swine fever virus particles showed that virus field isolates passaged in porcine macrophages changed antigenically more than a strain of a cell-adapted virus passaged in Vero cells. From seven clones isolated from the spleen of a field-infected pig, we found four clones that had the same antigenic properties, one clone that had large changes in proteins p150 and p27 and small changes in proteins p37 and p14, and two clones that had minor changes in proteins p150 and p27, respectively. An analysis of the binding properties of the monoclonal antibodies to 23 field isolates from Africa, Europe, and America showed that the African isolates differed among themselves more than the European and the American isolates; in this study we found changes in 8 of the 10 virus proteins tested. The most variable proteins in the African isolates were p150, p27, p14, and p12. In contrast to the African isolates, protein p12 from the non-African viruses did not change. The clustering of the field virus isolates in six antigenic homology groups indicated the existence of a complex variety of African swine fever virus serotypes. PMID:2422393

Cloning of cDNA of major antigen of foot and mouth disease virus and expression in E. coli

NASA Astrophysics Data System (ADS)

Küpper, Hans; Keller, Walter; Kurz, Christina; Forss, Sonja; Schaller, Heinz

1981-02-01

Double-stranded DNA copies of the single-stranded genomic RNA of foot and mouth disease virus have been cloned into the Escherichia coli plasmid pBR322. A restriction map of the viral genome was established and aligned with the biochemical map of foot and mouth disease virus. The coding sequence for structural protein VP1, the major antigen of the virus, was identified and inserted into a plasmid vector where the expression of this sequence is under control of the phage λ PL promoter. In an appropriate host the synthesis of antigenic polypeptide can be demonstrated by radioimmunoassay.

Study of rubella candidate vaccine based on a structurally modified plant virus.

PubMed

Trifonova, Ekaterina A; Zenin, Vladimir A; Nikitin, Nikolai A; Yurkova, Maria S; Ryabchevskaya, Ekaterina M; Putlyaev, Egor V; Donchenko, Ekaterina K; Kondakova, Olga A; Fedorov, Alexey N; Atabekov, Joseph G; Karpova, Olga V

2017-08-01

A novel rubella candidate vaccine based on a structurally modified plant virus - spherical particles (SPs) - was developed. SPs generated by the thermal remodelling of the tobacco mosaic virus are promising platforms for the development of vaccines. SPs combine unique properties: biosafety, stability, high immunogenicity and the effective adsorption of antigens. We assembled in vitro and characterised complexes (candidate vaccine) based on SPs and the rubella virus recombinant antigen. The candidate vaccine induced a strong humoral immune response against rubella. The IgG isotypes ratio indicated the predominance of IgG1 which plays a key role in immunity to natural rubella infection. The immune response was generally directed against the rubella antigen within the complexes. We suggest that SPs can act as a platform (depot) for the rubella antigen, enhancing specific immune response. Our results demonstrate that SPs-antigen complexes can be an effective and safe candidate vaccine against rubella. Copyright © 2017 Elsevier B.V. All rights reserved.

EFFECT OF PROFLAVINE ON THE SYNTHESIS OF ADENOVIRUS, TYPE 5, AND ASSOCIATED SOLUBLE ANTIGENS

PubMed Central

Wilcox, Wesley C.; Ginsberg, Harold S.

1962-01-01

Wilcox, Wesley C. (University of Pennsylvania, Philadelphia) and Harold S. Ginsberg. Effect of proflavine on the synthesis of adenovirus, type 5, and associated soluble antigens. J. Bacteriol. 84:526–533. 1962.—The synthesis of type 5 adenovirus in HeLa cells was suppressed to a considerable extent by low concentrations of proflavine, an acridine dye. In comparison, the processes leading to the production of soluble complement-fixing antigens and toxin were less sensitive to the action of this chemical. Addition of proflavine to infected cells at different times during the virus growth cycle revealed that the processes leading to the synthesis of soluble antigens began prior to the first appearance of newly synthesized virus. This observation is compatible with the hypothesis that the soluble antigens may represent virus subunits or precursor materials. In addition, these data indicate that it is possible to interrupt the latter stages of the virus synthetic process by addition of proflavine late in the eclipse period. PMID:14000661

Binding of HBGA-expressing bacteria does not protect Tulane virus from acute heat stress

USDA-ARS?s Scientific Manuscript database

Human noroviruses (HuNoVs) are the major cause of gastroenteritis outbreaks worldwide. Human noroviruses can interact with histo-blood group antigens (HBGAs) on the surface of mammalian cells as well as bacterial cells. HBGAs have been considered as putative receptors or co-receptors for HuNoVs in m...

Hepatitis B reactivation and timing for prophylaxis

PubMed Central

Tuna, Nazan; Karabay, Oguz

2015-01-01

It is known that immunotherapy and cancer chemotherapy may cause hepatitis B virus (HBV) reactivation in hepatitis B surface antigen carriers and inactive chronic hepatitis B patients. Guidelines recommend antiviral prophylaxis regardless of HBV DNA levels to prevent reactivation. We read from the article written by Liu et al that Lamivudine was given inadequate time for antiviral prophylaxis. PMID:25717269

A versatile SERS-based immunoassay for immunoglobulin detection using antigen-coated gold nanoparticles and malachite green-conjugated protein A/G

USDA-ARS?s Scientific Manuscript database

A surface enhanced Raman scattering (SERS) immunoassay for antibody detection in serum is described in the present work. The developed assay is conducted in solution and utilizes Au nanoparticles coated with the envelope (E) protein of West Nile Virus (WNV) as the SERS-active substrate and malachite...

Detection of intracellular canine distemper virus antigen in mink inoculated with an attenuated or a virulent strain of canine distemper virus.

PubMed

Blixenkrone-Møller, M

1989-09-01

Using an indirect immunofluorescence technique, the distribution of viral antigen in various tissues and blood mononuclear leukocytes was studied in wild mink, either vaccinated with an attenuated vaccine strain of canine distemper virus (CDV) or experimentally inoculated with the virulent Snyder-Hill strain of CDV. Viral antigen was detected in cells of the lymphoid system 6 to 12 days after vaccination. From 2 to 3 days after inoculation with the virulent strain, CDV antigen was demonstrated in cells of the lymphoid system and, during the incubation period, the antigen had spread to the epithelia and brain at days 6 and 12, respectively. In clinical cases of acute fatal canine distemper, the viral antigen was detected in a wide variety of tissues, including the cells of the lymphoid system, epithelial cells of skin, mucous membranes, lung, kidney, and cells of the CNS. The diagnostic importance of CDV antigen detection is discussed on the basis of these findings.

Inactivated Recombinant Rabies Viruses Displaying Canine Distemper Virus Glycoproteins Induce Protective Immunity against Both Pathogens

PubMed Central

da Fontoura Budaszewski, Renata; Hudacek, Andrew; Sawatsky, Bevan; Krämer, Beate; Yin, Xiangping

2017-01-01

ABSTRACT The development of multivalent vaccines is an attractive methodology for the simultaneous prevention of several infectious diseases in vulnerable populations. Both canine distemper virus (CDV) and rabies virus (RABV) cause lethal disease in wild and domestic carnivores. While RABV vaccines are inactivated, the live-attenuated CDV vaccines retain residual virulence for highly susceptible wildlife species. In this study, we developed recombinant bivalent vaccine candidates based on recombinant vaccine strain rabies virus particles, which concurrently display the protective CDV and RABV glycoprotein antigens. The recombinant viruses replicated to near-wild-type titers, and the heterologous glycoproteins were efficiently expressed and incorporated in the viral particles. Immunization of ferrets with beta-propiolactone-inactivated recombinant virus particles elicited protective RABV antibody titers, and animals immunized with a combination of CDV attachment protein- and fusion protein-expressing recombinant viruses were protected from lethal CDV challenge. However, animals that were immunized with only a RABV expressing the attachment protein of CDV vaccine strain Onderstepoort succumbed to infection with a more recent wild-type strain, indicating that immune responses to the more conserved fusion protein contribute to protection against heterologous CDV strains. IMPORTANCE Rabies virus and canine distemper virus (CDV) cause high mortality rates and death in many carnivores. While rabies vaccines are inactivated and thus have an excellent safety profile and high stability, live-attenuated CDV vaccines can retain residual virulence in highly susceptible species. Here we generated recombinant inactivated rabies viruses that carry one of the CDV glycoproteins on their surface. Ferrets immunized twice with a mix of recombinant rabies viruses carrying the CDV fusion and attachment glycoproteins were protected from lethal CDV challenge, whereas all animals that received recombinant rabies viruses carrying only the CDV attachment protein according to the same immunization scheme died. Irrespective of the CDV antigens used, all animals developed protective titers against rabies virus, illustrating that a bivalent rabies virus-based vaccine against CDV induces protective immune responses against both pathogens. PMID:28148801

Vaccine-associated enhanced respiratory disease is influenced by hemagglutinin and neuraminidase in whole inactivated influenza virus vaccines

USDA-ARS?s Scientific Manuscript database

Multiple subtypes and many antigenic variants of influenza A virus (IAV) co-circulate in swine in the USA, complicating effective use of commercial vaccines to control disease and transmission. Whole inactivated virus (WIV) vaccines may provide partial protection against IAV with substantial antigen...

Influenza A virus hemagglutinin glycosylation compensates for antibody escape fitness costs.

PubMed

Kosik, Ivan; Ince, William L; Gentles, Lauren E; Oler, Andrew J; Kosikova, Martina; Angel, Matthew; Magadán, Javier G; Xie, Hang; Brooke, Christopher B; Yewdell, Jonathan W

2018-01-01

Rapid antigenic evolution enables the persistence of seasonal influenza A and B viruses in human populations despite widespread herd immunity. Understanding viral mechanisms that enable antigenic evolution is critical for designing durable vaccines and therapeutics. Here, we utilize the primerID method of error-correcting viral population sequencing to reveal an unexpected role for hemagglutinin (HA) glycosylation in compensating for fitness defects resulting from escape from anti-HA neutralizing antibodies. Antibody-free propagation following antigenic escape rapidly selected viruses with mutations that modulated receptor binding avidity through the addition of N-linked glycans to the HA globular domain. These findings expand our understanding of the viral mechanisms that maintain fitness during antigenic evolution to include glycan addition, and highlight the immense power of high-definition virus population sequencing to reveal novel viral adaptive mechanisms.

A pathological study of the tongues of rabid dogs in the Philippines.

PubMed

Shiwa, Nozomi; Kimitsuki, Kazunori; Manalo, Daria Llenaresas; Inoue, Satoshi; Park, Chun-Ho

2018-06-01

During rabies virus infections, the minor salivary glands are one of the important organs for virus replication and excretion into the oral cavity. However, details of pathological findings and viral antigen distribution in the minor salivary glands remain poorly understood. In this study, we conducted pathological tests on the tongues of 71 rabid dogs in the Philippines; the minor salivary glands (von Ebner's glands, lingual glands), circumvallate papilla, autonomic ganglia, and skeletal muscles were evaluated. Inflammatory changes were observed in the von Ebner's glands of 20/71 dogs, in the circumvallate papilla of 10/71, and in the tongue muscle of 1/71. Conversely, no morphological changes were observed in the lingual glands and autonomic ganglia. Viral antigens were detected via immunohistochemistry-based methods in the cytoplasm of the acinar epithelium in the von Ebner's glands of all 71 dogs. Virus particles were confirmed in the intercellular canaliculi and acinar lumen via electron microscopy. In the autonomic ganglia, viral antigens were detected in 67/71 rabid dogs. Viral antigens were detected in the taste buds of all 71 dogs, and were distributed mainly in type II and III taste bud cells. In tongue muscle fibers, viral antigens were detected in 11/71 dogs. No virus antigens were detected in lingual glands. These findings suggest that rabies virus descends in the tongue along the glossopharyngeal nerve after proliferation in the brain, and von Ebner's glands and taste buds are one of the portals of virus excretion into the saliva in rabid dogs.

[Molecular analyses of human influenza viruses. Circulation of new variants since 1995/96].

PubMed

Biere, B; Schweiger, B

2008-09-01

The evolution of influenza viruses is increasingly pursued by molecular analyses that complement classical methods. The analyses focus on the two surface proteins hemagglutinin (HA) and neuraminidase (NA) which determine the viral antigenic profile. Influenza A(H3N2) viruses are exceptionally variable, so that usually at least two virus variants cocirculate at the same time. Together with influenza B viruses they caused approximately 90% of influenza virus infections in Germany during the last 12 seasons, while influenza A(H1N1) viruses only played a subordinate part. Unexpectedly, reassorted viruses of subtype A(H1N2) appeared during the seasons 2001/02 and 2002/03, but were isolated only rarely and gained no epidemiological significance. Furthermore, during the season 2001/02 influenza B viruses of the Victoria-lineage reappeared in Germany and other countries of the northern hemisphere after 10 years of absence. These viruses reassorted with the cocirculating Yamagata-like influenza B viruses, as could be seen by the appearance of viruses with a Victoria-like HA and a Yamagata-like NA.

In vitro stimulation of rabbit T lymphocytes by cells expressing herpes simplex antigens.

PubMed

Kapoor, A K; Ling, N R; Nash, A A; Bachan, A; Wildy, P

1982-04-01

Lymphocyte stimulation responses to herpes antigens were studied using virus-infected X-irradiated cells. Rabbits were immunized with herpes simplex virus type 1 (strain HFEM) grown in RK 13 cells. For in vitro stimulation assay BHK21 cells were X-irradiated (15 000 rad) and infected with a high m.o.i. of a temperature-sensitive (ts) mutant (N102) of HFEM strain at the non-permissive temperature (38.5 degrees C) of virus. Virus antigens were expressed on the infected cells and there was no leakage of infectious virus into the medium at 38.5 degrees C. T lymphocytes from rabbits immunized with herpes simplex virus were specifically activated by herpesvirus-infected X-irradiated cells; lymph node cells from rabbits immunized with RK13 cells and from non-immune rabbits showed no proliferative response.

Structural Basis of Preexisting Immunity to the 2009 H1N1 Pandemic Influenza Virus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Rui; Ekiert, Damian C.; Krause, Jens C.

The 2009 H1N1 swine flu is the first influenza pandemic in decades. The crystal structure of the hemagglutinin from the A/California/04/2009 H1N1 virus shows that its antigenic structure, particularly within the Sa antigenic site, is extremely similar to those of human H1N1 viruses circulating early in the 20th century. The cocrystal structure of the 1918 hemagglutinin with 2D1, an antibody from a survivor of the 1918 Spanish flu that neutralizes both 1918 and 2009 H1N1 viruses, reveals an epitope that is conserved in both pandemic viruses. Thus, antigenic similarity between the 2009 and 1918-like viruses provides an explanation for themore » age-related immunity to the current influenza pandemic.« less

Antigenic cartography of H9N2 virus and its impact on the vaccine efficacy in chickens

USDA-ARS?s Scientific Manuscript database

The H9 subtype of avian influenza virus (AIV) is wide-spread in Asia and the Middle East. The efficacy of vaccines is enhanced by the antigenic match of the hemagglutinin protein (HA) between the vaccine and the field strain. To determine how antigenic variations affect the vaccine efficacy, speci...

Hepatitis B virus core antigen: synthesis in Escherichia coli and application in diagnosis.

PubMed Central

Stahl, S; MacKay, P; Magazin, M; Bruce, S A; Murray, K

1982-01-01

Fragments of hepatitis B virus DNA cloned in plasmid pBR322 carrying the gene for the viral core antigen have been placed under the control of the lac promoter of Escherichia coli. Several of the new recombinants direct higher levels of synthesis of the antigen, but the degree of enhancement varies with the different structures of the plasmids and hence the mRNAs produced. The antigen in crude bacterial lysates is a satisfactory diagnostic reagent for antibodies to the core antigen in serum samples. Images PMID:7041126

Initial HIV-1 Antigen-Specific CD8+ T Cells in Acute HIV-1 Infection Inhibit Transmitted/Founder Virus Replication

PubMed Central

Freel, Stephanie A.; Picking, Ralph A.; Ferrari, Guido; Ding, Haitao; Ochsenbauer, Christina; Kappes, John C.; Kirchherr, Jennifer L.; Soderberg, Kelly A.; Weinhold, Kent J.; Cunningham, Coleen K.; Denny, Thomas N.; Crump, John A.; Cohen, Myron S.; McMichael, Andrew J.; Haynes, Barton F.

2012-01-01

CD8-mediated virus inhibition can be detected in HIV-1-positive subjects who naturally control virus replication. Characterizing the inhibitory function of CD8+ T cells during acute HIV-1 infection (AHI) can elucidate the nature of the CD8+ responses that can be rapidly elicited and that contribute to virus control. We examined the timing and HIV-1 antigen specificity of antiviral CD8+ T cells during AHI. Autologous and heterologous CD8+ T cell antiviral functions were assessed longitudinally during AHI in five donors from the CHAVI 001 cohort using a CD8+ T cell-mediated virus inhibition assay (CD8 VIA) and transmitted/founder (T/F) viruses. Potent CD8+ antiviral responses against heterologous T/F viruses appeared during AHI at the first time point sampled in each of the 5 donors (Fiebig stages 1/2 to 5). Inhibition of an autologous T/F virus was durable to 48 weeks; however, inhibition of heterologous responses declined concurrent with the resolution of viremia. HIV-1 viruses from 6 months postinfection were more resistant to CD8+-mediated virus inhibition than cognate T/F viruses, demonstrating that the virus escapes early from CD8+ T cell-mediated inhibition of virus replication. CD8+ T cell antigen-specific subsets mediated inhibition of T/F virus replication via soluble components, and these soluble responses were stimulated by peptide pools that include epitopes that were shown to drive HIV-1 escape during AHI. These data provide insights into the mechanisms of CD8-mediated virus inhibition and suggest that functional analyses will be important for determining whether similar antigen-specific virus inhibition can be induced by T cell-directed vaccine strategies. PMID:22514337

Performance of ELISA antigens prepared from 8 isolates of porcine reproductive and respiratory syndrome virus with homologous and heterologous antisera.

PubMed Central

Cho, H J; Entz, S C; Magar, R; Joo, H S

1997-01-01

Porcine reproductive and respiratory syndrome virus (PRRSV) ELISA antigens of high quality were produced using 8 different isolates of PRRSV: the European Lelystad virus (LV), the U.S. MN-1b, 89-46448, 93-44927, and 93-24025B, and the Canadian LHVA-93-3, PA-8 and GH-6 virus isolates. The performance of each of these 8 antigens and a commercial PRRSV antibody test kit (Idexx's HerdChek) were measured against antisera raised in 5 groups of 6 piglets inoculated with either LV, MN-1b, 89-46448, 93-44927, or 93-24025B. Among the 8 isolates, the 89-46448 isolate produced the broadest spectrum of antigen and resulted in earlier detection of antibodies to various North American PRRSV isolates, followed by MN-1b as the 2nd best ELISA antigen for the detection of North American PRRSV antibodies. The GH-6 and PA-8 viral antigens exhibited restricted detection of PRRSV antibodies. The LV and 89-46448 combined antigens produced the best performance for the detection of antibodies against both European and North American antigenic types of PRRSV. Using 173 panel samples collected at 11 to 60 d after intranasal inoculation with 1 of the 5 PRRSV isolates, the sensitivities of the indirect ELISA used were 73.4%, 98.3%, 90.8%, 98.3%, 83.2%, 93.1%, 77.1%, 64.2%, 98.8% and 95.9% for LV, MN-1b, LHVA-93-3, 89-46448, 93-44927, 93-24025B, PA-8, GH-6 antigens, 89-46448-LV combined antigens and Idexx's PRRSV antibody test kit, respectively. All 8 antigens gave negative results with preinfection porcine sera (n = 30); high background or nonspecific reactions were not observed with the antigens. PMID:9342455

Development and Evaluation of Single Domain Antibodies for Vaccinia and the L1 Antigen

PubMed Central

Walper, Scott A.; Liu, Jinny L.; Zabetakis, Daniel; Anderson, George P.; Goldman, Ellen R.

2014-01-01

There is ongoing interest to develop high affinity, thermal stable recognition elements to replace conventional antibodies in biothreat detection assays. As part of this effort, single domain antibodies that target vaccinia virus were developed. Two llamas were immunized with killed viral particles followed by boosts with the recombinant membrane protein, L1, to stimulate the immune response for envelope and membrane proteins of the virus. The variable domains of the induced heavy chain antibodies were selected from M13 phage display libraries developed from isolated RNA. Selection via biopanning on the L1 antigen produced single domain antibodies that were specific and had affinities ranging from 4×10−9 M to 7.0×10−10 M, as determined by surface plasmon resonance. Several showed good ability to refold after heat denaturation. These L1-binding single domain antibodies, however, failed to recognize the killed vaccinia antigen. Useful vaccinia binding single domain antibodies were isolated by a second selection using the killed virus as the target. The virus binding single domain antibodies were incorporated in sandwich assays as both capture and tracer using the MAGPIX system yielding limits of detection down to 4×105 pfu/ml, a four-fold improvement over the limit obtained using conventional antibodies. This work demonstrates the development of anti-vaccinia single domain antibodies and their incorporation into sandwich assays for viral detection. It also highlights the properties of high affinity and thermal stability that are hallmarks of single domain antibodies. PMID:25211488

Conserved epitope on influenza-virus hemagglutinin head defined by a vaccine-induced antibody

DOE Office of Scientific and Technical Information (OSTI.GOV)

Raymond, Donald D.; Bajic, Goran; Ferdman, Jack

Antigenic variation requires frequent revision of annual influenza vaccines. Next-generation vaccine design strategies aim to elicit a broader immunity by directing the human immune response toward conserved sites on the principal viral surface protein, the hemagglutinin (HA). We describe a group of antibodies that recognize a hitherto unappreciated, conserved site on the HA of H1 subtype influenza viruses. Mutations in that site, which required a change in the H1 component of the 2017 vaccine, had not previously “taken over” among circulating H1 viruses. Our results encourage vaccine design strategies that resurface a protein to focus the immune response on amore » specific region.« less

Modular MLV-VLPs co-displaying ovalbumin peptides and GM-CSF effectively induce expansion of CD11b+ APC and antigen-specific T cell responses in vitro.

PubMed

Gogesch, Patricia; Schülke, Stefan; Scheurer, Stephan; Mühlebach, Michael D; Waibler, Zoe

2018-05-28

The development of novel vaccination strategies is a persistent challenge to provide effective prophylactic treatments to encounter viral infections. In general, the physical conjugation of selected vaccine components, e.g. antigen and adjuvant, has been shown to enhance the immunogenicity and hence, can increase effectiveness of the vaccine. In our proof-of-concept study, we generated non-infectious, replication deficient Murine Leukemia Virus (MLV)-derived virus-like particles (VLPs) that physically link antigen and adjuvant in a modular fashion by co-displaying them on their surface. For this purpose, we selected the immunodominant peptides of the model antigen ovalbumin (OVA) and the cytokine granulocyte macrophage-colony stimulating factor (GM-CSF) as non-classical adjuvant. Our results show that murine GM-CSF displayed on MLV-VLPs mediates expansion and proliferation of CD11b + cells within murine bone marrow and total spleen cells. Moreover, we show increased immunogenicity of modular VLPs co-displaying OVA peptides and GM-CSF by their elevated capacity to induce OVA-specific T cell-activation and -proliferation within OT-I and OT-II splenocyte cultures. These enhanced effects were not achieved by using an equimolar mixture of VLPs displaying either OVA or GM-CSF. Taken together, OVA and GM-CSF co-displaying MLV-VLPs are able to target and expand antigen presenting cells which in turn results in enhanced antigen-specific T cell activation and proliferation in vitro. These data suggest MLV-VLPs to be an attractive platform to flexibly combine antigen and adjuvant for novel modular vaccination approaches. Copyright © 2018 Elsevier Ltd. All rights reserved.

Sensitive typing of reverse ABO blood groups with a waveguide-mode sensor.

PubMed

Uno, Shigeyuki; Tanaka, Torahiko; Ashiba, Hiroki; Fujimaki, Makoto; Tanaka, Mutsuo; Hatta, Yoshihiro; Takei, Masami; Awazu, Koichi; Makishima, Makoto

2018-07-01

Portable, on-site blood typing methods will help provide life-saving blood transfusions to patients during an emergency or natural calamity, such as significant earthquakes. We have previously developed waveguide-mode (WM) sensors for forward ABO and Rh(D) blood typing and detection of antibodies against hepatitis B virus and hepatitis C virus. In this study, we evaluated a WM-sensor for reverse ABO blood typing. Since reverse ABO blood typing is a method for detection of antibodies against type A and type B oligosaccharide antigens on the surface of red blood cells (RBCs), we fixed a synthetic type A or type B trisaccharide antigen on the sensor chip of the WM sensor. We obtained significant changes in the reflectance spectra from a WM sensor on type A antigen with type B plasma and type O plasma and on type B antigen with type A plasma and type O plasma, and no spectrum changes on type A antigen or type B antigen with type AB plasma. Signal enhancement with the addition of a peroxidase reaction failed to increase the sensitivity for detection on oligosaccharide chips. By utilizing hemagglutination detection using regent type A and type B RBCs, we successfully determined reverse ABO blood groups with higher sensitivity compared to a method using oligosaccharide antigens. Thus, functionality of a portable device utilizing a WM sensor can be expanded to include reverse ABO blood typing and, in combination with forward ABO typing and antivirus antibody detection, may be useful for on-site blood testing in emergency settings. Copyright © 2018 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Novel 6xHis tagged foot-and-mouth disease virus vaccine bound to nanolipoprotein adjuvant via metal ions provides antigenic distinction and effective protective immunity

USDA-ARS?s Scientific Manuscript database

Here, we engineered two FMD viruses and histidine residues inserted into or fused to the FMDV capsid. Both 6xHis viruses exhibited growth kinetics, plaque morphologies and antigenic characteristics similar to wild-type virus. The 6xHis tag allowed one-step purification of the mutant virions by Co2...

Trocara virus: a newly recognized Alphavirus (Togaviridae) isolated from mosquitoes in the Amazon Basin.

PubMed

Travassos da Rosa, A P; Turell, M J; Watts, D M; Powers, A M; Vasconcelos, P F; Jones, J W; Klein, T A; Dohm, D J; Shope, R E; Degallier, N; Popov, V L; Russell, K L; Weaver, S C; Guzman, H; Calampa, C; Brault, A C; Lemon, A P; Tesh, R B

2001-01-01

This report describes Trocara virus, a newly recognized member of the genus Alphavirus, that has been isolated from Aedes serratus mosquitoes collected at two widely separated sites in the Amazon Basin. Biological, antigenic and genetic characteristics of the new virus are given. Results of these studies indicate that Trocara virus is the first member of a newly discovered antigenic complex within the family Togaviridae genus Alphavirus. The public health and veterinary importance of Trocara virus is still unknown.

Antigenic characterization of H3 subtypes of avian influenza A viruses from North America

USGS Publications Warehouse

Bailey, Elizabeth; Long, Li-Pong; Zhao, Nan; Hall, Jeffrey S.; Baroch, John A; Nolting, Jaqueline; Senter, Lucy; Cunningham, Frederick L; Pharr, G Todd; Hanson, Larry; Slemons, Richard; DeLiberto, Thomas J.; Wan, Xiu-Feng

2016-01-01

Besides humans, H3 subtypes of influenza A viruses (IAVs) can infect various animal hosts, including avian, swine, equine, canine, and sea mammal species. These H3 viruses are both antigenically and genetically diverse. Here, we characterized the antigenic diversity of contemporary H3 avian IAVs recovered from migratory birds in North America. Hemagglutination inhibition (HI) assays were performed on 37 H3 isolates of avian IAVs recovered from 2007 to 2011 using generated reference chicken sera. These isolates were recovered from samples taken in the Atlantic, Mississippi, Central, and Pacific waterfowl migration flyways. Antisera to all the tested H3 isolates cross-reacted with each other and, to a lesser extent, with those to H3 canine and H3 equine IAVs. Antigenic cartography showed that the largest antigenic distance among the 37 avian IAVs is about four units, and each unit corresponds to a 2 log 2 difference in the HI titer. However, none of the tested H3 IAVs cross-reacted with ferret sera derived from contemporary swine and human IAVs. Our results showed that the H3 avian IAVs we tested lacked significant antigenic diversity, and these viruses were antigenically different from those circulating in swine and human populations. This suggests that H3 avian IAVs in North American waterfowl are antigenically relatively stable.

Antigenic mapping of an H9N2 avian influenza virus reveals two discrete antigenic sites and a novel mechanism of immune escape.

PubMed

Peacock, Thomas; Reddy, Kolli; James, Joe; Adamiak, Beata; Barclay, Wendy; Shelton, Holly; Iqbal, Munir

2016-01-07

H9N2 avian influenza virus is a major cause of poultry production loss across Asia leading to the wide use of vaccines. Efficacy of vaccines is often compromised due to the rapid emergence of antigenic variants. To improve the effectiveness of vaccines in the field, a better understanding of the antigenic epitopes of the major antigen, hemagglutinin, is required. To address this, a panel of nine monoclonal antibodies were generated against a contemporary Pakistani H9N2 isolate, which represents a major Asian H9N2 viral lineage. Antibodies were characterized in detail and used to select a total of 26 unique 'escape' mutants with substitutions across nine different amino acid residues in hemagglutinin including seven that have not been described as antigenic determinants for H9N2 viruses before. Competition assays and structural mapping revealed two novel, discrete antigenic sites "H9-A" and "H9-B". Additionally, a second subset of escape mutants contained amino acid deletions within the hemagglutinin receptor binding site. This constitutes a novel method of escape for group 1 hemagglutinins and could represent an alternative means for H9N2 viruses to overcome vaccine induced immunity. These results will guide surveillance efforts for arising antigenic variants as well as evidence based vaccine seed selection and vaccine design.

Antigenic mapping of an H9N2 avian influenza virus reveals two discrete antigenic sites and a novel mechanism of immune escape

PubMed Central

Peacock, Thomas; Reddy, Kolli; James, Joe; Adamiak, Beata; Barclay, Wendy; Shelton, Holly; Iqbal, Munir

2016-01-01

H9N2 avian influenza virus is a major cause of poultry production loss across Asia leading to the wide use of vaccines. Efficacy of vaccines is often compromised due to the rapid emergence of antigenic variants. To improve the effectiveness of vaccines in the field, a better understanding of the antigenic epitopes of the major antigen, hemagglutinin, is required. To address this, a panel of nine monoclonal antibodies were generated against a contemporary Pakistani H9N2 isolate, which represents a major Asian H9N2 viral lineage. Antibodies were characterized in detail and used to select a total of 26 unique ‘escape’ mutants with substitutions across nine different amino acid residues in hemagglutinin including seven that have not been described as antigenic determinants for H9N2 viruses before. Competition assays and structural mapping revealed two novel, discrete antigenic sites “H9-A” and “H9-B”. Additionally, a second subset of escape mutants contained amino acid deletions within the hemagglutinin receptor binding site. This constitutes a novel method of escape for group 1 hemagglutinins and could represent an alternative means for H9N2 viruses to overcome vaccine induced immunity. These results will guide surveillance efforts for arising antigenic variants as well as evidence based vaccine seed selection and vaccine design. PMID:26738561

Antigen-capture blocking enzyme-linked immunosorbent assay based on a baculovirus recombinant antigen to differentiate Transmissible gastroenteritis virus from Porcine respiratory coronavirus antibodies.

PubMed

López, Lissett; Venteo, Angel; García, Marga; Camuñas, Ana; Ranz, Ana; García, Julia; Sarraseca, Javier; Anaya, Carmen; Rueda, Paloma

2009-09-01

A new commercially available antigen-capture, blocking enzyme-linked immunosorbent assay (antigen-capture b-ELISA), based on baculovirus truncated-S recombinant protein of Transmissible gastroenteritis virus (TGEV) and 3 specific monoclonal antibodies, was developed and evaluated by examining a panel of 453 positive Porcine respiratory coronavirus (PRCoV), 31 positive TGEV, and 126 negative field sera by using another commercially available differential coronavirus b-ELISA as the reference technique to differentiate TGEV- from PRCoV-induced antibodies. The recombinant S protein-based ELISA appeared to be 100% sensitive for TGEV and PRCoV detection and highly specific for TGEV and PRCoV detection (100% and 92.06%, respectively), when qualitative results (positive or negative) were compared with those of the reference technique. In variability experiments, the ELISA gave consistent results when the same serum was evaluated on different wells and different plates. These results indicated that truncated recombinant S protein is a suitable alternative to the complete virus as antigen in ELISA assays. The use of recombinant S protein as antigen offers great advantages because it is an easy-to-produce, easy-to-standardize, noninfectious antigen that does not require further purification or concentration. Those advantages represent an important improvement for antigen preparation, in comparison with other assays in which an inactivated virus from mammalian cell cultures is used.

Antigenic Characterization of H3 Subtypes of Avian Influenza A Viruses from North America.

PubMed

Bailey, Elizabeth; Long, Li-Ping; Zhao, Nan; Hall, Jeffrey S; Baroch, John A; Nolting, Jacqueline; Senter, Lucy; Cunningham, Frederick L; Pharr, G Todd; Hanson, Larry; Slemons, Richard; DeLiberto, Thomas J; Wan, Xiu-Feng

2016-05-01

Besides humans, H3 subtypes of influenza A viruses (IAVs) can infect various animal hosts, including avian, swine, equine, canine, and sea mammal species. These H3 viruses are both antigenically and genetically diverse. Here, we characterized the antigenic diversity of contemporary H3 avian IAVs recovered from migratory birds in North America. Hemagglutination inhibition (HI) assays were performed on 37 H3 isolates of avian IAVs recovered from 2007 to 2011 using generated reference chicken sera. These isolates were recovered from samples taken in the Atlantic, Mississippi, Central, and Pacific waterfowl migration flyways. Antisera to all the tested H3 isolates cross-reacted with each other and, to a lesser extent, with those to H3 canine and H3 equine IAVs. Antigenic cartography showed that the largest antigenic distance among the 37 avian IAVs is about four units, and each unit corresponds to a 2 log 2 difference in the HI titer. However, none of the tested H3 IAVs cross-reacted with ferret sera derived from contemporary swine and human IAVs. Our results showed that the H3 avian IAVs we tested lacked significant antigenic diversity, and these viruses were antigenically different from those circulating in swine and human populations. This suggests that H3 avian IAVs in North American waterfowl are antigenically relatively stable.

Strategy for assessment of the colloidal and biological stability of H1N1 influenza A viruses.

PubMed

Hämmerling, Frank; Lorenz-Cristea, Oliver; Baumann, Pascal; Hubbuch, Jürgen

2017-01-30

Current influenza vaccines are mostly formulated as liquids which requires a continuous cold chain to maintain the stability of the antigen. For development of vaccines with an increased stability at ambient temperatures, manifold parameters and their influences on the colloidal stability and activity of the antigen have to be understood. This work presents a strategy to examine both, the colloidal stability and the remaining biological activity of H1N1 influenza viruses under various conditions after an incubation of 40 days. H1N1 phase diagrams were generated for several pH values and different initial H1N1 and NaCl concentrations. It was shown that the highest H1N1 recoveries were obtained for pH 6 and that moderate amounts of NaCl are favorable for increased recoveries. In contrast to colloidal stability, the highest remaining HA activity was observed at pH 9. The electrostatic and hydrophobic surface properties of H1N1 were investigated to reveal the mechanisms accounting for the decrease in stability. Secondly, the capability of virus precipitation by polyethylene glycol in combination with determination of surface hydrophobicity was proven to be useful as a predictive tool to rank stability under different conditions. This methodology enables the rapid assessment of aggregation propensity of H1N1 formulations and the influence on the activity of the virus particles and might become a standard tool during the development of vaccine formulations. Copyright © 2016 Elsevier B.V. All rights reserved.

Characterization of virus-like particles by atomic force microscopy in ambient conditions

NASA Astrophysics Data System (ADS)

Oropesa, Reinier; Ramos, Jorge R.; Falcón, Viviana; Felipe, Ariel

2013-06-01

Recombinant virus-like particles (VLPs) are attractive candidates for vaccine design since they resemble native viroids in size and morphology, but they are non-infectious due to the absence of a viral genome. The visualization of surface morphologies and structures can be used to deepen the understanding of physical, chemical, and biological phenomena. Atomic force microscopy (AFM) is a useful tool for the visualization of soft biological samples in a nanoscale resolution. In this work we have investigated the morphology of recombinant surface antigens of hepatitis B (rHBsAg) VLPs from Cuban vaccine against hepatitis B. The rHBsAg VLPs sizes estimated by AFM between 15 and 30 nm are similar to those reported on previous transmission electron microscopy (TEM) studies.

Crystal structure, conformational fixation and entry-related interactions of mature ligand-free HIV-1 Env

DOE Office of Scientific and Technical Information (OSTI.GOV)

Do Kwon, Young; Pancera, Marie; Acharya, Priyamvada

As the sole viral antigen on the HIV-1–virion surface, trimeric Env is a focus of vaccine efforts. In this paper, we present the structure of the ligand-free HIV-1–Env trimer, fix its conformation and determine its receptor interactions. Epitope analyses revealed trimeric ligand-free Env to be structurally compatible with broadly neutralizing antibodies but not poorly neutralizing ones. We coupled these compatibility considerations with binding antigenicity to engineer conformationally fixed Envs, including a 201C 433C (DS) variant specifically recognized by broadly neutralizing antibodies. DS-Env retained nanomolar affinity for the CD4 receptor, with which it formed an asymmetric intermediate: a closed trimer boundmore » by a single CD4 without the typical antigenic hallmarks of CD4 induction. Finally, antigenicity-guided structural design can thus be used both to delineate mechanism and to fix conformation, with DS-Env trimers in virus-like-particle and soluble formats providing a new generation of vaccine antigens.« less

Crystal structure, conformational fixation and entry-related interactions of mature ligand-free HIV-1 Env

DOE PAGES

Do Kwon, Young; Pancera, Marie; Acharya, Priyamvada; ...

2015-06-22

As the sole viral antigen on the HIV-1–virion surface, trimeric Env is a focus of vaccine efforts. In this paper, we present the structure of the ligand-free HIV-1–Env trimer, fix its conformation and determine its receptor interactions. Epitope analyses revealed trimeric ligand-free Env to be structurally compatible with broadly neutralizing antibodies but not poorly neutralizing ones. We coupled these compatibility considerations with binding antigenicity to engineer conformationally fixed Envs, including a 201C 433C (DS) variant specifically recognized by broadly neutralizing antibodies. DS-Env retained nanomolar affinity for the CD4 receptor, with which it formed an asymmetric intermediate: a closed trimer boundmore » by a single CD4 without the typical antigenic hallmarks of CD4 induction. Finally, antigenicity-guided structural design can thus be used both to delineate mechanism and to fix conformation, with DS-Env trimers in virus-like-particle and soluble formats providing a new generation of vaccine antigens.« less

Inhibition of CD1 antigen presentation by human cytomegalovirus.

PubMed

Raftery, Martin J; Hitzler, Manuel; Winau, Florian; Giese, Thomas; Plachter, Bodo; Kaufmann, Stefan H E; Schönrich, Günther

2008-05-01

The betaherpesvirus human cytomegalovirus (HCMV) encodes several molecules that block antigen presentation by the major histocompatibility complex (MHC) proteins. Humans also possess one other family of antigen-presenting molecules, the CD1 family; however, the effect of HCMV on CD1 expression is unknown. The majority of CD1 molecules are classified on the basis of homology as group 1 CD1 and are present almost exclusively on professional antigen-presenting cells such as dendritic cells, which are a major target for HCMV infection and latency. We have determined that HCMV encodes multiple blocking strategies targeting group 1 CD1 molecules. CD1 transcription is strongly inhibited by the HCMV interleukin-10 homologue cmvIL-10. HCMV also blocks CD1 antigen presentation posttranscriptionally by the inhibition of CD1 localization to the cell surface. This function is not performed by a known HCMV MHC class I-blocking molecule and is substantially stronger than the blockage induced by herpes simplex virus type 1. Antigen presentation by CD1 is important for the development of the antiviral immune response and the generation of mature antigen-presenting cells. HCMV present in antigen-presenting cells thus blunts the immune response by the blockage of CD1 molecules.

Dengue virus type 2: replication and tropisms in orally infected Aedes aegypti mosquitoes.

PubMed

Salazar, Ma Isabel; Richardson, Jason H; Sánchez-Vargas, Irma; Olson, Ken E; Beaty, Barry J

2007-01-30

To be transmitted by its mosquito vector, dengue virus (DENV) must infect midgut epithelial cells, replicate and disseminate into the hemocoel, and finally infect the salivary glands, which is essential for transmission. The extrinsic incubation period (EIP) is very relevant epidemiologically and is the time required from the ingestion of virus until it can be transmitted to the next vertebrate host. The EIP is conditioned by the kinetics and tropisms of virus replication in its vector. Here we document the virogenesis of DENV-2 in newly-colonized Aedes aegypti mosquitoes from Chetumal, Mexico in order to understand better the effect of vector-virus interactions on dengue transmission. After ingestion of DENV-2, midgut infections in Chetumal mosquitoes were characterized by a peak in virus titers between 7 and 10 days post-infection (dpi). The amount of viral antigen and viral titers in the midgut then declined, but viral RNA levels remained stable. The presence of DENV-2 antigen in the trachea was positively correlated with virus dissemination from the midgut. DENV-2 antigen was found in salivary gland tissue in more than a third of mosquitoes at 4 dpi. Unlike in the midgut, the amount of viral antigen (as well as the percent of infected salivary glands) increased with time. DENV-2 antigen also accumulated and increased in neural tissue throughout the EIP. DENV-2 antigen was detected in multiple tissues of the vector, but unlike some other arboviruses, was not detected in muscle. Our results suggest that the EIP of DENV-2 in its vector may be shorter that the previously reported and that the tracheal system may facilitate DENV-2 dissemination from the midgut. Mosquito organs (e.g. midgut, neural tissue, and salivary glands) differed in their response to DENV-2 infection.

Nuclear location of an endogenously expressed antigen, EBNA1, restricts access to macroautophagy and the range of CD4 epitope display.

PubMed

Leung, Carol S; Haigh, Tracey A; Mackay, Laura K; Rickinson, Alan B; Taylor, Graham S

2010-02-02

Whereas exogenously acquired proteins are the major source of antigens feeding the MHC class II pathway in antigen-presenting cells, some endogenously expressed antigens also access that pathway but the rules governing such access are poorly understood. Here we address this using Epstein-Barr virus (EBV)-coded nuclear antigen EBNA1, a protein naturally expressed in EBV-infected B lymphoblastoid cell lines (LCLs) and a source of multiple CD4(+) T cell epitopes. Using CD4(+) T cell clones against three indicator epitopes, we find that two epitopes are weakly displayed on the LCL surface whereas the third is undetectable, a pattern of limited epitope presentation that is maintained even when nuclear expression of EBNA1 is induced to high supraphysiological levels. Inhibitor and siRNA studies show that, of the two epitopes weakly presented under these conditions, one involves macroautophagy, and the second involves antigen delivery to the MHC II pathway by another endogenous route. In contrast, when EBNA1 is expressed as a cytoplasmic protein, all three CD4 epitopes are processed and presented much more efficiently, and all involve macroautophagy. We conclude that EBNA1's nuclear location limits its accessibility to the macroautophagy pathway and, in consequence, limits the level and range of EBNA1 CD4 epitopes naturally displayed on the infected cell surface.

A rapid and quantitative assay for measuring antibody-mediated neutralization of West Nile virus infection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pierson, Theodore C.; Sanchez, Melissa D.; Puffer, Bridget A.

2006-03-01

West Nile virus (WNV) is a neurotropic flavivirus within the Japanese encephalitis antigenic complex that is responsible for causing West Nile encephalitis in humans. The surface of WNV virions is covered by a highly ordered icosahedral array of envelope proteins that is responsible for mediating attachment and fusion with target cells. These envelope proteins are also primary targets for the generation of neutralizing antibodies in vivo. In this study, we describe a novel approach for measuring antibody-mediated neutralization of WNV infection using virus-like particles that measure infection as a function of reporter gene expression. These reporter virus particles (RVPs) aremore » produced by complementation of a sub-genomic replicon with WNV structural proteins provided in trans using conventional DNA expression vectors. The precision and accuracy of this approach stem from an ability to measure the outcome of the interaction between antibody and viral antigens under conditions that satisfy the assumptions of the law of mass action as applied to virus neutralization. In addition to its quantitative strengths, this approach allows the production of WNV RVPs bearing the prM-E proteins of different WNV strains and mutants, offering considerable flexibility for the study of the humoral immune response to WNV in vitro. WNV RVPs are capable of only a single round of infection, can be used under BSL-2 conditions, and offer a rapid and quantitative approach for detecting virus entry and its inhibition by neutralizing antibody.« less

Bicistronic DNA vaccines simultaneously encoding HIV, HSV and HPV antigens promote CD8⁺ T cell responses and protective immunity.

PubMed

Santana, Vinicius C; Diniz, Mariana O; Cariri, Francisco A M O; Ventura, Armando M; Cunha-Neto, Edécio; Almeida, Rafael R; Campos, Marco A; Lima, Graciela K; Ferreira, Luís C S

2013-01-01

Millions of people worldwide are currently infected with human papillomavirus (HPV), herpes simplex virus (HSV) or human immunodeficiency virus (HIV). For this enormous contingent of people, the search for preventive and therapeutic immunological approaches represents a hope for the eradication of latent infection and/or virus-associated cancer. To date, attempts to develop vaccines against these viruses have been mainly based on a monovalent concept, in which one or more antigens of a virus are incorporated into a vaccine formulation. In the present report, we designed and tested an immunization strategy based on DNA vaccines that simultaneously encode antigens for HIV, HSV and HPV. With this purpose in mind, we tested two bicistronic DNA vaccines (pIRES I and pIRES II) that encode the HPV-16 oncoprotein E7 and the HIV protein p24 both genetically fused to the HSV-1 gD envelope protein. Mice i.m. immunized with the DNA vaccines mounted antigen-specific CD8⁺ T cell responses, including in vivo cytotoxic responses, against the three antigens. Under experimental conditions, the vaccines conferred protective immunity against challenges with a vaccinia virus expressing the HIV-derived protein Gag, an HSV-1 virus strain and implantation of tumor cells expressing the HPV-16 oncoproteins. Altogether, our results show that the concept of a trivalent HIV, HSV, and HPV vaccine capable to induce CD8⁺ T cell-dependent responses is feasible and may aid in the development of preventive and/or therapeutic approaches for the control of diseases associated with these viruses.

Protection of chickens to antigenically variant avian influenza virus challenge after immunization with two antigenically unrelated strains of the same subtype

USDA-ARS?s Scientific Manuscript database

The antigenic diversity of avian influenza virus (AIV) within a subtype has been well established and is believed to be driven by the selection of immunologic escape mutants. In regions where vaccination against AIV has been implemented for prolonged periods (e.g. Vietnam and Egypt), vaccines which...

Dengue Type-2 Virus Envelope Protein Made Using Recombinant Baculovirus Protects Mice Against Virus Challenge

DTIC Science & Technology

1994-01-01

Spodoptera frugiperda (Sf9) cells, approximately I mg of recombinant E antigen was made per 10’ cells. This antigen reacted with polyclonal, anti...entry by fusion at acidic pH with host cell mem- in Spodoptera frugiperda (Sf9) cells brane.Ř The E antigen contains both T and B cell epitopes that

Flavivirus-induced antibody cross-reactivity

PubMed Central

Mansfield, Karen L.; Horton, Daniel L.; Johnson, Nicholas; Li, Li; Barrett, Alan D. T.; Smith, Derek J.; Galbraith, Sareen E.; Solomon, Tom

2011-01-01

Dengue viruses (DENV) cause countless human deaths each year, whilst West Nile virus (WNV) has re-emerged as an important human pathogen. There are currently no WNV or DENV vaccines licensed for human use, yet vaccines exist against other flaviviruses. To investigate flavivirus cross-reactivity, sera from a human cohort with a history of vaccination against tick-borne encephalitis virus (TBEV), Japanese encephalitis virus (JEV) and yellow fever virus (YFV) were tested for antibodies by plaque reduction neutralization test. Neutralization of louping ill virus (LIV) occurred, but no significant neutralization of Murray Valley encephalitis virus was observed. Sera from some individuals vaccinated against TBEV and JEV neutralized WNV, which was enhanced by YFV vaccination in some recipients. Similarly, some individuals neutralized DENV-2, but this was not significantly influenced by YFV vaccination. Antigenic cartography techniques were used to generate a geometric illustration of the neutralization titres of selected sera against WNV, TBEV, JEV, LIV, YFV and DENV-2. This demonstrated the individual variation in antibody responses. Most sera had detectable titres against LIV and some had titres against WNV and DENV-2. Generally, LIV titres were similar to titres against TBEV, confirming the close antigenic relationship between TBEV and LIV. JEV was also antigenically closer to TBEV than WNV, using these sera. The use of sera from individuals vaccinated against multiple pathogens is unique relative to previous applications of antigenic cartography techniques. It is evident from these data that notable differences exist between amino acid sequence identity and mapped antigenic relationships within the family Flaviviridae. PMID:21900425

Flavivirus-induced antibody cross-reactivity.

PubMed

Mansfield, Karen L; Horton, Daniel L; Johnson, Nicholas; Li, Li; Barrett, Alan D T; Smith, Derek J; Galbraith, Sareen E; Solomon, Tom; Fooks, Anthony R

2011-12-01

Dengue viruses (DENV) cause countless human deaths each year, whilst West Nile virus (WNV) has re-emerged as an important human pathogen. There are currently no WNV or DENV vaccines licensed for human use, yet vaccines exist against other flaviviruses. To investigate flavivirus cross-reactivity, sera from a human cohort with a history of vaccination against tick-borne encephalitis virus (TBEV), Japanese encephalitis virus (JEV) and yellow fever virus (YFV) were tested for antibodies by plaque reduction neutralization test. Neutralization of louping ill virus (LIV) occurred, but no significant neutralization of Murray Valley encephalitis virus was observed. Sera from some individuals vaccinated against TBEV and JEV neutralized WNV, which was enhanced by YFV vaccination in some recipients. Similarly, some individuals neutralized DENV-2, but this was not significantly influenced by YFV vaccination. Antigenic cartography techniques were used to generate a geometric illustration of the neutralization titres of selected sera against WNV, TBEV, JEV, LIV, YFV and DENV-2. This demonstrated the individual variation in antibody responses. Most sera had detectable titres against LIV and some had titres against WNV and DENV-2. Generally, LIV titres were similar to titres against TBEV, confirming the close antigenic relationship between TBEV and LIV. JEV was also antigenically closer to TBEV than WNV, using these sera. The use of sera from individuals vaccinated against multiple pathogens is unique relative to previous applications of antigenic cartography techniques. It is evident from these data that notable differences exist between amino acid sequence identity and mapped antigenic relationships within the family Flaviviridae.

Analysis of antigen conservation and inactivation of gamma-irradiated avian influenza virus subtype H9N2.

PubMed

Salehi, Bahareh; Motamedi-Sedeh, Farahnaz; Madadgar, Omid; Khalili, Iraj; Ghalyan Chi Langroudi, Arash; Unger, Hermann; Wijewardana, Viskam

2018-06-01

Avian influenza (AI) A subtype H9N2 virus belongs to Orthomyxoviridae family and causes low-pathogenic disease AI. The use of gamma-irradiated viral antigens has been developed in the production of effective vaccines. In this research, LPAIV H9N2 strain, A/Chicken/IRN/Ghazvin/2001, was multiplied on SPF eggs and irradiated by a Nordian gamma cell instrument. Irradiated and non-irradiated AI virus (AIV) samples were titrated by EID50 method and hemagglutinin (HA) antigen was analyzed by HA test as the WHO pattern method. Infectivity of irradiated virus was determined by egg inoculation method during four blind cultures. The results showed that after increasing the dose of gamma radiation, virus titer gradually decreased. D 10 value and optimum dose for complete virus inactivation were calculated by dose/response curve, 3.36 and 29.52 kGy, respectively. In addition, HA antigenicity of gamma-irradiated virus samples from 0 to 30 kGy was not changed. The results of safety test for gamma-irradiated AIV samples showed complete inactivation with gamma ray doses 30 and 35 kGy, without any multiplication on eggs after four blind cultures. According to the results of HA antigen assay and safety test, the gamma-irradiated and complete inactivated AIV subtype H9N2 is a good candidate as an inactivated immunogenic agent for poultry vaccination.

Virus-like particle expression and assembly in plants: hepatitis B and Norwalk viruses.

PubMed

Huang, Zhong; Elkin, Galina; Maloney, Bryan J; Beuhner, Norene; Arntzen, Charles J; Thanavala, Yasmin; Mason, Hugh S

2005-03-07

Expression of vaccine antigens in plants and delivery via ingestion of transgenic plant material has shown promise in numerous pre-clinical animal studies and in a few clinical trials. A number of different viral antigens have been tested, and among the most promising are those that can assemble virus-like particles (VLP), which mimic the form of authentic virions and display neutralizing antibody epitopes. We have extensively studied plant expression, VLP assembly, and immunogenicity of hepatitis B surface antigen (HBsAg) and Norwalk virus capsid protein (NVCP). The HBsAg small protein (S protein) was found by TEM to assemble tubular membrane complexes derived from endoplasmic reticulum in suspension cultured cells of tobacco and soybean, and in potato leaf and tuber tissues. The potato material was immunogenic in mice upon delivery by ingestion. Here we describe the plant expression and immunogenicity of HBsAg middle protein (M protein or pre-S2 + S) which contains additional 55 amino acid pre-S2 region at N-terminus of the S protein. Plant-derived recombinant M protein provoked stronger serum antibody responses against HBsAg than did S protein when injected systemically in mice. We discuss implications for use of fusion proteins for enhanced immunogenicity and mucosal targeting of HBsAg, as well as delivery of heterologous fused antigens. NVCP expressed in plants assembled 38 nm virion-size icosahedral (T = 3) VLP, similar to those produced in insect cells. The VLP stimulated serum IgG and IgA responses in mice and humans when they were delivered by ingestion of fresh potato tuber. Here we show that freeze-drying of transgenic NVCP tomato fruit yielded stable preparations that stimulated excellent IgG and IgA responses against NVCP when fed to mice. However, the predominant VLP form in tomato fruit was the small 23 nm particle also observed in insect cell-derived NVCP.

Identification of Amino Acid Substitutions Supporting Antigenic Change of Influenza A(H1N1)pdm09 Viruses

PubMed Central

Koel, Björn F.; Mögling, Ramona; Chutinimitkul, Salin; Fraaij, Pieter L.; Burke, David F.; van der Vliet, Stefan; de Wit, Emmie; Bestebroer, Theo M.; Rimmelzwaan, Guus F.; Osterhaus, Albert D. M. E.; Smith, Derek J.; Fouchier, Ron A. M.

2015-01-01

ABSTRACT The majority of currently circulating influenza A(H1N1) viruses are antigenically similar to the virus that caused the 2009 influenza pandemic. However, antigenic variants are expected to emerge as population immunity increases. Amino acid substitutions in the hemagglutinin protein can result in escape from neutralizing antibodies, affect viral fitness, and change receptor preference. In this study, we constructed mutants with substitutions in the hemagglutinin of A/Netherlands/602/09 in an attenuated backbone to explore amino acid changes that may contribute to emergence of antigenic variants in the human population. Our analysis revealed that single substitutions affecting the loop that consists of amino acid positions 151 to 159 located adjacent to the receptor binding site caused escape from ferret and human antibodies elicited after primary A(H1N1)pdm09 virus infection. The majority of these substitutions resulted in similar or increased replication efficiency in vitro compared to that of the virus carrying the wild-type hemagglutinin and did not result in a change of receptor preference. However, none of the substitutions was sufficient for escape from the antibodies in sera from individuals that experienced both seasonal and pandemic A(H1N1) virus infections. These results suggest that antibodies directed against epitopes on seasonal A(H1N1) viruses contribute to neutralization of A(H1N1)pdm09 antigenic variants, thereby limiting the number of possible substitutions that could lead to escape from population immunity. IMPORTANCE Influenza A viruses can cause significant morbidity and mortality in humans. Amino acid substitutions in the hemagglutinin protein can result in escape from antibody-mediated neutralization. This allows the virus to reinfect individuals that have acquired immunity to previously circulating strains through infection or vaccination. To date, the vast majority of A(H1N1)pdm09 strains remain antigenically similar to the virus that caused the 2009 influenza pandemic. However, antigenic variants are expected to emerge as a result of increasing population immunity. We show that single amino acid substitutions near the receptor binding site were sufficient to escape from antibodies specific for A(H1N1)pdm09 viruses but not from antibodies elicited in response to infections with seasonal A(H1N1) and A(H1N1)pdm09 viruses. This study identified substitutions in A(H1N1)pdm09 viruses that support escape from population immunity but also suggested that the number of potential escape variants is limited by previous exposure to seasonal A(H1N1) viruses. PMID:25609810

Antigen-specific T cell therapies for cancer

PubMed Central

Manzo, Teresa; Heslop, Helen E.; Rooney, Cliona M.

2015-01-01

Adoptively transferred antigen-specific T cells that recognize tumor antigens through their native receptors have many potential benefits as treatment for virus-associated diseases and malignancies, due to their ability to selectively recognize tumor antigens, expand and persist to provide long-term protection. Infusions of T cells targeting Epstein–Barr virus (EBV) antigens have shown encouraging response rates in patients with post-transplant lymphoproliferative disease as well as EBV-positive lymphomas and nasopharyngeal cancer, although a recent study also showed that human papilloma virus-reactive T cells can induce complete regression of metastatic cervical cancer. This strategy is also being evaluated to target non-viral tumor-associated antigens. Targeting these less immunogenic antigens is more challenging, as tumor antigens are generally weak, and high avidity T cells specific for self-antigens are deleted in the thymus, but tumor responses have been reported. Current research focusses on defining factors that promote in vivo persistence of transferred cells and ameliorate the immunosuppressive microenvironment. To this end, investigators are evaluating the effects of combining adoptive transfer of antigen-specific T cells with other immunotherapy moieties such as checkpoint inhibitors. Genetic modification of infused T cells may also be used to overcome tumor evasion mechanisms, and vaccines may be used to promote in vivo proliferation. PMID:26160910

Antigenic and Molecular Characterization of Avian Influenza A(H9N2) Viruses, Bangladesh

PubMed Central

Shanmuganatham, Karthik; Feeroz, Mohammed M.; Jones-Engel, Lisa; Smith, Gavin J.D.; Fourment, Mathieu; Walker, David; McClenaghan, Laura; Alam, S.M. Rabiul; Hasan, M. Kamrul; Seiler, Patrick; Franks, John; Danner, Angie; Barman, Subrata; McKenzie, Pamela; Krauss, Scott; Webby, Richard J.

2013-01-01

Human infection with avian influenza A(H9N2) virus was identified in Bangladesh in 2011. Surveillance for influenza viruses in apparently healthy poultry in live-bird markets in Bangladesh during 2008–2011 showed that subtype H9N2 viruses are isolated year-round, whereas highly pathogenic subtype H5N1 viruses are co-isolated with subtype H9N2 primarily during the winter months. Phylogenetic analysis of the subtype H9N2 viruses showed that they are reassortants possessing 3 gene segments related to subtype H7N3; the remaining gene segments were from the subtype H9N2 G1 clade. We detected no reassortment with subtype H5N1 viruses. Serologic analyses of subtype H9N2 viruses from chickens revealed antigenic conservation, whereas analyses of viruses from quail showed antigenic drift. Molecular analysis showed that multiple mammalian-specific mutations have become fixed in the subtype H9N2 viruses, including changes in the hemagglutinin, matrix, and polymerase proteins. Our results indicate that these viruses could mutate to be transmissible from birds to mammals, including humans. PMID:23968540

Oral delivery of plant-derived HIV-1 p24 antigen in low doses shows a superior priming effect in mice compared to high doses.

PubMed

Lindh, Ingrid; Bråve, Andreas; Hallengärd, David; Hadad, Ronza; Kalbina, Irina; Strid, Åke; Andersson, Sören

2014-04-25

During early infection with human immunodeficiency virus type 1 (HIV-1), there is a rapid depletion of CD4(+) T-cells in the gut-associated lymphoid tissue (GALT) in the gastrointestinal tract. Therefore, immediate protection at these surfaces is of high priority for the development of an HIV-1 vaccine. Thus, transgenic plants expressing HIV-1 antigens, which are exposed to immune competent cells in the GALT during oral administration, can be interesting as potential vaccine candidates. In the present study, we used two HIV-1 p24 antigen-expressing transgenic plant systems, Arabidopsis thaliana and Daucus carota, in oral immunization experiments. Both transgenic plant systems showed a priming effect in mice and induced humoral immune responses, which could be detected as anti-p24-specific IgG in sera after an intramuscular p24 protein boost. Dose-dependent antigen analyses using transgenic A. thaliana indicated that low p24 antigen doses were superior to high p24 antigen doses. Copyright © 2014. Published by Elsevier Ltd.

Antigenic and genetic evolution of contemporary swine H1 influenza viruses in the United States.

PubMed

Rajao, Daniela S; Anderson, Tavis K; Kitikoon, Pravina; Stratton, Jered; Lewis, Nicola S; Vincent, Amy L

2018-05-01

Several lineages of influenza A viruses (IAV) currently circulate in North American pigs. Genetic diversity is further increased by transmission of IAV between swine and humans and subsequent evolution. Here, we characterized the genetic and antigenic evolution of contemporary swine H1N1 and H1N2 viruses representing clusters H1-α (1A.1), H1-β (1A.2), H1pdm (1A.3.3.2), H1-γ (1A.3.3.3), H1-δ1 (1B.2.2), and H1-δ2 (1B.2.1) currently circulating in pigs in the United States. The δ1-viruses diversified into two new genetic clades, H1-δ1a (1B.2.2.1) and H1-δ1b (1B.2.2.2), which were also antigenically distinct from the earlier H1-δ1-viruses. Further characterization revealed that a few key amino acid changes were associated with antigenic divergence in these groups. The continued genetic and antigenic evolution of contemporary H1 viruses might lead to loss of vaccine cross-protection that could lead to significant economic impact to the swine industry, and represents a challenge to public health initiatives that attempt to minimize swine-to-human IAV transmission. Published by Elsevier Inc.

Genotyping of occult hepatitis B virus infection in Egyptian hemodialysis patients without hepatitis C virus infection.

PubMed

Esmail, Mona A; Mahdi, Wafaa K M; Khairy, Rasha M; Abdalla, Nilly H

2016-01-01

Occult hepatitis B viral infection is the presence of hepatitis B viral nucleic acids in the serum and/or liver in the absence of hepatitis B surface antigen. The study aimed to determine the prevalence of occult hepatitis B virus infection among hepatitis C virus-negative hemodialysis patients and to identify their genotypes. of 144 patients on maintenance hemodialysis, 50 hepatitis B surface antigen and hepatitis C virus nucleic acid-negative patients were selected according to strict inclusion criteria to avoid the effect of confounding variables. The following investigations were done: serum AST and ALT; HBsAg; HBcAb; HCV-Ab; HCV-RNA; and HBV-DNA. Positive hepatitis B viral nucleic acid was confirmed in 12/144 (8.3%) hemodialysis patients and 12/50 (24%) in our study group (occult infection). Mean hemodialysis periods for negative patients and occult hepatitis B virus patients were 27.3±18.8 and 38.4±8.14 months, respectively, and this difference was significant (p-value=0.02). Mean alanine transaminase levels were 20.27±5.5IU/L and 25.3±9.6 in negative patients and occult infection patients, respectively. This difference was non-significant. Aspartate transaminase levels were 21.4±10.2IU/L and 27.3±4.6IU/L, respectively, in negative patients and infected patients; this difference was significant (p-value=0.03). Half (6/12) of the positive samples belonged to genotype 'B', 33.3% (4/12) to 'C', and 16.6% (2/12) to genotype 'D'. OBI is likely among hemodialysis patients even without HCV coinfection (24%). Genotype D cannot be the only genotype distributed in Upper Egypt, as the current study reported relatively new results that 50% of the patients with occult B carry genotype B, 33.3% carry genotype C and only 16.6% carry genotype D. Copyright © 2015 King Saud Bin Abdulaziz University for Health Sciences. Published by Elsevier Ltd. All rights reserved.

Monitoring the antigenic evolution of human influenza A viruses to understand how and when viruses escape from existing immunity.

PubMed

Liao, Yu-Chieh; Lin, Hsin-Hung; Lin, Chieh-Hua

2013-06-11

The World Health Organization (WHO) organizes consultations in February and September of each year, spearheaded by an advisory group of experts to analyze influenza surveillance data generated by the WHO Global Influenza Surveillance and Response System (GISRS). The purpose of these consultations is to recommend the composition on influenza virus vaccines for the northern and southern hemispheres, respectively. The latest news of influenza viruses is made available to the public and updated on the WHO website. Although WHO discloses the manner in which it has made the recommendation, usually by considering epidemiological and clinical information to analyze the antigenic and genetic characteristics of seasonal influenza viruses, most individuals do not possess an understanding of antigenic drift and when it occurs. We have constructed a web server, named Fluctrl, and implemented a pipeline whereby HA sequence data is downloaded from the Influenza Virus Resource at NCBI along with their isolation information including isolation year and location, which are parsed and managed in MySQL database. By analyzing the frequency of each amino acid residue of the HA1 domain expressed by the viruses on annual basis, users are able to obtain evolutionary dynamics of human influenza viruses corresponding with epidemics. Users are able to upload and analyze their HA1 sequences for generating evolutionary dynamics. In addition, a distribution of amino acid residues at a particular site is represented geographically to trace the location where antigenic variants are seeded. Fluctrl is constructed for monitoring the antigenic evolution of human influenza A viruses. This tool is intended to inform the general public how and when influenza viruses evade the human body's immunity. Furthermore, leveraging the geographic information, the original locations of emerging influenza viruses can be traced. Fluctrl is freely accessible at http://sb.nhri.org.tw/fluctrl.

Virus-Based Nanoparticles as Versatile Nanomachines

PubMed Central

Koudelka, Kristopher J.; Pitek, Andrzej S.; Manchester, Marianne; Steinmetz, Nicole F.

2016-01-01

Nanoscale engineering is revolutionizing the way we prevent, detect, and treat diseases. Viruses have played a special role in these developments because they can function as prefabricated nanoscaffolds that have unique properties and are easily modified. The interiors of virus particles can encapsulate and protect sensitive compounds, while the exteriors can be altered to display large and small molecules in precisely defined arrays. These properties of viruses, along with their innate biocompatibility, have led to their development as actively targeted drug delivery systems that expand on and improve current pharmaceutical options. Viruses are naturally immunogenic, and antigens displayed on their surface have been used to create vaccines against pathogens and to break self-tolerance to initiate an immune response to dysfunctional proteins. Densely and specifically aligned imaging agents on viruses have allowed for high-resolution and noninvasive visualization tools to detect and treat diseases earlier than previously possible. These and future applications of viruses have created an exciting new field within the disciplines of both nanotechnology and medicine. PMID:26958921

Prediction of conformational changes by single mutation in the hepatitis B virus surface antigen (HBsAg) identified in HBsAg-negative blood donors.

PubMed

Ie, Susan I; Thedja, Meta D; Roni, Martono; Muljono, David H

2010-11-18

Selection of hepatitis B virus (HBV) by host immunity has been suggested to give rise to variants with amino acid substitutions at or around the 'a' determinant of the surface antigen (HBsAg), the main target of antibody neutralization and diagnostic assays. However, there have never been successful attempts to provide evidence for this hypothesis, partly because the 3 D structure of HBsAg molecules has not been determined. Tertiary structure prediction of HBsAg solely from its primary amino acid sequence may reveal the molecular energetic of the mutated proteins. We carried out this preliminary study to analyze the predicted HBsAg conformation changes of HBV variants isolated from Indonesian blood donors undetectable by HBsAg assays and its significance, compared to other previously-reported variants that were associated with diagnostic failure. Three HBV variants (T123A, M133L and T143M) and a wild type sequence were analyzed together with frequently emerged variants T123N, M133I, M133T, M133V, and T143L. Based on the Jameson-Wolf algorithm for calculating antigenic index, the first two amino acid substitutions resulted in slight changes in the antigenicity of the 'a' determinant, while all four of the comparative variants showed relatively more significant changes. In the pattern T143M, changes in antigenic index were more significant, both in its coverage and magnitude, even when compared to variant T143L. These data were also partially supported by the tertiary structure prediction, in which the pattern T143M showed larger shift in the HBsAg second loop structure compared to the others. Single amino acid substitutions within or near the 'a' determinant of HBsAg may alter antigenicity properties of variant HBsAg, which can be shown by both its antigenic index and predicted 3 D conformation. Findings in this study emphasize the significance of variant T143M, the prevalent isolate with highest degree of antigenicity changes found in Indonesian blood donors. This highlights the importance of evaluating the effects of protein structure alterations on the sensitivity of screening methods being used in detection of ongoing HBV infection, as well as the use of vaccines and immunoglobulin therapy in contributing to the selection of HBV variants.

[Circulation of the influenza A virus of H13 serosubtype among seagulls in the Northern Caspian (1979-1985)].

PubMed

Iamnikova, S S; Kovtun, T O; Dmitriev, G A; Aristova, V A; Krivonosov, G A; Rusanov, G M; Konechnyĭ, A G; L'vov, D K

1989-01-01

The results of seven-year ecologo-virological studies (1979-1985) of Laridae colonies on the island Zhemchuzhnyi, northern Kaspian Sea, showed annual isolation of influenza A viruses. Altogether, 95 hemagglutinating agent have been isolated. Strains with 4 different combinations of surface antigens were identified: H5N2, H13N2, H13N3, H13N6. The possibility of transovarial transmission is confirmed by the fact of isolation of an influenza virus strain A/black-headed herring gull/Astrakhan/458/85 (H13N6) from a nestling having no contacts with the environment. Simultaneous circulation of influenza A viruses (in 1983--H13N2 and H13N6, in 1985.--H13N3 and H13N6) and the presence in the virion of neuraminidase of human influenza virus (N2) allow to consider the isolates to be natural recombinants.

DNA vaccine encoding nucleocapsid and surface proteins of wild type canine distemper virus protects its natural host against distemper.

PubMed

Cherpillod, P; Tipold, A; Griot-Wenk, M; Cardozo, C; Schmid, I; Fatzer, R; Schobesberger, M; Zurbriggen, R; Bruckner, L; Roch, F; Vandevelde, M; Wittek, R; Zurbriggen, A

2000-07-01

Canine distemper virus (CDV), a member of the genus Morbillivirus induces a highly infectious, frequently lethal disease in dogs and other carnivores. Current vaccines against canine distemper consisting of attenuated viruses have been in use for many years and have greatly reduced the incidence of distemper in the dog population. However, certain strains may not guarantee adequate protection and others can induce post vaccinal encephalitis. We tested a DNA vaccine for its ability to protect dogs, the natural host of CDV, against distemper. We constructed plasmids containing the nucleocapsid, the fusion, and the attachment protein genes of a virulent canine distemper virus strain. Mice inoculated with these plasmids developed humoral and cellular immune responses against CDV antigens. Dogs immunized with the expression plasmids developed virus-neutralizing antibodies. Significantly, vaccinated dogs were protected against challenge with virulent CDV, whereas unvaccinated animals succumbed to distemper.

Production of Japanese encephalitis virus-like particles in insect cells.

PubMed

Yamaji, Hideki; Konishi, Eiji

2013-01-01

Virus-like particles (VLPs) are composed of one or several recombinant viral surface proteins that spontaneously assemble into particulate structures without the incorporation of virus DNA or RNA. The baculovirus-insect cell system has been used extensively for the production of recombinant virus proteins including VLPs. While the baculovirus-insect cell system directs the transient expression of recombinant proteins in a batch culture, stably transformed insect cells allow constitutive production. In our recent study, a secretory form of Japanese encephalitis (JE) VLPs was successfully produced by Trichoplusia ni BTI-TN-5B1-4 (High Five) cells engineered to coexpress the JE virus (JEV) premembrane (prM) and envelope (E) proteins. A higher yield of E protein was attained with recombinant High Five cells than with the baculovirus-insect cell system. This study demonstrated that recombinant insect cells offer a promising approach to the high-level production of VLPs for use as vaccines and diagnostic antigens.

Plasmodium knowlesi Sporozoite Antigen: Expression by Infectious Recombinant Vaccinia Virus

NASA Astrophysics Data System (ADS)

Smith, Geoffrey L.; Godson, G. Nigel; Nussenzweig, Victor; Nussenzweig, Ruth S.; Barnwell, John; Moss, Bernard

1984-04-01

The gene coding for the circumsporozoite antigen of the malaria parasite Plasmodium knowlesi was inserted into the vaccinia virus genome under the control of a defined vaccinia virus promoter. Cells infected with the recombinant virus synthesized polypeptides of 53,000 to 56,000 daltons that reacted with monoclonal antibody against the repeating epitope of the malaria protein. Furthermore, rabbits vaccinated with the recombinant virus produced antibodies that bound specifically to sporozoites. These data provide evidence for expression of a cloned malaria gene in mammalian cells and illustrate the potential of vaccinia virus recombinants as live malaria vaccines.

Structure and genetic variability of envelope glycoproteins of two antigenic variants of caprine arthritis-encephalitis lentivirus.

PubMed

Knowles, D P; Cheevers, W P; McGuire, T C; Brassfield, A L; Harwood, W G; Stem, T A

1991-11-01

To define the structure of the caprine arthritis-encephalitis virus (CAEV) env gene and characterize genetic changes which occur during antigenic variation, we sequenced the env genes of CAEV-63 and CAEV-Co, two antigenic variants of CAEV defined by serum neutralization. The deduced primary translation product of the CAEV env gene consists of a 60- to 80-amino-acid signal peptide followed by an amino-terminal surface protein (SU) and a carboxy-terminal transmembrane protein (TM) separated by an Arg-Lys-Lys-Arg cleavage site. The signal peptide cleavage site was verified by amino-terminal amino acid sequencing of native CAEV-63 SU. In addition, immunoprecipitation of [35S]methionine-labeled CAEV-63 proteins by sera from goats immunized with recombinant vaccinia virus expressing the CAEV-63 env gene confirmed that antibodies induced by env-encoded recombinant proteins react specifically with native virion SU and TM. The env genes of CAEV-63 and CAEV-Co encode 28 conserved cysteines and 25 conserved potential N-linked glycosylation sites. Nucleotide sequence variability results in 62 amino acid changes and one deletion within the SU and 34 amino acid changes within the TM.

Structure and genetic variability of envelope glycoproteins of two antigenic variants of caprine arthritis-encephalitis lentivirus.

PubMed Central

Knowles, D P; Cheevers, W P; McGuire, T C; Brassfield, A L; Harwood, W G; Stem, T A

1991-01-01

To define the structure of the caprine arthritis-encephalitis virus (CAEV) env gene and characterize genetic changes which occur during antigenic variation, we sequenced the env genes of CAEV-63 and CAEV-Co, two antigenic variants of CAEV defined by serum neutralization. The deduced primary translation product of the CAEV env gene consists of a 60- to 80-amino-acid signal peptide followed by an amino-terminal surface protein (SU) and a carboxy-terminal transmembrane protein (TM) separated by an Arg-Lys-Lys-Arg cleavage site. The signal peptide cleavage site was verified by amino-terminal amino acid sequencing of native CAEV-63 SU. In addition, immunoprecipitation of [35S]methionine-labeled CAEV-63 proteins by sera from goats immunized with recombinant vaccinia virus expressing the CAEV-63 env gene confirmed that antibodies induced by env-encoded recombinant proteins react specifically with native virion SU and TM. The env genes of CAEV-63 and CAEV-Co encode 28 conserved cysteines and 25 conserved potential N-linked glycosylation sites. Nucleotide sequence variability results in 62 amino acid changes and one deletion within the SU and 34 amino acid changes within the TM. Images PMID:1656067

Sero-prevalence of hepatitis B virus infection in Balochistan Province of Pakistan.

PubMed

Sheikh, Nadeem S; Sheikh, Azeem S; Sheikh, Aqleem A; Yahya, Samira; Lateef, Majid

2011-01-01

The objective was to evaluate the sero-prevalence of hepatitis B surface antigen (HBsAg) and IgM antibodies to hepatitis core antigen in Balochistan Province of Pakistan. A cross-sectional, population-based study. Place and time of the study: The study was conducted in Balochistan from 1 st January 2004 to 31 st December, 2008. The screening areas included Barkhan, Eashani, Khuzdar, Kodi Zikriani, Kohlu, Rakhni and Turbat. A total of 15,260 subjects were enrolled; 11,900 (78%) agreed to undergo screening. Fresh serum samples were tested for the presence of hepatitis B surface antigen and IgM antibodies to hepatitis B core antigen. HBsAg was detected in 1166 (9.8%) while anti-HBc IgM was found in 117 (10.0%). HBsAg positivity was seen in 875 (12.7%) males and 291 (5.8%) females. The prevalence of hepatitis B in Balochistan varies from 3.3% in Khuzdar to 17.0% in Kodi Zikriani. It is utmost important to educate the public, to take proper measures to control the spread of infection and vaccination in order to interrupt transmission of this threatening public health problem in Balochistan province of Pakistan.

Detection of varicella-zoster virus antigens in lesional skin of zosteriform lichen planus but not in that of linear lichen planus.

PubMed

Mizukawa, Y; Horie, C; Yamazaki, Y; Shiohara, T

2012-01-01

Distinctions between 'linear lichen planus' (LP) and 'zosteriform LP' are difficult to determine solely based on clinical findings. The aim of this study is to determine whether the presence of the varicella-zoster virus (VZV) antigens could be used to differentiate the zosteriform LP from the linear LP. We immunohistochemically investigated the presence of in vivo localization of VZV antigens in 8 LP lesions (zosteriform LP: n = 5, linear LP: n = 3). We describe 2 cases of zosteriform LP without apparent prior episodes of herpes zoster, in whom VZV antigens were detected in the eccrine epithelium. Further analysis showed that VZV antigens were exclusively detected in the eccrine epithelium in the zosteriform LP lesions, but not in the linear LP lesions. Etiological differences exist between zosteriform LP and linear LP. The presence of VZV antigens in lesional skin of the former indicates a possible triggering role of this virus in the pathogenesis of this variant. Copyright © 2012 S. Karger AG, Basel.

Antigenic, genetic, and pathogenic characterization of H5N1 highly pathogenic avian influenza viruses isolated from dead whooper swans (Cygnus cygnus) found in northern Japan in 2008.

PubMed

Okamatsu, Masatoshi; Tanaka, Tomohisa; Yamamoto, Naoki; Sakoda, Yoshihiro; Sasaki, Takashi; Tsuda, Yoshimi; Isoda, Norikazu; Kokumai, Norihide; Takada, Ayato; Umemura, Takashi; Kida, Hiroshi

2010-12-01

In April and May 2008, whooper swans (Cygnus cygnus) were found dead in Hokkaido in Japan. In this study, an adult whooper swan found dead beside Lake Saroma was pathologically examined and the identified H5N1 influenza virus isolates were genetically and antigenically analyzed. Pathological findings indicate that the swan died of severe congestive edema in the lungs. Phylogenetic analysis of the HA genes of the isolates revealed that they are the progeny viruses of isolates from poultry and wild birds in China, Russia, Korea, and Hong Kong. Antigenic analyses indicated that the viruses are distinguished from the H5N1 viruses isolated from wild birds and poultry before 2007. The chickens vaccinated with A/duck/Hokkaido/Vac-1/2004 (H5N1) survived for 14 days after challenge with A/whooper swan/Hokkaido/1/2008 (H5N1), although a small amount of the challenge virus was recovered from the tissues of the birds. These findings indicate that H5N1 highly pathogenic avian influenza viruses are circulating in wild birds in addition to domestic poultry in Asia and exhibit antigenic variation that may be due to vaccination.

Quantitative and qualitative features of heterologous virus-vector-induced antigen-specific CD8+ T cells against Trypanosoma cruzi infection.

PubMed

Takayama, Eiji; Ono, Takeshi; Carnero, Elena; Umemoto, Saori; Yamaguchi, Yoko; Kanayama, Atsuhiro; Oguma, Takemi; Takashima, Yasuhiro; Tadakuma, Takushi; García-Sastre, Adolfo; Miyahira, Yasushi

2010-11-01

We studied some aspects of the quantitative and qualitative features of heterologous recombinant (re) virus-vector-induced, antigen-specific CD8(+) T cells against Trypanosoma cruzi. We used three different, highly attenuated re-viruses, i.e., influenza virus, adenovirus and vaccinia virus, which all expressed a single, T. cruzi antigen-derived CD8(+) T-cell epitope. The use of two out of three vectors or the triple virus-vector vaccination regimen not only confirmed that the re-vaccinia virus, which was placed last in order for sequential immunisation, was an effective booster for the CD8(+) T-cell immunity in terms of the number of antigen-specific CD8(+) T cells, but also demonstrated that (i) the majority of cells exhibit the effector memory (T(EM)) phenotype, (ii) robustly secrete IFN-γ, (iii) express higher intensity of the CD122 molecule and (iv) present protective activity against T. cruzi infection. In contrast, placing the re-influenza virus last in sequential immunisation had a detrimental effect on the quantitative and qualitative features of CD8(+) T cells. The triple virus-vector vaccination was more effective at inducing a stronger CD8(+) T-cell immunity than using two re-viruses. The different quantitative and qualitative features of CD8(+) T cells induced by different immunisation regimens support the notion that the refinement of the best choice of multiple virus-vector combinations is indispensable for the induction of a maximum number of CD8(+) T cells of high quality. Copyright © 2010 Australian Society for Parasitology Inc. All rights reserved.

The impact of H9N2 avian influenza virus vaccine antigenic variation on virus infectious dose in chickens

USDA-ARS?s Scientific Manuscript database

The H9 subtype of avian influenza virus is wide-spread in the areas of Asia and Middle East. Selection of effective vaccines that provide effective protection mainly depends on the antigenic match of the hemagglutinin protein (HA), between the vaccine and the field strain. To determine how the ant...

Live attenuated influenza A virus vaccine protects against heterologous challenge with A(H1N1)pdm09 without inducing vaccine associated enhanced respiratory disease

USDA-ARS?s Scientific Manuscript database

Influenza A virus (IAV) vaccines that provide broad cross-protection against antigenic variants are necessary to prevent infection and shedding of the wide array of IAV cocirculating in swine. Whole inactivated virus (WIV) vaccines provide only partial protection against IAV with substantial antigen...

Immunization With Fc-Based Recombinant Epstein–Barr Virus gp350 Elicits Potent Neutralizing Humoral Immune Response in a BALB/c Mice Model

PubMed Central

Zhao, Bingchun; Zhang, Xiao; Krummenacher, Claude; Song, Shuo; Gao, Ling; Zhang, Haojiong; Xu, Miao; Feng, Lin; Feng, Qisheng; Zeng, Musheng; Xu, Yuting; Zeng, Yixin

2018-01-01

Epstein–Barr virus (EBV) was the first human virus proved to be closely associated with tumor development, such as lymphoma, nasopharyngeal carcinoma, and EBV-associated gastric carcinoma. Despite many efforts to develop prophylactic vaccines against EBV infection and diseases, no candidates have succeeded in effectively blocking EBV infection in clinical trials. Previous investigations showed that EBV gp350 plays a pivotal role in the infection of B-lymphocytes. Nevertheless, using monomeric gp350 proteins as antigens has not been effective in preventing infection. Multimeric forms of the antigen are more potently immunogenic than monomers; however, the multimerization elements used in previous constructs are not approved for human clinical trials. To prepare a much-needed EBV prophylactic vaccine that is potent, safe, and applicable, we constructed an Fc-based form of gp350 to serve as a dimeric antigen. Here, we show that the Fc-based gp350 antigen exhibits dramatically enhanced immunogenicity compared with wild-type gp350 protein. The complete or partial gp350 ectodomain was fused with the mouse IgG2a Fc domain. Fusion with the Fc domain did not impair gp350 folding, binding to a conformation-dependent neutralizing antibody (nAb) and binding to its receptor by enzyme-linked immunosorbent assay and surface plasmon resonance. Specific antibody titers against gp350 were notably enhanced by immunization with gp350-Fc dimers compared with gp350 monomers. Furthermore, immunization with gp350-Fc fusion proteins elicited potent nAbs against EBV. Our data strongly suggest that an EBV gp350 vaccine based on Fc fusion proteins may be an efficient candidate to prevent EBV infection in clinical applications. PMID:29765376

Elicitation of anti-vesicular stomatitis virus cytotoxic T lymphocytes by using purified viral and cellular antigens incorporated into phospholipid vesicles.

PubMed

Ruebush, M J; Hale, A H; Harris, D T

1981-05-01

We evaluated the minimal molecular and cellular requirements for elicitation of anti-vesicular stomatitis virus (VSV) cytotoxic T lymphocytes (CTL). The results indicated that lipid vesicles containing the purified major surface glyco-protein of VSV (G protein) and purified H-2K(k) glycoproteins elicited specific H-2K(k)-restricted anti-VSV CTL. These antiviral CTL were shown to be Ly 1(-),2(+). However, both Ly 1(+),2(-) and Ly1(-),2(+) T-cell subpopulations were shown to be required for elicitation of these CTL.

Primary Seronegative but Molecularly Evident Hepadnaviral Infection Engages Liver and Induces Hepatocarcinoma in the Woodchuck Model of Hepatitis B

PubMed Central

Mulrooney-Cousins, Patricia M.; Chauhan, Ranjit; Churchill, Norma D.; Michalak, Tomasz I.

2014-01-01

Hepadnavirus at very low doses establishes in woodchucks asymptomatic, serologically undetectable but molecularly evident persistent infection. This primary occult infection (POI) preferentially engages the immune system and initiates virus-specific T cell response in the absence of antiviral antibody induction. The current study aimed to determine whether POI with time may culminate in serologically identifiable infection and hepatitis, and what are, if any, its pathological consequences. Juvenile woodchucks were intravenously injected with inocula containing 10 or 100 virions of woodchuck hepatitis virus (WHV) to induce POI and followed for life or up to 5.5 years thereafter. All 10 animals established molecularly detectable infection with virus DNA in serum (<100–200 copies/mL) and in circulating lymphoid cells, but serum WHV surface antigen and antibodies to WHV core antigen remained undetectable for life. By approximately 2.5–3.5 years post-infection, circulating virus transiently increased to 103 copies/mL and virus replication became detectable in the livers, but serological markers of infection and biochemical or histological evidence of hepatitis remained undetectable. Nonetheless, typical hepatocellular carcinoma (HCC) developed in 2/10 animals. WHV DNA integration into hepatic and lymphatic system genomes was identified in 9/10 animals. Virus recovered from the liver virus-negative or virus-positive phases of POI displayed the wild-type sequence and transmitted infection to healthy woodchucks causing hepatitis and HCC. In summary, for the first time, our data demonstrate that an asymptomatic hepadnaviral persistence initiated by very small amounts of otherwise pathogenic virus, advancing in the absence of traditional serological markers of infection and hepatitis, coincides with virus DNA integration into the host's hepatic and immune system genomes, retains liver pro-oncogenic potency and is capable of transmitting liver pathogenic infection. This emphasizes the role for primary occult hepatitis B virus infection in the development of seemingly cyptogenic HCC in seronegative but virus DNA reactive patients. PMID:25165821

Genetic and Molecular Studies of the Phlebotomus Fever Group of Viruses.

DTIC Science & Technology

1980-08-01

unrelated rhabdovirus , VSV. Two sources of LAC viral antigens were used, those from virus grown in BHK-21 cells and those obtained from LAC virus grown in...KAR, SFS and CHG, as well as those of the bunyaviruses LAC, ORI and BUN, and the viral antigens of the unrelated rhabdovirus , VSV. Again two sources...and Shope, R.E., 1977b, Oligonucleotide fingerprints of RNA obtained from rhabdoviruses belonging to the vesicular stornatitis virus subgroup. J

THE ANTIGENIC POTENCY OF EPIDEMIC INFLUENZA VIRUS FOLLOWING INACTIVATION BY ULTRAVIOLET RADIATION

PubMed Central

Salk, Jonas E.; Lavin, G. I.; Francis, Thomas

1940-01-01

A study of the antigenic potency of influenza virus inactivated by ultraviolet radiation has been made. Virus so inactivated is still capable of functioning as an immunizing agent when given to mice by the intraperitoneal route. In high concentrations inactivated virus appears to be nearly as effective as active virus but when quantitative comparisons of the immunity induced by different dilutions are made, it is seen that a hundredfold loss in immunizing capacity occurs during inactivation. Virus in suspensions prepared from the lungs of infected mice is inactivated more rapidly than virus in tissue culture medium. A standard for the comparison of vaccines of epidemic influenza virus is proposed. PMID:19871057

TREATMENT OF VIRUSES

DOE Office of Scientific and Technical Information (OSTI.GOV)

Polley, J.R.

1963-04-01

A vaccine preparation method was developed for destroying the infectivity of live viruses while retaining the antigenicity. The method comprises exposing the virus to 0.5 to 6 x 10/sup 6/ rad of ionizing radiation (preferably gamma) in the presence of a protective agent. The protective agent is antioxidant in nature and should be used in amounts from 0.05to 0.3% (wt/vol). Histidine and sodium p-aminohippurate are preferred for influenza and mumps viruses respectively. The protective effects of various chemicals on the antigenicity of irradiated influenza A virus are illustrated. (D.L.C.)

Dengue-1 Virus Envelope Glycoprotein Gene Expressed in Recombinant Baculovirus Elicits Virus-Neutralizing Antibody in Mice and Protects them from Virus Challenge

DTIC Science & Technology

1991-01-01

8217 terminus of E. When the recombinant virus was grown in Spodoptera frugiperda cells. about I mg of E antigen was made per 10’ cells. Recombinant E antigen...assay with DEN-I virus coprotein gene and its expression in Spodoptera hyperimmune mouse ascitic fluid. This heat-in- frugiperda cells activated...immunization, S. frugiperda cells infected with tion with BstNI (cuts at nucleotides 801 and recombinant baculovirus were pelleted. lysed by 2150). The

A functional simian virus 40 origin of replication is required for the generation of a super T antigen with a molecular weight of 100,000 in transformed mouse cells.

PubMed Central

Chen, S; Grass, D S; Blanck, G; Hoganson, N; Manley, J L; Pollack, R E

1983-01-01

We used two recombinant plasmids, one containing wild-type simian virus 40 DNA (pSVR1) and the other containing a simian virus 40 genome with a defective origin of replication (pSVR1-origin-minus) to transfect NIH3T3 cells. Quantitation of T-antigen synthesis by indirect immunofluorescence at 48 h after transfection with either DNA revealed the same percentage of T-positive nuclei. The transformation frequencies observed were also similar with both plasmids. Immunoprecipitation of [35S]methionine-labeled cell extracts showed the expected 94,000-dalton (94K) T and 17K t antigens in all clones examined. In pSVR1-generated transformants, a 100K super T antigen was also detected. Transformants isolated from pSVR1-origin-minus transfection, however, never expressed this 100K super T antigen, and some of these clones originally also showed greatly reduced levels of 94K T antigen. However, after growth in culture for several generations, the levels of 94K T antigen synthesis in these underproducer clones were dramatically increased. A direct correlation between the amounts of T antigen synthesized and the ability to grow independently of anchorage was observed. The mechanism which brings about increasing levels of T-antigen synthesis in some of the clones is not clear, but it appears not to be due to changes in either the copy number or the methylation pattern of the integrated simian virus 40 DNA. Images PMID:6312105

Serological evidence of Ebola virus infection in Indonesian orangutans.

PubMed

Nidom, Chairul A; Nakayama, Eri; Nidom, Reviany V; Alamudi, Mohamad Y; Daulay, Syafril; Dharmayanti, Indi N L P; Dachlan, Yoes P; Amin, Mohamad; Igarashi, Manabu; Miyamoto, Hiroko; Yoshida, Reiko; Takada, Ayato

2012-01-01

Ebola virus (EBOV) and Marburg virus (MARV) belong to the family Filoviridae and cause severe hemorrhagic fever in humans and nonhuman primates. Despite the discovery of EBOV (Reston virus) in nonhuman primates and domestic pigs in the Philippines and the serological evidence for its infection of humans and fruit bats, information on the reservoirs and potential amplifying hosts for filoviruses in Asia is lacking. In this study, serum samples collected from 353 healthy Bornean orangutans (Pongo pygmaeus) in Kalimantan Island, Indonesia, during the period from December 2005 to December 2006 were screened for filovirus-specific IgG antibodies using a highly sensitive enzyme-linked immunosorbent assay (ELISA) with recombinant viral surface glycoprotein (GP) antigens derived from multiple species of filoviruses (5 EBOV and 1 MARV species). Here we show that 18.4% (65/353) and 1.7% (6/353) of the samples were seropositive for EBOV and MARV, respectively, with little cross-reactivity among EBOV and MARV antigens. In these positive samples, IgG antibodies to viral internal proteins were also detected by immunoblotting. Interestingly, while the specificity for Reston virus, which has been recognized as an Asian filovirus, was the highest in only 1.4% (5/353) of the serum samples, the majority of EBOV-positive sera showed specificity to Zaire, Sudan, Cote d'Ivoire, or Bundibugyo viruses, all of which have been found so far only in Africa. These results suggest the existence of multiple species of filoviruses or unknown filovirus-related viruses in Indonesia, some of which are serologically similar to African EBOVs, and transmission of the viruses from yet unidentified reservoir hosts into the orangutan populations. Our findings point to the need for risk assessment and continued surveillance of filovirus infection of human and nonhuman primates, as well as wild and domestic animals, in Asia.

Serological Evidence of Ebola Virus Infection in Indonesian Orangutans

PubMed Central

Nidom, Reviany V.; Alamudi, Mohamad Y.; Daulay, Syafril; Dharmayanti, Indi N. L. P.; Dachlan, Yoes P.; Amin, Mohamad; Igarashi, Manabu; Miyamoto, Hiroko; Yoshida, Reiko; Takada, Ayato

2012-01-01

Ebola virus (EBOV) and Marburg virus (MARV) belong to the family Filoviridae and cause severe hemorrhagic fever in humans and nonhuman primates. Despite the discovery of EBOV (Reston virus) in nonhuman primates and domestic pigs in the Philippines and the serological evidence for its infection of humans and fruit bats, information on the reservoirs and potential amplifying hosts for filoviruses in Asia is lacking. In this study, serum samples collected from 353 healthy Bornean orangutans (Pongo pygmaeus) in Kalimantan Island, Indonesia, during the period from December 2005 to December 2006 were screened for filovirus-specific IgG antibodies using a highly sensitive enzyme-linked immunosorbent assay (ELISA) with recombinant viral surface glycoprotein (GP) antigens derived from multiple species of filoviruses (5 EBOV and 1 MARV species). Here we show that 18.4% (65/353) and 1.7% (6/353) of the samples were seropositive for EBOV and MARV, respectively, with little cross-reactivity among EBOV and MARV antigens. In these positive samples, IgG antibodies to viral internal proteins were also detected by immunoblotting. Interestingly, while the specificity for Reston virus, which has been recognized as an Asian filovirus, was the highest in only 1.4% (5/353) of the serum samples, the majority of EBOV-positive sera showed specificity to Zaire, Sudan, Cote d’Ivoire, or Bundibugyo viruses, all of which have been found so far only in Africa. These results suggest the existence of multiple species of filoviruses or unknown filovirus-related viruses in Indonesia, some of which are serologically similar to African EBOVs, and transmission of the viruses from yet unidentified reservoir hosts into the orangutan populations. Our findings point to the need for risk assessment and continued surveillance of filovirus infection of human and nonhuman primates, as well as wild and domestic animals, in Asia. PMID:22815803

Serological markers of hepatitis B, C, and E viruses and human immunodeficiency virus type-1 infections in pregnant women in Bali, Indonesia.

PubMed

Surya, I Gede Putu; Kornia, Karkata; Suwardewa, Tjok Gde Agung; Mulyanto; Tsuda, Fumio; Mishiro, Shunji

2005-04-01

Except for hepatitis B virus (HBV), there have been few data on serological markers of hepatitis viruses such as hepatitis C virus (HCV) and E virus (HEV), and human immunodeficiency virus type-1 (HIV) in Bali, Indonesia. During 5 months from April to August 2003, sera were collected from 2,450 pregnant women at eight jurisdictions in Bali, and they were tested for markers of these viruses. Only one (0.04%) was positive for antibody to HCV, but none for antibody to HIV. Hepatitis B surface antigen (HBsAg) was detected in 46 (1.9%) at a prevalence significantly lower than that in 271 of the 10,526 (2.6%) pregnant women in Bali surveyed 10 years previously (P < 0.045). The prevalence of hepatitis B e antigen in pregnant women with HBsAg decreased, also, from 50% to 28% during the 10 years (P < 0.011). Antibody to HEV (anti-HEV) was examined in 819 pregnant women who had been randomly selected from the 2,450. The overall prevalence of anti-HEV was 18%, and there were substantial regional differences spanning from 5% at Tabanan district to 32% at Gianyar district. Furthermore, the prevalence of anti-HEV differed substantially by their religions. In the Sanglah area of Denpasar City, for instance, anti-HEV was detected in 20 of the 102 (20%) Hindus, significantly more frequently than in only 2 of the 101 (2.0%) Muslims (P < 0.001). Swine that are prohibited to Muslims, therefore, is likely to serve as a reservoir of HEV in Bali. In conclusion, HBV is decreasing, HCV and HIV have not prevailed, as yet, while HEV is endemic probably through zoonotic infection in Bali. (c) 2005 Wiley-Liss, Inc.

The impact of antigenic drift of influenza A virus on human herd immunity: Sero-epidemiological study of H1N1 in healthy Thai population in 2009.

PubMed

Kanai, Yuta; Boonsathorn, Naphatsawan; Chittaganpitch, Malinee; Bai, Guirong; Li, Yonggang; Kase, Tetsuo; Takahashi, Kazuo; Okuno, Yoshinobu; Jampangern, Wipawee; Ikuta, Kazuyoshi; Sawanpanyalert, Pathom

2010-07-26

To examine the effect of the antigenic drift of H1N1 influenza viruses on herd immunity, neutralization antibodies from 744 sera from Thai healthy volunteers in 2008-2009, who had not been vaccinated for at least the last 5 years, were investigated by microneutralization (MN) and hemagglutination inhibition (HI) assays. Significantly higher MN titers were observed for the H1N1 Thai isolate in 2006 than in 2008. The results indicate that the antigenically drifted virus effectively escaped herd immunity. Since the low neutralization activity of herd immunity against drifted viruses is an important factor for viruses to spread efficiently, continuous sero-epidemiological study is required for public health. Copyright 2010 Elsevier Ltd. All rights reserved.

Virus-mimetic nanovesicles as a versatile antigen-delivery system

PubMed Central

Zhang, Pengfei; Chen, Yixin; Zeng, Yun; Shen, Chenguang; Li, Rui; Guo, Zhide; Li, Shaowei; Zheng, Qingbing; Chu, Chengchao; Wang, Zhantong; Zheng, Zizheng; Tian, Rui; Ge, Shengxiang; Zhang, Xianzhong; Xia, Ning-Shao; Liu, Gang; Chen, Xiaoyuan

2015-01-01

It is a critically important challenge to rapidly design effective vaccines to reduce the morbidity and mortality of unexpected pandemics. Inspired from the way that most enveloped viruses hijack a host cell membrane and subsequently release by a budding process that requires cell membrane scission, we genetically engineered viral antigen to harbor into cell membrane, then form uniform spherical virus-mimetic nanovesicles (VMVs) that resemble natural virus in size, shape, and specific immunogenicity with the help of surfactants. Incubation of major cell membrane vesicles with surfactants generates a large amount of nano-sized uniform VMVs displaying the native conformational epitopes. With the diverse display of epitopes and viral envelope glycoproteins that can be functionally anchored onto VMVs, we demonstrate VMVs to be straightforward, robust and tunable nanobiotechnology platforms for fabricating antigen delivery systems against a wide range of enveloped viruses. PMID:26504197

Construction of a hepatitis B virus neutralizing chimeric monoclonal antibody recognizing escape mutants of the viral surface antigen (HBsAg).

PubMed

Golsaz-Shirazi, Forough; Amiri, Mohammad Mehdi; Farid, Samira; Bahadori, Motahareh; Bohne, Felix; Altstetter, Sebastian; Wolff, Lisa; Kazemi, Tohid; Khoshnoodi, Jalal; Hojjat-Farsangi, Mohammad; Chudy, Michael; Jeddi-Tehrani, Mahmood; Protzer, Ulrike; Shokri, Fazel

2017-08-01

Hepatitis B virus (HBV) infection is a global burden on the health-care system and is considered as the tenth leading cause of death in the world. Over 248 million patients are currently suffering from chronic HBV infection worldwide and annual mortality rate of this infection is 686000. The "a" determinant is a hydrophilic region present in all antigenic subtypes of hepatitis B surface antigen (HBsAg), and antibodies against this region can neutralize the virus and are protective against all subtypes. We have recently generated a murine anti-HBs monoclonal antibody (4G4), which can neutralize HBV infection in HepaRG cells and recognize most of the escape mutant forms of HBsAg. Here, we describe the production and characterization of the chimeric human-murine antibody 4G4 (c-4G4). Variable region genes of heavy and light chains of the m-4G4 were cloned and fused to constant regions of human kappa and IgG1 by splice overlap extension (SOE) PCR. The chimeric antibody was expressed in Chinese Hamster Ovary (CHO)-K1 cells and purified from culture supernatant. Competition ELISA proved that both antibodies bind the same epitope within HBsAg. Antigen-binding studies using ELISA and Western blot showed that c-4G4 has retained the affinity and specificity of the parental murine antibody, and displayed a similar pattern of reactivity to 13 escape mutant forms of HBsAg. Both, the parental and c-4G4 showed a comparably high HBV neutralization capacity in cell culture even at the lowest concentration (0.6μg/ml). Due to the ability of c-4G4 to recognize most of the sub-genotypes and escape mutants of HBsAg, this antibody either alone or in combination with other anti-HBs antibodies could be considered as a potent alternative for Hepatitis B immune globulin (HBIG) as an HBV infection prophylactic or for passive immunotherapy against HBV infection. Copyright © 2017 Elsevier B.V. All rights reserved.

Phylogenetic, antigenic and biological characterization of pigeon paramyxovirus type 1 circulating in China.

PubMed

Qiu, Xusheng; Meng, Chunchun; Zhan, Yuan; Yu, Shengqing; Li, Shichao; Ren, Tingting; Yuan, Weifeng; Xu, Shuqin; Sun, Yingjie; Tan, Lei; Song, Cuiping; Liao, Ying; Ding, Zhuang; Liu, Xiufan; Ding, Chan

2017-09-29

For many years, ND has been one of the most important infectious pigeon diseases in China. In recent years, a high mortality has been observed in ND-infected pigeons in China. Mortality is from 40% to 80% or 100% in some cases. The full-length genomes of four pigeon paramyxovirus type 1 (PPMV-1) strains, which were isolated from infected pigeons in China in 2012 and 2013, were sequenced and analyzed to determine the phylogenetic characteristics of PPMV-1 circulating in pigeons of China in recent years. Furthermore, cross hemagglutination inhibition and cross virus neutralization assays, as well as animal experiments were conducted to determine the antigenicity and pathogenicity of those viruses. Proteolytic cleavage sites (residues 112-117) of the F proteins were identified as the typical virulence motif, 112 RRQKR↓F 117 for all four PPMV-1 strains investigated. Phylogenetic analysis based on sequences of complete genomes and F gene revealed that the four PPMV-1 isolates and most of recent isolates in China were highly homologous to European isolates from 1998 to 2011. All those isolates were clustered in one clade of genotype VI NDV, termed as subgroup 4bii f. The R value was calculated based on cross hemagglutination inhibition and cross virus neutralization results, and confirmed antigenic difference of the PPMV-1 strains isolated in 2013 from the LaSota vaccine strain. Several mutations were identified in the surface glycoproteins F and HN, which probably gave rise to those antigenic differences. Our result suggested that the PPMV-1 strain prevailing in China in the last decade diverged from a common ancestor and was presumably transmitted from Europe. PPMV-1 isolates displayed obvious antigenic differences from vaccine strain LaSota. Even though PPMV-1 did not cause high mortality in experimental pigeons, the infected pigeons were exhibiting viral shedding for 3 weeks after infection, suggesting PPMV-1 is a potential threat to NDV control worldwide.

Performance characteristics of qualified cell lines for isolation and propagation of influenza viruses for vaccine manufacturing

PubMed Central

Donis, Ruben O.; Chen, i-Mei; Davis, C Todd; Foust, Angie; Hossain, M. Jaber; Johnson, Adam; Klimov, Alexander; Loughlin, Rosette; Xu, Xiyan; Tsai, Theodore; Blayer, Simone; Trusheim, Heidi; Colegate, Tony; Fox, John; Taylor, Beverly; Hussain, Althaf; Barr, Ian; Baas, Chantal; Louwerens, Jaap; Geuns, Ed; Lee, Min-Shi; Venhuizen, odewijk; Neumeier, Elisabeth; Ziegler, Thedi

2018-01-01

Cell culture is now available as a method for the production of influenza vaccines in addition to eggs. In accordance with currently accepted practice, viruses recommended as candidates for vaccine manufacture are isolated and propagated exclusively in hens' eggs prior to distribution to manufacturers. Candidate vaccine viruses isolated in cell culture are not available to support vaccine manufacturing in mammalian cell bioreactors so egg-derived viruses have to be used. Recently influenza A (H3N2) viruses have been difficult to isolate directly in eggs. As mitigation against this difficulty, and the possibility of no suitable egg-isolated candidate viruses being available, it is proposed to consider using mammalian cell lines for primary isolation of influenza viruses as candidates for vaccine production in egg and cell platforms. To investigate this possibility, we tested the antigenic stability of viruses isolated and propagated in cell lines qualified for influenza vaccine manufacture and subsequently investigated antigen yields of such viruses in these cell lines at pilot-scale. Twenty influenza A and B-positive, original clinical specimens were inoculated in three MDCK cell lines. The antigenicity of recovered viruses was tested by hemagglutination inhibition using ferret sera against contemporary vaccine viruses and the amino acid sequences of the hemagglutinin and neuraminidase were determined. MDCK cell lines proved to be highly sensitive for virus isolation. Compared to the virus sequenced from the original specimen, viruses passaged three times in the MDCK lines showed up to 2 amino acid changes in the hemagglutinin. Antigenic stability was also established by hemagglutination inhibition titers comparable to those of the corresponding reference virus. Viruses isolated in any of the three MDCK lines grew reasonably well but variably in three MDCK cells and in VERO cells at pilot-scale. These results indicate that influenza viruses isolated in vaccine certified cell lines may well qualify for use in vaccine production. PMID:24975811

Performance characteristics of qualified cell lines for isolation and propagation of influenza viruses for vaccine manufacturing.

PubMed

Donis, Ruben O; Davis, C Todd; Foust, Angie; Hossain, M Jaber; Johnson, Adam; Klimov, Alexander; Loughlin, Rosette; Xu, Xiyan; Tsai, Theodore; Blayer, Simone; Trusheim, Heidi; Colegate, Tony; Fox, John; Taylor, Beverly; Hussain, Althaf; Barr, Ian; Baas, Chantal; Louwerens, Jaap; Geuns, Ed; Lee, Min-Shi; Venhuizen, Odewijk; Neumeier, Elisabeth; Ziegler, Thedi

2014-11-12

Cell culture is now available as a method for the production of influenza vaccines in addition to eggs. In accordance with currently accepted practice, viruses recommended as candidates for vaccine manufacture are isolated and propagated exclusively in hens' eggs prior to distribution to manufacturers. Candidate vaccine viruses isolated in cell culture are not available to support vaccine manufacturing in mammalian cell bioreactors so egg-derived viruses have to be used. Recently influenza A (H3N2) viruses have been difficult to isolate directly in eggs. As mitigation against this difficulty, and the possibility of no suitable egg-isolated candidate viruses being available, it is proposed to consider using mammalian cell lines for primary isolation of influenza viruses as candidates for vaccine production in egg and cell platforms. To investigate this possibility, we tested the antigenic stability of viruses isolated and propagated in cell lines qualified for influenza vaccine manufacture and subsequently investigated antigen yields of such viruses in these cell lines at pilot-scale. Twenty influenza A and B-positive, original clinical specimens were inoculated in three MDCK cell lines. The antigenicity of recovered viruses was tested by hemagglutination inhibition using ferret sera against contemporary vaccine viruses and the amino acid sequences of the hemagglutinin and neuraminidase were determined. MDCK cell lines proved to be highly sensitive for virus isolation. Compared to the virus sequenced from the original specimen, viruses passaged three times in the MDCK lines showed up to 2 amino acid changes in the hemagglutinin. Antigenic stability was also established by hemagglutination inhibition titers comparable to those of the corresponding reference virus. Viruses isolated in any of the three MDCK lines grew reasonably well but variably in three MDCK cells and in VERO cells at pilot-scale. These results indicate that influenza viruses isolated in vaccine certified cell lines may well qualify for use in vaccine production. Published by Elsevier Ltd.

Prevalence of Hepatitis B surface antigen among biomedical students of African descent in Usmanu Danfodiyo University, Sokoto, Nigeria.

PubMed

Okwesili, A N; Onuigwe, F U; Ibrahim, K; Buhari, H; Ibrahim, A; Jafaru, H; Erhabor, O; Onuigwe, F U; Isaac, Z; Ahmed, M H; Mainasara, M Y; Adias, T C; Yeldu, M H; Uko, E K; Udoma, F

2015-12-23

Hepatitis B (HB) is a serious global public health problem that put health professionals particularly at risk. The aim of this study was to investigate the prevalence of Hepatitis B surface antigen (HBsAg) among Biomedical Students of African descent attending Usmanu Danfodiyo University Sokoto in North-Western Nigeria. The Onsite HBsAg (CTK Biotech, USA) was used to detect the presence of hepatitis B surface antigen. We tested 186 consecutively-recruited students consisting of 147 males and 39 females aged 18-35 years (mean age 26 ± 2.0 years). Of the 186 students tested, 25 (13.4%) were positive for HBsAg. The prevalence of HBsAg was significantly higher among students in the 21-25 years age group. Hepatitis B vaccination uptake among students was 7%. Majority of subjects were single 173(93.1%) compared to married 13 (6.9%). Ethnic distribution of the subjects indicated that 104(55.9%) were Hausa compared to Yoruba 32 (17.2%), other ethnic groups 21(11.3%), Fulani 20(10.8%) and Igbo 9(4.8%). This study indicates a high prevalence of hepatitis B virus infection among Biomedical students in Sokoto, North Western, Nigeria. Finding from this study is enough justification for the implementation of a policy to routinely test students entering into the biomedical professions for Hepatitis B virus infection. There is the need to provide hepatitis B vaccination universally to all those who are found negative prior to commencement of their biomedical training. There is also need to educate students entering biomedical professions and healthcare workers on the modes of transmission and prevention, importance of being compliant with protective vaccination as well as the need to observe universal precaution and infection control guidelines during their training and future professional practice.

Analysis of the role of tripeptidyl peptidase II in MHC class I antigen presentation in vivo1

PubMed Central

Kawahara, Masahiro; York, Ian A.; Hearn, Arron; Farfan, Diego; Rock, Kenneth L.

2015-01-01

Previous experiments using enzyme inhibitors and RNAi in cell lysates and cultured cells have suggested that tripeptidyl peptidase II (TPPII) plays a role in creating and destroying MHC class I-presented peptides. However, its precise contribution to these processes has been controversial. To elucidate the importance of TPPII in MHC class I antigen presentation, we analyzed TPPII-deficient gene-trapped mice and cell lines from these animals. In these mice, the expression level of TPPII was reduced by >90% compared to wild-type mice. Thymocytes from TPPII gene-trapped mice displayed more MHC class I on the cell surface, suggesting that TPPII normally limits antigen presentation by destroying peptides overall. TPPII gene-trapped mice responded as well as did wild-type mice to four epitopes from lymphocytic choriomeningitis virus (LCMV). The processing and presentation of peptide precursors with long N-terminal extensions in TPPII gene-trapped embryonic fibroblasts was modestly reduced, but in vivo immunization with recombinant lentiviral or vaccinia virus vectors revealed that such peptide precursors induced an equivalent CD8 T cell response in wild type and TPPII-deficient mice. These data indicate while TPPII contributes to the trimming of peptides with very long N-terminal extensions, TPPII is not essential for generating most MHC class I-presented peptides or for stimulating CTL responses to several antigens in vivo. PMID:19841172

Shifting Clade Distribution, Reassortment, and Emergence of New Subtypes of Highly Pathogenic Avian Influenza A(H5) Viruses Collected from Vietnamese Poultry from 2012 to 2015

PubMed Central

Jang, Yunho; Nguyen, Tho D.; Jones, Joyce; Shepard, Samuel S.; Yang, Hua; Gerloff, Nancy; Nguyen, Long V.; Inui, Ken; Yang, Genyan; Creanga, Adrian; Wang, Li; Mai, Duong T.; Thor, Sharmi; Stevens, James; To, Thanh L.; Wentworth, David E.; Nguyen, Tung; Pham, Dong V.; Bryant, Juliet E.

2016-01-01

ABSTRACT Whole-genome sequences of representative highly pathogenic avian influenza A(H5) viruses from Vietnam were generated, comprising samples from poultry outbreaks and active market surveillance collected from January 2012 to August 2015. Six hemagglutinin gene clades were characterized. Clade 1.1.2 was predominant in southern Mekong provinces throughout 2012 and 2013 but gradually disappeared and was not detected after April 2014. Clade 2.3.2.1c viruses spread rapidly during 2012 and were detected in the south and center of the country. A number of clade 1.1.2 and 2.3.2.1c interclade reassortant viruses were detected with different combinations of internal genes derived from 2.3.2.1a and 2.3.2.1b viruses, indicating extensive cocirculation. Although reassortment generated genetic diversity at the genotype level, there was relatively little genetic drift within the individual gene segments, suggesting genetic stasis over recent years. Antigenically, clade 1.1.2, 2.3.2.1a, 2.3.2.1b, and 2.3.2.1c viruses remained related to earlier viruses and WHO-recommended prepandemic vaccine strains representing these clades. Clade 7.2 viruses, although detected in only low numbers, were the exception, as indicated by introduction of a genetically and antigenically diverse strain in 2013. Clade 2.3.4.4 viruses (H5N1 and H5N6) were likely introduced in April 2014 and appeared to gain dominance across northern and central regions. Antigenic analyses of clade 2.3.4.4 viruses compared to existing clade 2.3.4 candidate vaccine viruses (CVV) indicated the need for an updated vaccine virus. A/Sichuan/26221/2014 (H5N6) virus was developed, and ferret antisera generated against this virus were demonstrated to inhibit some but not all clade 2.3.4.4 viruses, suggesting consideration of alternative clade 2.3.4.4 CVVs. IMPORTANCE Highly pathogenic avian influenza (HPAI) A(H5) viruses have circulated continuously in Vietnam since 2003, resulting in hundreds of poultry outbreaks and sporadic human infections. Despite a significant reduction in the number of human infections in recent years, poultry outbreaks continue to occur and the virus continues to diversify. Vaccination of poultry has been used as a means to control the spread and impact of the virus, but due to the diversity and changing distribution of antigenically distinct viruses, the utility of vaccines in the face of mismatched circulating strains remains questionable. This study assessed the putative amino acid changes in viruses leading to antigenic variability, underscoring the complexity of vaccine selection for both veterinary and public health purposes. Given the overlapping geographic distributions of multiple, antigenically distinct clades of HPAI A(H5) viruses in Vietnam, the vaccine efficacy of bivalent poultry vaccine formulations should be tested in the future. PMID:28003481

Shifting Clade Distribution, Reassortment, and Emergence of New Subtypes of Highly Pathogenic Avian Influenza A(H5) Viruses Collected from Vietnamese Poultry from 2012 to 2015.

PubMed

Nguyen, Diep T; Jang, Yunho; Nguyen, Tho D; Jones, Joyce; Shepard, Samuel S; Yang, Hua; Gerloff, Nancy; Fabrizio, Thomas; Nguyen, Long V; Inui, Ken; Yang, Genyan; Creanga, Adrian; Wang, Li; Mai, Duong T; Thor, Sharmi; Stevens, James; To, Thanh L; Wentworth, David E; Nguyen, Tung; Pham, Dong V; Bryant, Juliet E; Davis, C Todd

2017-03-01

Whole-genome sequences of representative highly pathogenic avian influenza A(H5) viruses from Vietnam were generated, comprising samples from poultry outbreaks and active market surveillance collected from January 2012 to August 2015. Six hemagglutinin gene clades were characterized. Clade 1.1.2 was predominant in southern Mekong provinces throughout 2012 and 2013 but gradually disappeared and was not detected after April 2014. Clade 2.3.2.1c viruses spread rapidly during 2012 and were detected in the south and center of the country. A number of clade 1.1.2 and 2.3.2.1c interclade reassortant viruses were detected with different combinations of internal genes derived from 2.3.2.1a and 2.3.2.1b viruses, indicating extensive cocirculation. Although reassortment generated genetic diversity at the genotype level, there was relatively little genetic drift within the individual gene segments, suggesting genetic stasis over recent years. Antigenically, clade 1.1.2, 2.3.2.1a, 2.3.2.1b, and 2.3.2.1c viruses remained related to earlier viruses and WHO-recommended prepandemic vaccine strains representing these clades. Clade 7.2 viruses, although detected in only low numbers, were the exception, as indicated by introduction of a genetically and antigenically diverse strain in 2013. Clade 2.3.4.4 viruses (H5N1 and H5N6) were likely introduced in April 2014 and appeared to gain dominance across northern and central regions. Antigenic analyses of clade 2.3.4.4 viruses compared to existing clade 2.3.4 candidate vaccine viruses (CVV) indicated the need for an updated vaccine virus. A/Sichuan/26221/2014 (H5N6) virus was developed, and ferret antisera generated against this virus were demonstrated to inhibit some but not all clade 2.3.4.4 viruses, suggesting consideration of alternative clade 2.3.4.4 CVVs. IMPORTANCE Highly pathogenic avian influenza (HPAI) A(H5) viruses have circulated continuously in Vietnam since 2003, resulting in hundreds of poultry outbreaks and sporadic human infections. Despite a significant reduction in the number of human infections in recent years, poultry outbreaks continue to occur and the virus continues to diversify. Vaccination of poultry has been used as a means to control the spread and impact of the virus, but due to the diversity and changing distribution of antigenically distinct viruses, the utility of vaccines in the face of mismatched circulating strains remains questionable. This study assessed the putative amino acid changes in viruses leading to antigenic variability, underscoring the complexity of vaccine selection for both veterinary and public health purposes. Given the overlapping geographic distributions of multiple, antigenically distinct clades of HPAI A(H5) viruses in Vietnam, the vaccine efficacy of bivalent poultry vaccine formulations should be tested in the future. Copyright © 2017 American Society for Microbiology.

Complete replication of hepatitis B virus and hepatitis C virus in a newly developed hepatoma cell line.

PubMed

Yang, Darong; Zuo, Chaohui; Wang, Xiaohong; Meng, Xianghe; Xue, Binbin; Liu, Nianli; Yu, Rong; Qin, Yuwen; Gao, Yimin; Wang, Qiuping; Hu, Jun; Wang, Ling; Zhou, Zebin; Liu, Bing; Tan, Deming; Guan, Yang; Zhu, Haizhen

2014-04-01

The absence of a robust cell culture system for hepatitis B virus (HBV) and hepatitis C virus (HCV) infection has limited the analysis of the virus lifecycle and drug discovery. We have established a hepatoma cell line, HLCZ01, the first cell line, to the authors' knowledge, supporting the entire lifecycle of both HBV and HCV. HBV surface antigen (HBsAg)-positive particles can be observed in the supernatant and the lumen of the endoplasmic reticulum of the cells via electron microscopy. Interestingly, HBV and HCV clinical isolates propagate in HLCZ01 cells. Both viruses replicate in the cells without evidence of overt interference. HBV and HCV entry are blocked by antibodies against HBsAg and human CD81, respectively, and the replication of HBV and HCV is inhibited by antivirals. HLCZ01 cells mount an innate immune response to virus infection. The cell line provides a powerful tool for exploring the mechanisms of virus entry and replication and the interaction between host and virus, facilitating the development of novel antiviral agents and vaccines.

Two complex, adenovirus-based vaccines that together induce immune responses to all four dengue virus serotypes.

PubMed

Holman, David H; Wang, Danher; Raviprakash, Kanakatte; Raja, Nicholas U; Luo, Min; Zhang, Jianghui; Porter, Kevin R; Dong, John Y

2007-02-01

Dengue virus infections can cause hemorrhagic fever, shock, encephalitis, and even death. Worldwide, approximately 2.5 billion people live in dengue-infested regions with about 100 million new cases each year, although many of these infections are believed to be silent. There are four antigenically distinct serotypes of dengue virus; thus, immunity from one serotype will not cross-protect from infection with the other three. The difficulties that hamper vaccine development include requirements of the natural conformation of the envelope glycoprotein to induce neutralizing immune responses and the necessity of presenting antigens of all four serotypes. Currently, the only way to meet these requirements is to use a mixture of four serotypes of live attenuated dengue viruses, but safety remains a major problem. In this study, we have developed the basis for a tetravalent dengue vaccine using a novel complex adenovirus platform that is capable of expressing multiple antigens de novo. This dengue vaccine is constructed as a pair of vectors that each expresses the premembrane and envelope genes of two different dengue virus serotypes. Upon vaccination, the vaccine expressed high levels of the dengue virus antigens in cells to mimic a natural infection and induced both humoral and cellular immune responses against multiple serotypes of dengue virus in an animal model. Further analyses show the humoral responses were indeed neutralizing against all four serotypes. Our studies demonstrate the concept of mimicking infections to induce immune responses by synthesizing dengue virus membrane antigens de novo and the feasibility of developing an effective tetravalent dengue vaccine by vector-mediated expression of glycoproteins of the four serotypes.

A new inositol triester from Taraxacum mongolicum.

PubMed

Liu, Jifeng; Zhang, Nenling; Liu, Mengqi

2014-01-01

One new inositol triester, 4,5,6-tri-O-p-hydroxyphenylacetyl-chiro-inositol (1), was isolated from the ethanolic extract of Taraxacum mongolicum, along with two known compounds, 11β,13-dihydrotaraxinic acid (2) and taraxinic acid β-d-glucopyranosyl ester (3). The isolates were tested for their anti-hepatitis B virus (HBV) activities; 11β,13-dihydrotaraxinic acid (2) exhibited an IC50 value of 0.91 mM inhibiting the secretion of the HBV surface antigen and an IC50 value of 0.34 mM inhibiting the secretion of the HBV e antigen using HBV transfected Hep G2.2.15 cell line.

Adeno-associated virus capsid antigen presentation is dependent on endosomal escape

PubMed Central

Li, Chengwen; He, Yi; Nicolson, Sarah; Hirsch, Matt; Weinberg, Marc S.; Zhang, Ping; Kafri, Tal; Samulski, R. Jude

2013-01-01

Adeno-associated virus (AAV) vectors are attractive for gene delivery-based therapeutics, but data from recent clinical trials have indicated that AAV capsids induce a cytotoxic T lymphocyte (CTL) response that eliminates transduced cells. In this study, we used traditional pharmacological agents and AAV mutants to elucidate the pathway of capsid cross-presentation in AAV-permissive cells. Endosomal acidification inhibitors blocked AAV2 antigen presentation by over 90%, while proteasome inhibitors completely abrogated antigen presentation. Using mutant viruses that are defective for nuclear entry, we observed a 90% decrease in capsid antigen presentation. Different antigen presentation efficiencies were achieved by selectively mutating virion nuclear localization signals. Low antigen presentation was demonstrated with basic region 1 (BR1) mutants, despite relatively high transduction efficiency, whereas there was no difference in antigen presentation between BR2 and BR3 mutants defective for transduction, as compared with wild-type AAV2. These results suggest that effective AAV2 capsid antigen presentation is dependent on AAV virion escape from the endosome/lysosome for antigen degradation by proteasomes, but is independent of nuclear uncoating. These results should facilitate the design of effective strategies to evade capsid-specific CTL-mediated elimination of AAV-transduced target cells in future clinical trials. PMID:23454772

A Review of Intra- and Extracellular Antigen Delivery Systems for Virus Vaccines of Finfish

PubMed Central

Munang'andu, Hetron Mweemba; Evensen, Øystein

2015-01-01

Vaccine efficacy in aquaculture has for a long time depended on evaluating relative percent survival and antibody responses after vaccination. However, current advances in vaccine immunology show that the route in which antigens are delivered into cells is deterministic of the type of adaptive immune response evoked by vaccination. Antigens delivered by the intracellular route induce MHC-I restricted CD8+ responses while antigens presented through the extracellular route activate MHC-II restricted CD4+ responses implying that the route of antigen delivery is a conduit to induction of B- or T-cell immune responses. In finfish, different antigen delivery systems have been explored that include live, DNA, inactivated whole virus, fusion protein, virus-like particles, and subunit vaccines although mechanisms linking these delivery systems to protective immunity have not been studied in detail. Hence, in this review we provide a synopsis of different strategies used to administer viral antigens via the intra- or extracellular compartments. Further, we highlight the differences in immune responses induced by antigens processed by the endogenous route compared to exogenously processed antigens. Overall, we anticipate that the synopsis put together in this review will shed insights into limitations and successes of the current vaccination strategies used in finfish vaccinology. PMID:26065009

Glycosylations in the globular head of the hemagglutinin protein modulate the virulence and antigenic properties of the H1N1 influenza viruses

PubMed Central

Medina, Rafael A.; Stertz, Silke; Manicassamy, Balaji; Zimmermann, Petra; Sun, Xiangjie; Albrecht, Randy A.; Uusi-Kerttula, Hanni; Zagordi, Osvaldo; Belshe, Robert B.; Frey, Sharon E.; Eggink, Dirk; Tumpey, Terrence M.; García-Sastre, Adolfo

2014-01-01

The global spread of the 2009 pandemic H1N1 (pH1N1) virus in humans increases the likelihood that this influenza virus strain could undergo antigenic drift in the coming years. Previous seasonal H1N1 and H3N2 influenza strains acquired additional glycosylations in the globular head of their hemagglutinin (HA) proteins as they evolved over time; these are believed to shield antigenically relevant regions. We used influenza A/Netherlands/602/2009 recombinant (rpH1N1) viruses to which we added additional HA glycosylation sites reflecting their temporal appearance in previous seasonal H1N1 viruses. Additional glycosylations resulted in substantial attenuation in mice and ferrets, while deleting HA glycosylation sites from a pre-pandemic 1991 seasonal H1N1 influenza virus resulted in increased pathogenicity in mice. Sera from mice infected with wild type (WT) rpH1N1 virus showed a considerable loss of HA inhibitory (HI) activity against rpH1N1 viruses glycosylated at sites 144 or 144-172, indicating that the polyclonal antibody response elicited by WT rpH1N1 HA seems to be directed against an immunodominant region, likely site Sa, shielded by glycosylation at 144. Sera from humans vaccinated with the pH1N1 inactivated vaccine also showed reduced activity against the 144 and 144-172 mutant viruses. Remarkably, the HI activity of sera from virus-infected mice demonstrated that glycosylation at position 144 resulted in the induction of a broader polyclonal response able to cross-neutralize all WT and glycosylation mutant pH1N1 viruses. Mice infected with a recent seasonal virus in which glycosylation sites 71, 142 and 177 were removed, elicited antibodies that protected against challenge with the antigenically distant pH1N1 virus. Thus, acquisition of glycosylation sites in the HA of H1N1 human influenza viruses not only affects their pathogenicity and ability to escape from polyclonal antibodies elicited by previous influenza virus strains, but also their ability to induce cross-reactive antibodies against drifted antigenic variants. These findings provide the basis for designing improved vaccines and immunization strategies capable of protecting against a broader range of influenza virus strains. PMID:23720581

ViroSpot microneutralization assay for antigenic characterization of human influenza viruses.

PubMed

van Baalen, Carel A; Jeeninga, Rienk E; Penders, Germaine H W M; van Gent, Brenda; van Beek, Ruud; Koopmans, Marion P G; Rimmelzwaan, Guus F

2017-01-03

The hemagglutination inhibition (HI) assay has been used for the antigenic characterization of influenza viruses for decades. However, the majority of recent seasonal influenza A viruses of the H3N2 subtype has lost the capacity to agglutinate erythrocytes of various species. The hemagglutination (HA) activity of other A(H3N2) strains is generally sensitive to the action of the neuraminidase inhibitor oseltamivir, which indicates that the neuraminidase and not the hemagglutinin is responsible for the HA activity. These findings complicate the antigenic characterization and selection of A(H3N2) vaccine strains, calling for alternative antigenic characterization assays. Here we describe the development and use of the ViroSpot microneutralization (MN) assay as a reliable and robust alternative for the HI assay. Serum neutralization of influenza A(H3N2) reference virus strains and epidemic isolates was determined by automated readout of immunostained cell monolayers, in a format designed to minimize the influence of infectious virus doses on serum neutralization titers. Neutralization of infection was largely independent from rates of viral replication and cell-to-cell transmission, facilitating the comparison of different virus isolates. Other advantages of the ViroSpot MN assay include its relative insensitivity to variation in test dose of infectious virus, automated capture and analyses of residual infection patterns, and compatibility with standardized large scale analyses. Using this assay, a number of epidemic influenza A(H3N2) strains that failed to agglutinate erythrocytes, were readily characterized antigenically. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Effect of Shock Waves Generated by Pulsed Electric Discharges in Water on Yeast Cells and Virus Particles

NASA Astrophysics Data System (ADS)

Girdyuk, A. E.; Gorshkov, A. N.; Egorov, V. V.; Kolikov, V. A.; Snetov, V. N.; Shneerson, G. A.

2018-02-01

The aim of this study is to determine the optimal parameters of the electric pulses and shock waves generated by them for the soft destruction of the virus and yeast envelopes with no changes in the structure of antigenic surface albumin and in the cell morphology in order to use them to produce antivirus vaccines and in biotechnology. The pulse electric discharges in water have been studied for different values of amplitude, pulse duration and the rate of the rise in the current. A mathematical model has been developed to estimate the optimal parameters of pulsed electric charges and shock waves for the complete destruction of the yeast cell envelopes and virus particles at a minimum of pulses.

Genetic and Antigenic Typing of Seasonal Influenza Virus Breakthrough Cases from a 2008-2009 Vaccine Efficacy Trial

PubMed Central

Durviaux, Serge; Treanor, John; Beran, Jiri; Duval, Xavier; Esen, Meral; Feldman, Gregory; Frey, Sharon E.; Launay, Odile; Leroux-Roels, Geert; McElhaney, Janet E.; Nowakowski, Andrzej; Ruiz-Palacios, Guillermo M.; van Essen, Gerrit A.; Oostvogels, Lidia; Devaster, Jeanne-Marie

2014-01-01

Estimations of the effectiveness of vaccines against seasonal influenza virus are guided by comparisons of the antigenicities between influenza virus isolates from clinical breakthrough cases with strains included in a vaccine. This study examined whether the prediction of antigenicity using a sequence analysis of the hemagglutinin (HA) gene-encoded HA1 domain is a simpler alternative to using the conventional hemagglutination inhibition (HI) assay, which requires influenza virus culturing. Specimens were taken from breakthrough cases that occurred in a trivalent influenza virus vaccine efficacy trial involving >43,000 participants during the 2008-2009 season. A total of 498 influenza viruses were successfully subtyped as A(H3N2) (380 viruses), A(H1N1) (29 viruses), B(Yamagata) (23 viruses), and B(Victoria) (66 viruses) from 603 PCR- or culture-confirmed specimens. Unlike the B strains, most A(H3N2) (377 viruses) and all A(H1N1) viruses were classified as homologous to the respective vaccine strains based on their HA1 domain nucleic acid sequence. HI titers relative to the respective vaccine strains and PCR subtyping were determined for 48% (182/380) of A(H3N2) and 86% (25/29) of A(H1N1) viruses. Eighty-four percent of the A(H3N2) and A(H1N1) viruses classified as homologous by sequence were matched to the respective vaccine strains by HI testing. However, these homologous A(H3N2) and A(H1N1) viruses displayed a wide range of relative HI titers. Therefore, although PCR is a sensitive diagnostic method for confirming influenza virus cases, HA1 sequence analysis appeared to be of limited value in accurately predicting antigenicity; hence, it may be inappropriate to classify clinical specimens as homologous or heterologous to the vaccine strain for estimating vaccine efficacy in a prospective clinical trial. PMID:24371255

Differentiation of West Nile and St. Louis Encephalitis Virus Infections by Use of Noninfectious Virus-Like Particles with Reduced Cross-Reactivity▿ †

PubMed Central

Roberson, Jill A.; Crill, Wayne D.; Chang, Gwong-Jen J.

2007-01-01

Differential diagnosis of St. Louis encephalitis virus (SLEV) and West Nile virus (WNV) infections can be complicated due to the high degree of cross-reactivity observed in most serodiagnostic assays. In an effort to provide a more specific diagnostic test, we developed virus-like particle (VLP) antigens with reduced cross-reactivity for both SLEV and WNV by identifying and mutating envelope protein amino acids within the cross-reactive epitopes of VLP expression plasmids. To determine the serodiagnostic discriminatory ability of the antigens with reduced cross-reactivity, a panel of 134 human serum samples collected predominately from North American patients with SLEV or WNV infections was used to evaluate the performance of these novel antigens in imunoglobulin M antibody-capture enzyme-linked immunosorbent assays. Positive/negative ratios and the resulting diagnostic classifications were compared between the mutant and the wild-type (WT) VLPs. The mutant VLP antigens were more specific, with higher positive predictive values and higher likelihood ratios than the WT VLP antigens. Both the SLEV and WNV mutant VLPs greatly reduced the observed cross-reactivity, significantly increasing the specificity and sensitivity of the assay. The use of these novel VLP antigens with reduced cross-reactivity in these serodiagnostic assays and others should lead to more accurate diagnoses of current infections, thereby reducing the need for time-consuming and cumbersome confirmatory plaque-reduction neutralization tests to differentiate between SLEV and WNV infections in North America. PMID:17715375

Microbially synthesized modular virus-like particles and capsomeres displaying group A streptococcus hypervariable antigenic determinants.

PubMed

Chuan, Yap P; Wibowo, Nani; Connors, Natalie K; Wu, Yang; Hughes, Fiona K; Batzloff, Michael R; Lua, Linda H L; Middelberg, Anton P J

2014-06-01

Effective and low-cost vaccines are essential to control severe group A streptococcus (GAS) infections prevalent in low-income nations and the Australian aboriginal communities. Highly diverse and endemic circulating GAS strains mandate broad-coverage and customized vaccines. This study describes an approach to deliver cross-reactive antigens from endemic GAS strains using modular virus-like particle (VLP) and capsomere systems. The antigens studied were three heterologous N-terminal peptides (GAS1, GAS2, and GAS3) from the GAS surface M-protein that are specific to endemic strains in Australia Northern Territory Aboriginal communities. In vivo data presented here demonstrated salient characteristics of the modular delivery systems in the context of GAS vaccine design. First, the antigenic peptides, when delivered by unadjuvanted modular VLPs or adjuvanted capsomeres, induced high titers of peptide-specific IgG antibodies (over 1 × 10(4) ). Second, delivery by capsomere was superior to VLP for one of the peptides investigated (GAS3), demonstrating that the delivery system relative effectiveness was antigen-dependant. Third, significant cross-reactivity of GAS2-induced IgG with GAS1 was observed using either VLP or capsomere, showing the possibility of broad-coverage vaccine design using these delivery systems and cross-reactive antigens. Fourth, a formulation containing three pre-mixed modular VLPs, each at a low dose of 5 μg (corresponding to <600 ng of each GAS peptide), induced significant titers of IgGs specific to each peptide, demonstrating that a multivalent, broad-coverage VLP vaccine formulation was possible. In summary, the modular VLPs and capsomeres reported here demonstrate, with promising preliminary data, innovative ways to design GAS vaccines using VLP and capsomere delivery systems amenable to microbial synthesis, potentially adoptable by developing countries. © 2013 Wiley Periodicals, Inc.

Effects of dendritic cells from hepatitis B virus transgenic mice-stimulated autologous lymphocytes on hepatitis B virus replication: a study on the impact of specific sensitized effector cells on in vitro virus replication.

PubMed

Shen, Zhong-Yang; Zheng, Wei-Ping; Liu, Tao; Yang, Yang; Song, Hong-Li

2015-03-01

The objective of this study was to explore the effects of dendritic cells (DCs) from hepatitis B virus (HBV) transgenic mice-stimulated autologous lymphocytes on in vitro HBV replication. DCs from HBV transgenic mice were induced to maturity by lipopolysaccharide, followed by incubation with hepatitis B surface antigen (HBsAg) and hepatitis B core antigen (HBcAg) in vitro. Mature DCs and autologous lymphocytes were co-stimulated to form specific sensitized immune effector cells (IEC), which were then co-cultured with the human hepatoma cell line HepG2.2.15. Changes in morphology and activity of hepatocytes were then observed, as well as analysis of changes in liver enzyme, and HBV DNA and inflammatory cytokine levels in the culture supernatant. Intracellular HBV DNA and covalently closed circular DNA (cccDNA) concentration were measured by real-time polymerase chain reaction. Co-stimulation by mature DCs and IEC showed no impact on the morphology and liver enzyme expression level of HepG2.2.15 cells, but the supernatant HBV DNA and intracellular HBV DNA and cccDNA levels decreased significantly compared with those cells co-cultured with immature DCs. Secretion of inflammatory cytokines in the supernatant showed that when HBV DNA was highly expressed, the concentration of IFN-γ and IL-2 decreased, while IL-10 increased. Contrastingly, when HBV DNA had low expression, the concentration of IFN-γ and IL-2 increased and IL-10 decreased. Co-stimulation of HBV-related antigen-induced mature DCs and autologous lymphocytes showed inhibitory effects on ex vivo HBV replication, and cytokines were suggested to mediate this effect.

Both Genome Segments Contribute to the Pathogenicity of Very Virulent Infectious Bursal Disease Virus

PubMed Central

Escaffre, Olivier; Le Nouën, Cyril; Amelot, Michel; Ambroggio, Xavier; Ogden, Kristen M.; Guionie, Olivier; Toquin, Didier; Müller, Hermann; Islam, Mohammed R.

2013-01-01

Infectious bursal disease virus (IBDV) causes an economically significant disease of chickens worldwide. Very virulent IBDV (vvIBDV) strains have emerged and induce as much as 60% mortality. The molecular basis for vvIBDV pathogenicity is not understood, and the relative contributions of the two genome segments, A and B, to this phenomenon are not known. Isolate 94432 has been shown previously to be genetically related to vvIBDVs but exhibits atypical antigenicity and does not cause mortality. Here the full-length genome of 94432 was determined, and a reverse genetics system was established. The molecular clone was rescued and exhibited the same antigenicity and reduced pathogenicity as isolate 94432. Genetically modified viruses derived from 94432, whose vvIBDV consensus nucleotide sequence was restored in segment A and/or B, were produced, and their pathogenicity was assessed in specific-pathogen-free chickens. We found that a valine (position 321) that modifies the most exposed part of the capsid protein VP2 critically modified the antigenicity and partially reduced the pathogenicity of 94432. However, a threonine (position 276) located in the finger domain of the virus polymerase (VP1) contributed even more significantly to attenuation. This threonine is partially exposed in a hydrophobic groove on the VP1 surface, suggesting possible interactions between VP1 and another, as yet unidentified molecule at this amino acid position. The restored vvIBDV-like pathogenicity was associated with increased replication and lesions in the thymus and spleen. These results demonstrate that both genome segments influence vvIBDV pathogenicity and may provide new targets for the attenuation of vvIBDVs. PMID:23269788

The detection of African horse sickness virus antigens and antibodies in young Equidae.

PubMed Central

Hamblin, C.; Anderson, E. C.; Mellor, P. S.; Graham, S. D.; Mertens, P. P.; Burroughs, J. N.

1992-01-01

Four ponies were each inoculated with a different serotype of African horse sickness virus (AHSV) which had been passaged through cell culture in order to achieve attenuation. Three of the ponies died suddenly after showing mild clinical signs, the fourth pony remained clinically normal and was killed at day 38. Infectious AHSV was isolated from blood samples collected at intervals from all four ponies. Positive antigen ELISA reactions were only observed with blood samples from two of the ponies on the two days preceding death. Specific AHSV antibodies were detected by ELISA in serum samples from the other two ponies although one eventually died. African horse sickness viral antigens were detected by ELISA in post-mortem tissue samples collected from all four ponies. No infectious virus could be detected in tissue samples taken post-mortem from the pony which survived African horse sickness (AHS) infection. In the event of a suspected outbreak of AHS it is recommended that sera and heparinized blood should be tested for specific antibodies and AHSV antigen respectively. When available, post-mortem tissues, including spleen, heart, lung and liver, should also be tested for AHSV antigen. Although the ELISA used for the detection of AHSV antigen is highly sensitive and specific, negative ELISA results should be confirmed by virus isolation attempts. PMID:1547837

Vaccinia virus decreases major histocompatibility complex (MHC) class II antigen presentation, T-cell priming, and peptide association with MHC class II

PubMed Central

Rehm, Kristina E; Connor, Ramsey F; Jones, Gwendolyn J B; Yimbu, Kenneth; Mannie, Mark D; Roper, Rachel L

2009-01-01

Vaccinia virus (VACV) is the current live virus vaccine used to protect humans against smallpox and monkeypox, but its use is contraindicated in several populations because of its virulence. It is therefore important to elucidate the immune evasion mechanisms of VACV. We found that VACV infection of antigen-presenting cells (APCs) significantly decreased major histocompatibility complex (MHC) II antigen presentation and decreased synthesis of 13 chemokines and cytokines, suggesting a potent viral mechanism for immune evasion. In these model systems, responding T cells were not directly affected by virus, indicating that VACV directly affects the APC. VACV significantly decreased nitric oxide production by peritoneal exudate cells and the RAW macrophage cell line in response to lipopolysaccharide (LPS) and interferon (IFN)-γ, decreased class II MHC expression on APCs, and induced apoptosis in macrophages and dendritic cells. However, VACV decreased antigen presentation by 1153 B cells without apparent apoptosis induction, indicating that VACV differentially affects B lymphocytes and other APCs. We show that the key mechanism of VACV inhibition of antigen presentation may be its reduction of antigenic peptide loaded into the cleft of MHC class II molecules. These data indicate that VACV evades the host immune response by impairing critical functions of the APC. PMID:20067538

Limited Antigenic Diversity in Contemporary H7 Avian-Origin Influenza A Viruses from North America

PubMed Central

Xu, Yifei; Bailey, Elizabeth; Spackman, Erica; Li, Tao; Wang, Hui; Long, Li-Ping; Baroch, John A.; Cunningham, Fred L.; Lin, Xiaoxu; Jarman, Richard G.; DeLiberto, Thomas J.; Wan, Xiu-Feng

2016-01-01

Subtype H7 avian–origin influenza A viruses (AIVs) have caused at least 500 confirmed human infections since 2003 and culling of >75 million birds in recent years. Here we antigenically and genetically characterized 93 AIV isolates from North America (85 from migratory waterfowl [1976–2010], 7 from domestic poultry [1971–2012], and 1 from a seal [1980]). The hemagglutinin gene of these H7 viruses are separated from those from Eurasia. Gradual accumulation of nucleotide and amino acid substitutions was observed in the hemagglutinin of H7 AIVs from waterfowl and domestic poultry. Genotype characterization suggested that H7 AIVs in wild birds form diverse and transient internal gene constellations. Serologic analyses showed that the 93 isolates cross-reacted with each other to different extents. Antigenic cartography showed that the average antigenic distance among them was 1.14 units (standard deviation [SD], 0.57 unit) and that antigenic diversity among the H7 isolates we tested was limited. Our results suggest that the continuous genetic evolution has not led to significant antigenic diversity for H7 AIVs from North America. These findings add to our understanding of the natural history of IAVs and will inform public health decision-making regarding the threat these viruses pose to humans and poultry. PMID:26858078

Human immunodeficiency virus coinfection with hepatitis B virus leads to a decrease in extracellular and intracellular hepatitis B antigen.

PubMed

Pan, Wei; Wu, Zuoqiao; Wu, Shuwen; Guo, Deyin; Gong, Xiaoyan; Po, Tien

2015-04-01

Chronic hepatitis B virus (HBV) infection could cause severe liver disease including cirrhosis, hepatocellular carcinoma, and end-stage liver failure in HIV-positive individuals. The available data from clinical studies suggest that HIV infection modulates the HBV-specific T cell response. However, the virological and molecular aspects of HIV-HBV coinfection are currently poorly understood due to the lack of appropriate model systems. In this study, the effect of HIV infection on the life cycle of HBV was explored using an in vitro model system. The present data show that the extracellular and intracellular hepatitis B surface antigen (HBsAg) and e antigen (HBeAg) decrease significantly in HepG2 cells cotransfected with HIV NL4-3 and pHBV1.3 as compared to those cells transfected only with pHBV1.3. Moreover, a significant decrease in HBV DNA and mRNA expression was also observed in the cotransfected cells. HIV Rev protein, an RNA-bound regulatory protein, could significantly decrease the expression levels of extracellular and intracellular HBsAg and HBeAg by mediating the expression of HBV mRNA in cells cotransfected with plasmids containing HIV-1 Rev and pHBV1.3. Further experiments demonstrate that HIV Rev manipulated neither the promoters of HBV nor the nuclear export of HBV mRNA. These results from the in vitro model system might provide clues to further understand the rapid progression of liver disease in HIV-HBV-coinfected patients.

Immune Escape Variants of H9N2 Influenza Viruses Containing Deletions at the Hemagglutinin Receptor Binding Site Retain Fitness In Vivo and Display Enhanced Zoonotic Characteristics.

PubMed

Peacock, Thomas P; Benton, Donald J; James, Joe; Sadeyen, Jean-Remy; Chang, Pengxiang; Sealy, Joshua E; Bryant, Juliet E; Martin, Stephen R; Shelton, Holly; Barclay, Wendy S; Iqbal, Munir

2017-07-15

H9N2 avian influenza viruses are enzootic in poultry across Asia and North Africa, where they pose a threat to human health as both zoonotic agents and potential pandemic candidates. Poultry vaccination against H9N2 viruses has been employed in many regions; however, vaccine effectiveness is frequently compromised due to antigenic drift arising from amino acid substitutions in the major influenza virus antigen hemagglutinin (HA). Using selection with HA-specific monoclonal antibodies, we previously identified H9N2 antibody escape mutants that contained deletions of amino acids in the 220 loop of the HA receptor binding sites (RBSs). Here we analyzed the impact of these deletions on virus zoonotic infection characteristics and fitness. We demonstrated that mutant viruses with RBS deletions are able to escape polyclonal antiserum binding and are able to infect and be transmitted between chickens. We showed that the deletion mutants have increased binding to human-like receptors and greater replication in primary human airway cells; however, the mutant HAs also displayed reduced pH and thermal stability. In summary, we infer that variant influenza viruses with deletions in the 220 loop could arise in the field due to immune selection pressure; however, due to reduced HA stability, we conclude that these viruses are unlikely to be transmitted from human to human by the airborne route, a prerequisite for pandemic emergence. Our findings underscore the complex interplay between antigenic drift and viral fitness for avian influenza viruses as well as the challenges of predicting which viral variants may pose the greatest threats for zoonotic and pandemic emergence. IMPORTANCE Avian influenza viruses, such as H9N2, cause disease in poultry as well as occasionally infecting humans and are therefore considered viruses with pandemic potential. Many countries have introduced vaccination of poultry to try to control the disease burden; however, influenza viruses are able to rapidly evolve to escape immune pressure in a process known as "antigenic drift." Previously, we experimentally generated antigenic-drift variants in the laboratory, and here, we test our "drifted" viruses to assess their zoonotic infection characteristics and transmissibility in chickens. We found that the drifted viruses were able to infect and be transmitted between chickens and showed increased binding to human-like receptors. However, the drift mutant viruses displayed reduced stability, and we predict that they are unlikely to be transmitted from human to human and cause an influenza pandemic. These results demonstrate the complex relationship between antigenic drift and the potential of avian influenza viruses to infect humans. Copyright © 2017 Peacock et al.

A tetravalent virus-like particle vaccine designed to display domain III of dengue envelope proteins induces multi-serotype neutralizing antibodies in mice and macaques which confer protection against antibody dependent enhancement in AG129 mice

PubMed Central

Shukla, Rahul; Poddar, Ankur; Shanmugam, Rajgokul K.; White, Laura J.; Mattocks, Melissa M.; Raut, Rajendra; Perween, Ashiya; Tyagi, Poornima; de Silva, Aravinda M.; Bhaumik, Siddhartha K.; Kaja, Murali Krishna; Villinger, François; Ahmed, Rafi; Johnston, Robert E.; Khanna, Navin

2018-01-01

Background Dengue is one of the fastest spreading vector-borne diseases, caused by four antigenically distinct dengue viruses (DENVs). Antibodies against DENVs are responsible for both protection as well as pathogenesis. A vaccine that is safe for and efficacious in all people irrespective of their age and domicile is still an unmet need. It is becoming increasingly apparent that vaccine design must eliminate epitopes implicated in the induction of infection-enhancing antibodies. Methodology/principal findings We report a Pichia pastoris-expressed dengue immunogen, DSV4, based on DENV envelope protein domain III (EDIII), which contains well-characterized serotype-specific and cross-reactive epitopes. In natural infection, <10% of the total neutralizing antibody response is EDIII-directed. Yet, this is a functionally relevant domain which interacts with the host cell surface receptor. DSV4 was designed by in-frame fusion of EDIII of all four DENV serotypes and hepatitis B surface (S) antigen and co-expressed with unfused S antigen to form mosaic virus-like particles (VLPs). These VLPs displayed EDIIIs of all four DENV serotypes based on probing with a battery of serotype-specific anti-EDIII monoclonal antibodies. The DSV4 VLPs were highly immunogenic, inducing potent and durable neutralizing antibodies against all four DENV serotypes encompassing multiple genotypes, in mice and macaques. DSV4-induced murine antibodies suppressed viremia in AG129 mice and conferred protection against lethal DENV-4 virus challenge. Further, neither murine nor macaque anti-DSV4 antibodies promoted mortality or inflammatory cytokine production when passively transferred and tested in an in vivo dengue disease enhancement model of AG129 mice. Conclusions/significance Directing the immune response to a non-immunodominant but functionally relevant serotype-specific dengue epitope of the four DENV serotypes, displayed on a VLP platform, can help minimize the risk of inducing disease-enhancing antibodies while eliciting effective tetravalent seroconversion. DSV4 has a significant potential to emerge as a safe, efficacious and inexpensive subunit dengue vaccine candidate. PMID:29309412

Detection of genome, antigen, and antibodies in oral fluids from pigs infected with foot-and-mouth disease virus.

PubMed

Senthilkumaran, Chandrika; Yang, Ming; Bittner, Hilary; Ambagala, Aruna; Lung, Oliver; Zimmerman, Jeffrey; Giménez-Lirola, Luis G; Nfon, Charles

2017-04-01

Virus nucleic acids and antibody response to pathogens can be measured using swine oral fluids (OFs). Detection of foot-and-mouth disease virus (FMDV) genome in swine OFs has previously been demonstrated. Virus isolation and viral antigen detection are additional confirmatory assays for diagnosing FMDV, but these methods have not been evaluated using swine OF. The objectives of this study were to further validate the molecular detection of FMDV in oral fluids, evaluate antigen detection and FMDV isolation from swine OFs, and develop an assay for isotypic anti-FMDV antibody detection in OFs. Ribonucleic acid (RNA) from FMDV was detected in OFs from experimentally infected pigs by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) from 1 day post-infection (dpi) to 21 dpi. Foot-and-mouth disease virus (FMDV) was isolated from OFs at 1 to 5 dpi. Additionally, FMDV antigens were detected in OFs from 1 to 6 dpi using a lateral flow immunochromatographic strip test (LFIST), which is a rapid pen-side test, and from 2 to 3 dpi using a double-antibody sandwich enzyme-linked immunosorbent assay (DAS ELISA). Furthermore, FMDV-specific immunoglobulin A (IgA) was detected in OFs using an isotype-specific indirect ELISA starting at dpi 14. These results further demonstrated the potential use of oral fluids for detecting FMDV genome, live virus, and viral antigens, as well as for quantifying mucosal IgA antibody response.

Relationship of envelope antigens of animal influenza viruses to human A2 influenza strains isolated in the years 1957-68*

PubMed Central

Tumova, Bela; Easterday, Bernard C.

1969-01-01

This study demonstrates relationships in envelope antigens of 4 human influenza A2 strains isolated during the period 1957-68 (including A2/Hong Kong/68), 2 strains of A/Equi-2/63 and 7 avian influenza viruses isolated in Europe, North America, and the Ukraine in the years 1960-67. Antigenic relationships among the strains were determined on the basis of haemagglutination-inhibition, virus-neutralization, strain-specific complement-fixation, and neuraminidase-inhibition tests. North American avian influenza strains, Turkey/California/64, Turkey/Massachusetts/65, Turkey/Wisconsin/66, Turkey/Ontario/6828/67, and the Italian strain Duck/Italy/574/66 are antigenically related to human A2 influenza viruses by haemagglutinin and/or neuraminidase. None of these viruses is antigenically related to the A/Equi-2/63 strains, Duck/Ukraine/2/60, Duck/Ukraine/1/63 or A2/Hong Kong/68. However, A2/Hong Kong/68 has neuraminidase similar to other A2 strains from previous years. A definite relationship was shown between the haemagglutinin of A/Equi-2/63, A2/Hong Kong/68, Duck/Ukraine/2/60 and Duck/Ukraine/1/63 strains by the use of hyperimmune sera in 3 different serological tests. Related neuraminidase was demonstrated only between A/Equi-2/63 and both duck strains from the Ukraine. The significance of these findings and their interpretation with respect to the ecology of influenza viruses are discussed. PMID:5309452

Elicitation of anti-Sendai virus cytotoxic T lymphocytes by viral and H-2 antigens incorporated into the same lipid bilayer by membrane fusion and by reconstitution into liposomes.

PubMed

Hale, A H; Lyles, D S; Fan, D P

1980-02-01

We have investigated the minimal molecular requirements for elicitation of anti-Sendai virus cytotoxic T lymphocytes (CTL), and the minimal molecular requirements for the recognition and lysis processes associated with anti-Sendai virus CTL-target cell interactions. This report demonstrates a) that the hemagglutinin-neuraminidase and/or fusion glycoproteins of Sendai virus can elicit anti-Sendai virus CTL and b) that these glycoproteins and H-2 antigens must be within the same membrane lipid bilayer for effective elicitation of anti-Sendai-virus CTL and for effective recognition and lysis of target cells by anti-Sendai virus CTL.

Mutations in the S gene region of hepatitis B virus genotype D in Turkish patients.

PubMed

Ozaslan, Mehmet; Ozaslan, Ersan; Barsgan, Arzu; Koruk, Mehmet

2007-12-01

The S gene region of the hepatitis B virus (HBV) is responsible for the expression of surface antigens and includes the 'a'-determinant region. Thus, mutation(s) in this region would afford HBV variants a distinct survival advantage, permitting the mutant virus to escape from the immune system. The aim of this study was to search for mutations of the S gene region in different patient groups infected with genotype D variants of HBV, and to analyse the biological significance of these mutations. Moreover, we investigated S gene mutation inductance among family members. Forty HBV-DNA-positive patients were determined among 132 hepatitis B surface antigen (HbsAg) carriers by the first stage of seminested PCR. Genotypes and subtypes were established by sequencing of the amplified S gene regions. Variants were compared with original sequences of these serotypes, and mutations were identified. All variants were designated as genotype D and subtype ayw3. Ten kinds of point mutations were identified within the S region. The highest rates of mutation were found in chronic hepatitis patients and their family members. The amino acid mutations 125 (M -> T) and 127 (T -> P) were found on the first loop of 'a'-determinant. The other consequence was mutation inductance in a family member. We found some mutations in the S gene region known to be stable and observed that some of these mutations affected S gene expression.

Development of electrochemical immunosensors based on different serum antibody immobilization methods for detection of Japanese encephalitis virus

NASA Astrophysics Data System (ADS)

Tran, Quang Huy; Hanh Nguyen, Thi Hong; Mai, Anh Tuan; Thuy Nguyen, Thi; Khue Vu, Quang; Nga Phan, Thi

2012-03-01

This paper describes the development of electrochemical immunosensors based on human serum antibodies with different immobilization methods for detection of Japanese encephalitis virus (JEV). Human serum containing anti-JEV antibodies was used to immobilize onto the surface of silanized interdigitated electrodes by four methods: direct adsorption (APTES-serum), covalent binding with a cross linker of glutaraldehyde (APTES-GA-serum), covalent binding with a cross linker of glutaraldehyde combined with anti-human IgG (APTES-GA-anti-HIgG-serum) and covalent binding with a cross linker of glutaraldehyde combined with a bioaffinity of protein A (APTES-GA-PrA-serum). Atomic force microscopy was used to verify surface characteristics of the interdigitated electrodes before and after treatment with serum antibodies. The output signal of the immunosensors was measured by the change of conductivity resulting from the specific binding of JEV antigens and serum antibodies immobilized on the electrodes, with the help of horseradish peroxidase (HRP)-labeled secondary antibody against JEV. The results showed that the APTES-GA-PrA-serum method provided the highest signal of the electrochemical immunosensor for detection of JEV antigens, with the linear range from 25 ng ml-1 to 1 μg ml-1, and the limit of detection was about 10 ng ml-1. This study shows a potential development of novel electrochemical immunosensors applied for virus detection in clinical samples in case of possible outbreaks.

In vivo analysis of replication and immunogenicity of proviral clones of human T-lymphotropic virus type 1 with selective envelope surface-unit mutations

PubMed Central

Silverman, Lee R.; Phipps, Andrew J.; Montgomery, Andy; Fernandez, Soledad; Tsukahara, Tomonori; Ratner, Lee; Lairmore, Michael D.

2005-01-01

Human T-cell leukemia virus type 1 (HTLV-1) is the causative agent of adult T-cell lymphoma/leukemia (ATL). The HTLV-1 envelope gene exhibits limited variability when examined from infected individuals, but has not been tested using infectious clones of the virus in animal models. In vitro assays indicate that HTLV-1 envelope (Env) Ser75Ile, Asn95Asp, and Asn195Asp surface unit (SU) mutants are able to replicate in and immortalize lymphocytes. Herein, we examined the effects of these Env mutants in rabbits inoculated with HTLV-1 immortalized ACH.75, ACH.95, or ACH.195 cell lines (expressing full-length molecular clones with the SU mutations) or the ACH.1 cell line (expressing wild-type SU). All rabbits became infected, and the fidelity of the mutations was maintained throughout the 8-week study. However, SU point mutations resulted in decreased antibody responses to viral group-associated antigen (Gag) and Env antigens. ACH.195 rabbits had a selective decreased antibody response to SU, and one ACH.195 rabbit had an antibody response to both HTLV-1 and HTLV-2 SUs. Some mutant inoculation groups had altered proviral loads. However, peripheral-blood mononuclear cell (PBMC) proviral loads did not correlate with antibody responses. Our data are the first to demonstrate that mutations in critical determinants of HTLV-1 Env SU altered antibody responses and proviral loads, but do not prevent viral replication in vivo. PMID:16046523

ANTIGENIC VARIANTS OF INFLUENZA A VIRUS (PR8 STRAIN)

PubMed Central

Hamre, Dorothy; Loosli, Clayton G.; Gerber, Paul

1958-01-01

Seven variant strains of influenza A PR8-S virus, each derived from the previous one by serial passage in the lungs of mice immunized with the homologous agent have been produced. With the H.I. and neutralization procedures these variants showed a progressive serological deviation from the parent PR8-S virus. The seven variants provoked antibodies in varying titers to the preceding variants and the parent virus but not in relation to their position in the series. Thus, the seventh variant provoked significantly more antibody to the PR8-S virus than did the fifth variant. A possible explanation for this is presented. The first four variant viruses showed progressively less ability to react with antisera of the preceding variants and the PR8-S virus, and the three most recently derived variants showed essentially no ability to react with PR8-S and first variant antisera. The variant viruses remained antigenically stable through numerous lung passages in normal mice. Cross absorption tests revealed common antigenic components among the variant viruses and also individual characteristics which classify them as being different from one another. The implications of these findings in relation to studies by others have been discussed. PMID:13539308

Assessment of pathogenicity and antigenicity of American lineage influenza H5N2 viruses in Taiwan.

PubMed

Lin, Chun-Yang; Chia, Min-Yuan; Chen, Po-Ling; Yeh, Chia-Tsui; Cheng, Ming-Chu; Su, Ih-Jen; Lee, Min-Shi

2017-08-01

During December 2003 and March 2004, large scale epidemics of low-pathogenic avian influenza (LPAI) H5N2 occurred in poultry farms in central and southern Taiwan. Based on genomic analysis, these H5N2 viruses contain HA and NA genes of American-lineage H5N2 viruses and six internal genes from avian influenza A/H6N1 viruses endemic in poultry in Taiwan. After disappearing for several years, these novel influenza H5N2 viruses caused outbreaks in poultry farms again in 2008, 2010 and 2012, and have evolved into high pathogenic AI (HPAI) since 2010. Moreover, asymptomatic infections of influenza H5N2 were detected serologically in poultry workers in 2012. Therefore, we evaluated antigenicity and pathogenicity of the novel H5N2 viruses in ferrets. We found that no significant antigenic difference was detected among the novel H5N2 viruses isolated from 2003 to 2014 and the novel H5N2 viruses could cause mild infections in ferrets. Monitoring zoonotic transmission of the novel H5N2 viruses is necessary. Copyright © 2017 Elsevier Inc. All rights reserved.

Simultaneous surface display and cargo loading of encapsulin nanocompartments and their use for rational vaccine design.

PubMed

Lagoutte, Priscillia; Mignon, Charlotte; Stadthagen, Gustavo; Potisopon, Supanee; Donnat, Stéphanie; Mast, Jan; Lugari, Adrien; Werle, Bettina

2018-05-11

In the past decades protein nanoparticles have successfully been used for vaccine applications. Their particulate nature and dense repetitive subunit organization makes them perfect carriers for antigen surface display and confers high immunogenicity. Nanoparticles have emerged as excellent candidates for vectorization of biological and immunostimulating molecules. Nanoparticles and biomolecular nanostructures such as ferritins or virus like particles have been used as diagnostic and therapeutic delivery systems, in vaccine development, as nanoreactors, etc. Recently, a new class of bacterial protein compartment has been discovered referred to as encapsulin nanocompartment. These compartments have been used for targeted diagnostics, as therapeutic delivery systems and as nanoreactors. Their biological origin makes them conveniently biocompatible and allows genetic functionalization. The aim of our study was to implement encapsulin nanocompartements for simultaneous epitope surface display and heterologous protein loading for rational vaccine design. For this proof-of-concept-study, we produced Thermotoga maritima encapsulin nanoparticles in E. coli. We demonstrated the ability of simultaneous display in our system by inserting Matrix protein 2 ectodomain (M2e) of influenza A virus at the nanoparticle surface and by packaging of a fluorescent reporter protein (GFP) into the internal cavity. Characterization of the nanoparticles by electronic microscopy confirmed homogenously shaped particles of 24 nm diameter in average. The results further show that engineering of the particle surface improved the loading capacity of the heterologous reporter protein suggesting that surface display may induce a critical elastic deformation resulting in improved stiffness. In Balb/c mice, nanoparticle immunization elicited antibody responses against both the surface epitope and the loaded cargo protein. These results confirm the potential of encapsulin nanocompartments for customized vaccine design and antigen delivery. Copyright © 2018 Elsevier Ltd. All rights reserved.

Potent neutralizing monoclonal antibodies against Ebola virus infection

PubMed Central

Zhang, Qi; Gui, Miao; Niu, Xuefeng; He, Shihua; Wang, Ruoke; Feng, Yupeng; Kroeker, Andrea; Zuo, Yanan; Wang, Hua; Wang, Ying; Li, Jiade; Li, Chufang; Shi, Yi; Shi, Xuanling; Gao, George F.; Xiang, Ye; Qiu, Xiangguo; Chen, Ling; Zhang, Linqi

2016-01-01

Ebola virus infections cause a deadly hemorrhagic disease for which no vaccines or therapeutics has received regulatory approval. Here we show isolation of three (Q206, Q314 and Q411) neutralizing monoclonal antibodies (mAbs) against the surface glycoprotein (GP) of Ebola virus identified in West Africa in 2014 through sequential immunization of Chinese rhesus macaques and antigen-specific single B cell sorting. These mAbs demonstrated potent neutralizing activities against both pseudo and live Ebola virus independent of complement. Biochemical, single particle EM, and mutagenesis analysis suggested Q206 and Q411 recognized novel epitopes in the head while Q314 targeted the glycan cap in the GP1 subunit. Q206 and Q411 appeared to influence GP binding to its receptor NPC1. Treatment with these mAbs provided partial but significant protection against disease in a mouse model of Ebola virus infection. These novel mAbs could serve as promising candidates for prophylactic and therapeutic interventions against Ebola virus infection. PMID:27181584

Potent neutralizing monoclonal antibodies against Ebola virus infection.

PubMed

Zhang, Qi; Gui, Miao; Niu, Xuefeng; He, Shihua; Wang, Ruoke; Feng, Yupeng; Kroeker, Andrea; Zuo, Yanan; Wang, Hua; Wang, Ying; Li, Jiade; Li, Chufang; Shi, Yi; Shi, Xuanling; Gao, George F; Xiang, Ye; Qiu, Xiangguo; Chen, Ling; Zhang, Linqi

2016-05-16

Ebola virus infections cause a deadly hemorrhagic disease for which no vaccines or therapeutics has received regulatory approval. Here we show isolation of three (Q206, Q314 and Q411) neutralizing monoclonal antibodies (mAbs) against the surface glycoprotein (GP) of Ebola virus identified in West Africa in 2014 through sequential immunization of Chinese rhesus macaques and antigen-specific single B cell sorting. These mAbs demonstrated potent neutralizing activities against both pseudo and live Ebola virus independent of complement. Biochemical, single particle EM, and mutagenesis analysis suggested Q206 and Q411 recognized novel epitopes in the head while Q314 targeted the glycan cap in the GP1 subunit. Q206 and Q411 appeared to influence GP binding to its receptor NPC1. Treatment with these mAbs provided partial but significant protection against disease in a mouse model of Ebola virus infection. These novel mAbs could serve as promising candidates for prophylactic and therapeutic interventions against Ebola virus infection.

Wesselsbron virus antibody in domestic animals in Nigeria: retrospective and prospective studies.

PubMed

Baba, S S; Fagbami, A H; Ojeh, C K; Olaleye, O D; Omilabu, S A

1995-04-01

Retrospective and prospective serological surveys to determine the prevalence of Wesslsbron (WSL) virus infections in animal populations were carried out in different vegetational zones in Nigeria. Sera from 1,492 animals comprising 292 camels, 81 horses, 4 donkeys, 320 cattle, 235 sheep, 260 goats, 114 pigs, 101 dogs and 85 domestic fowls were assayed by haemagglutination-inhibition (HI) test for presence of antibodies to WSL virus and other flavivirus antigens: Yellow Fever (YF), Potiskum (POT), Banzi (BAN), Uganda S (UGS) and West Nile (WN) viruses. Four hundred and eighty one (32%) of the total sera tested were positive for the presence of flavivirus antibodies. The prevalence rates among animals varied with species and vegetational zones of the country. The highest prevalence was noted in animals from a swamp forest zone and was higher among camels, horses, donkeys and sheep when compared with goats, pigs and fowls in different zones. Although monotypic reactions with WSL virus antigen were observed in positive sera, the majority of the WSL virus positive sera cross-reacted with more than two other flavivirus antigens. Serological cross-reactions were most extensive in WSL virus positive horse sera. A ten month sentinel survey among 28 cattle, 68 sheep and 30 goats revealed considerable activity of WSL virus in Nigeria. Of these, 11 cattle and 12 sheep showed antibody conversion to WSL virus antigen. None of the goats seroconverted. Although, there are no records of outbreak of WSL disease in Nigeria, this study revealed that WSL virus is actively circulating among livestock populations in this environment. Flavivirus nucleotide data are needed for final determination of genetic relatedness in this group of viruses.

WHO recommendations for the viruses used in the 2013-2014 Northern Hemisphere influenza vaccine: Epidemiology, antigenic and genetic characteristics of influenza A(H1N1)pdm09, A(H3N2) and B influenza viruses collected from October 2012 to January 2013.

PubMed

Barr, Ian G; Russell, Colin; Besselaar, Terry G; Cox, Nancy J; Daniels, Rod S; Donis, Ruben; Engelhardt, Othmar G; Grohmann, Gary; Itamura, Shigeyuki; Kelso, Anne; McCauley, John; Odagiri, Takato; Schultz-Cherry, Stacey; Shu, Yuelong; Smith, Derek; Tashiro, Masato; Wang, Dayan; Webby, Richard; Xu, Xiyan; Ye, Zhiping; Zhang, Wenqing

2014-08-20

In February the World Health Organisation (WHO) recommends influenza viruses to be included in influenza vaccines for the forthcoming winter in the Northern Hemisphere. These recommendations are based on data collected by National Influenza Centres (NICs) through the WHO Global Influenza Surveillance and Response System (GISRS) and a more detailed analysis of representative and potential antigenically variant influenza viruses from the WHO Collaborating Centres for Influenza (WHO CCs) and Essential Regulatory Laboratories (ERLs). This article provides a detailed summary of the antigenic and genetic properties of viruses and additional background data used by WHO experts during development of the recommendations of the 2013-2014 Northern Hemisphere influenza vaccine composition. Published by Elsevier Ltd.

Update on occult hepatitis B virus infection

PubMed Central

Makvandi, Manoochehr

2016-01-01

The event of mutations in the surface antigen gene of hepatitis B virus (HBV) results in undetectable hepatitis B surface antigen with positive/negative anti-hepatitis B core (anti-HBc) antibody status in serum and this phenomenon is named occult hepatitis B infection (OBI). The presence of anti-HBc antibody in serum is an important key for OBI tracking, although about 20% of OBI cases are negative for anti-HBc antibody. The diagnosis of OBI is mainly based on polymerase chain reaction (PCR) and real-time PCR assays. However, real-time PCR is a more reliable method than PCR. OBI is a great issue for the public health problem and a challenge for the clinical entity worldwide. The persistence of OBI may lead to the development of cirrhosis and hepatocellular carcinoma. With regard to OBI complications, the screening of HBV DNA by the highly sensitive molecular means should be implemented for: (1) patients with a previous history of chronic or acute HBV infection; (2) patients co-infected with hepatitis C virus/human immunodeficiency virus; (3) patients undergoing chemotherapy or anti-CD20 therapy; (4) recipients of organ transplant; (5) blood donors; (6) organ transplant donors; (7) thalassemia and hemophilia patients; (8) health care workers; (9) patients with liver related disease (cryptogenic); (10) hemodialysis patients; (11) patients undergoing lamivudine or interferon therapy; and (12) children in time of HBV vaccination especially in highly endemic areas of HBV. Active HBV vaccination should be implemented for the close relatives of patients who are negative for OBI markers. Thus, the goal of this review is to evaluate the rate of OBI with a focus on status of high risk groups in different regions of the world. PMID:27818588

Epitope Mapping of Avian Influenza M2e Protein: Different Species Recognise Various Epitopes

PubMed Central

Hasan, Noor Haliza; Ignjatovic, Jagoda; Tarigan, Simson; Peaston, Anne; Hemmatzadeh, Farhid

2016-01-01

A common approach for developing diagnostic tests for influenza virus detection is the use of mouse or rabbit monoclonal and/or polyclonal antibodies against a target antigen of the virus. However, comparative mapping of the target antigen using antibodies from different animal sources has not been evaluated before. This is important because identification of antigenic determinants of the target antigen in different species plays a central role to ensure the efficiency of a diagnostic test, such as competitive ELISA or immunohistochemistry-based tests. Interest in the matrix 2 ectodomain (M2e) protein of avian influenza virus (AIV) as a candidate for a universal vaccine and also as a marker for detection of virus infection in vaccinated animals (DIVA) is the rationale for the selection of this protein for comparative mapping evaluation. This study aimed to map the epitopes of the M2e protein of avian influenza virus H5N1 using chicken, mouse and rabbit monoclonal or monospecific antibodies. Our findings revealed that rabbit antibodies (rAbs) recognized epitope 6EVETPTRN13 of the M2e, located at the N-terminal of the protein, while mouse (mAb) and chicken antibodies (cAbs) recognized epitope 10PTRNEWECK18, located at the centre region of the protein. The findings highlighted the difference between the M2e antigenic determinants recognized by different species that emphasized the importance of comparative mapping of antibody reactivity from different animals to the same antigen, especially in the case of multi-host infectious agents such as influenza. The findings are of importance for antigenic mapping, as well as diagnostic test and vaccine development. PMID:27362795

[The phenomenon of antigenic defectiveness in naturally circulating strains of the tick-borne encephalitis virus and its possible connection to seronegative forms of the disease].

PubMed

Pogodina, V V; Bochkova, N G; Dzhivanian, T I; Levina, L S; Karganova, G G; Riasova, R A; Sergeeva, V A; Lashkevich, V A

1992-01-01

Ten strains of tick-borne encephalitis (TBE) virus isolated from single specimens of I. persulcatus ticks were studied. The strains were divided into antigenically complete (AC) and antigenically defective (AD), depending on the presence or absence of some virus antigens in concentrated virus preparations, characteristics in rocket immune electrophoresis (RIEP), rate and intensity of humoral immune response in monkeys and rabbits, and plaque size in SPEV cell culture. The AC-strain markers include high activities of precipitating, hemagglutinating (HA), and complement-fixing (CF) antigens, formation of precipitates moving in rocket shape towards anode and cathode in RIEP, rapid development of antihemagglutinins and virus-neutralizing antibodies, large plaques (3-5 mm). The AD variants are characterized by the lack of HA and precipitating activity, low titres of CF antigen, slow and poor immune response, the lack of cathode precipitate "rocket", very small plaques. The antigenic defectiveness is transitory and shows in early passages; after 10-11 passages in SPEV cell cultures or in white mice, transformation AD----AC occurs. A transformed strain is neutralized, like standard TBE strains, by blood sera of a typical patient with poliomyelitis-like form of TBE. Examinations of blood sera from the population of an endemic zone (Yaroslavl Province) and 67 TBE patients (Kurgan Province) demonstrated the association of AC and AD variants with the formation of immune portion of the population and TBE etiology. Cases of the disease confirmed by seroconversion in HI with commercial diagnosticum are associated with AC variants, whereas AD variants are associated with those TBE cases which are difficult to diagnose using the commercial diagnosticum.

Mutation analysis of the fusion domain region of St. Louis encephalitis virus envelope protein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Trainor, Nicole B.; Crill, Wayne D.; Roberson, Jill A.

2007-04-10

The immune response to flavivirus infections produces both species-specific and flavivirus cross-reactive antibodies. The presence of cross-reactive antibodies complicates serodiagnosis of flavivirus infections, especially secondary infections caused by a heterologous virus. A successful public health response to the growing global threat posed by flaviviruses necessitates the development of virus-specific diagnostic antigens. The flavivirus envelope (E) glycoprotein is the principle antigen stimulating protective immunity during infection. Using recombinant St. Louis encephalitis virus-like particles (VLPs), we have identified amino acid residues involved in flavivirus cross-reactive epitope determinants. Most significant among the residues studied are three highly conserved amino acids in the fusionmore » peptide: Gly104, Gly106, and Leu107. Substitutions of these residues dramatically influenced VLP secretion and cross-reactive monoclonal antibody reactivity. These results provide critical insight into the antigenic structure of the flaviviral E protein and toward development of species-specific diagnostic antigens that should improve both flavivirus diagnosis and estimates of disease burden.« less

Breadth of neutralizing antibodies elicited by stable, homogeneous clade A and clade C HIV-1 gp140 envelope trimers in guinea pigs.

PubMed

Nkolola, Joseph P; Peng, Hanqin; Settembre, Ethan C; Freeman, Michael; Grandpre, Lauren E; Devoy, Colleen; Lynch, Diana M; La Porte, Annalena; Simmons, Nathaniel L; Bradley, Ritu; Montefiori, David C; Seaman, Michael S; Chen, Bing; Barouch, Dan H

2010-04-01

The native envelope (Env) spike on the surface of human immunodeficiency virus type 1 (HIV-1) is trimeric, and thus trimeric Env vaccine immunogens are currently being explored in preclinical immunogenicity studies. Key challenges have included the production and purification of biochemically homogeneous and stable trimers and the evaluation of these immunogens utilizing standardized virus panels for neutralization assays. Here we report the binding and neutralizing antibody (NAb) responses elicited by clade A (92UG037.8) and clade C (CZA97.012) Env gp140 trimer immunogens in guinea pigs. These trimers have been selected and engineered for optimal biochemical stability and have defined antigenic properties. Purified gp140 trimers with Ribi adjuvant elicited potent, cross-clade NAb responses against tier 1 viruses as well as detectable but low-titer NAb responses against select tier 2 viruses from clades A, B, and C. In particular, the clade C trimer elicited NAbs that neutralized 27%, 20%, and 47% of tier 2 viruses from clades A, B, and C, respectively. Heterologous DNA prime, protein boost as well as DNA prime, recombinant adenovirus boost regimens expressing these antigens, however, did not result in an increased magnitude or breadth of NAb responses in this system. These data demonstrate the immunogenicity of stable, homogeneous clade A and clade C gp140 trimers and exemplify the utility of standardized tier 1 and tier 2 virus panels for assessing the NAb responses of candidate HIV-1 Env immunogens.

Development of a highly sensitive bioluminescent enzyme immunoassay for hepatitis B virus surface antigen capable of detecting divergent mutants.

PubMed

Minekawa, Takayuki; Takehara, Shizuka; Takahashi, Masaharu; Okamoto, Hiroaki

2013-08-01

Hepatitis B virus (HBV) infections are sometimes overlooked when using commercial kits to measure hepatitis B virus surface antigen (HBsAg) due to their low sensitivities and reactivities to mutant strains of various genotypes. We developed an ultrasensitive bioluminescent enzyme immunoassay (BLEIA) for HBsAg using firefly luciferase, which is adaptable to a variety of HBsAg mutants, by combining four monoclonal antibodies with a polyclonal antibody against HBsAg. The measurement of seroconversion panels showed trace amounts of HBsAg during the early infection phase by the BLEIA because of its high sensitivity of 5 mIU/ml. The BLEIA detected HBsAg as early as did PCR in five of seven series and from 2.1 to 9.4 days earlier than commercial immunoassay methods. During the late infection phase, the BLEIA successfully detected HBsAg even 40 days after the disappearance of HBV DNA and the emergence of antibodies against HBsAg. The HBsAg BLEIA successfully detected all 13 recombinant HBsAg and 45 types of HBsAg mutants with various mutations within amino acids 90 to 164 in the S gene product. Some specimens had higher values determined by the BLEIA than those by a commercial chemiluminescent immunoassay; this suggests that such discrepancies were caused by the dissociation of preS1/preS2 peptides from the particle surface. With its highly sensitive detection of low-titer HBsAg, including various mutants, the HBsAg BLEIA is considered to be useful for the early diagnosis and prevention of HBV infection because of the shorter window of infection prior to detection, which facilitates early prediction of recurrence in HBV-infected individuals.

Dengue virus type 2: replication and tropisms in orally infected Aedes aegypti mosquitoes

PubMed Central

Salazar, Ma Isabel; Richardson, Jason H; Sánchez-Vargas, Irma; Olson, Ken E; Beaty, Barry J

2007-01-01

Background To be transmitted by its mosquito vector, dengue virus (DENV) must infect midgut epithelial cells, replicate and disseminate into the hemocoel, and finally infect the salivary glands, which is essential for transmission. The extrinsic incubation period (EIP) is very relevant epidemiologically and is the time required from the ingestion of virus until it can be transmitted to the next vertebrate host. The EIP is conditioned by the kinetics and tropisms of virus replication in its vector. Here we document the virogenesis of DENV-2 in newly-colonized Aedes aegypti mosquitoes from Chetumal, Mexico in order to understand better the effect of vector-virus interactions on dengue transmission. Results After ingestion of DENV-2, midgut infections in Chetumal mosquitoes were characterized by a peak in virus titers between 7 and 10 days post-infection (dpi). The amount of viral antigen and viral titers in the midgut then declined, but viral RNA levels remained stable. The presence of DENV-2 antigen in the trachea was positively correlated with virus dissemination from the midgut. DENV-2 antigen was found in salivary gland tissue in more than a third of mosquitoes at 4 dpi. Unlike in the midgut, the amount of viral antigen (as well as the percent of infected salivary glands) increased with time. DENV-2 antigen also accumulated and increased in neural tissue throughout the EIP. DENV-2 antigen was detected in multiple tissues of the vector, but unlike some other arboviruses, was not detected in muscle. Conclusion Our results suggest that the EIP of DENV-2 in its vector may be shorter that the previously reported and that the tracheal system may facilitate DENV-2 dissemination from the midgut. Mosquito organs (e.g. midgut, neural tissue, and salivary glands) differed in their response to DENV-2 infection. PMID:17263893

Biomimetic structural engineering of P22 virus-like particles for catalysis and immune modulation

NASA Astrophysics Data System (ADS)

Schwarz, Benjamin

Within biology molecules are arranged in hierarchical structures that coordinate and control the many processes that allow for complex organisms to exist. Proteins and other functional macromolecules are often studied outside their natural nanostructural context because it remains difficult to create controlled arrangements of proteins at this size scale. Viruses are elegantly simple nano-systems that exist at the interface of living organisms and non-living biological machines. Studied and viewed primarily as pathogens to be combatted, viruses have emerged as models of structural efficiency at the nanoscale and have spurred the development of biomimetic nanoparticle systems. Virus-like particles (VLPs) are noninfectious protein cages derived from viruses or other cage-forming systems. VLPs provide incredibly regular scaffolds for building at the nanoscale. In this work I have utilized the VLP derived from the bacteriophage P22 as a platform for the organization of enzymes, antigens, and immune-stimulating proteins inside and outside the capsid through purely genetic means. In the case of enzymes, encapsulation of a two-enzyme pathway has led to the development of metabolic nanoparticle catalysts and an expanded understanding of the control that structure exerts on metabolic flux. These same structural elements applied to the delivery of protein subunit antigens directed at cytotoxic T cell immunity result in drastically enhanced antigen processing and lasting immunological memory. Lastly, presentation of immune-stimulating proteins from the Tumor Necrosis Factor Super Family on the surface of the P22 VLP enhances the cell signaling efficiency of these compounds 50-fold and provides strategies for the application of these proteins as immune modulatory oncology therapeutics. In all of these cases, the reintroduction of nanostructure to these protein systems, reminiscent of their natural environment, has led to both new technologies and a better understanding of the role of structure in biological processes.

Inactivated Recombinant Rabies Viruses Displaying Canine Distemper Virus Glycoproteins Induce Protective Immunity against Both Pathogens.

PubMed

da Fontoura Budaszewski, Renata; Hudacek, Andrew; Sawatsky, Bevan; Krämer, Beate; Yin, Xiangping; Schnell, Matthias J; von Messling, Veronika

2017-04-15

The development of multivalent vaccines is an attractive methodology for the simultaneous prevention of several infectious diseases in vulnerable populations. Both canine distemper virus (CDV) and rabies virus (RABV) cause lethal disease in wild and domestic carnivores. While RABV vaccines are inactivated, the live-attenuated CDV vaccines retain residual virulence for highly susceptible wildlife species. In this study, we developed recombinant bivalent vaccine candidates based on recombinant vaccine strain rabies virus particles, which concurrently display the protective CDV and RABV glycoprotein antigens. The recombinant viruses replicated to near-wild-type titers, and the heterologous glycoproteins were efficiently expressed and incorporated in the viral particles. Immunization of ferrets with beta-propiolactone-inactivated recombinant virus particles elicited protective RABV antibody titers, and animals immunized with a combination of CDV attachment protein- and fusion protein-expressing recombinant viruses were protected from lethal CDV challenge. However, animals that were immunized with only a RABV expressing the attachment protein of CDV vaccine strain Onderstepoort succumbed to infection with a more recent wild-type strain, indicating that immune responses to the more conserved fusion protein contribute to protection against heterologous CDV strains. IMPORTANCE Rabies virus and canine distemper virus (CDV) cause high mortality rates and death in many carnivores. While rabies vaccines are inactivated and thus have an excellent safety profile and high stability, live-attenuated CDV vaccines can retain residual virulence in highly susceptible species. Here we generated recombinant inactivated rabies viruses that carry one of the CDV glycoproteins on their surface. Ferrets immunized twice with a mix of recombinant rabies viruses carrying the CDV fusion and attachment glycoproteins were protected from lethal CDV challenge, whereas all animals that received recombinant rabies viruses carrying only the CDV attachment protein according to the same immunization scheme died. Irrespective of the CDV antigens used, all animals developed protective titers against rabies virus, illustrating that a bivalent rabies virus-based vaccine against CDV induces protective immune responses against both pathogens. Copyright © 2017 American Society for Microbiology.

Seroprevalence of human herpes simplex, hepatitis B and epstein-barr viruses in children with acute lymphoblastic leukemia in southern iran.

PubMed

Mahjour, Seyed Babak; Ghaffarpasand, Fariborz; Fattahi, Mohammad Javad; Ghaderi, Abbas; Fotouhi Ghiam, Alireza; Karimi, Mehran

2010-12-01

To investigate the seroprevalence of Herpes Simplex Viruses (HSV1 and HSV2), Ebstein-Barr Virus (EBV) and Hepatitis B Virus (HBV) in children with acute lymphoblastic leukemia (ALL) in southern Iran. 90 patients with ALL and 90 age-sex matched healthy participants were enrolled in this study. Antibodies (IgG) against HBsAg, HSV1, HSV2, EBV different antigens including Epstein-Barr nuclear antigen-1 (EBNA-1), viral capsid antigen (VCA) and early antigen (EA) were measured by enzyme-linked immunosorbent assay (ELISA). There were 54 (60%) male and 36 (40%) female in both study groups with mean age of 8.47 ± 3.61 and 8.61 ± 2.84 years in case and control group respectively (P = 0.812). The prevalence of antibodies against HBsAg (P = 0.002), HSV1 (P < 0.0001), VCA (P = 0.021) and EA (P < 0.0001) antigens of EBV were significantly higher in ALL patients. With the results of this study, we could not exclude a connection between these viral infections and later leukemogenesis in childhood ALL, although nor latent infection nor congenital infection cannot be excluded by this method.

Strategies to alleviate original antigenic sin responses to influenza viruses.

PubMed

Kim, Jin Hyang; Davis, William G; Sambhara, Suryaprakash; Jacob, Joshy

2012-08-21

Original antigenic sin is a phenomenon wherein sequential exposure to closely related influenza virus variants reduces antibody (Ab) response to novel antigenic determinants in the second strain and, consequently, impairs the development of immune memory. This could pose a risk to the development of immune memory in persons previously infected with or vaccinated against influenza. Here, we explored strategies to overcome original antigenic sin responses in mice sequentially exposed to two closely related hemagglutinin 1 neuraminidase 1 (H1N1) influenza strains A/PR/8/34 and A/FM/1/47. We found that dendritic cell-activating adjuvants [Bordetella pertussis toxin (PT) or CpG ODN or a squalene-based oil-in-water nanoemulsion (NE)], upon administration during the second viral exposure, completely protected mice from a lethal challenge and enhanced neutralizing-Ab titers against the second virus. Interestingly, PT and NE adjuvants when administered during the first immunization even prevented original antigenic sin in subsequent immunization without any adjuvants. As an alternative to using adjuvants, we also found that repeated immunization with the second viral strain relieved the effects of original antigenic sin. Taken together, our studies provide at least three ways of overcoming original antigenic sin.

Simian virus 40 small t antigen is not required for the maintenance of transformation but may act as a promoter (cocarcinogen) during establishment of transformation in resting rat cells.

PubMed Central

Seif, R; Martin, R G

1979-01-01

Simian virus 40 deletion mutants affecting the 20,000-dalton (20K) t antigen and tsA mutants rendering the 90K T antigen temperature sensitive, as well as double mutants containing both mutations, induced host DNA synthesis in resting rat cells at the restrictive temperature. Nonetheless, the deletion mutants and double mutants did not induce transformation in resting cells even at the permissive temperature. On the other hand, the deletion mutants did induce full transformants when actively growing rat cells were infected; the transformants grew efficiently in agar and to high saturation densities on platic. The double mutants did not induce T-antigen-independent (temperature-insensitive) transformants which were shown previously to arise preferentially from resting cells. Thus, small t antigen was dispensable for the maintenance of the transformed phenotype in T-antigen-dependent rat transformants (transformants derived from growing cells) and may play a role in the establishment of T-antigen-independent transformants. We attempt to establish a parallel between transformation induced by chemical carcinogens and simian virus 40-induced transformation. Images PMID:229274

Simian virus 40 small t antigen is not required for the maintenance of transformation but may act as a promoter (cocarcinogen) during establishment of transformation in resting rat cells.

PubMed

Seif, R; Martin, R G

1979-12-01

Simian virus 40 deletion mutants affecting the 20,000-dalton (20K) t antigen and tsA mutants rendering the 90K T antigen temperature sensitive, as well as double mutants containing both mutations, induced host DNA synthesis in resting rat cells at the restrictive temperature. Nonetheless, the deletion mutants and double mutants did not induce transformation in resting cells even at the permissive temperature. On the other hand, the deletion mutants did induce full transformants when actively growing rat cells were infected; the transformants grew efficiently in agar and to high saturation densities on platic. The double mutants did not induce T-antigen-independent (temperature-insensitive) transformants which were shown previously to arise preferentially from resting cells. Thus, small t antigen was dispensable for the maintenance of the transformed phenotype in T-antigen-dependent rat transformants (transformants derived from growing cells) and may play a role in the establishment of T-antigen-independent transformants. We attempt to establish a parallel between transformation induced by chemical carcinogens and simian virus 40-induced transformation.

Oral Fluids as a Live-Animal Sample Source for Evaluating Cross-Reactivity and Cross-Protection following Intranasal Influenza A Virus Vaccination in Pigs

PubMed Central

Hughes, Holly R.; Vincent, Amy L.; Brockmeier, Susan L.; Gauger, Phillip C.; Pena, Lindomar; Santos, Jefferson; Braucher, Douglas R.

2015-01-01

In North American swine, there are numerous antigenically distinct H1 influenza A virus (IAV) variants currently circulating, making vaccine development difficult due to the inability to formulate a vaccine that provides broad cross-protection. Experimentally, live-attenuated influenza virus (LAIV) vaccines demonstrate increased cross-protection compared to inactivated vaccines. However, there is no standardized assay to predict cross-protection following LAIV vaccination. Hemagglutination-inhibiting (HI) antibody in serum is the gold standard correlate of protection following IAV vaccination. LAIV vaccination does not induce a robust serum HI antibody titer; however, a local mucosal antibody response is elicited. Thus, a live-animal sample source that could be used to evaluate LAIV immunogenicity and cross-protection is needed. Here, we evaluated the use of oral fluids (OF) and nasal wash (NW) collected after IAV inoculation as a live-animal sample source in an enzyme-linked immunosorbent assay (ELISA) to predict cross-protection in comparison to traditional serology. Both live-virus exposure and LAIV vaccination provided heterologous protection, though protection was greatest against more closely phylogenetically related viruses. IAV-specific IgA was detected in NW and OF samples and was cross-reactive to representative IAV from each H1 cluster. Endpoint titers of cross-reactive IgA in OF from pigs exposed to live virus was associated with heterologous protection. While LAIV vaccination provided significant protection, LAIV immunogenicity was reduced compared to live-virus exposure. These data suggest that OF from pigs inoculated with wild-type IAV, with surface genes that match the LAIV seed strain, could be used in an ELISA to assess cross-protection and the antigenic relatedness of circulating and emerging IAV in swine. PMID:26291090

Oral Fluids as a Live-Animal Sample Source for Evaluating Cross-Reactivity and Cross-Protection following Intranasal Influenza A Virus Vaccination in Pigs.

PubMed

Hughes, Holly R; Vincent, Amy L; Brockmeier, Susan L; Gauger, Phillip C; Pena, Lindomar; Santos, Jefferson; Braucher, Douglas R; Perez, Daniel R; Loving, Crystal L

2015-10-01

In North American swine, there are numerous antigenically distinct H1 influenza A virus (IAV) variants currently circulating, making vaccine development difficult due to the inability to formulate a vaccine that provides broad cross-protection. Experimentally, live-attenuated influenza virus (LAIV) vaccines demonstrate increased cross-protection compared to inactivated vaccines. However, there is no standardized assay to predict cross-protection following LAIV vaccination. Hemagglutination-inhibiting (HI) antibody in serum is the gold standard correlate of protection following IAV vaccination. LAIV vaccination does not induce a robust serum HI antibody titer; however, a local mucosal antibody response is elicited. Thus, a live-animal sample source that could be used to evaluate LAIV immunogenicity and cross-protection is needed. Here, we evaluated the use of oral fluids (OF) and nasal wash (NW) collected after IAV inoculation as a live-animal sample source in an enzyme-linked immunosorbent assay (ELISA) to predict cross-protection in comparison to traditional serology. Both live-virus exposure and LAIV vaccination provided heterologous protection, though protection was greatest against more closely phylogenetically related viruses. IAV-specific IgA was detected in NW and OF samples and was cross-reactive to representative IAV from each H1 cluster. Endpoint titers of cross-reactive IgA in OF from pigs exposed to live virus was associated with heterologous protection. While LAIV vaccination provided significant protection, LAIV immunogenicity was reduced compared to live-virus exposure. These data suggest that OF from pigs inoculated with wild-type IAV, with surface genes that match the LAIV seed strain, could be used in an ELISA to assess cross-protection and the antigenic relatedness of circulating and emerging IAV in swine. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Freeze-Drying of Plant Tissue Containing HBV Surface Antigen for the Oral Vaccine against Hepatitis B

PubMed Central

Milczarek, Magdalena; Pajtasz-Piasecka, Elżbieta; Wietrzyk, Joanna

2014-01-01

The aim of this study was to develop a freeze-drying protocol facilitating successful processing of plant material containing the small surface antigen of hepatitis B virus (S-HBsAg) while preserving its VLP structure and immunogenicity. Freeze-drying of the antigen in lettuce leaf tissue, without any isolation or purification step, was investigated. Each process step was consecutively evaluated and the best parameters were applied. Several drying profiles and excipients were tested. The profile of 20°C for 20 h for primary and 22°C for 2 h for secondary drying as well as sucrose expressed efficient stabilisation of S-HBsAg during freeze-drying. Freezing rate and postprocess residual moisture were also analysed as important factors affecting S-HBsAg preservation. The process was reproducible and provided a product with VLP content up to 200 µg/g DW. Assays for VLPs and total antigen together with animal immunisation trials confirmed preservation of antigenicity and immunogenicity of S-HBsAg in freeze-dried powder. Long-term stability tests revealed that the stored freeze-dried product was stable at 4°C for one year, but degraded at elevated temperatures. As a result, a basis for an efficient freeze-drying process has been established and a suitable semiproduct for oral plant-derived vaccine against HBV was obtained. PMID:25371900

Equine infectious anemia virus-infected dendritic cells retain antigen presentation capability

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rivera, Julie A.; McGuire, Travis C.

2005-05-10

To determine if equine monocyte-derived dendritic cells (DC) were susceptible to equine infectious anemia virus (EIAV) infection, ex vivo-generated DC were infected with virus in vitro. EIAV antigen was detected by immunofluorescence 3 days post-infection with maximum antigen being detected on day 4, whereas there was no antigen detected in DC incubated with the same amount of heat-inactivated EIAV. No cytolytic activity was observed after EIAV{sub WSU5} infection of DC. These monocyte-derived DC were more effective than macrophages and B cells in stimulating allogenic T lymphocytes. Both infected macrophages and DC stimulated similar levels of memory CTL responses in mixturesmore » of CD8+ and CD4+ cells as detected with {sup 51}Cr-release assays indicating that EIAV infection of DC did not alter antigen presentation. However, EIAV-infected DC were more effective than infected macrophages when used to stimulate memory CTL in isolated CD8+ cells. The maintenance of antigen processing and presenting function by EIAV-infected DC in vitro suggests that this function is maintained during in vivo infection.« less

The effects of a deleterious mutation load on patterns of influenza A/H3N2's antigenic evolution in humans

PubMed Central

Koelle, Katia; Rasmussen, David A

2015-01-01

Recent phylogenetic analyses indicate that RNA virus populations carry a significant deleterious mutation load. This mutation load has the potential to shape patterns of adaptive evolution via genetic linkage to beneficial mutations. Here, we examine the effect of deleterious mutations on patterns of influenza A subtype H3N2's antigenic evolution in humans. By first analyzing simple models of influenza that incorporate a mutation load, we show that deleterious mutations, as expected, act to slow the virus's rate of antigenic evolution, while making it more punctuated in nature. These models further predict three distinct molecular pathways by which antigenic cluster transitions occur, and we find phylogenetic patterns consistent with each of these pathways in influenza virus sequences. Simulations of a more complex phylodynamic model further indicate that antigenic mutations act in concert with deleterious mutations to reproduce influenza's spindly hemagglutinin phylogeny, co-circulation of antigenic variants, and high annual attack rates. DOI: http://dx.doi.org/10.7554/eLife.07361.001 PMID:26371556

Chitosan-based nanoparticles for improving immunization against hepatitis B infection.

PubMed

Prego, Cecilia; Paolicelli, Patrizia; Díaz, Belen; Vicente, Sara; Sánchez, Alejandro; González-Fernández, Africa; Alonso, María José

2010-03-19

The design of effective vaccine delivery vehicles is opening up new possibilities for making immunization more equitable, safe and efficient. In this work, we purpose polysaccharidic-based nanoparticles as delivery structures for virus-like particle antigens, using recombinant hepatitis B surface antigen (rHBsAg) as a model. Polysaccharidic-based nanoparticles were prepared using a very mild ionic gelation technique, by cross-linking the polysaccharide chitosan (CS) with a counter ion. The resulting nanoparticles could be easily isolated with a size in the nanometric range (160-200 nm) and positive surface charge (+6 to +10 mV). More importantly, CS-based nanoparticles allowed the efficient association of the antigen (>60%) while maintaining the antigenic epitope intact, as determined by ELISA and Western blot. The entrapped antigen was further released in vitro from the nanoparticles in a sustained manner without compromising its antigenicity. In addition, loaded CS-based nanoparticles were stable, and protected the associated antigen during storage, either as an aqueous suspension under different temperature conditions (+4 degrees C and -20 degrees C), or as a dried form after freeze-drying the nanoparticles. Finally, immunization studies showed the induction of important seroprotection rates after intramuscular administration of the nanoparticles, indicating their adjuvant capacity. In fact, CS-based nanoparticles were able to induce anti-HBsAg IgG levels up to 5500 mIU/ml, values 9-fold the conventional alum-adsorbed vaccine. In conclusion, we report here a polysaccharidic nanocarrier which exhibits a number of in vitro and in vivo features that make it a promising adjuvant for vaccine delivery of subunit antigens. Copyright 2010 Elsevier Ltd. All rights reserved.

Antigenic characterization of bovine ephemeral fever rhabdovirus G and GNS glycoproteins expressed from recombinant baculoviruses.

PubMed

Johal, Jasjit; Gresty, Karryn; Kongsuwan, Kritaya; Walker, Peter J

2008-01-01

Recombinant baculoviruses expressing the BEFV envelope glycoprotein G and non-structural glycoprotein G(NS) were constructed. The G protein expressed in insect cells was located on the cell surface and induced spontaneous cell fusion at mildly acidic pH. The expressed G protein reacted with MAbs to continuous and conformational neutralization sites (G1, G2, G3b and G4), but not to conformational site G3a. The expressed G(NS) protein was also located on the cell surface but did not exhibit fusogenic activity. The G(NS) protein reacted with polyclonal antiserum produced from vaccinia-virus-expressed recombinant G(NS) but did not react with G protein antibodies. A His(6)-tagged, soluble form of the G protein was expressed and purified by Ni(2+)-NTA chromatography. The purified G protein reacted with BEFV-neutralizing MAbs to all continuous and conformational antigenic sites. The highly protective characteristics of the native BEFV G protein suggest that the secreted, baculovirus-expressed product may be a useful vaccine antigen.

Computational identification of epitopes in the glycoproteins of novel bunyavirus (SFTS virus) recognized by a human monoclonal antibody (MAb 4-5)

NASA Astrophysics Data System (ADS)

Zhang, Wenshuai; Zeng, Xiaoyan; Zhang, Li; Peng, Haiyan; Jiao, Yongjun; Zeng, Jun; Treutlein, Herbert R.

2013-06-01

In this work, we have developed a new approach to predict the epitopes of antigens that are recognized by a specific antibody. Our method is based on the "multiple copy simultaneous search" (MCSS) approach which identifies optimal locations of small chemical functional groups on the surfaces of the antibody, and identifying sequence patterns of peptides that can bind to the surface of the antibody. The identified sequence patterns are then used to search the amino-acid sequence of the antigen protein. The approach was validated by reproducing the binding epitope of HIV gp120 envelop glycoprotein for the human neutralizing antibody as revealed in the available crystal structure. Our method was then applied to predict the epitopes of two glycoproteins of a newly discovered bunyavirus recognized by an antibody named MAb 4-5. These predicted epitopes can be verified by experimental methods. We also discuss the involvement of different amino acids in the antigen-antibody recognition based on the distributions of MCSS minima of different functional groups.

Generation and characterization of high affinity humanized fab against hepatitis B surface antigen.

PubMed

Tiwari, Ashutosh; Dutta, Durgashree; Khanna, Navin; Acharya, Subrat K; Sinha, Subrata

2009-09-01

5S is a mouse monoclonal IgG1 that binds to the 'a' epitope of the Hepatitis B surface antigen (HBsAg) and tested positive in an in vitro test for virus neutralization. We have earlier reported the generation of humanized single chain variable fragment (scFv) from the same. In this article we report the generation of a recombinant Fab molecule by fusing humanized variable domains of 5S with the constant domains of human IgG1. The humanized Fab expressed in E. coli and subsequently purified, retained a high binding affinity (K(D) = 3.63 nmol/L) to HBsAg and bound to the same epitope of HBsAg as the parent molecule. The humanized Fab also maintained antigen binding in the presence of various destabilizing agents like 3 M NaCl, 30% DMSO, 8 M urea, and extreme pH. This high affinity humanized Fab provides a basis for the development of therapeutic molecules that can be safely utilized for the prophylaxis and treatment for Hepatitis B infection.

Hepatitis B Vaccination, Screening, and Linkage to Care: Best Practice Advice From the American College of Physicians and the Centers for Disease Control and Prevention.

PubMed

Abara, Winston E; Qaseem, Amir; Schillie, Sarah; McMahon, Brian J; Harris, Aaron M

2017-12-05

Vaccination, screening, and linkage to care can reduce the burden of chronic hepatitis B virus (HBV) infection. However, recommendations vary among organizations, and their implementation has been suboptimal. The American College of Physicians' High Value Care Task Force and the Centers for Disease Control and Prevention developed this article to present best practice statements for hepatitis B vaccination, screening, and linkage to care. A narrative literature review of clinical guidelines, systematic reviews, randomized trials, and intervention studies on hepatitis B vaccination, screening, and linkage to care published between January 2005 and June 2017 was conducted. Clinicians should vaccinate against hepatitis B virus (HBV) in all unvaccinated adults (including pregnant women) at risk for infection due to sexual, percutaneous, or mucosal exposure; health care and public safety workers at risk for blood exposure; adults with chronic liver disease, end-stage renal disease (including hemodialysis patients), or HIV infection; travelers to HBV-endemic regions; and adults seeking protection from HBV infection. Clinicians should screen (hepatitis B surface antigen, antibody to hepatitis B core antigen, and antibody to hepatitis B surface antigen) for HBV in high-risk persons, including persons born in countries with 2% or higher HBV prevalence, men who have sex with men, persons who inject drugs, HIV-positive persons, household and sexual contacts of HBV-infected persons, persons requiring immunosuppressive therapy, persons with end-stage renal disease (including hemodialysis patients), blood and tissue donors, persons infected with hepatitis C virus, persons with elevated alanine aminotransferase levels (≥19 IU/L for women and ≥30 IU/L for men), incarcerated persons, pregnant women, and infants born to HBV-infected mothers. Clinicians should provide or refer all patients identified with HBV (HBsAg-positive) for posttest counseling and hepatitis B-directed care.

Molecular Evolution and Genetic Analysis of the Major Capsid Protein VP1 of Duck Hepatitis A Viruses: Implications for Antigenic Stability

PubMed Central

Ma, Xiuli; Sheng, Zizhang; Huang, Bing; Qi, Lihong; Li, Yufeng; Yu, Kexiang; Liu, Cunxia; Qin, Zhuoming; Wang, Dan; Song, Minxun; Li, Feng

2015-01-01

The duck hepatitis A virus (DHAV), a member of the family Picornaviridae, is the major cause of outbreaks with high mortality rates in young ducklings. It has three distinctive serotypes and among them, serotypes 1 (DHAV-1) and 3 (DHAV-3) were recognized in China. To investigate evolutionary and antigenic properties of the major capsid protein VP1 of these two serotypes, a primary target of neutralizing antibodies, we determined the VP1 coding sequences of 19 DHAV-1 (spanning 2000-2012) and 11 DHAV-3 isolates (spanning 2008-2014) associated with disease outbreaks. By bioinformatics analysis of VP1 sequences of these isolates and other DHAV strains reported previously, we demonstrated that DHAV-1 viruses evolved into two genetic lineages, while DHAV-3 viruses exhibited three distinct lineages. The rate of nucleotide substitution for DHAV-1 VP1 genes was estimated to be 5.57 x 10-4 per site per year, which was about one-third times slower than that for DHAV-3 VP1 genes. The population dynamics analysis showed an upward trend for infection of DHAV-1 viruses over time with little change observed for DHAV-3 viruses. Antigenic study of representative DHAV-1 and DHAV-3 strains covering all observed major lineages revealed no detectable changes in viral neutralization properties within the serotype, despite the lack of cross-neutralization between serotypes 1 and 3 strains. Structural analysis identified VP1 mutations in DHAV-1 and DHAV-3 viruses that underpin the observed antigenic phenotypes. Results of our experiments described here shall give novel insights into evolution and antigenicity of duck picornaviruses. PMID:26173145

Whole-genome sequencing and antigenic analysis of the first equine influenza virus identified in Turkey.

PubMed

Gahan, Jacinta; Garvey, Marie; Gildea, Sarah; Gür, Emre; Kagankaya, Anil; Cullinane, Ann

2018-05-01

In 2013, there was an outbreak of acute respiratory disease in racehorses in Turkey. The clinical signs were consistent with equine influenza (EI). The aim was to confirm the cause of the outbreak and characterise the causal virus. A pan-reactive influenza type A real-time RT-PCR and a rapid antigen detection kit were used for confirmatory diagnosis of equine influenza virus (EIV). Immunological susceptibility to EIV was examined using single radial haemolysis and ELISA. Antigenic characterisation was completed by haemagglutinin inhibition using a panel of specific ferret antisera. Genetic characterisation was achieved by whole-genome sequencing using segment-specific primers with M13 tags. A H3N8 EIV of the Florida clade 2 sublineage (FC2) was confirmed as the causal agent. The index cases were unvaccinated and immunologically susceptible. Phylogenetic analysis of the HA1 and NA genes demonstrated that A/equine/Ankara/1/2013 clustered with the FC2 strains circulating in Europe. Antigenic characterisation confirmed the FC2 classification and demonstrated the absence of significant drift. Whole-genome sequencing indicated that A/equine/Ankara/1/2013 is most closely related to the viruses described as the 179 group based on the substitution I179V in HA1, for example A/equine/East Renfrewshire/2/2011, A/equine/Cambremer/1/2012 and A/equine/Saone et Loire/1/2015. The greatest diversity was observed in the NS1 segment and the polymerase complex. The first recorded outbreak of EI in Turkey was caused by an FC2 virus closely related to viruses circulating in Europe. Antigenic and genetic characterisation gave no indication that the current OIE recommendations for EI vaccine composition require modification. © 2017 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.

Species-Specific Elements in the Large T-Antigen J Domain Are Required for Cellular Transformation and DNA Replication by Simian Virus 40

PubMed Central

Sullivan, Christopher S.; Tremblay, James D.; Fewell, Sheara W.; Lewis, John A.; Brodsky, Jeffrey L.; Pipas, James M.

2000-01-01

The J domain of simian virus 40 (SV40) large T antigen is required for efficient DNA replication and transformation. Despite previous reports demonstrating the promiscuity of J domains in heterologous systems, results presented here show the requirement for specific J-domain sequences in SV40 large-T-antigen-mediated activities. In particular, chimeric-T-antigen constructs in which the SV40 T-antigen J domain was replaced with that from the yeast Ydj1p or Escherichia coli DnaJ proteins failed to replicate in BSC40 cells and did not transform REF52 cells. However, T antigen containing the JC virus J domain was functional in these assays, although it was less efficient than the wild type. The inability of some large-T-antigen chimeras to promote DNA replication and elicit cellular transformation was not due to a failure to interact with hsc70, since a nonfunctional chimera, containing the DnaJ J domain, bound hsc70. However, this nonfunctional chimeric T antigen was reduced in its ability to stimulate hsc70 ATPase activity and unable to liberate E2F from p130, indicating that transcriptional activation of factors required for cell growth and DNA replication may be compromised. Our data suggest that the T-antigen J domain harbors species-specific elements required for viral activities in vivo. PMID:10891510

Different cross protection scopes of two avian influenza H5N1 vaccines against infection of layer chickens with a heterologous highly pathogenic virus.

PubMed

Poetri, Okti Nadia; Van Boven, Michiel; Koch, Guus; Stegeman, Arjan; Claassen, Ivo; Wayan Wisaksana, I; Bouma, Annemarie

2017-10-01

Avian influenza (AI) virus strains vary in antigenicity, and antigenic differences between circulating field virus and vaccine virus will affect the effectiveness of vaccination of poultry. Antigenic relatedness can be assessed by measuring serological cross-reactivity using haemagglutination inhibition (HI) tests. Our study aims to determine the relation between antigenic relatedness expressed by the Archetti-Horsfall ratio, and reduction of virus transmission of highly pathogenic H5N1 AI strains among vaccinated layers. Two vaccines were examined, derived from H5N1 AI virus strains A/Ck/WJava/Sukabumi/006/2008 and A/Ck/CJava/Karanganyar/051/2009. Transmission experiments were carried out in four vaccine and two control groups, with six sets of 16 specified pathogen free (SPF) layer chickens. Birds were vaccinated at 4weeks of age with one strain and challenge-infected with the homologous or heterologous strain at 8weeks of age. No transmission or virus shedding occurred in groups challenged with the homologous strain. In the group vaccinated with the Karanganyar strain, high cross-HI responses were observed, and no transmission of the Sukabumi strain occurred. However, in the group vaccinated with the Sukabumi strain, cross-HI titres were low, virus shedding was not reduced, and multiple transmissions to contact birds were observed. This study showed large differences in cross-protection of two vaccines based on two different highly pathogenic H5N1 virus strains. This implies that extrapolation of in vitro data to clinical protection and reduction of virus transmission might not be straightforward. Copyright © 2017 Elsevier Ltd. All rights reserved.

H1N1 seasonal influenza virus evolutionary rate changed over time.

PubMed

Suptawiwat, Ornpreya; Kongchanagul, Alita; Boonarkart, Chompunuch; Auewarakul, Prasert

2018-05-02

It was previously shown that the seasonal H1N1 influenza virus antigenic drift occurred at a slower rate than the seasonal H3N2 virus during the first decade of the 21th century. It was hypothesized that the slower antigenic evolution led to a decrease in average ages of infection, which in turn resulted in lower level of global viral circulation. It is unclear what caused the difference between the two viruses, but a plausible explanation may be related to the fact that the H1N1 virus had been in human population for much longer than the H3N2 virus. This would suggest that H1N1 antigenic drift in an earlier period may have been different from a more recent period. To test this hypothesis, we analyzed seasonal H1N1 influenza sequences during various time periods. In comparison to more recent H1N1 virus, the older H1N1 virus during the first half of the 20th century showed evidences of higher nonsynnonymous/synonymous ration (dN/dS) in its hemagglutinin (HA) gene. We compared amino acid sequence changes in the HA epitopes for each outbreak season and found that there were less changes in later years. Amino acid sequence diversity in the epitopes as measured by sequence entropy became smaller for each passing decade. These suggest that there might be some limit to the antigenic drift. The longer an influenza virus has drifted in human population, the less flexibility it may become. With less flexibility to adapt and escape the host immunity, the virus may have to rely more on younger naïve population. Copyright © 2018 Elsevier B.V. All rights reserved.

Recombinant glycoprotein E produced in mammalian cells in large-scale as an antigen for varicella-zoster-virus serology.

PubMed

Thomsson, Elisabeth; Persson, Linn; Grahn, Anna; Snäll, Johanna; Ekblad, Maria; Brunhage, Eva; Svensson, Frida; Jern, Christina; Hansson, Gunnar C; Bäckström, Malin; Bergström, Tomas

2011-07-01

A recombinant glycoprotein E (gE) from varicella-zoster virus (VZV) was generated and produced in Chinese Hamster Ovary (CHO) cells, in the development of a specific antigen for analysis of IgG antibodies to VZV. Several stable gE-secreting clones were established and one clone was adapted to growth in serum-free suspension culture. When the cells were cultured in a perfusion bioreactor, gE was secreted into the medium, from where it could be easily purified. The recombinant gE was then evaluated as a serological antigen in ELISA. When compared to a conventional whole virus antigen, the VZV gE showed similar results in ELISA-based seroprevalence studies of 854 samples derived from blood donors, students, ischemic stroke patients and their controls, including samples with border-line results in previous analyses. Eight samples (0.9%) were discordant, all being IgG-negative by the VZV gE ELISA and positive by the whole virus ELISA. The sensitivity and specificity of the VZV gE ELISA were 99.9% and 100%, respectively, compared to 100% and 88.9% for the VZV whole virus ELISA. The elderly subjects showed similar reactivities to both antigens, while VZV gE gave lower signals in the younger cohorts, suggesting that antibodies to gE may increase with age. It was concluded that the recombinant VZV gE from CHO cells was suitable as a serological antigen for the detection of IgG antibodies specific for VZV. Copyright © 2011 Elsevier B.V. All rights reserved.

Poly-ϵ-caprolactone/chitosan nanoparticles provide strong adjuvant effect for hepatitis B antigen.

PubMed

Jesus, Sandra; Soares, Edna; Borchard, Gerrit; Borges, Olga

2017-10-01

This work aims to investigate the adjuvant effect of poly-ϵ-caprolactone/chitosan nanoparticles (NPs) for hepatitis B surface antigen (HBsAg) and the plasmid DNA encoding HBsAg (pRC/CMV-HBs). Both antigens were adsorbed onto preformed NPs. Vaccination studies were performed in C57BL/6 mice. Transfection efficiency was investigated in A549 cell line. HBsAg-adsorbed NPs generated strong anti-HBsAg IgG titers, mainly of IgG1 isotype, and induced antigen-specific IFN-γ and IL-17 secretion by spleen cells. The addition of pRC/CMV-HBs to the HBsAg-adsorbed NPs inhibited IL-17 secretion but had minor effect on IFN-γ levels. Lastly, pRC/CMV-HBs-loaded NPs generated a weak serum antibody response. Poly-ϵ-caprolactone/chitosan NPs provide a strong humoral adjuvant effect for HBsAg and induce a Th1/Th17-mediated cellular immune responses worth explore for hepatitis B virus vaccination.

Prediction of common epitopes on hemagglutinin of the influenza A virus (H1 subtype).

PubMed

Guo, Chunyan; Xie, Xin; Li, Huijin; Zhao, Penghua; Zhao, Xiangrong; Sun, Jingying; Wang, Haifang; Liu, Yang; Li, Yan; Hu, Qiaoxia; Hu, Jun; Li, Yuan

2015-02-01

Influenza A virus infection is a persistent threat to public health worldwide due to hemagglutinin (HA) variation. Current vaccines against influenza A virus provide immunity to viral isolates similar to vaccine strains. Antibodies against common epitopes provide immunity to diverse influenza virus strains and protect against future pandemic influenza. Therefore, it is vital to analyze common HA antigenic epitopes of influenza virus. In this study, 14 strains of monoclonal antibodies with high sensitivity to common epitopes of influenza virus antigens identified in our previous study were selected as the tool to predict common HA epitopes. The common HA antigenic epitopes were divided into four categories by ELISA blocking experiments, and separately, into three categories according to the preliminary results of computer simulation. Comparison between the results of computer simulations and ELISA blocking experiments indicated that at least two classes of common epitopes are present in influenza virus HA. This study provides experimental data for improving the prediction of HA epitopes of influenza virus (H1 subtype) and the development of a potential universal vaccine as well as a novel approach for the prediction of epitopes on other pathogenic microorganisms. Copyright © 2014 Elsevier Inc. All rights reserved.

Progressive antigenic drift and phylogeny of human influenza A(H3N2) virus over five consecutive seasons (2009-2013) in Hangzhou, China.

PubMed

Shao, Tie-Juan; Li, Jun; Yu, Xin-Fen; Kou, Yu; Zhou, Yin-Yan; Qian, Xin

2014-12-01

Vaccine efficacy (VE) can be affected by progressive antigenic drift or any new reassortment of influenza viruses. To effectively track the evolution of human influenza A(H3N2) virus circulating in Hangzhou, China, a total of 65 clinical specimens were selected randomly from outpatients infected by A(H3N2) viruses during the study period from November 2009 to December 2013. The results of reduced VE and antigenic drift of the correspondent epitopes (C-D-E to A-B) suggest that the current vaccine provides suboptimal protection against the A(H3N2) strains circulating recently. Phylogenetic analysis of the entire HA and NA sequences demonstrated that these two genes underwent independent evolutionary pathways during recent seasons. The H3-based phylogenetic tree showed that a special strain A/Hangzhou/A289/2012 fell in a cluster among viruses with reduced VE predominantly circulating in 2013. Our findings underscore a possible early warning for the circulation of A(H3N2) variants with antigenic drift during the previous seasons. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

Initial report of a hepatitis investigation in rural Belize.

PubMed

Hoffman, K J; Gaydos, J C; Krieg, R E; Duncan, J F; MacArthy, P O; Ticehurst, J R; Jaramillo, R; Reyes, L G; Sjogren, M H; Legters, L J

1993-01-01

In spring 1991, Belizian health officials expressed concern about a possible hepatitis outbreak in a banana farming district. A study was designed to identify cases and to address the serological prevalence of hepatitis virus markers. Three populations were studied: (i) persons meeting a clinical case definition for hepatitis; (ii) designated banana workers; and (iii) people in a random sample of households in the community. Information was collected using questionnaires and sera were collected for laboratory testing. This report presents the preliminary results of a study conducted in June 1991. Among people who met the clinical case definition, 24% of 42 tested had immunoglobulin M antibody to hepatitis B virus (HBV) core antigen (anti-HBc IgM). In the worker and household survey populations, 284 and 280 people, respectively, were tested for anti-HBc IgM. In each group, 4% were positive. HBV surface antigen was found in 37% of 43 clinical cases, 18% of workers, and 13% of people in the household survey. Among the 3 study populations, the prevalence of HBV core antibody (anti-HBc) ranged from 73% to 81%. Almost all tested persons had evidence of prior hepatitis A virus infection. Evidence of prior infection with hepatitis viruses A and B was widespread, but an aetiology could not be established for most of the clinical cases. However, the prevalence of hepatitis B markers in this population was very high compared to other reports from the Caribbean.

Core protein: a pleiotropic keystone in the HBV lifecycle

PubMed Central

Zlotnick, Adam; Venkatakrishnan, Balasubramanian; Tan, Zhenning; Lewellyn, Eric; Turner, William; Francis, Samson

2015-01-01

Hepatitis B Virus (HBV) is a small virus whose genome has only four open reading frames. We argue that the simplicity of the virion correlates with a complexity of functions for viral proteins. We focus on the HBV core protein (Cp), a small (183 residue) protein that self-assembles to form the viral capsid. However, its functions are a little more complicated than that. In an infected cell Cp modulates every step of the viral lifecycle. Cp is bound to nuclear viral DNA and affects its epigenetics. Cp correlates with RNA specificity. Cp assembles specifically on a reverse transcriptase-viral RNA complex or, apparently, nothing at all. Indeed Cp has been one of the model systems for investigation of virus self-assembly. Cp participates in regulation of reverse transcription. Cp signals completion of reverse transcription to support virus secretion. Cp carries both nuclear localization signals and HBV surface antigen (HBsAg) binding sites; both of these functions appear to be regulated by contents of the capsid. Cp can be targeted by antivirals -- while self-assembly is the most accessible of Cp activities, we argue that it makes sense to engage the broader spectrum of Cp function. This article forms part of a symposium in Antiviral Research on “From the discovery of the Australia antigen to the development of new curative therapies for hepatitis B: an unfinished story.” PMID:26129969

Hepatitis A as an etiologic agent of acute liver failure in Latin America.

PubMed

Ciocca, Mirta; Moreira-Silva, Sandra Fagundes; Alegría, Sylvia; Galoppo, Maria Cristina; Ruttiman, Ricardo; Porta, Gilda; Da Silvera, Themis Reverbel; Rubio, Pilar; Macias, Mercedes; Cervantes, Yolanda; Avila-Aguero, Maria Luisa; Clemens, Sue Anne Costa; Clemens, Ralf; Weil, John

2007-08-01

This prospective, multicenter study examined the importance of hepatitis viruses as etiological agents of acute liver failure (ALF) and the outcome of ALF cases in Latin American children and adolescents. The study was conducted for minimum 12 months in 9 centers in Argentina, Brazil, Chile, Colombia, Costa Rica, and Mexico during 2001-2002. Hospitalized patients aged 1-20 years with a suspected diagnosis of ALF were included in the study and tested for serologic markers for hepatitis A, B, and C viruses. Of the 106 patients enrolled, 88 were included in the analysis. Median age was 5 years, and 55% with ALF were aged 1-5 years. A total of 37 individuals (43%) tested positive for anti-hepatitis A virus (HAV) immunoglobulin M (IgM) as marker of acute HAV infection; one was positive for anti-hepatitis B core antigen IgM and negative for hepatitis B surface antigen. None had markers of hepatitis C virus infection. Mortality rates in the overall study cohort (45%) and for those who tested anti-HAV IgM positive (41%) were similar. Forty-one percent of all patients and 46% of those positive for anti-HAV IgM underwent transplantation. The mortality rate in those with liver transplantation was half of that in patients who were not transplanted (28% versus 57%). HAV was the main etiologic agent of ALF in the population studied.

Inhibition of allergic encephalomyelitis in marmosets by vaccination with recombinant vaccinia virus encoding for myelin basic protein.

PubMed

Genain, C P; Gritz, L; Joshi, N; Panicali, D; Davis, R L; Whitaker, J N; Letvin, N L; Hauser, S L

1997-11-01

A primary demyelinating form of experimental allergic encephalomyelitis (EAE) resembling human multiple sclerosis (MS) occurs in Callithrix jacchus marmosets following immunization with human white matter. Participation of a T-cell immune response against myelin basic protein (MBP) in this disease model is supported by observations of increased reactivity against MBP in PBMC and of adoptive transfer of an inflammatory form of EAE by MBP-reactive T-cells. To evaluate the effects of ectopic presentation of MBP on marmoset EAE, animals were vaccinated prior to induction of EAE by subcutaneous injection of attenuated strains of vaccinia virus genetically engineered to contain either the entire coding sequence for human MBP (vT15) or the equine herpes virus glycoprotein gH gene (vAbT249). Vaccination with vT15 was followed by transient cytoplasmic and surface membrane expression of MBP in circulating PBMC (15-45 days). The onset of clinical EAE after immunization (pi) was markedly delayed in vT15-vaccinated animals (37-97 days pi, n = 4) compared to vAbT249-vaccinated controls (14-18 days pi, n = 3). Proliferative responses against MBP but not against vaccinia antigens or phytohemagglutinin were suppressed in protected animals. Thus, development of attenuated live viruses carrying genes for myelin antigens could be useful for induction of immunologic tolerance and for modulation of autoimmune demyelination.

Epidemiological study of hepatitis A, B and C in the largest Afro-Brazilian isolated community.

PubMed

Matos, Márcia A D; Reis, Nádia Rúbia S; Kozlowski, Aline G; Teles, Sheila A; Motta-Castro, Ana Rita C; Mello, Francisco C A; Gomes, Selma A; Martins, Regina M B

2009-09-01

This study was conducted to estimate the prevalence and molecular epidemiological features of viral hepatitis A, B and C in the Kalunga population, which represents the largest Afro-Brazilian isolated community. Among 878 individuals studied, the overall prevalence of anti-hepatitis A virus antibodies was 80.9%, with a significant rise from 44.8% to near 100% between the first and fourth decade of life. Rates for hepatitis B surface antigen (HBsAg) and antibody to hepatitis B core antigen (anti-HBc) of 1.8% and 35.4%, respectively, were found. Increasing age, male gender, illiteracy and history of multiple sexual partners were associated with hepatitis B virus (HBV) infection. An occult HBV infection rate of 1.7% (5/295) was found among anti-HBc-positive individuals. HBV genotype A (subtype Aa) was dominant in this community. Only 5/878 individuals (0.6%) were positive for anti-hepatitis C virus (HCV). HCV RNA was detected in three of them, who were infected with genotype 1 (subtype 1a). These findings point out high, intermediate and low endemicity for hepatitis A, B and C, respectively, in the Kalunga community in Brazil. Circulation of HBV genotype A (subtype Aa) in this Afro-Brazilian isolated community indicates the introduction of this virus during the slave trade from Africa to Brazil.

Transformation of Mouse Macrophages by Simian Virus 40

PubMed Central

Stone, Lawrence B.; Takemoto, Kenneth K.

1970-01-01

Studies were undertaken to prove that simian virus 40 (SV40) can transform the mouse macrophage, a cell type naturally restricted from deoxyribonucleic acid (DNA) replication. Balb/C macrophages infected with SV40 demonstrated T-antigen production and induced DNA synthesis simultaneously. In the absence of apparent division, these cells remained T antigen-positive for at least 45 days. SV40 could be rescued from nondividing, unaltered macrophages during the T antigen-producing period. Proliferating transformants appeared at an average of 66 days post-SV40 infection. Established cell lines were T antigen-positive and were negative for infectious virus, but yielded SV40 after fusion with African green monkey kidney cells. Their identity as transformed macrophages was substantiated by evaluation of cellular morphology, phagocytosis, acid phosphatase, β1c synthesis, and aminoacridine incorporation. Images PMID:4320698

Statistical analysis of nucleotide sequences of the hemagglutinin gene of human influenza A viruses.

PubMed Central

Ina, Y; Gojobori, T

1994-01-01

To examine whether positive selection operates on the hemagglutinin 1 (HA1) gene of human influenza A viruses (H1 subtype), 21 nucleotide sequences of the HA1 gene were statistically analyzed. The nucleotide sequences were divided into antigenic and nonantigenic sites. The nucleotide diversities for antigenic and nonantigenic sites of the HA1 gene were computed at synonymous and nonsynonymous sites separately. For nonantigenic sites, the nucleotide diversities were larger at synonymous sites than at nonsynonymous sites. This is consistent with the neutral theory of molecular evolution. For antigenic sites, however, the nucleotide diversities at nonsynonymous sites were larger than those at synonymous sites. These results suggest that positive selection operates on antigenic sites of the HA1 gene of human influenza A viruses (H1 subtype). PMID:8078892

BST2/CD317 counteracts human coronavirus 229E productive infection by tethering virions at the cell surface

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Shiu-Mei; Institute of Clinical Medicine, National Yang-Ming University School of Medicine, Taipei, Taiwan; Huang, Kuo-Jung

2014-01-20

Bone marrow stromal antigen 2 (BST2), an interferon-inducible antiviral factor, has been shown to block the release of various enveloped viruses from cells. It has also been identified as an innate immune system component. Most enveloped viruses subject to BST2 restriction bud at the plasma membrane. Here we report our findings that (a) the production of human coronavirus 229E (HCoV-229E) progeny viruses, whose budding occurs at the ER-Golgi intermediate compartment (ERGIC), markedly decreases in the presence of BST2; and (b) BST2 knockdown expression results in enhanced HCoV-229E virion production. Electron microscopy analyses indicate that HCoV-229E virions are tethered to cellmore » surfaces or intracellular membranes by BST2. Our results suggest that BST2 exerts a broad blocking effect against enveloped virus release, regardless of whether budding occurs at the plasma membrane or intracellular compartments. - Highlights: • BST2 knockdown expression results in enhanced HCoV-229E egress. • HCoV-229E virions are tethered to cell surfaces or intracellular membranes by BST2. • HCoV-229E infection at high MOI can significantly downregulate HeLa BST2 and rescue HIV-1 egress.« less

Genetically Diverse Low Pathogenicity Avian Influenza A Virus Subtypes Co-Circulate among Poultry in Bangladesh.

PubMed

Gerloff, Nancy A; Khan, Salah Uddin; Zanders, Natosha; Balish, Amanda; Haider, Najmul; Islam, Ausraful; Chowdhury, Sukanta; Rahman, Mahmudur Ziaur; Haque, Ainul; Hosseini, Parviez; Gurley, Emily S; Luby, Stephen P; Wentworth, David E; Donis, Ruben O; Sturm-Ramirez, Katharine; Davis, C Todd

2016-01-01

Influenza virus surveillance, poultry outbreak investigations and genomic sequencing were assessed to understand the ecology and evolution of low pathogenicity avian influenza (LPAI) A viruses in Bangladesh from 2007 to 2013. We analyzed 506 avian specimens collected from poultry in live bird markets and backyard flocks to identify influenza A viruses. Virus isolation-positive specimens (n = 50) were subtyped and their coding-complete genomes were sequenced. The most frequently identified subtypes among LPAI isolates were H9N2, H11N3, H4N6, and H1N1. Less frequently detected subtypes included H1N3, H2N4, H3N2, H3N6, H3N8, H4N2, H5N2, H6N1, H6N7, and H7N9. Gene sequences were compared to publicly available sequences using phylogenetic inference approaches. Among the 14 subtypes identified, the majority of viral gene segments were most closely related to poultry or wild bird viruses commonly found in Southeast Asia, Europe, and/or northern Africa. LPAI subtypes were distributed over several geographic locations in Bangladesh, and surface and internal protein gene segments clustered phylogenetically with a diverse number of viral subtypes suggesting extensive reassortment among these LPAI viruses. H9N2 subtype viruses differed from other LPAI subtypes because genes from these viruses consistently clustered together, indicating this subtype is enzootic in Bangladesh. The H9N2 strains identified in Bangladesh were phylogenetically and antigenically related to previous human-derived H9N2 viruses detected in Bangladesh representing a potential source for human infection. In contrast, the circulating LPAI H5N2 and H7N9 viruses were both phylogenetically and antigenically unrelated to H5 viruses identified previously in humans in Bangladesh and H7N9 strains isolated from humans in China. In Bangladesh, domestic poultry sold in live bird markets carried a wide range of LPAI virus subtypes and a high diversity of genotypes. These findings, combined with the seven year timeframe of sampling, indicate a continuous circulation of these viruses in the country.

Genetically Diverse Low Pathogenicity Avian Influenza A Virus Subtypes Co-Circulate among Poultry in Bangladesh

PubMed Central

Gerloff, Nancy A.; Khan, Salah Uddin; Zanders, Natosha; Balish, Amanda; Haider, Najmul; Islam, Ausraful; Chowdhury, Sukanta; Rahman, Mahmudur Ziaur; Haque, Ainul; Hosseini, Parviez; Gurley, Emily S.; Luby, Stephen P.; Wentworth, David E.; Donis, Ruben O.; Sturm-Ramirez, Katharine; Davis, C. Todd

2016-01-01

Influenza virus surveillance, poultry outbreak investigations and genomic sequencing were assessed to understand the ecology and evolution of low pathogenicity avian influenza (LPAI) A viruses in Bangladesh from 2007 to 2013. We analyzed 506 avian specimens collected from poultry in live bird markets and backyard flocks to identify influenza A viruses. Virus isolation-positive specimens (n = 50) were subtyped and their coding-complete genomes were sequenced. The most frequently identified subtypes among LPAI isolates were H9N2, H11N3, H4N6, and H1N1. Less frequently detected subtypes included H1N3, H2N4, H3N2, H3N6, H3N8, H4N2, H5N2, H6N1, H6N7, and H7N9. Gene sequences were compared to publicly available sequences using phylogenetic inference approaches. Among the 14 subtypes identified, the majority of viral gene segments were most closely related to poultry or wild bird viruses commonly found in Southeast Asia, Europe, and/or northern Africa. LPAI subtypes were distributed over several geographic locations in Bangladesh, and surface and internal protein gene segments clustered phylogenetically with a diverse number of viral subtypes suggesting extensive reassortment among these LPAI viruses. H9N2 subtype viruses differed from other LPAI subtypes because genes from these viruses consistently clustered together, indicating this subtype is enzootic in Bangladesh. The H9N2 strains identified in Bangladesh were phylogenetically and antigenically related to previous human-derived H9N2 viruses detected in Bangladesh representing a potential source for human infection. In contrast, the circulating LPAI H5N2 and H7N9 viruses were both phylogenetically and antigenically unrelated to H5 viruses identified previously in humans in Bangladesh and H7N9 strains isolated from humans in China. In Bangladesh, domestic poultry sold in live bird markets carried a wide range of LPAI virus subtypes and a high diversity of genotypes. These findings, combined with the seven year timeframe of sampling, indicate a continuous circulation of these viruses in the country. PMID:27010791

Patterns of amino acid conservation in human and animal immunodeficiency viruses.

PubMed

Voitenko, Olga S; Dhroso, Andi; Feldmann, Anna; Korkin, Dmitry; Kalinina, Olga V

2016-09-01

Due to their high genomic variability, RNA viruses and retroviruses present a unique opportunity for detailed study of molecular evolution. Lentiviruses, with HIV being a notable example, are one of the best studied viral groups: hundreds of thousands of sequences are available together with experimentally resolved three-dimensional structures for most viral proteins. In this work, we use these data to study specific patterns of evolution of the viral proteins, and their relationship to protein interactions and immunogenicity. We propose a method for identification of two types of surface residues clusters with abnormal conservation: extremely conserved and extremely variable clusters. We identify them on the surface of proteins from HIV and other animal immunodeficiency viruses. Both types of clusters are overrepresented on the interaction interfaces of viral proteins with other proteins, nucleic acids or low molecular-weight ligands, both in the viral particle and between the virus and its host. In the immunodeficiency viruses, the interaction interfaces are not more conserved than the corresponding proteins on an average, and we show that extremely conserved clusters coincide with protein-protein interaction hotspots, predicted as the residues with the largest energetic contribution to the interaction. Extremely variable clusters have been identified here for the first time. In the HIV-1 envelope protein gp120, they overlap with known antigenic sites. These antigenic sites also contain many residues from extremely conserved clusters, hence representing a unique interacting interface enriched both in extremely conserved and in extremely variable clusters of residues. This observation may have important implication for antiretroviral vaccine development. A Python package is available at https://bioinf.mpi-inf.mpg.de/publications/viral-ppi-pred/ voitenko@mpi-inf.mpg.de or kalinina@mpi-inf.mpg.de Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Poly I:C adjuvanted inactivated swine influenza vaccine induces heterologous protective immunity in pigs.

PubMed

Thomas, Milton; Wang, Zhao; Sreenivasan, Chithra C; Hause, Ben M; Gourapura J Renukaradhya; Li, Feng; Francis, David H; Kaushik, Radhey S; Khatri, Mahesh

2015-01-15

Swine influenza is widely prevalent in swine herds in North America and Europe causing enormous economic losses and a public health threat. Pigs can be infected by both avian and mammalian influenza viruses and are sources of generation of reassortant influenza viruses capable of causing pandemics in humans. Current commercial vaccines provide satisfactory immunity against homologous viruses; however, protection against heterologous viruses is not adequate. In this study, we evaluated the protective efficacy of an intranasal Poly I:C adjuvanted UV inactivated bivalent swine influenza vaccine consisting of Swine/OH/24366/07 H1N1 and Swine/CO/99 H3N2, referred as PAV, in maternal antibody positive pigs against an antigenic variant and a heterologous swine influenza virus challenge. Groups of three-week-old commercial-grade pigs were immunized intranasally with PAV or a commercial vaccine (CV) twice at 2 weeks intervals. Three weeks after the second immunization, pigs were challenged with the antigenic variant Swine/MN/08 H1N1 (MN08) and the heterologous Swine/NC/10 H1N2 (NC10) influenza virus. Antibodies in serum and respiratory tract, lung lesions, virus shedding in nasal secretions and virus load in lungs were assessed. Intranasal administration of PAV induced challenge viruses specific-hemagglutination inhibition- and IgG antibodies in the serum and IgA and IgG antibodies in the respiratory tract. Importantly, intranasal administration of PAV provided protection against the antigenic variant MN08 and the heterologous NC10 swine influenza viruses as evidenced by significant reductions in lung virus load, gross lung lesions and significantly reduced shedding of challenge viruses in nasal secretions. These results indicate that Poly I:C or its homologues may be effective as vaccine adjuvants capable of generating cross-protective immunity against antigenic variants/heterologous swine influenza viruses in pigs. Copyright © 2014 Elsevier Ltd. All rights reserved.

An Overview of Live Attenuated Recombinant Pseudorabies Viruses for Use as Novel Vaccines

PubMed Central

Dong, Bo; Zarlenga, Dante S.; Ren, Xiaofeng

2014-01-01

Pseudorabies virus (PRV) is a double-stranded, DNA-based swine virus with a genome approximating 150 kb in size. PRV has many nonessential genes which can be replaced with genes encoding heterologous antigens but without deleterious effects on virus propagation. Recombinant PRVs expressing both native and foreign antigens are able to stimulate immune responses. In this paper, we review the current status of live attenuated recombinant PRVs and live PRV-based vector vaccines with potential for controlling viral infections in animals. PMID:24995348

Characterization of the disassembly and reassembly of the HBV glycoprotein surface antigen, a pliable nanoparticle vaccine platform

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gallagher, John R.; Torian, Udana; McCraw, Dustin

While nanoparticle vaccine technology is gaining interest due to the success of vaccines like those for the human papillomavirus that is based on viral capsid nanoparticles, little information is available on the disassembly and reassembly of viral surface glycoprotein-based nanoparticles. One such particle is the hepatitis B virus surface antigen (sAg) that exists as nanoparticles. Here we show, using biochemical analysis coupled with electron microscopy, that sAg nanoparticle disassembly requires both reducing agent to disrupt intermolecular disulfide bonds, and detergent to disrupt hydrophobic interactions that stabilize the nanoparticle. Particles were otherwise resistant to salt and urea, suggesting the driving mechanismmore » of particle formation involves hydrophobic interactions. We reassembled isolated sAg protein into nanoparticles by detergent removal and reassembly resulted in a wider distribution of particle diameters. Knowledge of these driving forces of nanoparticle assembly and stability should facilitate construction of epitope-displaying nanoparticles that can be used as immunogens in vaccines.« less

Characterization of the disassembly and reassembly of the HBV glycoprotein surface antigen, a pliable nanoparticle vaccine platform.

PubMed

Gallagher, John R; Torian, Udana; McCraw, Dustin M; Harris, Audray K

2017-02-01

While nanoparticle vaccine technology is gaining interest due to the success of vaccines like those for the human papillomavirus that is based on viral capsid nanoparticles, little information is available on the disassembly and reassembly of viral surface glycoprotein-based nanoparticles. One such particle is the hepatitis B virus surface antigen (sAg) that exists as nanoparticles. Here we show, using biochemical analysis coupled with electron microscopy, that sAg nanoparticle disassembly requires both reducing agent to disrupt intermolecular disulfide bonds, and detergent to disrupt hydrophobic interactions that stabilize the nanoparticle. Particles were otherwise resistant to salt and urea, suggesting the driving mechanism of particle formation involves hydrophobic interactions. We reassembled isolated sAg protein into nanoparticles by detergent removal and reassembly resulted in a wider distribution of particle diameters. Knowledge of these driving forces of nanoparticle assembly and stability should facilitate construction of epitope-displaying nanoparticles that can be used as immunogens in vaccines. Published by Elsevier Inc.

Emergence of Influenza A Virus Variants after Prolonged Shedding from Pheasants▿

PubMed Central

Humberd, Jennifer; Boyd, Kelli; Webster, Robert G.

2007-01-01

We previously demonstrated the susceptibility of pheasants to infection with influenza A viruses of 15 hemagglutinin (HA) subtypes: 13/23 viruses tested were isolated for ≥14 days, all in the presence of serum-neutralizing antibodies; one virus (H10) was shed for 45 days postinfection. Here we confirmed that 20% of pheasants shed low-pathogenic influenza viruses for prolonged periods. We aimed to determine why the antibody response did not clear the virus in the usual 3 to 10 days, because pheasants serve as a long-term source of influenza viruses in poultry markets. We found evidence of virus replication and histological changes in the large intestine, bursa of Fabricius, and cecal tonsil. The virus isolated 41 days postinfection was antigenically distinct from the parental H10 virus, with corresponding changes in the HA and neuraminidase. Ten amino acid differences were found between the parental H10 and the pheasant H10 virus; four were in potential antigenic sites of the HA molecule. Prolonged shedding of virus by pheasants results from a complex interplay between the diversity of virus variants and the host response. It is often argued that vaccination pressure is a mechanism that contributes to the generation of antigenic-drift variants in poultry. This study provided evidence that drift variants can occur naturally in pheasants after prolonged shedding of virus, thus strengthening our argument for the removal of pheasants from live-bird retail markets. PMID:17267493

Expression of a foot-and-mouth disease virus immunodominant epitope by a filamentous bacteriophage vector.

PubMed

Kim, Y J; Lebreton, F; Kaiser, C; Crucière, C; Rémond, M

2004-02-01

We described the construction of a recombinant filamentous phage displaying on its surface the immunodominant site of VP1 protein of foot-and-mouth disease virus (FMDV). The coding sequence was inserted at the amino-terminus of the major coat protein pVIII via a spacer. The hybrid phage proved to be antigenic as it was recognized by polyclonal and monoclonal anti FMDV sera. In two experiments involving immunisation of guinea-pigs with the recombinant phage, a low antibody response was generated. This suggests a possible role for phage displayed peptides in inducing anti FMDV immunity and the possibility of further development is discussed.

Genogroup IV and VI Canine Noroviruses Interact with Histo-Blood Group Antigens

PubMed Central

Breiman, Adrien; le Pendu, Jacques

2014-01-01

ABSTRACT Human noroviruses (HuNV) are a significant cause of viral gastroenteritis in humans worldwide. HuNV attaches to cell surface carbohydrate structures known as histo-blood group antigens (HBGAs) prior to internalization, and HBGA polymorphism among human populations is closely linked to susceptibility to HuNV. Noroviruses are divided into 6 genogroups, with human strains grouped into genogroups I (GI), II, and IV. Canine norovirus (CNV) is a recently discovered pathogen in dogs, with strains classified into genogroups IV and VI. Whereas it is known that GI to GIII noroviruses bind to HBGAs and GV noroviruses recognize terminal sialic acid residues, the attachment factors for GIV and GVI noroviruses have not been reported. This study sought to determine the carbohydrate binding specificity of CNV and to compare it to the binding specificities of noroviruses from other genogroups. A panel of synthetic oligosaccharides were used to assess the binding specificity of CNV virus-like particles (VLPs) and identified α1,2-fucose as a key attachment factor. CNV VLP binding to canine saliva and tissue samples using enzyme-linked immunosorbent assays (ELISAs) and immunohistochemistry confirmed that α1,2-fucose-containing H and A antigens of the HBGA family were recognized by CNV. Phenotyping studies demonstrated expression of these antigens in a population of dogs. The virus-ligand interaction was further characterized using blockade studies, cell lines expressing HBGAs, and enzymatic removal of candidate carbohydrates from tissue sections. Recognition of HBGAs by CNV provides new insights into the evolution of noroviruses and raises concerns regarding the potential for zoonotic transmission of CNV to humans. IMPORTANCE Infections with human norovirus cause acute gastroenteritis in millions of people each year worldwide. Noroviruses can also affect nonhuman species and are divided into 6 different groups based on their capsid sequences. Human noroviruses in genogroups I and II interact with histo-blood group antigen carbohydrates, bovine noroviruses (genogroup III) interact with alpha-galactosidase (α-Gal) carbohydrates, and murine norovirus (genogroup V) recognizes sialic acids. The canine-specific strains of norovirus are grouped into genogroups IV and VI, and this study is the first to characterize which carbohydrate structures they can recognize. Using canine norovirus virus-like particles, this work shows that representative genogroup IV and VI viruses can interact with histo-blood group antigens. The binding specificity of canine noroviruses is therefore very similar to that of the human norovirus strains classified into genogroups I and II. This raises interesting questions about the evolution of noroviruses and suggests it may be possible for canine norovirus to infect humans. PMID:25008923

Improve T Cell Therapy in Neuroblastoma

DTIC Science & Technology

2012-07-01

Epstein - Barr - virus (EBV)-specific cytotoxic T lymphocytes (EBV-CTLs) genetically modified to express a chimeric antigen receptor (CAR-GD2) targeting the...A. Krance, M. K. Brenner, and C. M. Rooney. 1996. Long-term restoration of immunity against Epstein - Barr virus infection by adoptive transfer of gene... Barr - virus (EBV)- specific cytotoxic T l ymphocytes (EBV-CTLs) genetically modified to express a c himeric antigen receptor (CAR-GD2) targeting the GD2

Gold nanorod vaccine for respiratory syncytial virus

NASA Astrophysics Data System (ADS)

Stone, John W.; Thornburg, Natalie J.; Blum, David L.; Kuhn, Sam J.; Wright, David W.; Crowe, James E., Jr.

2013-07-01

Respiratory syncytial virus (RSV) is a major cause of pneumonia and wheezing in infants and the elderly, but to date there is no licensed vaccine. We developed a gold nanorod construct that displayed the major protective antigen of the virus, the fusion protein (F). Nanorods conjugated to RSV F were formulated as a candidate vaccine preparation by covalent attachment of viral protein using a layer-by-layer approach. In vitro studies using ELISA, electron microscopy and circular dichroism revealed that conformation-dependent epitopes were maintained during conjugation, and transmission electron microscopy studies showed that a dispersed population of particles could be achieved. Human dendritic cells treated with the vaccine induced immune responses in primary human T cells. These results suggest that this vaccine approach may be a potent method for immunizing against viruses such as RSV with surface glycoproteins that are targets for the human immune response.

Rapid Detection of Ebola Virus with a Reagent-Free, Point-of-Care Biosensor

PubMed Central

Baca, Justin T.; Severns, Virginia; Lovato, Debbie; Branch, Darren W.; Larson, Richard S.

2015-01-01

Surface acoustic wave (SAW) sensors can rapidly detect Ebola antigens at the point-of-care without the need for added reagents, sample processing, or specialized personnel. This preliminary study demonstrates SAW biosensor detection of the Ebola virus in a concentration-dependent manner. The detection limit with this methodology is below the average level of viremia detected on the first day of symptoms by PCR. We observe a log-linear sensor response for highly fragmented Ebola viral particles, with a detection limit corresponding to 1.9 × 104 PFU/mL prior to virus inactivation. We predict greatly improved sensitivity for intact, infectious Ebola virus. This point-of-care methodology has the potential to detect Ebola viremia prior to symptom onset, greatly enabling infection control and rapid treatment. This biosensor platform is powered by disposable AA batteries and can be rapidly adapted to detect other emerging diseases in austere conditions. PMID:25875186

Mimicry of the immunodominant conformation-dependent antigenic site of hepatitis A virus by motifs selected from synthetic peptide libraries.

PubMed

Mattioli, S; Imberti, L; Stellini, R; Primi, D

1995-09-01

Hepatitis A virus (HAV) is a positive-strand RNA virus with a genome length of approximately 7,480 nucleotides. Although HAV morphogenesis is thought to be similar to that of poliovirus, the prototype picornavirus, the complete characterization of the antigenic structure of this virus remains elusive. All the available evidences, however, support the existence, on HAV virions and empty capsids, of an immunodominant neutralization antigenic site which is conformation dependent and whose structure involves residues of both VP1 and VP3 capsid proteins. This particular feature and the difficulty of obtaining high virus yield in tissue cultures make HAV an ideal target for developing synthetic peptides that simulate the structure of its main antigenic determinant. To this end we utilized, in the present work, the divide-couple-recombine approach to generate a random library composed of millions of different hexapeptides. This vast library was screened with a well-characterized anti-HAV monoclonal antibody. By this strategy we identified a peptide that reacted specifically with monoclonal and polyclonal anti-HAV antibodies and, in mice, induced a specific anti-virus immune response. Furthermore, the peptide could also be used in an enzyme-linked immunosorbent assay for revealing a primary immunoglobulin M immune response in sera of acutely infected human patients. Interestingly, no sequence homology was found between the identified peptide and the HAV capsid proteins VP1 and VP3. Collectively, these data represent an additional important paradigm of a mimotope capable of mimicking an antigenic determinant with unknown tertiary structure.

Hepatitis B surface antigen levels during natural history of chronic hepatitis B: a Chinese perspective study.

PubMed

Zeng, Lin-Yan; Lian, Jiang-Shan; Chen, Jian-Yang; Jia, Hong-Yu; Zhang, Yi-Min; Xiang, Dai-Rong; Yu, Liang; Hu, Jian-Hua; Lu, Ying-Feng; Zheng, Lin; Li, Lan-Juan; Yang, Yi-Da

2014-07-21

To determine the baseline hepatitis B surface antigen (HBsAg) levels during the different phases of chronic hepatitis B (CHB) patients in China. Six hundred and twenty-three hepatitis B virus or un-infected patients not receiving antiviral therapy were analyzed in a cross-sectional study. The CHB patients were classified into five phases: immune-tolerant (IT, n = 108), immune-clearance (IC, n = 161), hepatitis B e antigen negative hepatitis (ENH, n = 149), low-replicative (LR, n = 135), and liver cirrhosis (LC, n = 70). HBsAg was quantified (Abbott ARCHITECT assay) and correlated with hepatitis B virus (HBV) DNA, and serum alanine aminotransferase/aspartate aminotransferase (ALT/AST) in each phase of CHB was also determined. Median HBsAg titers were different in each phase of CHB (P < 0.001): IT (4.85 log10 IU/mL), IC (4.36 log10 IU/mL), ENH (2.95 log10 IU/mL), LR (3.18 log10 IU/mL) and LC (2.69 log10 IU/mL). HBsAg titers were highest in the IT phase and lowest in the LC phase. Serum HBsAg titers showed a strong correlation with HBV viral load in the IC phase (r = 0.683, P < 0.001). No correlation between serum HBsAg level and ALT/AST was observed. The mean baseline HBsAg levels differ significantly during the five phases of CHB, providing evidence on the natural history of HBV infection. HBsAg quantification may predict the effects of immune-modulator or oral nucleos(t)ide analogue therapy.

Isolation of circulating immune complexes using Raji cells. Separation of antigens from immune complexes and production of antiserum.

PubMed Central

Theofilopoulos, A N; Eisenberg, R A; Dixon, F J

1978-01-01

Raji cells were used for the isolation of complement-fixing antigen-antibody complexes from serum. Immune complexes bound to these cells were radiolabeled at the cell surface with lactoperoxidase. The complexes were then eluted from the cells with isotonic citrate buffer pH 3.2 or recovered by immunoprecipitation of cell lysates. The antigen and antibody moieties of the complexes were isolated by dissociating sucrose density gradient centrifugation or by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A variety of preformed immune complexes were successfully isolated from serum with this approach. In addition, these techniques were used to isolate and identify the antigens in immune complexes in the serum of rabbits with chronic serum sickness and rats with Moloney virus-induced sarcomas. Methods were also developed for the production of antisera against the antigenic moiety of immune complexes isolated from serum. Repeated challenge of rabbits with whole Raji cells with bound complexes or eluates from such cells resulted in antibody production against the antigens of the immune complexes, although reactivity against cellular and serum components was also elicited. Monospecific antisera against the antigens in immune complexes were produced by immunizing rabbits with the alum-precipitated antigen isolated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These techniques may be useful in isolating antigens in immune complex-associated diseases of unknown etiology. Images PMID:659616

Gravisensitivity of various host plant -virus systems in simulated microgravity

NASA Astrophysics Data System (ADS)

Mishchenko, Lidiya; Taran, Oksana; Gordejchyk, Olga

In spite of considerable achievements in the study of gravity effects on plant development, some issues of gravitropism, like species-specificity and gravitation response remain unclear. The so-lution of such problems is connected with the aspects of life supply, in piloted space expeditions. The role of microgravity remains practically unstudied in the development of relations in the system host plant-virus, which are important for biotechnologies in crop production. It is ev-ident that the conditions of space flight can act as stressors, and the stress inducted by them favors the reactivation of latest herpes viruses in humans (satish et al., 2009) Viral infections of plants, which also can be in a latest state at certain stages of plant organism development, cause great damage to the growth and development of a host plant. Space flight conditions may cause both reactivation of latent viral infection in plants and its elimination, as it has been found by us for the system WSMW -wheat (Mishchenko et al., 2004). Our further research activities were concentrated on the identification of gravisensitivity in the system virus -potato plant to find out whether there was any species -related specificity of the reaction. In our research we used potato plants of Krymska Rosa, Zhuravushka, Agave, Belarosa, Kupalinka, and Zdubytok varieties. Simulated microgravity was ensured by clinostats KG-8 and Cycle -2. Gravisensitiv-ity has been studied the systems including PVX, PVM and PVY. Virus concentrations have been determined by ELISA using LOEWE reagents (placecountry-regionGermany). Virus iden-tification by morphological features were done by electron microscopy. For the system PVX -potato plant, we found the reduction in virus antigens content with prolonged clinostating. On the 18th day of cultivation, the plants showed a high level of X-virus antigen content on both stationary (control) and clinostated variants. On 36th and 47th day, depending plant variety, clinostated plants had lower X-virus antigen content, compared with negative control. In plants, cultivated without clinostating, PVX antigen content was 5-10 times greater than on negative control variants. Prolonged (over 43 days) clinostating, depending on potato plant genotype, may cause both simulation and impeding of the accumulation of Y-virus antigens in potato plants. Studying the interaction between the host plant and PVM, we found that prolonged clinorotation at first reduced the antigen content by 25-30% compared with stationary control. Further on after 44 days of experimentation, the antigen content increased with more intensive increase in non-clinostated plants. Thus, prolonged clinostating reduced the intensity of anti-gen accumulation but did not stop it completely. We admit that proves a low sensitivity of the system PVM -potato plant to simulated microgravity. The phenomena of PVX reproduction in simulated microgravity may find on employment in present-day biotechnologies.

Sero-prevalence of Hepatitis B Virus Infection in Balochistan Province of Pakistan

PubMed Central

Sheikh, Nadeem S.; Sheikh, Azeem S.; Sheikh, Aqleem A.; Yahya, Samira; Rafi-U-Shan; Lateef, Majid

2011-01-01

Background/Aim: The objective was to evaluate the sero-prevalence of hepatitis B surface antigen (HBsAg) and IgM antibodies to hepatitis core antigen in Balochistan Province of Pakistan. Design of the study: A cross-sectional, population-based study. Place and time of the study: The study was conducted in Balochistan from 1st January 2004 to 31st December, 2008. The screening areas included Barkhan, Eashani, Khuzdar, Kodi Zikriani, Kohlu, Rakhni and Turbat. Materials and Methods: A total of 15,260 subjects were enrolled; 11,900 (78%) agreed to undergo screening. Fresh serum samples were tested for the presence of hepatitis B surface antigen and IgM antibodies to hepatitis B core antigen. Results: HBsAg was detected in 1166 (9.8%) while anti-HBc IgM was found in 117 (10.0%). HBsAg positivity was seen in 875 (12.7%) males and 291 (5.8%) females. The prevalence of hepatitis B in Balochistan varies from 3.3% in Khuzdar to 17.0% in Kodi Zikriani. Conclusions: It is utmost important to educate the public, to take proper measures to control the spread of infection and vaccination in order to interrupt transmission of this threatening public health problem in Balochistan province of Pakistan. PMID:21546720

Ebola Virus Disease in Pregnancy: Clinical, Histopathologic, and Immunohistochemical Findings.

PubMed

Muehlenbachs, Atis; de la Rosa Vázquez, Olimpia; Bausch, Daniel G; Schafer, Ilana J; Paddock, Christopher D; Nyakio, Jean Paul; Lame, Papys; Bergeron, Eric; McCollum, Andrea M; Goldsmith, Cynthia S; Bollweg, Brigid C; Prieto, Miriam Alía; Lushima, Robert Shongo; Ilunga, Benoit Kebela; Nichol, Stuart T; Shieh, Wun-Ju; Ströher, Ute; Rollin, Pierre E; Zaki, Sherif R

2017-01-01

Here we describe clinicopathologic features of Ebola virus disease in pregnancy. One woman infected with Sudan virus in Gulu, Uganda, in 2000 had a stillbirth and survived, and another woman infected with Bundibugyo virus had a live birth with maternal and infant death in Isiro, the Democratic Republic of the Congo in 2012. Ebolavirus antigen was seen in the syncytiotrophoblast and placental maternal mononuclear cells by immunohistochemical analysis, and no antigen was seen in fetal placental stromal cells or fetal organs. In the Gulu case, ebolavirus antigen localized to malarial parasite pigment-laden macrophages. These data suggest that trophoblast infection may be a mechanism of transplacental ebolavirus transmission. Published by Oxford University Press for the Infectious Diseases Society of America 2016. This work is written by (a) US Government employee(s) and is in the public domain in the US.

Antibody to hepatitis B surface antigen among employees in the National Hospital, Oslo, Norway: a prevalence study.

PubMed

Hovig, B; Rollag, H; Dahl, O

1985-07-01

During the last decade, several studies of serologic markers of hepatitis B virus infections in hospital personnel have demonstrated an increased prevalence of antibodies to hepatitis B virus (anti-HB) compared with the general population. Norway has a very low incidence rate of hepatitis B as seen on a global scale, and this study was performed to evaluate the infection risk by hospital workers in such environments. The employees, 2,546 (94.7% of the population), in the 800-bed National Hospital in Oslo were tested for antibody to hepatitis B surface antigen (anti-HBs) in serum. Five per cent (128 persons) were anti-HBs-positive; this was only slightly higher than that in the general Norwegian population. Male employees were more often positive than females (7.0% vs. 4.4%). Staff more than 50 years of age or with 16 or more years of employment in the health services had a rate twice as high as the rest of the employees. Staff in the porter services (mostly men) had a higher rate than others, whereas the rates in the different professional groups showed no statistical differences. Contrary to many other studies, significant differences in prevalence according to frequency of patient contact or blood handling were not found.

DOE Office of Scientific and Technical Information (OSTI.GOV)

McLellan, Jason S.; Chen, Man; Chang, Jung-San

Respiratory syncytial virus (RSV) is a major cause of pneumonia and bronchiolitis in infants and elderly people. Currently there is no effective vaccine against RSV, but passive prophylaxis with neutralizing antibodies reduces hospitalizations. To investigate the mechanism of antibody-mediated RSV neutralization, we undertook structure-function studies of monoclonal antibody 101F, which binds a linear epitope in the RSV fusion glycoprotein. Crystal structures of the 101F antigen-binding fragment in complex with peptides from the fusion glycoprotein defined both the extent of the linear epitope and the interactions of residues that are mutated in antibody escape variants. The structure allowed for modeling ofmore » 101F in complex with trimers of the fusion glycoprotein, and the resulting models suggested that 101F may contact additional surfaces located outside the linear epitope. This hypothesis was supported by surface plasmon resonance experiments that demonstrated 101F bound the peptide epitope {approx}16,000-fold more weakly than the fusion glycoprotein. The modeling also showed no substantial clashes between 101F and the fusion glycoprotein in either the pre- or postfusion state, and cell-based assays indicated that 101F neutralization was not associated with blocking virus attachment. Collectively, these results provide a structural basis for RSV neutralization by antibodies that target a major antigenic site on the fusion glycoprotein.« less

Genetically Engineered Natural Killer Cells as a Means for Adoptive Tumor Immunotherapy.

PubMed

Michen, Susanne; Temme, Achim

2016-01-01

Natural killer (NK) cells are lymphoid cells of the innate immune system; they stand at the first defense line against viruses and transformed cells. NK cells use an array of germline-encoded activating and inhibitory receptors that sense virus-infected cells or malignant cells displaying altered surface expression of activating and inhibitory NK cell ligands. They exert potent cytotoxic responses to cellular targets and thus are candidate effector cells for immunotherapy of cancer. In particular, the genetic engineering of NK cells with chimeric antigen receptors (CARs) against surface-expressed tumor-associated antigens (TAAs) seems promising. In the allogeneic context, gene-modified NK cells compared to T cells may be superior because they are short-lived effector cells and do not cause graft-versus-host disease. Furthermore, their anti-tumoral activity can be augmented by combinatorial use with therapeutic antibodies, chemotherapeutics, and radiation. Today, efforts are being undertaken for large-scale NK-cell expansion and their genetic engineering for adoptive cell transfer. With the recent advances in understanding the complex biological interactions that regulate NK cells, it is expected that the genetic engineering of NK cells and a combinatorial blockade of immune evasion mechanisms are required to exploit the full potential of NK-cell-based immunotherapies.

Use of PCR in resolving diagnostic difficulties potentially caused by genetic variation of hepatitis B virus.

PubMed Central

van Deursen, F J; Hino, K; Wyatt, D; Molyneaux, P; Yates, P; Wallace, L A; Dow, B C; Carman, W F

1998-01-01

AIMS: To assess the relevance of genetic variants of hepatitis B virus (HBV) and to demonstrate the usefulness of the polymerase chain reaction (PCR) in cases of HBV diagnostic difficulty. METHODS: Five serum samples from patients that presented diagnostic difficulty in routine laboratories were sent to a research laboratory for PCR, and if appropriate, S gene sequencing, in vitro expression, and antigenic analysis. RESULTS: The demonstration of HBV in serum by PCR allowed a definitive diagnosis of current infection. One serum sample with poor reactivity in a diagnostic assay had a minor hepatitis B surface antigen (HBsAg) variant and another with very poor reactivity had multiple variants of HBsAg. Transient HBsAg reactivity was observed in a recently vaccinated patient. A hepatitis Be antigen (HBeAg) false positive reaction was noted in a patient from a well defined risk group for HBV. One patient who was strongly HBsAg/HBeAg positive, but anti-hepatitis B core antibody negative, was viraemic. CONCLUSIONS: PCR may become the gold standard for the diagnosis of current HBV infection. HBV variants are responsible for a proportion of diagnostically difficult cases. Modification of commercial assays is necessary to increase the sensitivity of detection of such variants. PMID:9602690

Effects of amino acid substitutions in hepatitis B virus surface protein on virion secretion, antigenicity, HBsAg and viral DNA.

PubMed

Xiang, Kuan-Hui; Michailidis, Eleftherios; Ding, Hai; Peng, Ya-Qin; Su, Ming-Ze; Li, Yao; Liu, Xue-En; Dao Thi, Viet Loan; Wu, Xian-Fang; Schneider, William M; Rice, Charles M; Zhuang, Hui; Li, Tong

2017-02-01

As important virological markers, serum hepatitis B surface antigen (HBsAg) and hepatitis B virus (HBV) DNA levels show large fluctuations among chronic hepatitis B patients. The aim of this study was to reveal the potential impact and mechanisms of amino acid substitutions in small hepatitis B surface proteins (SHBs) on serum HBsAg and HBV DNA levels. Serum samples from 230 untreated chronic hepatitis B patients with genotype C HBV were analyzed in terms of HBV DNA levels, serological markers of HBV infection and SHBs sequences. In vitro functional analysis of the identified SHBs mutants was performed. Among 230 SHBs sequences, there were 39 (16.96%) sequences with no mutation detected (wild-type) and 191 (83.04%) with single or multiple mutations. SHBs consist of 226 amino acids, of which 104 (46.02%) had mutations in our study. Some mutations (e.g., sE2G, sL21S, sR24K, sT47A/K, sC69stop (sC69∗), sL95W, sL98V, and sG145R) negatively correlated with serum HBsAg levels. HBsAg and HBV DNA levels from this group of patients had a positive correlation (r=0.61, p<0.001). In vitro analysis showed that these mutations reduced extracellular HBsAg and HBV DNA levels by restricting virion secretion and antibody binding capacity. Virion secretion could be rescued for sE2G, sC69∗, and sG145R by co-expression of wild-type HBsAg. The serum HBsAg levels were lower in untreated CHB patients with novel SHBs mutations outside the major antigenic region than those without mutations. Underlying mechanisms include impairment of virion secretion and lower binding affinity to antibodies used for HBsAg measurements. The hepatitis B surface antigen (HBsAg) is a major viral protein of the hepatitis B virus (HBV) secreted into patient blood serum and its quantification value serves as an important marker for the evaluation of chronic HBV infection and antiviral response. We found a few new amino acid substitutions in HBsAg associated with lower serum HBsAg and HBV DNA levels. These different substitutions might impair virion secretion, change the ability of HBsAg to bind to antibodies, or impact HBV replication. These could all result in decreased detectable levels of serum HBsAg. The factors affecting circulating HBsAg level and HBsAg detection are varied and caution is needed when interpreting clinical significance of serum HBsAg levels. Clinical trial number: NCT01088009. Copyright © 2016 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

A host-restricted viral vector for antigen-specific immunization against Lyme disease pathogen.

PubMed

Xiao, Sa; Kumar, Manish; Yang, Xiuli; Akkoyunlu, Mustafa; Collins, Peter L; Samal, Siba K; Pal, Utpal

2011-07-18

Newcastle disease virus (NDV) is an avian virus that is attenuated in primates and is a potential vaccine vector for human use. We evaluated NDV as a vector for expressing selected antigens of the Lyme disease pathogen Borrelia burgdorferi. A series of recombinant NDVs were generated that expressed intracellular or extracellular forms of two B. burgdorferi antigens: namely, the basic membrane protein A (BmpA) and the outer surface protein C (OspC). Expression of the intracellular and extracellular forms of these antigens was confirmed in cultured chicken cells. C3H or Balb/C mice that were immunized intranasally with the NDV vectors mounted vigorous serum antibody responses against the NDV vector, but failed to mount a robust response against either the intracellular or extracellular forms of BmpA or OspC. By contrast, a single immunization of hamsters with the NDV vectors via the intranasal, intramuscular, or intraperitoneal route resulted in rapid and rigorous antibody responses against the intracellular or extracellular forms of BmpA and OspC. When groups of hamsters were separately inoculated with various NDV vectors and challenged with B. burgdorferi (10(8)cells/animal), immunization with vector expressing either intracellular or extracellular BmpA was associated with a significant reduction of the pathogen load in the joints. Taken together, our studies highlighted the importance of NDV as vaccine vector that can be used for simple yet effective immunization of hosts against bacterial infections including Lyme disease. Copyright © 2011 Elsevier Ltd. All rights reserved.

Characterization of a variant strain of Norwalk virus from a food-borne outbreak of gastroenteritis on a cruise ship in Hawaii.

PubMed Central

Herwaldt, B L; Lew, J F; Moe, C L; Lewis, D C; Humphrey, C D; Monroe, S S; Pon, E W; Glass, R I

1994-01-01

A gastroenteritis outbreak affecting at least 217 (41%) of 527 passengers on a cruise ship was caused by a variant strain of Norwalk virus (NV) that is related to but distinct from the prototype NV strain. Consumption of fresh-cut fruit served at two buffets was significantly associated with illness (P < or = 0.01), and a significant dose-response relationship was evident between illness and the number of various fresh-cut fruit items eaten. Seven (58%) of 12 paired serum specimens from ill persons demonstrated at least fourfold rises in antibody response to recombinant NV capsid antigen. A 32-nm small round-structured virus was visualized by electron microscopy in 4 (29%) of 14 fecal specimens, but none of the 8 specimens that were examined by an enzyme immunoassay for NV antigen demonstrated antigen. Four (40%) of 10 fecal specimens were positive by reverse transcriptase-PCR by using primer pairs selected from the polymerase region of NV. In a 145-bp region, the PCR product shared only 72% nucleotide sequence identity with the reference NV strain and 77% nucleotide sequence identity with Southampton virus but shared 95% nucleotide sequence identity with UK2 virus, a United Kingdom reference virus strain. In addition, the outbreak virus was serotyped as UK2 virus by solid-phase immune electron microscopy. The genetic and antigenic divergence of the outbreak strain from the reference NV strain highlights the need for more broadly reactive diagnostic assays and for improved understanding of the relatedness of the NV group of agents. Images PMID:8027335

Enzyme-linked immunosorbent assay for detection of antibodies to Epstein-Barr virus antigens.

PubMed

Voevodin, A F; Pácsa, A S

1983-01-01

Enzyme-Linked Immunosorbent Assay (ELISA) was standardized for measurement of antibody activity of reference human and baboon (Papio hamadryas) sera to soluble Epstein-Barr virus (EBV) antigens. A comparison with the immunofluorescent (IF) method showed that ELISA detects antibody specifically and sensitivity. In ELISA, Herpesvirus Papio (HVP) nuclear antigen (HUPNA) positive baboon serum reacted with EBV nuclear antigen (EBNA), as a further indication of the antigenic similarity between HVP and EBV. Forty-two baboon sera were tested with EBV antigens in both ELISA and IF test. The results showed an agreement between the two methods and also that by the use of EBV antigens, ELISA measures anti-HVP activity of baboon sera. ELISA did not reveal significant difference in antibody activity of 23 baboons with lymphoma and that of 24 healthy baboons. Results provide further data that ELISA can be used effectively in the field of EBV serology.

A label-free optical biosensor for serotyping "unknown" influenza viruses

NASA Astrophysics Data System (ADS)

Zhang, Hanyuan; Henry Dunand, Carole; Wilson, Patrick; Miller, Benjamin L.

2016-05-01

The ability to accurately classify influenza viruses is critical to understanding patterns of infection, vaccine efficacy, and to the process of developing new vaccines. Unfortunately, this task is hampered both by the virus' ability to undergo antigenic drift and shift (rendering it a "previously unknown" strain), and by technological limitations. In an effort to overcome these challenges, we have developed a label-free human monoclonal antibody array for flu serology, using a pattern recognition approach to assign virus serotype. The array is built on the Arrayed Imaging Reflectometry (AIR) platform. AIR relies on the creation of a near-perfect antireflective condition on the surface of a silicon chip. When this antireflective condition is perturbed because of binding to an antibody spot (or other immobilized probe molecule), binding may be sensitively and quantitatively detected as an increase in reflected light. We describe fabrication and characterization of the array, and preliminary testing with isolated influenza hemagglutinin. We anticipate that this approach may be extended to other viruses by expansion of the array.

Antigenic profile of African horse sickness virus serotype 4 VP5 and identification of a neutralizing epitope shared with bluetongue virus and epizootic hemorrhagic disease virus.

PubMed

Martínez-Torrecuadrada, J L; Langeveld, J P; Venteo, A; Sanz, A; Dalsgaard, K; Hamilton, W D; Meloen, R H; Casal, J I

1999-05-10

African horse sickness virus (AHSV) causes a fatal disease in horses. The virus capsid is composed of a double protein layer, the outermost of which is formed by two proteins: VP2 and VP5. VP2 is known to determine the serotype of the virus and to contain the neutralizing epitopes. The biological function of VP5, the other component of the capsid, is unknown. In this report, AHSV VP5, expressed in insect cells alone or together with VP2, was able to induce AHSV-specific neutralizing antibodies. Moreover, two VP5-specific monoclonal antibodies (MAbs) that were able to neutralize the virus in a plaque reduction assay were generated. To dissect the antigenic structure of AHSV VP5, the protein was cloned in Escherichia coli using the pET3 system. The immunoreactivity of both MAbs, and horse and rabbit polyclonal antisera, with 17 overlapping fragments from VP5 was analyzed. The most immunodominant region was found in the N-terminal 330 residues of VP5, defining two antigenic regions, I (residues 151-200) and II (residues 83-120). The epitopes were further defined by PEPSCAN analysis with 12mer peptides, which determined eight antigenic sites in the N-terminal half of the molecule. Neutralizing epitopes were defined at positions 85-92 (PDPLSPGE) for MAb 10AE12 and at 179-185 (EEDLRTR) for MAb 10AC6. Epitope 10AE12 is highly conserved between the different orbiviruses. MAb 10AE12 was able to recognize bluetongue virus VP5 and epizootic hemorrhagic disease virus VP5 by several techniques. These data will be especially useful for vaccine development and diagnostic purposes. Copyright 1999 Academic Press.

Improved impression cytology techniques for the immunopathological diagnosis of superficial viral infections

PubMed Central

Thiel, M; Bossart, W; Bernauer, W

1997-01-01

BACKGROUND—For epidemiological and therapeutic reasons early diagnosis of superficial viral infections is crucial. Conventional microbiological techniques are expensive, time consuming, and not sufficiently sensitive. In this study impression cytology techniques were evaluated to analyse their diagnostic potential in viral infections of the ocular surface.
METHOD—A Biopore membrane device instead of the original impression cytology technique was used to allow better quality and handling of the specimens. The impressions were processed, using monoclonal antibodies and immunoperoxidase or immunofluorescence techniques to assess the presence of herpes simplex virus, varicella zoster virus, or adenovirus antigens. Ocular surface specimens from healthy individuals (n=10) and from patients with suspected viral surface disease (n=19) were studied. Infected and non-infected cell cultures served as controls.
RESULTS—This modified technique of impression cytology allowed the collection of large conjunctival and corneal epithelial cell layers with excellent morphology. Immunocytological staining of these samples provided diagnostic results for all three viruses in patients with viral surface disease.
CONCLUSIONS—The use of Biopore membrane devices for the collection of ocular surface epithelia offers new diagnostic possibilities for external eye diseases. Immunopathological methods that are applied directly on these membrane devices can provide virological results within 1-4 hours. This contributes considerably to the clinical management of patients with infectious diseases of the ocular surface.

 PMID:9505824

[Immunotherapy for refractory viral infections].

PubMed

Morio, Tomohiro; Fujita, Yuriko; Takahashi, Satoshi

Various antiviral agents have been developed, which are sometimes associated with toxicity, development of virus-resistant strain, and high cost. Virus-specific T-cell (VST) therapy provides an alternative curative therapy that can be effective for a prolonged time without eliciting drug resistance. VSTs can be directly separated using several types of capture devices and can be obtained by stimulating peripheral blood mononuclear cells with viral antigens (virus, protein, or peptide) loaded on antigen-presenting cells (APC). APC can be transduced with virus-antigen coding plasmid or pulsed with overlapping peptides. VST therapy has been studied in drug non-responsive viral infections after hematopoietic cell transplantation (HCT). Several previous studies have demonstrated the efficacy of VST therapy without significant severe GVHD. In addition, VSTs from a third-party donor have been prepared and administered for post-HCT viral infection. Although target viruses of VSTs include herpes virus species and polyomavirus species, a wide variety of pathogens, such as papillomavirus, intracellular bacteria, and fungi, can be treated by pathogen-specific T-cells. Perhaps, these specific T-cells could be used for opportunistic infections in other immunocompromised hosts in the near future.

Amino Acid Variation in HLA Class II Proteins Is a Major Determinant of Humoral Response to Common Viruses

PubMed Central

Hammer, Christian; Begemann, Martin; McLaren, Paul J.; Bartha, István; Michel, Angelika; Klose, Beate; Schmitt, Corinna; Waterboer, Tim; Pawlita, Michael; Schulz, Thomas F.; Ehrenreich, Hannelore; Fellay, Jacques

2015-01-01

The magnitude of the human antibody response to viral antigens is highly variable. To explore the human genetic contribution to this variability, we performed genome-wide association studies of the immunoglobulin G response to 14 pathogenic viruses in 2,363 immunocompetent adults. Significant associations were observed in the major histocompatibility complex region on chromosome 6 for influenza A virus, Epstein-Barr virus, JC polyomavirus, and Merkel cell polyomavirus. Using local imputation and fine mapping, we identified specific amino acid residues in human leucocyte antigen (HLA) class II proteins as the most probable causal variants underlying these association signals. Common HLA-DRβ1 haplotypes showed virus-specific patterns of humoral-response regulation. We observed an overlap between variants affecting the humoral response to influenza A and EBV and variants previously associated with autoimmune diseases related to these viruses. The results of this study emphasize the central and pathogen-specific role of HLA class II variation in the modulation of humoral immune response to viral antigens in humans. PMID:26456283

Antigenic and genetic evolution of contemporary swine H1 influenza viruses in the United States

USDA-ARS?s Scientific Manuscript database

Several lineages of influenza A viruses (IAV) currently circulate in North American pigs. Genetic diversity is further increased by the bidirectional transmission of IAV between swine and humans and the subsequent processes of antigenic shift and drift. Such evolution can be the basis for changes in...

African swine fever virus serotype-specific proteins are significant protective antigens for African swine fever

USDA-ARS?s Scientific Manuscript database

African swine fever (ASF) is an emerging disease threat for the swine industry worldwide. No ASF vaccine is available and progress is hindered by lack of knowledge concerning the extent of African swine fever virus (ASFV) strain diversity and the viral antigens conferring type specific protective im...

Characterization of a novel oil-in-water emulsion adjuvant for swine influenza virus and Mycoplasma hyopneumoniae vaccines

USDA-ARS?s Scientific Manuscript database

Vaccines consisting of subunit or inactivated bacteria/virus and potent adjuvants are widely used to control and prevent infectious diseases. Because inactivated and subunit antigens are often less antigenic than live microbes, a growing need exists for the development of new and improved vaccine ad...

Limited antigenic diversity in contemporary H7 avian-origin influenza A viruses from North America

USDA-ARS?s Scientific Manuscript database

Subtype H7 avian–origin influenza A viruses (AIVs) have caused at least 500 confirmed human infections since 2003 and culling of >75 million birds in recent years. Understanding the antigenic diversity and genetic evolution of H7 AIVs is critical for developing effective strategies for disease prev...

Genomic and antigenic characterization of bovine parainfluenze-3 viruses in the United States including modified live virus vaccine (MLV) strains and field strains from cattle

USDA-ARS?s Scientific Manuscript database

This study investigated the genetic and antigenic characterization of parainfluenza-3 virus (PI3V) of cattle. Using molecular tests including real time PCR and viral genome sequencing, PI3V strains could be separated into PI3V types, including PI3V A, PI3V B, and PI3V C. Isolates from cattle with bo...

Challenges and opportunities of using liquid chromatography and mass spectrometry methods to develop complex vaccine antigens as pharmaceutical dosage forms.

PubMed

Hickey, John M; Sahni, Neha; Toth, Ronald T; Kumru, Ozan S; Joshi, Sangeeta B; Middaugh, C Russell; Volkin, David B

2016-10-01

Liquid chromatographic methods, combined with mass spectrometry, offer exciting and important opportunities to better characterize complex vaccine antigens including recombinant proteins, virus-like particles, inactivated viruses, polysaccharides, and protein-polysaccharide conjugates. The current abilities and limitations of these physicochemical methods to complement traditional in vitro and in vivo vaccine potency assays are explored in this review through the use of illustrative case studies. Various applications of these state-of-the art techniques are illustrated that include the analysis of influenza vaccines (inactivated whole virus and recombinant hemagglutinin), virus-like particle vaccines (human papillomavirus and hepatitis B), and polysaccharide linked to protein carrier vaccines (pneumococcal). Examples of utilizing these analytical methods to characterize vaccine antigens in the presence of adjuvants, which are often included to boost immune responses as part of the final vaccine dosage form, are also presented. Some of the challenges of using chromatographic and LC-MS as physicochemical assays to routinely test complex vaccine antigens are also discussed. Copyright © 2016 Elsevier B.V. All rights reserved.

A comparative analysis on the physicochemical properties of tick-borne encephalitis virus envelope protein residues that affect its antigenic properties.

PubMed

Bukin, Yu S; Dzhioev, Yu P; Tkachev, S E; Kozlova, I V; Paramonov, A I; Ruzek, D; Qu, Z; Zlobin, V I

2017-06-15

This work is dedicated to the study of the variability of the main antigenic envelope protein E among different strains of tick-borne encephalitis virus at the level of physical and chemical properties of the amino acid residues. E protein variants were extracted from then NCBI database. Four amino acid residues properties in the polypeptide sequences were investigated: the average volume of the amino acid residue in the protein tertiary structure, the number of amino acid residue hydrogen bond donors, the charge of amino acid residue lateral radical and the dipole moment of the amino acid residue. These physico-chemical properties are involved in antigen-antibody interactions. As a result, 103 different variants of the antigenic determinants of the tick-borne encephalitis virus E protein were found, significantly different by physical and chemical properties of the amino acid residues in their structure. This means that some strains among the natural variants of tick-borne encephalitis virus can potentially escape the immune response induced by the standard vaccine. Copyright © 2017 Elsevier B.V. All rights reserved.

Newcastle disease virus vectored vaccines as bivalent or antigen delivery vaccines

PubMed Central

2017-01-01

Recent advances in reverse genetics techniques make it possible to manipulate the genome of RNA viruses such as Newcastle disease virus (NDV). Several NDV vaccine strains have been used as vaccine vectors in poultry, mammals, and humans to express antigens of different pathogens. The safety, immunogenicity, and protective efficacy of these NDV-vectored vaccines have been evaluated in pre-clinical and clinical studies. The vaccines are safe in mammals, humans, and poultry. Bivalent NDV-vectored vaccines against pathogens of economic importance to the poultry industry have been developed. These bivalent vaccines confer solid protective immunity against NDV and other foreign antigens. In most cases, NDV-vectored vaccines induce strong local and systemic immune responses against the target foreign antigen. This review summarizes the development of NDV-vectored vaccines and their potential use as a base for designing other effective vaccines for veterinary and human use. PMID:28775971

Varicella zoster virus in the temporal artery of a patient with giant cell arteritis.

PubMed

Nagel, Maria A; Khmeleva, Nelly; Boyer, Philip J; Choe, Alexander; Bert, Robert; Gilden, Don

2013-12-15

We recently detected varicella zoster virus (VZV) in the temporal arteries (TA) of 5/24 patients with clinically suspect giant cell arteritis (GCA) whose TAs were GCA-negative pathologically; in those GCA-negative, VZV+TAs, virus antigen predominated in the arterial adventitia, but without medial necrosis and multinucleated giant cells. During our continuing search for VZV antigen in GCA-negative TAs, in the TA of one subject, we found abundant VZV antigen, as well as VZV DNA, in multiple regions (skip areas) of the TA spanning 350 μm, as well as in skeletal muscle adjacent to the infected TA. Additional pathological analysis of sections adjacent to those containing viral antigen revealed inflammation involving the arterial media and abundant multinucleated giant cells characteristic of GCA. Detection of VZV in areas of the TA with pathological features of GCA warrants further correlative pathological-virological analysis of VZV in GCA. © 2013.

Detection of peste des petits ruminants virus antigen using immunofiltration and antigen-competition ELISA methods.

PubMed

Raj, G Dhinakar; Rajanathan, T M C; Kumar, C Senthil; Ramathilagam, G; Hiremath, Geetha; Shaila, M S

2008-06-22

Peste des petits ruminants (PPR) is one of the most economically important diseases affecting sheep and goats in India. An immunofiltration-based test has been developed using either mono-specific serum/monoclonal antibodies (mAb) prepared against a recombinant truncated nucleocapsid protein of rinderpest virus (RPV) cross-reactive with PPR virus. This method consists of coating ocular swab eluate from suspected animals onto a nitrocellulose membrane housed in a plastic module, which is allowed to react with suitable dilutions of a mAb or a mono-specific polyclonal antibody. The antigen-antibody complex formed on the membrane is then detected by protein A-colloidal gold conjugate, which forms a pink colour. In the immunofiltration test, concordant results were obtained using either PPRV mAb or mono-specific serum. Another test, an antigen-competition ELISA which relies on the competition between plate-coated recombinant truncated 'N' protein of RPV and the PPRV 'N' protein present in ocular swab eluates (sample) for binding to the mono-specific antibody against N protein of RPV (in liquid phase) was developed. The cut-off value for this test was established using reverse transcription polymerase chain reaction (RT-PCR) positive and negative oculo-nasal swab samples. Linear correlation between percent inhibition (PI) values in antigen-competition ELISA and virus infectivity titres was 0.992. Comparison of the immunofiltration test with the antigen-competition ELISA yielded a sensitivity of 80% and specificity of 100%. These two tests can serve as a screening (immunofiltration) and confirmatory (antigen-competition ELISA) test, respectively, in the diagnosis of PPR in sheep or goats.

Immunity to herpes simplex virus type 2. Suppression of virus-induced immune responses in ultraviolet B-irradiated mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yasumoto, S.; Hayashi, Y.; Aurelian, L.

1987-10-15

Ultraviolet B irradiation (280 to 320 nm) of mice at the site of intradermal infection with herpes simplex virus type 2 increased the severity of the herpes simplex virus type 2 disease and decreased delayed-type hypersensitivity (DTH) responses to viral antigen. Decrease in DTH resulted from the induction of suppressor T cells, as evidenced by the ability of spleen cells from UV-irradiated mice to inhibit DTH and proliferative responses after adoptive transfer. Lymph node cells from UV-irradiated animals did not transfer suppression. DTH was suppressed at the induction but not the expression phase. Suppressor T cells were Lyt-1+, L3T4+, andmore » their activity was antigen-specific. However, after in vitro culture of spleen cells from UV-irradiated mice with herpes simplex virus type 2 antigen, suppressor activity was mediated by Lyt-2+ cells. Culture supernatants contained soluble nonantigen-specific suppressive factors.« less

Phylodynamics of H1N1/2009 influenza reveals the transition from host adaptation to immune-driven selection

PubMed Central

Su, Yvonne C. F.; Bahl, Justin; Joseph, Udayan; Butt, Ka Man; Peck, Heidi A.; Koay, Evelyn S. C.; Oon, Lynette L. E.; Barr, Ian G.; Vijaykrishna, Dhanasekaran; Smith, Gavin J. D.

2015-01-01

Influenza A H1N1/2009 virus that emerged from swine rapidly replaced the previous seasonal H1N1 virus. Although the early emergence and diversification of H1N1/2009 is well characterized, the ongoing evolutionary and global transmission dynamics of the virus remain poorly investigated. To address this we analyse >3,000 H1N1/2009 genomes, including 214 full genomes generated from our surveillance in Singapore, in conjunction with antigenic data. Here we show that natural selection acting on H1N1/2009 directly after introduction into humans was driven by adaptation to the new host. Since then, selection has been driven by immunological escape, with these changes corresponding to restricted antigenic diversity in the virus population. We also show that H1N1/2009 viruses have been subject to regular seasonal bottlenecks and a global reduction in antigenic and genetic diversity in 2014. PMID:26245473

Prevalence and risk factors of hepatitis B in Spanish prostitutes.

PubMed Central

Requena Caballero, L.; Requena Caballero, C.; Requena Caballero, I.; Sánchez López, M.; Vázquez López, F.; Romero Guerrero, J.; Casado Jiménez, M.

1987-01-01

Eighty prostitutes were tested by solid-phase radioimmunoassay for serum markers of hepatitis B virus (HBV). Of 8 (10%) with hepatitis B surface antigen (HBsAg), 6 (75%) also had hepatitis Be antigen (HBeAg). Antibodies to HBsAg (anti-HBs) and to hepatitis B core antigen (anti-HBc) were found in 52 (65%). Antibodies to HBeAg (anti-HBe) were positive in 32 (40%). Anti-HBc alone was found in 5 (6%) and anti-HBs alone in 2 (2%). Sixty-seven (84%) were positive for at least one HBV marker and 13 (16%) were still susceptible to infection. Hepatitis B markers were more prevalent in prostitutes than in the normal Spanish population. Age, a history of sexually transmitted diseases (STD), drug abuse and promiscuity are factors which were highly related to hepatitis B markers. We concluded that screening prostitutes for the presence of markers and vaccinating those who are negative would be worth while. PMID:3428379

Prior infection of pigs with a recent human H3N2 influenza virus confers minimal cross-protection against a European swine H3N2 virus.

PubMed

Qiu, Yu; van der Meulen, Karen; Van Reeth, Kristien

2013-11-01

H3N2 influenza viruses circulating in humans and European pigs originate from the pandemic A/Hong Kong/68 virus. Because of slower antigenic drift in swine, the antigenic divergence between swine and human viruses has been increasing. It remains unknown to what extent this results in a reduced cross-protection between recent human and swine H3N2 influenza viruses. We examined whether prior infection of pigs with an old [A/Victoria/3/75 (A/Vic/75)] or a more recent [A/Wisconsin/67/05 (A/Wis/05)] human H3N2 virus protected against a European swine H3N2 virus [sw/Gent/172/08 (sw/Gent/08)]. Genetic and antigenic relationships between sw/Gent/08 and a selection of human H3N2 viruses were also assessed. After challenge with sw/Gent/08, all challenge controls had high virus titers in the entire respiratory tract at 3 days post-challenge and nasal virus excretion for 5-6 days. Prior infection with sw/Gent/08 or A/Vic/75 offered complete virological protection against challenge. Pigs previously inoculated with A/Wis/05 showed similar virus titers in the respiratory tract as challenge controls, but the mean duration of nasal shedding was 1·3 days shorter. Unlike sw/Gent/08- and A/Vic/75-inoculated pigs, A/Wis/05-inoculated pigs lacked cross-reactive neutralizing antibodies against sw/Gent/08 before challenge, but they showed a more rapid antibody response to sw/Gent/08 than challenge controls after challenge. Cross-protection and serological responses correlated with genetic and antigenic differences. Infection immunity to a recent human H3N2 virus confers minimal cross-protection against a European swine H3N2 virus. We discuss our findings with regard to the recent zoonotic infections of humans in the United States with a swine-origin H3N2 variant virus. © 2013 John Wiley & Sons Ltd.

Definition of epitopes and antigens recognized by vaccinia specific immune responses: their conservation in variola virus sequences, and use as a model system to study complex pathogens.

PubMed

Sette, Alessandro; Grey, Howard; Oseroff, Carla; Peters, Bjoern; Moutaftsi, Magdalini; Crotty, Shane; Assarsson, Erika; Greenbaum, Jay; Kim, Yohan; Kolla, Ravi; Tscharke, David; Koelle, David; Johnson, R Paul; Blum, Janice; Head, Steven; Sidney, John

2009-12-30

In the last few years, a wealth of information has become available relating to the targets of vaccinia virus (VACV)-specific CD4(+) T cell, CD8(+) T cell and antibody responses. Due to the large size of its genome, encoding more than 200 different proteins, VACV represents a useful model system to study immunity to complex pathogens. Our data demonstrate that both cellular and humoral responses target a large number of antigens and epitopes. This broad spectrum of targets is detected in both mice and humans. CD4(+) T cell responses target late and structural antigens, while CD8(+) T cells preferentially recognize early antigens. While both CD4(+) and CD8(+) T cell responses target different types of antigens, the antigens recognized by T(H) cells are highly correlated with those recognized by antibody responses. We further show that protein abundance and antibody recognition can be used to predict antigens recognized by CD4(+) T cell responses, while early expression at the mRNA level predicts antigens targeted by CD8(+) T cells. Finally, we find that the vast majority of VACV epitopes are conserved in variola virus (VARV), thus suggesting that the epitopes defined herein also have relevance for the efficacy of VACV as a smallpox vaccine.

Update: Influenza Activity - United States, October 1-November 25, 2017.

PubMed

Dugan, Vivien G; Blanton, Lenee; Elal, Anwar Isa Abd; Alabi, Noreen; Barnes, John; Brammer, Lynnette; Burns, Erin; Cummings, Charisse N; Davis, Todd; Flannery, Brendan; Fry, Alicia M; Garg, Shikha; Garten, Rebecca; Gubareva, Larisa; Jang, Yunho; Kniss, Krista; Kramer, Natalie; Lindstrom, Stephen; Mustaquim, Desiree; O'Halloran, Alissa; Olsen, Sonja J; Sessions, Wendy; Taylor, Calli; Trock, Susan; Xu, Xiyan; Wentworth, David E; Katz, Jacqueline; Jernigan, Daniel

2017-12-08

Influenza activity in the United States was low during October 2017, but has been increasing since the beginning of November. Influenza A viruses have been most commonly identified, with influenza A(H3N2) viruses predominating. Several influenza activity indicators were higher than is typically seen for this time of year. The majority of influenza viruses characterized during this period were genetically or antigenically similar to the 2017-18 Northern Hemisphere cell-grown vaccine reference viruses. These data indicate that currently circulating viruses have not undergone significant antigenic drift; however, circulating A(H3N2) viruses are antigenically less similar to egg-grown A(H3N2) viruses used for producing the majority of influenza vaccines in the United States. It is difficult to predict which influenza viruses will predominate in the 2017-18 influenza season; however, in recent past seasons in which A(H3N2) viruses predominated, hospitalizations and deaths were more common, and the effectiveness of the vaccine was lower. Annual influenza vaccination is recommended for all persons aged ≥6 months who do not have contraindications. Multiple influenza vaccines are approved and recommended for use during the 2017-18 season, and vaccination should continue to be offered as long as influenza viruses are circulating and unexpired vaccine is available. This report summarizes U.S. influenza activity* during October 1-November 25, 2017 (surveillance weeks 40-47). † .

Evaluation of an enzyme immunoassay for detection of dengue virus NS1 antigen in human serum.

PubMed

Dussart, Philippe; Labeau, Bhety; Lagathu, Gisèle; Louis, Philippe; Nunes, Marcio R T; Rodrigues, Sueli G; Storck-Herrmann, Cécile; Cesaire, Raymond; Morvan, Jacques; Flamand, Marie; Baril, Laurence

2006-11-01

We evaluated a one-step sandwich-format microplate enzyme immunoassay for detecting dengue virus NS1 antigen (Ag) in human serum by use of Platelia Dengue NS1 Ag kits (Bio-Rad Laboratories, Marnes La Coquette, France). We collected 299 serum samples from patients with dengue disease and 50 serum samples from patients not infected with dengue virus. For the 239 serum samples from patients with acute infections testing positive by reverse transcription-PCR and/or virus isolation for one of the four dengue virus serotypes, the sensitivity of the Platelia Dengue NS1 Ag kit was 88.7% (95% confidence interval, 84.0% to 92.4%). None of the serum samples from patients not infected with dengue virus tested positive with the Platelia Dengue NS1 Ag kit. A diagnostic strategy combining the Platelia Dengue NS1 Ag test for acute-phase sera and immunoglobulin M capture enzyme-linked immunosorbent assay for early-convalescent-phase sera increased sensitivity only from 88.7% to 91.9%. Thus, NS1 antigen detection with the Platelia Dengue NS1 Ag kit could be used for first-line testing for acute dengue virus infection in clinical diagnostic laboratories.