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Sample records for evaluating viral antigens

  1. Radioimmunoassays of hidden viral antigens

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    Neurath, A.R. (Lindsley F. Kimbell Research Inst., New York, NY); Strick, N.; Baker, L.; Krugman, S.

    1982-07-01

    Antigens corresponding to infectious agents may be present in biological specimens only in a cryptic form bound to antibodies and, thus, may elude detection. We describe a solid-phase technique for separation of antigens from antibodies. Immune complexes are precipitated from serum by polyethylene glycol, dissociated with NaSCN, and adsorbed onto nitrocellulose or polystyrene supports. Antigens remain topographically separated from antibodies after removal of NaSCN and can be detected with radiolabeled antibodies. Genomes from viruses immobilized on nitrocellulose can be identified by nucleic acid hybridization. Nanogram quantities of sequestered hepatitis B surface and core antigens and picogram amounts of hepatitis B virus DNA were detected. Antibody-bound adenovirus, herpesvirus, and measles virus antigens were discerned by the procedure.

  2. Evaluation of the abbot Architect(™) epstein-barr virus viral capsid antigen IgM, viral capsid antigen IgG and nuclear antigen IgG assays in a pediatric and adult population.

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    Grandjean Lapierre, Simon; Vallières, Emilie; Rabaamad, Leila; Labrecque, Manon; Chartrand, Caroline; Renaud, Christian

    2016-08-01

    The detection of antibodies against Epstein-Barr viral capsid (VCA) and nuclear (EBNA) antigens is routinely performed with different commercially available immunoassays. In this study, we evaluated the concordance and performance of the Architect(™) chemiluminescent microparticle immunoassays (CMIAs) using Captia(™) enzyme linked immunosorbent assays (ELISA) for VCA IgM, and standard immunofluorescence (IF) assays for VCA IgG and EBNA IgG as comparative techniques. Sera were selected from a heterogeneous population including pediatric and adult patients. Concordance between CMIAs and comparative assays was high with total agreement percentages of 84,1% (95% CI: 77.8-88.9) for VCA IgM, 90,6% (95% CI: 84.2-94.7) for EBNA IgG and 98,0% (95% CI: 93.9-99.6) for VCA IgG. Moreover, kappa statistic values showed good to excellent correlation with values of 0.68 (95% CI: 0.57-0.79) for VCA IgM, 0.73 (95% CI: 0.58-0.87) for EBNA IgG and 0.95 (95% CI: 0.89-1.00) for VCA IgG. A correlation was observed between positivity levels on CMIAs and semi-quantitative fluorescence intensity on IF for VCA IgG and EBNA IgG assays. With regard to an accepted gold standard IF assays, CMIA was 98,1% (95% CI: 93.3-99.8) sensitive and 97,4% (95% CI: 86.5-99.9) specific for the detection of VCA IgG. For the detection of EBNA IgG, it was 92,2% (95% CI: 85.1-96.6) sensitive and 84,6% (95% CI: 65.1-95.6) specific. In summary, we demonstrated that the CMIA EBV antibody detection panel has high performance and high concordance with other commercially available immunoassays. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Expression of VP7, a Bluetongue Virus Group Specific Antigen by Viral Vectors: Analysis of the Induced Immune Responses and Evaluation of Protective Potential in Sheep

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    Bouet-Cararo, Coraline; Contreras, Vanessa; Caruso, Agathe; Top, Sokunthea; Szelechowski, Marion; Bergeron, Corinne; Viarouge, Cyril; Desprat, Alexandra; Relmy, Anthony; Guibert, Jean-Michel; Dubois, Eric; Thiery, Richard; Bréard, Emmanuel; Bertagnoli, Stephane; Richardson, Jennifer; Foucras, Gilles; Meyer, Gilles; Schwartz-Cornil, Isabelle; Zientara, Stephan; Klonjkowski, Bernard

    2014-01-01

    Bluetongue virus (BTV) is an economically important Orbivirus transmitted by biting midges to domestic and wild ruminants. The need for new vaccines has been highlighted by the occurrence of repeated outbreaks caused by different BTV serotypes since 1998. The major group-reactive antigen of BTV, VP7, is conserved in the 26 serotypes described so far, and its role in the induction of protective immunity has been proposed. Viral-based vectors as antigen delivery systems display considerable promise as veterinary vaccine candidates. In this paper we have evaluated the capacity of the BTV-2 serotype VP7 core protein expressed by either a non-replicative canine adenovirus type 2 (Cav-VP7 R0) or a leporipoxvirus (SG33-VP7), to induce immune responses in sheep. Humoral responses were elicited against VP7 in almost all animals that received the recombinant vectors. Both Cav-VP7 R0 and SG33-VP7 stimulated an antigen-specific CD4+ response and Cav-VP7 R0 stimulated substantial proliferation of antigen-specific CD8+ lymphocytes. Encouraged by the results obtained with the Cav-VP7 R0 vaccine vector, immunized animals were challenged with either the homologous BTV-2 or the heterologous BTV-8 serotype and viral burden in plasma was followed by real-time RT-PCR. The immune responses triggered by Cav-VP7 R0 were insufficient to afford protective immunity against BTV infection, despite partial protection obtained against homologous challenge. This work underscores the need to further characterize the role of BTV proteins in cross-protective immunity. PMID:25364822

  4. Expression of VP7, a Bluetongue virus group specific antigen by viral vectors: analysis of the induced immune responses and evaluation of protective potential in sheep.

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    Coraline Bouet-Cararo

    Full Text Available Bluetongue virus (BTV is an economically important Orbivirus transmitted by biting midges to domestic and wild ruminants. The need for new vaccines has been highlighted by the occurrence of repeated outbreaks caused by different BTV serotypes since 1998. The major group-reactive antigen of BTV, VP7, is conserved in the 26 serotypes described so far, and its role in the induction of protective immunity has been proposed. Viral-based vectors as antigen delivery systems display considerable promise as veterinary vaccine candidates. In this paper we have evaluated the capacity of the BTV-2 serotype VP7 core protein expressed by either a non-replicative canine adenovirus type 2 (Cav-VP7 R0 or a leporipoxvirus (SG33-VP7, to induce immune responses in sheep. Humoral responses were elicited against VP7 in almost all animals that received the recombinant vectors. Both Cav-VP7 R0 and SG33-VP7 stimulated an antigen-specific CD4+ response and Cav-VP7 R0 stimulated substantial proliferation of antigen-specific CD8+ lymphocytes. Encouraged by the results obtained with the Cav-VP7 R0 vaccine vector, immunized animals were challenged with either the homologous BTV-2 or the heterologous BTV-8 serotype and viral burden in plasma was followed by real-time RT-PCR. The immune responses triggered by Cav-VP7 R0 were insufficient to afford protective immunity against BTV infection, despite partial protection obtained against homologous challenge. This work underscores the need to further characterize the role of BTV proteins in cross-protective immunity.

  5. Immunization with viral antigens: Infectious haematopoietic necrosis

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    Winton, J.R.; Midtlyng, Paul J.; Brown, F.

    1997-01-01

    Infectious haematopoietic necrosis (IHN) is one of the most important viral diseases of salmonids, especially among juvenile fish where losses can be high. For over 20 years, researchers have tested a variety of preparations for control of IHN. Early vaccines consisted of killed virus and were effective when delivered by injection, but too costly to be practical on a large scale. Attenuated vaccines were developed by serial passage in cell culture and by monoclonal antibody selection. These offered excellent protection and were cost-effective, but residual virulence and uncertainty about their effects on other aquatic species made them poor candidates for licensing. Subunit vaccines using part of the IHNV glycoprotein gene cloned into E. coli or into an attenuated strain of A. salmonicida have been tested, appeared safe and were inexpensive. These vaccines were reported to provide some protection when delivered by immersion. Information on the location of antigenic sites on the glycoprotein led to trials using synthetic peptides, but these did not seem to be economically viable. Recently, plasmid vectors encoding the glycoprotein gene under control of a cytomegalovirus promoter were developed for genetic immunization. The constructs were highly protective when delivered by injection, but a more practical delivery system is needed. Thus, while several vaccine strategies have been tried in order to stimulate specific immunity against IHN, more research is needed to develop a commercially viable product for control of this important disease.

  6. [Australia antigen and viral hepatitis. Clinico-statistical considerations].

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    Menicagli, V; Pardelli, G C

    1977-03-10

    204 cases of viral hepatitis, 192 related to the MS1 form and 12 to MS2, were studied for the presence of Au. antigen and possible relations between this and the course of the disease. The antigen was encountered in 12 cases of the first group (6.2%) and in 7 of the second (58.3%). The disease was more serious and developed more slowly in Au.-positive cases compared to negative cases, it was also observed that only after the disappearance of the antigen from the serum was clinical cure possible.

  7. Viral immune evasion: Lessons in MHC class I antigen presentation.

    Science.gov (United States)

    van de Weijer, Michael L; Luteijn, Rutger D; Wiertz, Emmanuel J H J

    2015-03-01

    The MHC class I antigen presentation pathway enables cells infected with intracellular pathogens to signal the presence of the invader to the immune system. Cytotoxic T lymphocytes are able to eliminate the infected cells through recognition of pathogen-derived peptides presented by MHC class I molecules at the cell surface. In the course of evolution, many viruses have acquired inhibitors that target essential stages of the MHC class I antigen presentation pathway. Studies on these immune evasion proteins reveal fascinating strategies used by viruses to elude the immune system. Viral immunoevasins also constitute great research tools that facilitate functional studies on the MHC class I antigen presentation pathway, allowing the investigation of less well understood routes, such as TAP-independent antigen presentation and cross-presentation of exogenous proteins. Viral immunoevasins have also helped to unravel more general cellular processes. For instance, basic principles of ER-associated protein degradation via the ubiquitin-proteasome pathway have been resolved using virus-induced degradation of MHC class I as a model. This review highlights how viral immunoevasins have increased our understanding of MHC class I-restricted antigen presentation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Hepatitis B viral antigenic structure: signature analysis by monoclonal radioimmunoassays.

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    Wands, J R; Wong, M A; Shorey, J; Brown, R D; Marciniak, R A; Isselbacher, K J

    1984-04-01

    An approach has been developed for the analysis of hepatitis B viral (HBV) antigenic structure that creates numerical "signatures" of HBV strains. This technique employs high-affinity IgM and IgG monoclonal antibodies (anti-HBsAg) directed toward distinct and separate determinants on hepatitis B surface antigen (HBsAg). Such antibodies have been used to develop sensitive and specific radioimmuno-assays for measurement of HBsAg-associated determinants in serum. In performing "signature" analysis separate binding curves for each monoclonal anti-HBsAg are generated by measuring immunoreactivity in serial dilutions of HBsAg-positive serum. Since the HBsAg concentration in serum is unknown, the binding profiles of groups of samples from the same "classic" HBV subtype are aligned by an iterative maximum likelihood procedure to give the numerical signature of that HBV subtype. By using this approach, HBsAg shows far more antigenic heterogeneity than previously recognized by polyvalent anti-HBsAg antibodies. Indeed, there are subgroups within the classic HBsAg subtypes. In addition, the a domain (common to all known subtypes or strains of HBV) has been shown to be multideterminant. Thus, these studies have demonstrated heretofore unrecognized differences in HBV subtypes. This approach also has broader significance for the study of subtle or major antigenic changes among other viral agents since it is not necessary to know the concentration of virus or viral protein in complex protein mixtures.

  9. Evaluation of different heterologous prime-boost immunization strategies against Babesia bovis using viral vectored and protein-adjuvant vaccines based on a chimeric multi-antigen.

    Science.gov (United States)

    Jaramillo Ortiz, José Manuel; Molinari, María Paula; Gravisaco, María José; Paoletta, Martina Soledad; Montenegro, Valeria Noely; Wilkowsky, Silvina Elizabeth

    2016-07-19

    Protection against the intraerythrocytic bovine parasite Babesia bovis requires both humoral and cellular immune responses. Therefore, tailored combinations of immunogens targeted at both arms of the immune system are strategies of choice to pursue sterilizing immunity. In this study, different heterologous prime-boost vaccination schemes were evaluated in mice to compare the immunogenicity induced by a recombinant adenovirus, a modified vaccinia Ankara vector or a subunit vaccine all expressing a chimeric multi-antigen. This multi-antigen includes the immunodominant B and T cell epitopes of three B. bovis proteins: Merozoite Surface Antigen - 2c (MSA-2c), Rhoptry Associated Protein - 1 (RAP-1) and Heat Shock Protein 20 (HSP20). Both priming with the adenovirus or recombinant multi-antigen and boosting with the modified vaccinia Ankara vector achieved a high degree of activation of TNFα and IFNγ-secreting CD4(+) and CD8(+) specific T cells 60days after the first immunization. High titers of specific IgG antibodies were also detected at the same time point and lasted up to day 120 of the first immunization. Only the adenovirus - MVA combination triggered a marked isotype skew for the IgG2a antibody subclass meanwhile for the other immune traits analyzed here, both vaccination schemes showed similar performances. The immunological characterization in the murine model of these rationally designed immunogens led us to propose that adenoviruses as well as the bacterially expressed multi-antigen are highly reliable primer candidates to be considered in future experiments in cattle to test protection against bovine babesiosis. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Potentiation of anthrax vaccines using protective antigen-expressing viral replicon vectors.

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    Wang, Hai-Chao; An, Huai-Jie; Yu, Yun-Zhou; Xu, Qing

    2015-02-01

    DNA vaccines require improvement for human use because they are generally weak stimulators of the immune system in humans. The efficacy of DNA vaccines can be improved using a viral replicon as vector to administer antigen of pathogen. In this study, we comprehensively evaluated the conventional non-viral DNA, viral replicon DNA or viral replicon particles (VRP) vaccines encoding different forms of anthrax protective antigen (PA) for specific immunity and protective potency against anthrax. Our current results clearly suggested that these viral replicon DNA or VRP vaccines derived from Semliki Forest virus (SFV) induced stronger PA-specific immune responses than the conventional non-viral DNA vaccines when encoding the same antigen forms, which resulted in potent protection against challenge with the Bacillus anthracis strain A16R. Additionally, the naked PA-expressing SFV replicon DNA or VRP vaccines without the need for high doses or demanding particular delivery regimens elicited robust immune responses and afforded completely protective potencies, which indicated the potential of the SFV replicon as vector of anthrax vaccines for use in clinical application. Therefore, our results suggest that these PA-expressing SFV replicon DNA or VRP vaccines may be suitable as candidate vaccines against anthrax. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Evaluation of Viral Meningoencephalitis Cases

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    Handan Ilhan

    2012-08-01

    Full Text Available AIM: To evaluate retrospectively adult cases of viral encephalitis. METHOD: Fifteen patients described viral encephalitis hospitalized between the years 2006-2011 follow-up and treatment at the infectious diseases clinic were analyzed retrospectively. RESULTS: Most of the patients (%60 had applied in the spring. Fever (87%, confusion (73%, neck stiffness (73%, headache (73%, nausea-vomiting (33%, loss of consciousness (33%, amnesia (33%, agitation (20%, convulsion (%20, focal neurological signs (13%, Brudzinski-sign (13% were most frequently encountered findings. Electroencephalography test was applied to 13 of 14 patients, and pathological findings compatible with encephalitis have been found. Radiological imaging methods such as CT and MRI were performed in 9 of the 14 patients, and findings consistent with encephalitis were reported. All of initial cerebrospinal fluid (CSF samples were abnormal. The domination of the first examples was lymphocytes in 14 patients; only one patient had an increase in neutrophilic cells have been found. CSF protein level was high in nine patients, and low glucose level was detected in two patients. Herpes simplex virus polymerized chain reaction (PCR analyze was performed to fourteen patients CSF. Only two of them (14% were found positive. One of the patients sample selectively examined was found to be Parvovirus B19 (+, the other patient urine sample Jacobs-creutzfeld virus PCR was found to be positively. Empiric acyclovir therapy was given to all patients. Neuropsychiatric squeal developed at the one patient. CONCLUSION: The cases in the forefront of change in mental status viral meningoencephalitis should be considered and empirical treatment with acyclovir should be started. [TAF Prev Med Bull 2012; 11(4.000: 447-452

  12. Development of recombinant antigen array for simultaneous detection of viral antibodies.

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    Yi Liu

    Full Text Available Protein microarrays have been developed to study antibody reactivity against a large number of antigens, demonstrating extensive perspective for clinical application. We developed a viral antigen array by spotting four recombinant antigens and synthetic peptide, including glycoprotein G of herpes simplex virus (HSV type 1 and 2, phosphoprotein 150 of cytomegalovirus (CMV, Rubella virus (RV core plus glycoprotein E1 and E2 as well as a E1 peptide with the optimal concentrations on activated glass slides to simultaneously detect IgG and IgM against HSV1, HSV2, CMV and RV in clinical specimens of sera and cerebrospinal fluids (CSFs. The positive reference sera were initially used to measure the sensitivity and specificity of the array with the optimal conditions. Then clinical specimens of 144 sera and 93 CSFs were tested for IgG and IgM antibodies directed against HSV1, HSV2, CMV and RV by the antigen array. Specificity of the antigen array for viral antibodies detection was satisfying compared to commercial ELISA kits but sensitivity of the array varied relying on quality and antigenic epitopes of the spotting antigens. In short, the recombinant antigen array has potential to simultaneous detect multiple viral antibodies using minute amount (3 µl of samples, which holds the particularly advantage to detect viral antibodies in clinical CSFs being suspicious of neonatal meningitis and encephalitis.

  13. Evaluation of the Architect Epstein-Barr Virus (EBV) viral capsid antigen (VCA) IgG, VCA IgM, and EBV nuclear antigen 1 IgG chemiluminescent immunoassays for detection of EBV antibodies and categorization of EBV infection status using immunofluorescence assays as the reference method.

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    Corrales, Isabel; Giménez, Estela; Navarro, David

    2014-05-01

    Commercial immunoassays for detecting IgG and IgM antibodies against Epstein-Barr virus (EBV), viral capsid antigens (VCA), and IgGs toward EBV nuclear antigen-1 (EBNA-1) are routinely used in combination to categorize EBV infection status. In this study, we evaluated the performances of the Architect EBV VCA IgG, VCA IgM, and EBNA-1 IgG chemiluminescent microparticle assays (CMIAs) in EBV serological analyses using indirect immunofluorescence assays and anticomplement immunofluorescence assays as the reference methods for VCA IgG, VCA IgM, and EBNA-1 IgG antibody detection, respectively. A total of 365 serum samples representing different EBV serological profiles were included in this study. The κ values (concordances between the results) obtained in the Architect CMIA and those in the reference assays were 0.905 (P Architect CMIA panel were, respectively, 99.15% and 98.6% for diagnosing a primary infection, 97.62% and 93.39% for diagnosing a past EBV infection, and 92.42% and 97.82% for diagnosing the absence of an EBV infection. In summary, we demonstrated that the Architect EBV antibody panel performs very well for EBV antibody detection and correctly categorizes clinically relevant EBV infection states.

  14. Improving dengue viral antigens detection in dengue patient serum specimens using a low pH glycine buffer treatment.

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    Shen, Wen-Fan; Galula, Jedhan Ucat; Chang, Gwong-Jen J; Wu, Han-Chung; King, Chwan-Chuen; Chao, Day-Yu

    2017-04-01

    Early diagnosis of dengue virus (DENV) infection to monitor the potential progression to hemorrhagic fever can influence the timely management of dengue-associated severe illness. Nonstructural protein 1 (NS1) antigen detection in acute serum specimens has been widely accepted as an early diagnostic assay for dengue infection; however, lower sensitivity of the NS1 antigen-capture enzyme-linked immunosorbent assay (Ag-ELISA) in secondary dengue viral infection has been reported. In this study, we developed two forms of Ag-ELISA capable of detecting E-Ag containing virion and virus-like particles, and secreted NS1 (sNS1) antigens, respectively. The temporal kinetics of viral RNA, sNS1, and E-Ag were evaluated based on the in vitro infection experiment. Meanwhile, a panel of 62 DENV-2 infected patients' sera was tested. The sensitivity was 3.042 ng/mL and 3.840 ng/mL for sNS1 and E, respectively. The temporal kinetics of the appearance of viral RNA, E, NS1, and infectious virus in virus-infected tissue culture media suggested that viral RNAs and NS1 antigens could be detected earlier than E-Ag and infectious virus. Furthermore, a panel of 62 sera from patients infected by DENV Serotype 2 was tested. Treating clinical specimens with the dissociation buffer increased the detectable level of E from 13% to 92% and NS1 antigens from 40% to 85%. Inclusion of a low-pH glycine buffer treatment step in the commercially available Ag-ELISA is crucial for clinical diagnosis and E-containing viral particles could be a valuable target for acute DENV diagnosis, similar to NS1 detection. Copyright © 2015. Published by Elsevier B.V.

  15. Evaluation of an Antigen-Antibody

    African Journals Online (AJOL)

    GB

    detect samples with high viral loads have a significant implication for this assay designed to detect both antigen and antibody proteins. Arguably, HCV as a member of Flaviviridae family has an icosahedral capsid of T3 or T4 symmetry with 180 or 240 core protein subunits. Since one RNA genome contains 240 subunits of.

  16. Viral immune evasion : Lessons in MHC class I antigen presentation

    NARCIS (Netherlands)

    van de Weijer, Michael L.; Luteijn, Rutger D.; Wiertz, EJHJ

    2015-01-01

    The MHC class I antigen presentation pathway enables cells infected with intracellular pathogens to signal the presence of the invader to the immune system. Cytotoxic T lymphocytes are able to eliminate the infected cells through recognition of pathogen-derived peptides presented by MHC class I

  17. Viral hepatitis and hepatitis B antigen: recent advances

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    Krugman, Saul

    1974-01-01

    Recent advances in hepatitis research have shed new light on the etiology, pathogenesis, epidemiology and prevention of type B hepatitis infection. The so-called ‘Dane’ particle is probably the complete hepatitis B virion; its outer coat is the hepatitis B (Australia) antigen (HB Ag) and its inner core is an immunologically distinct particle. Subtypes of HB Ag (a, d, y, w and r) are useful indices for epidemiological surveys. Concepts of epidemiology have changed: type B hepatitis is transmissible by contact as well as by inoculation. The presence of HB Ag in blood is indicative of the presence of hepatitis B virus. Tests to detect antigen and use of voluntary blood donors have played a major role in the decreased incidence of post transfusion hepatitis. A special hepatitis B gammaglobulin preparation and a heat-inactivated hepatitis B vaccine have proved to be effective in preliminary studies. PMID:4219230

  18. Viral sequestration of antigen subverts cross presentation to CD8(+ T cells.

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    Eric F Tewalt

    2009-05-01

    Full Text Available Virus-specific CD8(+ T cells (T(CD8+ are initially triggered by peptide-MHC Class I complexes on the surface of professional antigen presenting cells (pAPC. Peptide-MHC complexes are produced by two spatially distinct pathways during virus infection. Endogenous antigens synthesized within virus-infected pAPC are presented via the direct-presentation pathway. Many viruses have developed strategies to subvert direct presentation. When direct presentation is blocked, the cross-presentation pathway, in which antigen is transferred from virus-infected cells to uninfected pAPC, is thought to compensate and allow the generation of effector T(CD8+. Direct presentation of vaccinia virus (VACV antigens driven by late promoters does not occur, as an abortive infection of pAPC prevents production of these late antigens. This lack of direct presentation results in a greatly diminished or ablated T(CD8+ response to late antigens. We demonstrate that late poxvirus antigens do not enter the cross-presentation pathway, even when identical antigens driven by early promoters access this pathway efficiently. The mechanism mediating this novel means of viral modulation of antigen presentation involves the sequestration of late antigens within virus factories. Early antigens and cellular antigens are cross-presented from virus-infected cells, as are late antigens that are targeted to compartments outside of the virus factories. This virus-mediated blockade specifically targets the cross-presentation pathway, since late antigen that is not cross-presented efficiently enters the MHC Class II presentation pathway. These data are the first to describe an evasion mechanism employed by pathogens to prevent entry into the cross-presentation pathway. In the absence of direct presentation, this evasion mechanism leads to a complete ablation of the T(CD8+ response and a potential replicative advantage for the virus. Such mechanisms of viral modulation of antigen presentation

  19. Pathology and viral antigen distribution following experimental infection of sheep and goats with capripoxvirus.

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    Embury-Hyatt, C; Babiuk, S; Manning, L; Ganske, S; Bowden, T R; Boyle, D B; Copps, J

    2012-01-01

    Current understanding of capripoxvirus pathogenesis is limited since there have been no detailed studies examining cell tropism at well-defined intervals following infection. We undertook time-course studies in sheep and goats following inoculation of sheeppox or goatpox viruses in their respective homologous hosts, and examined tissues by light microscopy. A monoclonal antibody generated to a sheeppox virus core protein was used for immunohistochemical detection of viral antigen in tissue sections. Lesions and virus antigen were observed consistently in the skin, lung and lymph nodes. Antigen was detected at 6 and 8 days post inoculation for skin and lung, respectively, within cells which appeared to be of monocyte/macrophage lineage. In sheep skin capripoxvirus immunoreactivity was detected within previously unreported large multinucleated cells. In the lung, double immunolabelling detected the simultaneous expression of capripoxvirus antigen and cytokeratin indicating the presence of virus within pneumocytes. Lung double immunolabelling also detected the expression of capripoxvirus antigen in CD68(+) cells, confirming the presence of viral antigen within macrophages. Based on early detection of infected macrophages, dissemination of virus within the host and localization to tissues likely occurred through cells of the monocyte/macrophage lineage. Histological findings revealed similarities with both monkeypox and smallpox, thus capripoxvirus infection in sheep and goats may represent useful models with which to study strategies for poxvirus-specific virus vaccine concepts and therapeutics. Crown Copyright © 2011. Published by Elsevier Ltd. All rights reserved.

  20. Understanding MHC class I presentation of viral antigens by human dendritic cells as a basis for rational design of therapeutic vaccines

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    Nadine eVan Montfoort

    2014-04-01

    Full Text Available 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 allows 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 DC subsets involved, however, these results cannot be readily translated towards 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.

  1. Detection of Avian Antigen-Specific T Cells Induced by Viral Vaccines

    DEFF Research Database (Denmark)

    Dalgaard, Tina Sørensen; Norup, Liselotte Rothmann; Juul-Madsen, Helle Risdahl

    2016-01-01

    Live attenuated viral vaccines are widely used in commercial poultry production, but the development of new effective inactivated/subunit vaccines is needed. Studies of avian antigen-specific T cells are primarily based on analyses ex vivo after activating the cells with recall antigen....... There is a particular interest in developing robust high-throughput assays as chicken vaccine trials usually comprise many individuals. In many respects, the avian immune system differs from the mammalian, and T cell assessment protocols must be adjusted accordingly to account for, e.g., differences in leukocyte...

  2. Effect of BSA Antigen Sensitization during the Acute Phase of Influenza A Viral Infection on CD11c+ Pulmonary Antigen Presenting Cells

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    Fumitaka Sato

    2009-01-01

    Conclusions: BSA antigen sensitization during the acute phase of influenza A viral infection enhanced IL-10 production from naive CD4+ T cell interaction with CD11c+ pulmonary APCs. The IL-10 secretion evoked Th2 responses in the lungs with downregulation of Th1 responses and was important for the eosinophil recruitment into the lungs after BSA antigen challenge.

  3. Norovirus drug candidates that inhibit viral capsid attachment to human histo-blood group antigens

    OpenAIRE

    Ali, Eunüs S.; Rajapaksha, Harinda; Carr, Jillian M.; Petrovsky, Nikolai

    2016-01-01

    Human noroviruses are the leading causative agents of epidemic and sporadic viral gastroenteritis and childhood diarrhoea worldwide. Human histo-blood group antigens (HBGA) serve as receptors for norovirus capsid protein attachment and play a critical role in infection. This makes HBGA-norovirus binding a promising target for drug development. Recently solved crystal structures of norovirus bound to HBGA have provided a structural basis for identification of potential anti-norovirus drugs and...

  4. KIR content genotypes associate with carriage of hepatitis B surface antigen, e antigen and HBV viral load in Gambians.

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    Louis-Marie Yindom

    Full Text Available Hepatocellular carcinoma (HCC causes over 800,000 deaths worldwide annually, mainly in low income countries, and incidence is rising rapidly in the developed world with the spread of hepatitis B (HBV and C (HCV viruses. Natural Killer (NK cells protect against viral infections and tumours by killing abnormal cells recognised by Killer-cell Immunoglobulin-like Receptors (KIR. Thus genes and haplotypes encoding these receptors may be important in determining both outcome of initial hepatitis infection and subsequent chronic liver disease and tumour formation. HBV is highly prevalent in The Gambia and the commonest cause of liver disease. The Gambia Liver Cancer Study was a matched case-control study conducted between September 1997 and January 2001 where cases with liver disease were identified in three tertiary referral hospitals and matched with out-patient controls with no clinical evidence of liver disease.We typed 15 KIR genes using the polymerase chain reaction with sequence specific primers (PCR-SSP in 279 adult Gambians, 136 with liver disease (HCC or Cirrhosis and 143 matched controls. We investigated effects of KIR genotypes and haplotypes on HBV infection and associations with cirrhosis and HCC.Homozygosity for KIR group A gene-content haplotype was associated with HBsAg carriage (OR 3.7, 95% CI 1.4-10.0 whilst telomeric A genotype (t-AA was associated with reduced risk of e antigenaemia (OR 0.2, 95% CI 0.0-0.6 and lower viral loads (mean log viral load 5.2 vs. 6.9, pc = 0.022. One novel telomeric B genotype (t-ABx2 containing KIR3DS1 (which is rare in West Africa was also linked to e antigenaemia (OR 8.8, 95% CI 1.3-60.5. There were no associations with cirrhosis or HCC.Certain KIR profiles may promote clearance of hepatitis B surface antigen whilst others predispose to e antigen carriage and high viral load. Larger studies are necessary to quantify the effects of individual KIR genes, haplotypes and KIR/HLA combinations on long

  5. Viral Escape Mutant Epitope Maintains TCR Affinity for Antigen yet Curtails CD8 T Cell Responses.

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    Shayla K Shorter

    Full Text Available T cells have the remarkable ability to recognize antigen with great specificity and in turn mount an appropriate and robust immune response. Critical to this process is the initial T cell antigen recognition and subsequent signal transduction events. This antigen recognition can be modulated at the site of TCR interaction with peptide:major histocompatibility (pMHC or peptide interaction with the MHC molecule. Both events could have a range of effects on T cell fate. Though responses to antigens that bind sub-optimally to TCR, known as altered peptide ligands (APL, have been studied extensively, the impact of disrupting antigen binding to MHC has been highlighted to a lesser extent and is usually considered to result in complete loss of epitope recognition. Here we present a model of viral evasion from CD8 T cell immuno-surveillance by a lymphocytic choriomeningitis virus (LCMV escape mutant with an epitope for which TCR affinity for pMHC remains high but where the antigenic peptide binds sub optimally to MHC. Despite high TCR affinity for variant epitope, levels of interferon regulatory factor-4 (IRF4 are not sustained in response to the variant indicating differences in perceived TCR signal strength. The CD8+ T cell response to the variant epitope is characterized by early proliferation and up-regulation of activation markers. Interestingly, this response is not maintained and is characterized by a lack in IL-2 and IFNγ production, increased apoptosis and an abrogated glycolytic response. We show that disrupting the stability of peptide in MHC can effectively disrupt TCR signal strength despite unchanged affinity for TCR and can significantly impact the CD8+ T cell response to a viral escape mutant.

  6. HIV-1 Activates T Cell Signaling Independently of Antigen to Drive Viral Spread.

    Science.gov (United States)

    Len, Alice C L; Starling, Shimona; Shivkumar, Maitreyi; Jolly, Clare

    2017-01-24

    HIV-1 spreads between CD4 T cells most efficiently through virus-induced cell-cell contacts. To test whether this process potentiates viral spread by activating signaling pathways, we developed an approach to analyze the phosphoproteome in infected and uninfected mixed-population T cells using differential metabolic labeling and mass spectrometry. We discovered HIV-1-induced activation of signaling networks during viral spread encompassing over 200 cellular proteins. Strikingly, pathways downstream of the T cell receptor were the most significantly activated, despite the absence of canonical antigen-dependent stimulation. The importance of this pathway was demonstrated by the depletion of proteins, and we show that HIV-1 Env-mediated cell-cell contact, the T cell receptor, and the Src kinase Lck were essential for signaling-dependent enhancement of viral dissemination. This study demonstrates that manipulation of signaling at immune cell contacts by HIV-1 is essential for promoting virus replication and defines a paradigm for antigen-independent T cell signaling. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  7. Overcoming viral escape with vaccines that generate and display antigen diversity in vivo

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    García-Quintanilla Albert

    2007-11-01

    Full Text Available Abstract Background Viral diversity is a key problem for the design of effective and universal vaccines. Virtually, a vaccine candidate including most of the diversity for a given epitope would force the virus to create escape mutants above the viability threshold or with a high fitness cost. Presentation of the hypothesis Therefore, I hypothesize that priming the immune system with polyvalent vaccines where each single vehicle generates and displays multiple antigen variants in vivo, will elicit a broad and long-lasting immune response able to avoid viral escape. Testing the hypothesis To this purpose, I propose the use of yeasts that carry virus-like particles designed to pack the antigen-coding RNA inside and replicate it via RNA-dependent RNA polymerase. This would produce diversity in vivo limited to the target of interest and without killing the vaccine vehicle. Implications of the hypothesis This approach is in contrast with peptide cocktails synthesized in vitro and polyvalent strategies where every cell or vector displays a single or definite number of mutants; but similarly to all them, it should be able to overcome original antigenic sin, avoid major histocompatibility complex restriction, and elicit broad cross-reactive immune responses. Here I discuss additional advantages such as minimal global antagonism or those derived from using a yeast vehicle, and potential drawbacks like autoimmunity. Diversity generated by this method could be monitored both genotypically and phenotypically, and therefore selected or discarded before use if needed.

  8. Chronic hepatitis B in pregnant women: is hepatitis B surface antigen quantification useful for viral load prediction?

    Science.gov (United States)

    Fujiko, Masita; Chalid, Maisuri T; Turyadi; Ie, Susan I; Maghfira; Syafri; Wahyuni, Ridha; Roni, Martono; Patellongi, Ilhamjaya; Massi, M Nasrum; Muljono, David H

    2015-12-01

    New cases of hepatitis B virus (HBV) infection continue to occur worldwide. Most of these are due to mother-to-child transmission (MTCT), with maternal viraemia as the most important contributing factor. The hepatitis B surface antigen (HBsAg) level, which correlates positively with viral load, has been used for treatment monitoring in chronic hepatitis B. This study evaluated the usefulness of quantitative HBsAg for viral load prediction in HBsAg-positive pregnant women. A total of 943 pregnant women in Makassar, Indonesia, were screened for HBsAg. Sixty-four women were HBsAg-positive and investigated. HBsAg level and hepatitis B e antigen (HBeAg)/hepatitis B e antibody (anti-HBe) status were determined serologically. Viral load was measured by real-time PCR. HBV DNA was sequenced and analysed for identification of genotype and basal core promoter (BCP)/precore (PC) mutations. Of 64 subjects, 12 (18.8%) were HBeAg-positive and 52 (81.3%) were HBeAg-negative. HBsAg and HBV DNA levels were significantly higher in the HBeAg-positive group (ppregnant women. The HBeAg-negative group had a frequent occurrence of BCP/PC variants, which may have contributed to the lack of correlation observed. Samples with a low HBsAg level, which is associated with a low risk of MTCT, do not require HBV DNA measurement. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  9. Treating cancer as an infectious disease--viral antigens as novel targets for treatment and potential prevention of tumors of viral etiology.

    Directory of Open Access Journals (Sweden)

    Xing Guo Wang

    2007-10-01

    Full Text Available Nearly 20% of human cancers worldwide have an infectious etiology with the most prominent examples being hepatitis B and C virus-associated hepatocellular carcinoma and human papilloma virus-associated cervical cancer. There is an urgent need to find new approaches to treatment and prevention of virus-associated cancers.Viral antigens have not been previously considered as targets for treatment or prevention of virus-associated cancers. We hypothesized that it was possible to treat experimental HPV16-associated cervical cancer (CC and Hepatitis B-associated hepatocellular carcinoma (HCC by targeting viral antigens expressed on cancer cells with radiolabeled antibodies to viral antigens. Treatment of experimental CC and HCC tumors with (188Re-labeled mAbs to E6 and HBx viral proteins, respectively, resulted in significant and dose-dependent retardation of tumor growth in comparison with untreated mice or mice treated with unlabeled antibodies.This strategy is fundamentally different from the prior uses of radioimmunotherapy in oncology, which targeted tumor-associated human antigens and promises increased specificity and minimal toxicity of treatment. It also raises an exciting possibility to prevent virus-associated cancers in chronically infected patients by eliminating cells infected with oncogenic viruses before they transform into cancer.

  10. Norovirus drug candidates that inhibit viral capsid attachment to human histo-blood group antigens.

    Science.gov (United States)

    Ali, Eunüs S; Rajapaksha, Harinda; Carr, Jillian M; Petrovsky, Nikolai

    2016-09-01

    Human noroviruses are the leading causative agents of epidemic and sporadic viral gastroenteritis and childhood diarrhoea worldwide. Human histo-blood group antigens (HBGA) serve as receptors for norovirus capsid protein attachment and play a critical role in infection. This makes HBGA-norovirus binding a promising target for drug development. Recently solved crystal structures of norovirus bound to HBGA have provided a structural basis for identification of potential anti-norovirus drugs and subsequently performed in silico and in vitro drug screens have identified compounds that block norovirus binding and may thereby serve as structural templates for design of therapeutic norovirus inhibitors. This review explores norovirus therapeutic options based on the strategy of blocking norovirus-HBGA binding. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Dose and timing requirements for immunogenicity of viral poultry vaccine antigen: investigations of emulsion-based depot function.

    Science.gov (United States)

    Jansen, Theo; Hofmans, Marij P M; Theelen, Marc J G; Manders, Frans G A; Schijns, Virgil E J C

    2007-10-01

    The release requirements for vaccine antigens delivered by adjuvants with presumed depot function are poorly understood. Water-in-oil (W/O) emulsions are routinely used in many poultry vaccines. They strongly activate antibody production, and are regarded as a depot from which antigens are slowly released, resulting in prolonged antigen residence. However, from earlier studies we concluded that W/O adjuvant activity is partly based on the immunostimulatory activity of the oil phase. Here we assess the dose and regimen requirements for viral antigen in immunization experiments in chickens. Three-week-old to 4-week-old White Leghorn chickens were repeatedly injected with inactivated infectious bursal disease virus antigen over 48 days. Our aim was to compare the antibody responses in repeatedly injected animals, receiving fractioned doses of antigen, with the responses in animals receiving only one injection of the full dose of antigen formulated in either a W/O emulsion or in saline. We observed that repeated administration of small amounts of antigen results in a gradual increase of specific humoral immune responses during the immunization regimen. Immunization with a higher first dose evoked an early higher antibody response, which, however, reached a similar plateau level at the end of the regimen. When compared with lower first-dose regimens, a slow decline of serum antibody titre 2 weeks after the end of antigen injections indicated that repeated injection of small doses of antigen indeed mimics the efficacy of depot-forming adjuvants. All regimens of fractioned antigen in saline, however, proved less effective, when compared with a single-dose vaccination of the cumulative amount of antigen formulated in a W/O emulsion. From our data we confirm that W/O emulsions are very effective vaccine vehicles for improving antigen-specific humoral responses in chickens, owing to a combination of antigen residence-prolonging activity and direct immune stimulation.

  12. Chagasic patients are able to respond against a viral antigen from influenza virus

    Directory of Open Access Journals (Sweden)

    Lasso Paola

    2012-08-01

    Full Text Available Abstract Background Trypanosoma cruzi, the etiological agent of Chagas’ disease, is an obligate intracellular parasite which induces a CD8+ T cell immune response with secretion of cytokines and release of cytotoxic granules. Although an immune-suppressive effect of T. cruzi on the acute phase of the disease has been described, little is known about the capacity of CD8+ T cell from chronic chagasic patients to respond to a non-T. cruzi microbial antigen. Methods In the present paper, the frequency, phenotype and the functional activity of the CD8+ T cells specific from Flu-MP*, an influenza virus epitope, were determined in 13 chagasic patients and 5 healthy donors. Results The results show that Flu-MP* peptide specific CD8+ T cells were found with similar frequencies in both groups. In addition, Flu-MP* specific CD8+ T cells were distributed in the early or intermediate/late differentiation stages without showing enrichment of a specific sub-population. The mentioned Flu-MP* specific CD8+ T cells from chagasic patients were predominately TEM (CCR7- CD62L-, producing IL-2, IFNγ, CD107a/b and perforin, and did not present significant differences when compared with those from healthy donors. Conclusions Our results support the hypothesis that there is no CD8+ T cell nonspecific immune-suppression during chronic Chagas disease infection. Nonetheless, other viral antigens must be studied in order to confirm our findings.

  13. First report of Bovine Viral Diarrhea Virus antigen from pneumonic cattle in Sudan

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    Intisar Kamil Saeed

    2015-06-01

    Full Text Available To explore the expected role of Bovine Viral Diarrhea Virus (BVDV in pneumonia in cattle, cattle lungs (n=242 showing signs of pneumonia were collected from slaughter houses of three different localities located at Northern, Central and Western Sudan during 2010–2013. The collected samples were tested for the presence of BVDV antigen using Enzyme-Linked Immunosorbent Assay (ELISA, and Fluorescent Antibody Test (FAT. Twenty six (10.7% out of 242 samples were found to be positive for BVDV. Positive results were seen in all the three studied areas, with the highest prevalence (16.7%; n=4/24 at Gezira State in Central Sudan. BVDV genome could be detected in all ELISA positive samples. The results indicated the existence of BVDV infection in cattle in different areas in Sudan, and its possible association with respiratory infections in cattle. Analysis using BLAST indicated that the sequence was identical to the previously reported BVDV-1 (GenBank accession AF220247.1.; nucleotide A was found in our study at position 9 of our sequence, whereas T was present instead in the reference virus. This is the first report of detecting BVDV antigen, genome, and its sequence analysis collected from cattle lungs in Sudan.

  14. Pelacakan Secara Imunohistokimiawi Antigen Virus pada Ayam yang Diinfeksi dengan Virus Penyakit Tetelo (IMMUNOHISTOCHEMICAL DETECTION OF VIRAL ANTIGEN IN TISSUE OF CHICKENS EXPERIMENTALLY INFECTED WITH NEWCASTLE DISEASE VIRUS

    Directory of Open Access Journals (Sweden)

    Anak Agung Ayu Mirah Adi

    2013-07-01

    Full Text Available In order to study the distribution of Newcastle disease virus (NDV following infection, chickenswere experimentally infected with visceretropic velogenic NDV isolate. Monoclonal antibodies (mAbsagainst the NDV LaSota vaccine strain were then produced to detect viral antigen in the infectedorgans. The mAbs were firstly tested for their specificity by enzyme linked immunosorbent assay(ELISA using NDV and normal allantoic fluids as antigens. Eight mAbs specific against NDVwere isolated and two mAbs were used for immunodetection of NDV antigen in chicken’s tissues.By immunohistochemistry labeled streptavidin-biotin (LSAB staining NDV–antigen was detectedin paraffin embedded tissues of NDV-infected chickens. NDV antigen was not detected in noninfected chickens. In the infected chickens, high intensity of NDV antigen was detected in thelymphoid tissues, lung and intestine. The NDV antigen with a lesser intensity was detected in thebrain, trachea, liver and myocardium. This study shows that although viscerotropic velogenicNDV isolate can infect almost all organs, the main target of infection are lung, intestine andlymphoids tissues

  15. Immunological Detection of Newcastle Disease Viral Antigen in the Naturally Infected Chickens by Monoclonal Antibodies against Fusion-2 Protein

    Directory of Open Access Journals (Sweden)

    Nyoman Mantik Astawa

    2014-08-01

    Full Text Available Monoclonal antibodies (mAbs against Fusion (F2 protein of Newcastle disease virus (NDV wereproduced for the detection of the viral antigen in infected chickens. Cells derived from spleen of Balb/c miceimmunized with the virus were fused with mouse myeloma cells to generate hybridomas capable ofproducing mAbs against the virus. The hybridomas were screened by enzyme-linked immunosorbent assay(ELISA for anti-NDV specific mAbs using crude viral antigen (allantoic fluid of NDV-infected fertile eggsand normal uninfected allantoic fluid of fertile eggs as negative control. The NDV proteins reactive withmAbs were then determined by Western Blotting using purified NDV as antigen. The mAbs reactive withF2 (12.5 KDa protein of NDV were then used for the detection of NDV antigen in both the allantoic fluidof NDV- infected chicken embryos and in organs of naturally infected chickens. The results showed that 2out of 5 mAbs produced were against F2 protein of NDV. By indirect ELISA, the mAbs were able to detectthe viral antigen in allantoic fluid of NDV infected fertile chicken eggs at the titre as low as 2-2 to 2-4 HAunits per 0.1 mL. NDV–antigen was also detected by immunoperoxidase staining in paraffin-embeddedtissues of NDV-infected chickens but not in normal uninfected chickens. The most prominent infection wasdetected in the gastrointestinal tract and the lung. The NDV antigen was also detected in other organssuch as the brain, spleen, and several other tissues. It is evident that mAbs produced against F2 proteinof NDV were applicable for use in the detection of NDV antigen in infected chickens.

  16. Avaliação de três cepas de vírus rábico, antigenicamente distintas, em camundongos: II - Estudo da disseminação viral por diferentes órgãos Evaluation of three antigenically different rabies virus strains in mice: II -Study of the viral dissemination in different organs

    Directory of Open Access Journals (Sweden)

    Pedro Manuel Leal Germano

    1988-12-01

    Full Text Available Estudou-se, comparativamente, o grau de disseminação de três cepas de vírus rábico, duas de origem de cão, Jales e Nigéria, e uma de origem de morcego, DR 19, com perfis antigênicos do nucleocapside distintos. Estas cepas foram inoculadas por via intramuscular, na face interna da coxa, em dois grupos de camundongos, com 21 e 28 dias de idade. Os animais foram mantidos em observação por um período total de 30 dias, e dos animais vitimados pela infecção, foram coletados diferentes órgãos, músculo lingual, coração, pulmão, rim e fígado, além do cérebro e da medula espinal, para avaliar-se o grau de disseminação de cada cepa viral, através da prova de imunofluorescência direta (IFD. Os resultados obtidos evidenciaram que os decalques de cérebro e de medula espinal apresentaram total concordância na prova de IFD, constatando-se as maiores diferenças com as cepas Jales e Nigéria, situando-se a cepa DR 19, intermediariamente, a estas duas. O músculo lingual foi o órgão que apresentou maior freqüência de positividade para ambos os grupos etários e para as três cepas virais.A study was conducted to compare three strains of rabies virus, two of them, Jales and Nigeria, isolated from dogs, and the other DR 19, from vampire bats, with different nucleo-capside antigenic characteristics. These strains were intramuscularly inoculated in the inner side of the thigh of 21 and 28 day-old mice. The animals were observed for 30 days and different organs: brain, spinal cord, tongue, heart, lung, kidney and liver were collected from the animals which died of rabies for the dissemination study by the immunofluorescent antibody technique (IFA. This technique showed complete agreement between brain and spinal cord. The greatest differences in dissemination were observed between Jales and Nigeria strains. The results observed for the DR 19 strain were situated between those of the two dog strains. The greates degree of dissemination

  17. Viral antigen production in cell cultures on microcarriers Bovine parainfluenza 3 virus and MDBK cells.

    Science.gov (United States)

    Conceição, M M; Tonso, A; Freitas, C B; Pereira, C A

    2007-11-07

    Viral antigens can be obtained from infected mammalian cells cultivated on microcarriers. We have worked out parameters for the production of bovine parainfluenza 3 (PI-3) virus by Mandin-Darby Bovine Kidney (MDBK) cells cultivated on Cytodex 1 microcarriers (MCs) in spinners flasks and bioreactor using fetal bovine serum (FBS) supplemented Eagle minimal essential medium (Eagle-MEM). Medium renewal during the cell culture was shown to be crucial for optimal MCs loading (>90% MCs with confluent cell monolayers) and cell growth (2.5 x 10(6)cells/mL and a micro(x) (h(-1)) 0.05). Since cell cultures performed with lower amount of MCs (1g/L), showed good performances in terms of cell loading, we designed batch experiments with a lower concentration of MCs in view of optimizing the cell growth and virus production. Studies of cell growth with lower concentrations of MCs (0.85 g/L) showed that an increase in the initial cell seeding (from 7 to 40 cells/MC) led to a different kinetic of initial cell growth but to comparable final cell concentrations ((8-10)x10(5)cells/mL at 120 h) and cell loading (210-270 cells/MC). Upon infection with PI-3 virus, cultures showed a decrease in cell growth and MC loading directly related to the multiplicity of infection (moi) used for virus infection. Infected cultures showed also a higher consumption of glucose and production of lactate. The PI-3 virus and PI-3 antigen production among the cultures was not significantly different and attained values ranging from, respectively, 7-9 log(10) TCID(50)/mL and 1.5-2.2 OD. The kinetics of PI-3 virus production showed a sharp increase during the first 24h and those of PI-3 antigen increased after 24h. The differential kinetics of PI-3 virus and PI-3 antigen can be explained by the virus sensitivity to temperature. In view of establishing a protocol of virus production and based on the previous experiments, MDBK cell cultures performed under medium perfusion in a bioreactor of 1.2L were infected

  18. Interleukin 10 (IL-10) and viral IL-10 strongly reduce antigen-specific human T cell proliferation by diminishing the antigen-presenting capacity of monocytes via downregulation of class II major histocompatibility complex expression

    NARCIS (Netherlands)

    de Waal Malefyt, R.; Haanen, J.; Spits, H.; Roncarolo, M. G.; te Velde, Anje; Figdor, C.; Johnson, K.; Kastelein, R.; Yssel, H.; de Vries, J. E.

    1991-01-01

    Interleukin 10 (IL-10) and viral IL-10 (v-IL-10) strongly reduced antigen-specific proliferation of human T cells and CD4+ T cell clones when monocytes were used as antigen-presenting cells. In contrast, IL-10 and v-IL-10 did not affect the proliferative responses to antigens presented by autologous

  19. Psoralen Inactivation of Viruses: A Process for the Safe Manipulation of Viral Antigen and Nucleic Acid

    Directory of Open Access Journals (Sweden)

    Katherine Schneider

    2015-11-01

    Full Text Available High consequence human pathogenic viruses must be handled at biosafety level 2, 3 or 4 and must be rendered non-infectious before they can be utilized for molecular or immunological applications at lower biosafety levels. Here we evaluate psoralen-inactivated Arena-, Bunya-, Corona-, Filo-, Flavi- and Orthomyxoviruses for their suitability as antigen in immunological processes and as template for reverse transcription PCR and sequencing. The method of virus inactivation using a psoralen molecule appears to have broad applicability to RNA viruses and to leave both the particle and RNA of the treated virus intact, while rendering the virus non-infectious.

  20. First report of immunohistochemical detection of Peste des petit ruminants, parainfluenza 3 and respiratory syncytial viral antigens in lungs of Nigerian goats.

    Science.gov (United States)

    Jarikre, Theophilus Aghogho; Emikpe, Benjamin Obukowho

    2017-01-01

    This study determined the of involvement of PPR, PI3, and RS viruses in the pathology of caprine pneumonia across Nigeria. 150 goats were selected randomly. PI3 and RSV monoclonal antibodies and PPR polyclonal antibody were used for the immunolocalization of the antigens. Histologically, 61 of the goats had broncho-interstitial pneumonia, 25 had interstitial pneumonia, 42 had bronchopneumonia, 12 had bronchiolitis, and 10 were normal. PPR, PI3, and RS viral antigens were demonstrated in: intact and desquamated bronchial, bronchiolar epithelial cells, macrophages, leukocytes, pneumocytes, and giant cells. 23% of the caprine lungs had positive immuno-staining to PI3 viral antigen, 10% were positive for RSV antigen while 34% were positive for PPR viral antigen. 8% showed immunostaining for the two and or three respiratory viral antigens in the goats. PI3 and RSV antigens were more in the young goats, red sokoto breed and during the dry season. This is the first report of immunohistochemical detection of PPR, PI3 and RS viral antigens in caprine lungs in Nigeria. These findings underscore the importance of PI3 and RSV viruses in the control of caprine pneumonia in Nigeria.

  1. Short Communication: False Recent Ratio of the Limiting-Antigen Avidity Assay and Viral Load Testing Algorithm Among Cameroonians with Long-Term HIV Infection.

    Science.gov (United States)

    Lynch, Briana A; Patel, Eshan U; Courtney, Colleen R; Nanfack, Aubin J; Bimela, Jude; Wang, Xiaohong; Eid, Issa; Quinn, Thomas C; Laeyendecker, Oliver; Nyambi, Phillipe N; Duerr, Ralf; Redd, Andrew D

    2017-11-01

    Current serological assays that are used for cross-sectional HIV incidence estimation have been shown to misclassify individuals with chronic infection. Limited information exists on the performance of cross-sectional incidence assays in Central Africa. HIV-positive individuals from Cameroon who were infected for at least 1 or 2 years were evaluated to determine the false recent ratio (FRR) of a two-assay algorithm, which includes the Limiting Antigen Avidity (LAg-Avidity) assay (normalized optical density units, ODn HIV viral load (>1000 copies/ml). The subject-level FRR was 5.3% (95% confidence interval [CI], 2.1-10.5) for individuals infected for ≥1 year and 3.9% (95% CI, 0.8-11.0) for individuals infected for ≥2 years. These data suggest that the LAg-Avidity plus viral load incidence algorithm may overestimate HIV incidence rates in Central Africa.

  2. Stability of Bovine viral diarrhea virus antigen in ear punch samples collected from bovine fetuses.

    Science.gov (United States)

    Ridpath, Julia F; Chiang, Yu-Wei; Waldbillig, Jill; Neill, John D

    2009-05-01

    Fourteen first-calf heifers were tested free of antibodies against Bovine viral diarrhea viruses (BVDV) by serum neutralization and free of BVDV by polymerase chain reaction. Twelve were exposed to BVDV-1b strain CA0401186a at 84-86 days of gestation, and 2 were exposed to mock inoculum and served as negative controls. Fetuses were harvested by cesarean section at 115-117 days of gestation. The 12 fetuses removed from the BVDV-exposed heifers were BVDV positive based on virus isolation from kidney, thymus, cerebellum, and spleen. It can be assumed that these fetuses would have developed into persistently infected calves had they been allowed to go to term. Virus was not isolated from the fetuses of control animals. Ear punch samples were collected from all fetuses at time of harvest. Antigen capture enzyme-linked immunosorbent assay (ACE), using a commercial kit, was performed on ear punch samples that were frozen within 5 hr of collection and stored at -20 degrees C until tested, tested after storage for 7 days at room temperature (18-25 degrees C), or tested after storage for 7 days at 37 degrees C. Samples stored for 7 days at room temperature or 37 degrees C lost an average of 34% of their starting weight. All samples from BVDV isolation-positive fetuses tested positive by ACE, whereas samples from nonexposed fetuses tested negative, regardless of storage conditions. These results suggest that ACE testing of skin samples collected from aborted fetuses and stillborn calves found in the field may represent a practical surveillance method for BVDV-induced reproductive disease.

  3. Mucosal vaccination with heterologous viral vectored vaccine targeting subdominant SIV accessory antigens strongly inhibits early viral replication

    DEFF Research Database (Denmark)

    Xu, Huanbin; Andersson, Anne-Marie Carola; Ragonnaud, Emeline

    2017-01-01

    Conventional HIV T cell vaccine strategies have not been successful in containing acute peak viremia, nor in providing long-term control. We immunized rhesus macaques intramuscularly and rectally using a heterologous adenovirus vectored SIV vaccine regimen encoding normally weakly immunogenic tat...... immune hyperactivation as measured by naïve T cell depletion, Ki-67 and PD-1 expression on T cells. These results indicate that vaccination towards SIV accessory antigens vaccine can provide a level of acute control of SIV replication with a suggestion of beneficial immunological consequences in infected...... animals of unknown long-term significance. In conclusion, our studies demonstrate that a vaccine encoding subdominant antigens not normally associated with virus control can exert a significant impact on acute peak viremia....

  4. Role of very late antigen-1 in T-cell-mediated immunity to systemic viral infection

    DEFF Research Database (Denmark)

    Ørding Kauffmann, Susanne; Thomsen, Allan Randrup; Christensen, Jan Pravsgaard

    2006-01-01

    The T-cell response to lymphocytic choriomeningitis virus was studied in mice lacking very late antigen-1 (VLA-1). The generation of virus-specific effector T cells was unimpaired in VLA-1(-/-) mice. In the memory phase, VLA-1 deficiency did not influence the number of memory CD8(+) T cells...... significant differences between the two strains. Expression of VLA-1 was also found to be redundant regarding the ability of effector T cells to eliminate virus from internal organs of i.v. infected mice. Using delayed-type hypersensitivity (DTH) assays to evaluate subdermal CD8(+) T......, the current findings indicate that the expression of VLA-1 is not pivotal for T-cell-mediated antiviral immunity to a systemic infection....

  5. Membranoproliferative glomerulonephritis and mixed cryoglobulinemia after hepatitis C virus infection secondary to glomerular NS3 viral antigen deposits.

    Science.gov (United States)

    Bataille, Stanislas; Kaplanski, Gilles; Boucraut, José; Halfon, Philippe; Camus, Claire; Daniel, Laurent; Burtey, Stéphane; Berland, Yvon; Dussol, Bertrand

    2012-01-01

    We report on 3 cases of membranoproliferative glomerulonephritis associated with mixed cryoglobulin in patients with hepatitis C virus (HCV) antibodies but a negative blood viral load. These cases explore the pathogenesis of the renal disease. We searched for occult HCV infection in peripheral blood mononuclear cells, cryoprecipitate, bone marrow cells, and glomeruli using ultrasensitive PCR assays and immunohistochemistry. We also looked for infraclinical B cell lymphoma by computed tomodensitometry, bone marrow aspiration and biopsy, and lymphocyte typing. By PCR assays, we did not evidence occult hepatitis C infection in peripheral blood mononuclear cells, bone marrow cells, or cryoprecipitates. In the only patient with available kidney specimen, we evidenced HCV-NS3 antigen in glomeruli. HCV-associated lymphoma was excluded, but mild polyclonal B lymphocytosis was present in the 3 patients. Remission occurred spontaneously in 1 patient, and in another patient it occurred after rituximab treatment. The third patient was lost to follow-up. In patients with hepatitis C-negative viral load, membranoproliferative glomerulonephritis could be induced by the persistence of HCV antigen in the kidney but not in hematopoietic cells. Nonlymphomatous B cell proliferation may also be induced by chronic viral stimulation. Copyright © 2012 S. Karger AG, Basel.

  6. RNA Sequencing of Murine Norovirus-Infected Cells Reveals Transcriptional Alteration of Genes Important to Viral Recognition and Antigen Presentation

    Directory of Open Access Journals (Sweden)

    Daniel Enosi Tuipulotu

    2017-08-01

    Full Text Available Viruses inherently exploit normal cellular functions to promote replication and survival. One mechanism involves transcriptional control of the host, and knowledge of the genes modified and their molecular function can aid in understanding viral-host interactions. Norovirus pathogenesis, despite the recent advances in cell cultivation, remains largely uncharacterized. Several studies have utilized the related murine norovirus (MNV to identify innate response, antigen presentation, and cellular recognition components that are activated during infection. In this study, we have used next-generation sequencing to probe the transcriptomic changes of MNV-infected mouse macrophages. Our in-depth analysis has revealed that MNV is a potent stimulator of the innate response including genes involved in interferon and cytokine production pathways. We observed that genes involved in viral recognition, namely IFIH1, DDX58, and DHX58 were significantly upregulated with infection, whereas we observed significant downregulation of cytokine receptors (Il17rc, Il1rl1, Cxcr3, and Cxcr5 and TLR7. Furthermore, we identified that pathways involved in protein degradation (including genes Psmb3, Psmb4, Psmb5, Psmb9, and Psme2, antigen presentation, and lymphocyte activation are downregulated by MNV infection. Thus, our findings illustrate that MNV induces perturbations in the innate immune transcriptome, particularly in MHC maturation and viral recognition that can contribute to disease pathogenesis.

  7. Localization of viral antigens in leaf protoplasts and plants by immunogold labelling

    NARCIS (Netherlands)

    Lent, van J.W.M.

    1988-01-01

    This thesis describes the application of an immunocytochemical technique, immunogold labelling, new in the light and electron microscopic study of the plant viral infection. In Chapter 1 the present state of knowledge of the plant viral infection process, as revealed by

  8. Cloning, expression, purification and antigenic evaluation of ...

    African Journals Online (AJOL)

    Streptococcus pyogenes produce an extracellular hyaluronidase which is associated with the spread of the organism during infection. Enzyme hyaluronidase is capable of degrading hyaluronic acid. The aim of the present study was to clone and express antigenic regions of the hylA of S.pyogenes in Escherichia coli.

  9. Cloning, expression, purification and antigenic evaluation of ...

    African Journals Online (AJOL)

    Yomi

    2012-01-31

    Jan 31, 2012 ... Key word: Hyaluronidase gene, cloning, expression of recombinant gene, antigenic region. INTRODUCTION. Group A streptococcus (Streptococcus pyogenes) is an important species of gram-positive pathogenic extra- cellular bacteria. This bacteria can produce wide range of infectious diseases like ...

  10. Persistence of airway hyperresponsiveness and viral antigen following respiratory syncytial virus bronchiolitis in young guinea-pigs.

    Science.gov (United States)

    Riedel, F; Oberdieck, B; Streckert, H J; Philippou, S; Krusat, T; Marek, W

    1997-03-01

    Respiratory syncytial virus (RSV) bronchiolitis in infancy is known to be followed by chronic respiratory symptoms and airway hyperresponsiveness in a subgroup of patients. To further investigate the pathogenesis of RSV-induced chronic airway pathology, we infected young guinea-pigs at 4 weeks of age with RSV applied as an aerosol (n=30), and control guinea-pigs with virus-free culture medium (n=24). Infection was confirmed by positive antibody titre to RSV after 6 weeks, and by typical pathological changes of bronchiolitis after 1 week in six animals from each group. Airway hyperresponsiveness was measured weekly for 5 weeks by histamine challenge, using body-plethysmographic measurement of compressed air (CA). The provocative concentration of histamine producing significant airway obstruction (i.e. CA = 0.1 mL) (PC0.1 mL CA in mg x mL(-1)) was calculated from dose-response curves. Six weeks postinfection, the lungs were investigated for the presence of inflammation and of viral antigen by immunofluorescence and immunohistochemistry using a rabbit hyperimmune serum and monoclonal antibodies. Airway responsiveness was increased in the RSV group 1 week postinfection compared to the control group (PC0.1 mL CA median 2.50 vs >10 mg x mL(-1); p10 mg x mL(-1); p<0.001). During the same period, viral antigen persisted in the lungs of infected animals, although there was less inflammation at 6 weeks postinfection than at 1 week postinfection. In guinea-pigs, respiratory syncytial virus infection of the airways causes persistent airway hyperresponsiveness over a period of at least 5 weeks. During this time, viral antigen, but not inflammation, remains detectable in the lungs and might be responsible for ongoing airway hyperresponsiveness.

  11. Viral inhibition of the transporter associated with antigen processing (TAP: a striking example of functional convergent evolution.

    Directory of Open Access Journals (Sweden)

    Marieke C Verweij

    2015-04-01

    Full Text Available 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

  12. Combination of autoantibodies against NY-ESO-1 and viral capsid antigen immunoglobulin A for improved detection of nasopharyngeal carcinoma.

    Science.gov (United States)

    Peng, Yu-Hui; Xu, Yi-Wei; Qiu, Si-Qi; Hong, Chao-Qun; Zhai, Tian-Tian; Li, En-Min; Xu, Li-Yan

    2014-09-01

    Nasopharyngeal carcinoma (NPC) is one of the most common malignant tumors in Southern China and Southeast Asia, and early detection remains a challenge. Autoantibodies have been found to precede the manifestations of symptomatic cancer by several months to years, making their identification of particular relevance for early detection. In the present study, the diagnostic value of serum autoantibodies against NY-ESO-1 in NPC patients was evaluated. The study included 112 patients with NPC and 138 normal controls. Serum levels of autoantibodies against NY-ESO-1 and classical Epstein-Barr virus marker, viral capsid antigen immunoglobulin A (VCA-IgA), were measured by enzyme-linked immunosorbent assay. Measurement of autoantibodies against NY-ESO-1 and VCA-IgA demonstrated a sensitivity/specificity of 42.9/94.9% [95% confidence interval (CI), 33.7-52.6/89.4-97.8%] and 55.4/95.7% (95% CI, 45.7-64.7/90.4-98.2%), respectively. The area under receiver operating characteristic curve for autoantibodies against NY-ESO-1 (0.821; 95% CI, 0.771-0.871) was marginally lower than that for VCA-IgA (0.860; 95% CI, 0.810-0.910) in NPC. The combination of autoantibodies against NY-ESO-1 and VCA-IgA yielded an enhanced sensitivity of 80.4% (95% CI, 71.6-87.0%) and a specificity of 90.6% (95% CI, 84.1-94.7%). Moreover, detection of autoantibodies against NY-ESO-1 could differentiate early-stage NPC patients from normal controls. Our results suggest that autoantibodies against NY-ESO-1 may serve as a potential biomarker, as a supplement to VCA-IgA, for the screening and diagnosis of NPC.

  13. Cross-presentation of viral antigens in dribbles leads to efficient activation of virus-specific human memory T cells.

    Science.gov (United States)

    Ye, Wei; Xing, Yun; Paustian, Christopher; van de Ven, Rieneke; Moudgil, Tarsem; Hilton, Traci L; Fox, Bernard A; Urba, Walter J; Zhao, Wei; Hu, Hong-Ming

    2014-04-16

    Autophagy regulates innate and adaptive immune responses to pathogens and tumors. We have reported that autophagosomes derived from tumor cells after proteasome inhibition, DRibbles (Defective ribosomal products in blebs), were excellent sources of antigens for efficient cross priming of tumor-specific CD8⁺ T cells, which mediated regression of established tumors in mice. But the activity of DRibbles in human has not been reported. DRibbles or cell lysates derived from HEK293T or UbiLT3 cell lines expressing cytomegalovirus (CMV) pp65 protein or transfected with a plasmid encoding dominant HLA-A2 restricted CMV, Epstein-Barr virus (EBV), and Influenza (Flu) epitopes (CEF) were loaded onto human monocytes or PBMCs and the response of human CMV pp65 or CEF antigen-specific CD4⁺ and CD8⁺ memory T cells was detected by intracellular staining. The effect of cytokines (GM-CSF, IL-4, IL-12, TNF-α, IFN-α and IFN-γ) TLR agonists (Lipopolysaccharide, Polyinosinic-polycytidylic acid (poly(I:C), M52-CpG, R848, TLR2 ligand) and CD40 ligand on the cross-presentation of antigens contained in DRibbles or cell lysates was explored. In this study we showed that purified monocytes, or human PBMCs, loaded with DRibbles isolated from cells expressing CMV or CEF epitopes, could activate CMV- or CEF-specific memory T cells. DRibbles were significantly more efficient at stimulating CD8⁺ memory T cells compared to cell lysates expressing the same antigenic epitopes. We optimized the conditions for T-cell activation and IFN-γ production following direct loading of DRibbles onto PBMCs. We found that the addition of Poly(I:C), CD40 ligand, and GM-CSF to the PBMCs together with DRibbles significantly increased the level of CD8⁺ T cell responses. DRibbles containing specific viral antigens are an efficient ex vivo activator of human antigen-specific memory T cells specific for those antigens. This function could be enhanced by combining with Poly(I:C), CD40 ligand, and GM

  14. Presence of p24-Antigen Associated to Erythrocyte in HIV-Positive Individuals Even in Patients with Undetectable Plasma Viral Load

    Science.gov (United States)

    Garcia, Maria Noé; dos Ramos Farias, Maria Sol; Ávila, Maria Mercedes; Rabinovich, Roberto Daniel

    2011-01-01

    Background HIV adherence to erythrocytes has been demonstrated in vitro, and it has been suggested that erythrocytes may be carriers of the virus. However, the association between HIV particles or viral proteins and erythrocytes in HIV-infected individuals is still to be elucidated. Methodology/Principal Findings HIV-positive participants (n = 112) were classified into two groups according to values of three plasma viral loads (pVL) determined during the 12-month period prior to the study. The first group included 71 individuals with detectable pVL, whereas the second group included 41 individuals with undetectable pVL. Plasma viral load, erythrocyte-associated p24-antigen and p24-antigen in plasma were determined at the moment of the study. A total of 51 out of the 71 patients with detectable pVL showed erythrocyte-associated p24-antigen whereas 13 showed p24-antigen in plasma. Twenty-two out of the 51 patients with erythrocyte-associated p24-antigen showed pVLerythrocyte-associated p24-antigen was not related to p24-antigen in plasma or pVL levels in this group. Among the 41 patients with prior undetectable pVL, eight presented detectable pVL and erythrocyte-associated p24-antigen at the moment of the study. The other 33 showed undetectable pVL and five of these presented erythrocyte-associated p24-antigen. A positive relationship was found between the presence of erythrocyte-associated p24-antigen and the detectable pVL at the moment of the study (perythrocytes was determined and it was always accompanied by erythrocyte-associated p24-antigen detection. Conclusions/Significance This study demonstrates the presence of erythrocyte-associated p24-antigen in HIV-infected individuals. Since erythrocyte-associated p24-antigen is not always related to pVL or p24-antigen in plasma, erythrocyte-associated p24-antigen showed viral expression not represented in plasma. Therefore, the determination of erythrocyte-associated p24-antigen may contribute to better

  15. High-level HIV-1 viremia suppresses viral antigen-specific CD4+ T cell proliferation

    OpenAIRE

    McNeil, Andrew C.; Shupert, W. Lesley; Iyasere, Christiana A.; Hallahan, Claire W.; Mican, JoAnn; Davey, Richard T.; Connors, Mark

    2001-01-01

    In chronic viral infections of humans and experimental animals, virus-specific CD4+ T cell function is believed to be critical for induction and maintenance of host immunity that mediates effective restriction of viral replication. Because in vitro proliferation of HIV-specific memory CD4+ T cells is only rarely demonstrable in HIV-infected individuals, it is presumed that HIV-specific CD4+ T cells are killed upon encountering the virus, and maintenance of CD4+ T cell responses in some patien...

  16. An Evaluation of Usefulness of Prostate Specific Antigen and Digital ...

    African Journals Online (AJOL)

    Objective: To evaluate the usefulness of prostate specific antigen (PSA) and digital rectal examination (DRE) in the diagnosis of cancer of the prostate (CaP) amongst unscreened patients. Patients, Materials ans Methods: A prospective study168 unscreened men who were referred for evaluation for CaP. They all had a ...

  17. Targeting of non-dominant antigens as a vaccine strategy to broaden T-cell responses during chronic viral infection

    DEFF Research Database (Denmark)

    Holst, Peter Johannes; Jensen, Benjamin Anderschou Holbech; Ragonnaud, Emeline

    2015-01-01

    In this study, we compared adenoviral vaccine vectors with the capacity to induce equally potent immune responses against non-dominant and immunodominant epitopes of murine lymphocytic choriomeningitis virus (LCMV). Our results demonstrate that vaccination targeting non-dominant epitopes...... challenge with live virus, the CD8+ T cells specific for vaccine-encoded epitopes, displayed a phenotype typically associated with prolonged/persistent antigenic stimulation marked by high levels of KLRG-1, as compared to T cells reacting to epitopes not included in the vaccine. Notably, this association...... facilitates potent virus-induced T-cell responses against immunodominant epitopes during subsequent challenge with highly invasive virus. In contrast, when an immunodominant epitope was included in the vaccine, the T-cell response associated with viral challenge remained focussed on that epitope. Early after...

  18. [The isolation and evaluation of Aspergillus fumigatus antigens].

    Science.gov (United States)

    Lirio, V de S; de Assis, C M; Cano, M I; Lacaz, C da S

    1992-01-01

    Antigens from three strains of Aspergillus fumigatus (354, 356, and JIG) and an antiserum against the mixing of these antigens have been produced, and evaluated immunochemically. The antigens were obtained through a modified Coleman & Kaufman technique (culture filtrate concentrated by acetone). Analysis by the immunodiffusion test (ID) against homologous serum has yielded 100% sensitivity (with the studied sera). Concerning heterologous sera we found reactivity with a serum of a patient of candidiasis and another with histoplasmosis. The same result was obtained with a reference antigen in immunodiffusion, showing similar standards of response. Titration of the antiserum by ID and counterimmunoelectrophoresis showed a title of 1:32, and by complement fixation (micro-technique) a title of 1:128. Using immunoelectrophoresis (IEF), the produced antiserum yielded 8 lines of precipitation (5 in the anodic pole and 3 in the cathodic one). In SDS-PAGE at 12.5% the antigen has presented a rather complex electrophoretic profile (26 proteic subunits with a molecular weight ranging from 18 a > 100 kDa). Immunogenicity of the antigen was observed in all fractions of SDS-PAGE when the immunoblotting against the antiserum was carried out.

  19. Evaluation of three enzyme immunoassays for HIV-1 antigen detection.

    Science.gov (United States)

    Willoughby, P B; Lisker, A; Folds, J D

    1989-01-01

    Three enzyme immunoassay (EIA) methods for the detection of human immunodeficiency virus (HIV-1) were evaluated. Serum or plasma samples from 22 individuals seropositive for HIV-1 antibodies were tested with the Abbott, Coulter, and DuPont kits for presence of HIV-1 p24 antigen. Another 12 samples were tested with two kits only. Discordant results were obtained with 9 of 34 (26%) HIV-1-antibody-positive patient samples tested. Most of these discrepancies were found in samples containing less than 30 pg/ml of HIV-1 p24 core antigen. A sampling of sera from normal blood donors and patients with infectious or autoimmune diseases revealed a low level of false positive reactions, especially with sera containing antinuclear antibodies or rheumatoid factor. Noteworthy is the frequency of false positive reactions seen with the DuPont EIA for HIV-1 p24 antigen. 18/111 sera (16.2%) containing auto-antibodies tested positively with the DuPont HIV-1 p24 antigen EIA. The nonspecific nature of the test reactivity for 9/10 of these samples was confirmed using an HIV-1 p24 antigen inhibition assay. These findings are discussed in light of the need for HIV-1 antigen detection in the clinical laboratory and of other methods for HIV-1 detection: the polymerase chain reaction and measurements of reverse transcriptase activity.

  20. Asymmetric Assembly of Merkel Cell Polyomavirus Large T-Antigen Origin Binding Domains at the Viral Origin

    Energy Technology Data Exchange (ETDEWEB)

    C Harrison; G Meinke; H Kwun; H Rogalin; P Phelan; P Bullock; Y Chang; P Moore; A Bohm

    2011-12-31

    The double-stranded DNA polyomavirus Merkel cell polyomavirus (MCV) causes Merkel cell carcinoma, an aggressive but rare human skin cancer that most often affects immunosuppressed and elderly persons. As in other polyomaviruses, the large T-antigen of MCV recognizes the viral origin of replication by binding repeating G(A/G)GGC pentamers. The spacing, number, orientation, and necessity of repeats for viral replication differ, however, from other family members such as SV40 and murine polyomavirus. We report here the 2.9 {angstrom} crystal structure of the MCV large T-antigen origin binding domain (OBD) in complex with a DNA fragment from the MCV origin of replication. Consistent with replication data showing that three of the G(A/G)GGC-like binding sites near the center of the origin are required for replication, the crystal structure contains three copies of the OBD. This stoichiometry was verified using isothermal titration calorimetry. The affinity for G(A/G)GGC-containing double-stranded DNA was found to be {approx} 740 nM, approximately 8-fold weaker than the equivalent domain in SV40 for the analogous region of the SV40 origin. The difference in affinity is partially attributable to DNA-binding residue Lys331 (Arg154 in SV40). In contrast to SV40, a small protein-protein interface is observed between MCV OBDs when bound to the central region of the origin. This protein-protein interface is reminiscent of that seen in bovine papilloma virus E1 protein. Mutational analysis indicates, however, that this interface contributes little to DNA binding energy.

  1. Priming Cross-Protective Bovine Viral Diarrhea Virus-Specific Immunity Using Live-Vectored Mosaic Antigens.

    Directory of Open Access Journals (Sweden)

    Shehnaz Lokhandwala

    Full Text Available Bovine viral diarrhea virus (BVDV plays a key role in bovine respiratory disease complex, which can lead to pneumonia, diarrhea and death of calves. Current vaccines are not very effective due, in part, to immunosuppressive traits and failure to induce broad protection. There are diverse BVDV strains and thus, current vaccines contain representative genotype 1 and 2 viruses (BVDV-1 & 2 to broaden coverage. BVDV modified live virus (MLV vaccines are superior to killed virus vaccines, but they are susceptible to neutralization and complement-mediated destruction triggered by passively acquired antibodies, thus limiting their efficacy. We generated three novel mosaic polypeptide chimeras, designated NproE2123; NS231; and NS232, which incorporate protective determinants that are highly conserved among BVDV-1a, 1b, and BVDV-2 genotypes. In addition, strain-specific protective antigens from disparate BVDV strains were included to broaden coverage. We confirmed that adenovirus constructs expressing these antigens were strongly recognized by monoclonal antibodies, polyclonal sera, and IFN-γ-secreting T cells generated against diverse BVDV strains. In a proof-of-concept efficacy study, the multi-antigen proto-type vaccine induced higher, but not significantly different, IFN-γ spot forming cells and T-cell proliferation compared to a commercial MLV vaccine. In regards to the humoral response, the prototype vaccine induced higher BVDV-1 specific neutralizing antibody titers, whereas the MLV vaccine induced higher BVDV-2 specific neutralizing antibody titers. Following BVDV type 2a (1373 challenge, calves immunized with the proto-type or the MLV vaccine had lower clinical scores compared to naïve controls. These results support the hypothesis that a broadly protective subunit vaccine can be generated using mosaic polypeptides that incorporate rationally selected and validated protective determinants from diverse BVDV strains. Furthermore, regarding

  2. Evaluation of an Antigen-Antibody “Combination” Enzyme Linked ...

    African Journals Online (AJOL)

    Failure to detect the two samples with viral loads considered above threshold of detection for antigen proteins suggested a lack of sensitivity by this assay to discover viral capsid protein in patient samples. Genotyping of these samples revealed genotype 1b, a HCV-subtype which is widespread and should thus be easily ...

  3. Tracking serum antibody response to viral antigens with arrayed imaging reflectometry

    Science.gov (United States)

    Mace, Charles R.; Rose, Robert C.; Miller, Benjamin L.

    2009-02-01

    Arrayed Imaging Reflectometry, or "AIR", is a new label-free technique for detecting proteins that relies on bindinginduced changes in the response of an antireflective coating on the surface of a silicon ship. Because the technique provides high sensitivity, excellent dynamic range, and readily integrates with standard silicon wafer processing technology, it is an exceptionally attractive platform on which to build systems for detecting proteins in complex solutions. In our early research, we used AIR chips bearing secreted receptor proteins from enteropathogenic E. coli to develop sensors for this pathogen. Recently, we have been exploring an alternative strategy: Rather than detecting the pathogen directly, can one immobilize antigens from a pathogen, and employ AIR to detect antibody responses to those antigens? Such a strategy would provide enhanced sensitivity for pathogen detection (as the immune system essentially amplifies the "signal" caused by the presence of an organism to which it responds), and would also potentially prove useful in the process of vaccine development. We describe herein preliminary results in the application of such a strategy to the detection of antibodies to human papillomavirus (HPV).

  4. The level of viral infection of antigen-presenting cells correlates with the level of development of Theiler's murine encephalomyelitis virus-induced demyelinating disease.

    Science.gov (United States)

    Jin, Young Hee; Kang, Hyun Seok; Hou, Wanqiu; Meng, Liping; Kim, Byung S

    2015-02-01

    Intracerebral infection with Theiler's murine encephalomyelitis virus (TMEV) induces immune-mediated demyelinating disease in susceptible SJL/J mice but not in resistant C57BL/6 mice. Previous studies have indicated that the major histocompatibility complex (MHC) genes play the most prominent role in the development of TMEV-induced demyelinating disease. In this study, we used C57BL/6.S (B6.S) congenic mice, which carry H-2(s) MHC genes instead of H-2(b) MHC genes in conjunction with the C57BL/6 (B6) background genes. Our data show that virus-infected B6.S mice are free from disease and have significantly lower viral loads than susceptible SJL mice, particularly in the spinal cord. A strong protective Th1-type T helper response with virtually no pathogenic Th17 response was detected in B6.S mice, in contrast to the reduced Th1- and robust Th17-type responses in SJL mice. Notably, lower levels of viral infectivity in B6.S antigen-presenting cells (APCs) correlated with the disease resistance and T-cell-type response. In vitro studies using APCs from B6.S and SJL mice show that TLR2, -3, -4, and -7, but not TLR9, signaling can replace viral infection and augment the effect of viral infection in the differentiation of the pathogenic Th17 cell type. Taken together, these results strongly suggest that the viral replication levels in APCs critically affect the induction of protective versus pathogenic Th cell types via the signaling of pattern recognition receptors for innate immune responses. Our current findings further imply that the levels of viral infectivity/replication and TLR-mediated signaling play critical roles in the pathogenesis of chronic viral diseases. This study indicates that innate immune cytokines produced in antigen-presenting cells stimulating the T cell immune responses during early viral infection play a critical role in determining the susceptibility of mice to the development of demyelinating disease. The level of innate immune cytokines

  5. The Level of Viral Infection of Antigen-Presenting Cells Correlates with the Level of Development of Theiler's Murine Encephalomyelitis Virus-Induced Demyelinating Disease

    Science.gov (United States)

    Jin, Young Hee; Kang, Hyun Seok; Hou, Wanqiu; Meng, Liping

    2014-01-01

    ABSTRACT Intracerebral infection with Theiler's murine encephalomyelitis virus (TMEV) induces immune-mediated demyelinating disease in susceptible SJL/J mice but not in resistant C57BL/6 mice. Previous studies have indicated that the major histocompatibility complex (MHC) genes play the most prominent role in the development of TMEV-induced demyelinating disease. In this study, we used C57BL/6.S (B6.S) congenic mice, which carry H-2s MHC genes instead of H-2b MHC genes in conjunction with the C57BL/6 (B6) background genes. Our data show that virus-infected B6.S mice are free from disease and have significantly lower viral loads than susceptible SJL mice, particularly in the spinal cord. A strong protective Th1-type T helper response with virtually no pathogenic Th17 response was detected in B6.S mice, in contrast to the reduced Th1- and robust Th17-type responses in SJL mice. Notably, lower levels of viral infectivity in B6.S antigen-presenting cells (APCs) correlated with the disease resistance and T-cell-type response. In vitro studies using APCs from B6.S and SJL mice show that TLR2, -3, -4, and -7, but not TLR9, signaling can replace viral infection and augment the effect of viral infection in the differentiation of the pathogenic Th17 cell type. Taken together, these results strongly suggest that the viral replication levels in APCs critically affect the induction of protective versus pathogenic Th cell types via the signaling of pattern recognition receptors for innate immune responses. Our current findings further imply that the levels of viral infectivity/replication and TLR-mediated signaling play critical roles in the pathogenesis of chronic viral diseases. IMPORTANCE This study indicates that innate immune cytokines produced in antigen-presenting cells stimulating the T cell immune responses during early viral infection play a critical role in determining the susceptibility of mice to the development of demyelinating disease. The level of innate immune

  6. Evaluating viral marketing: isolating the key criteria in insurance industry

    National Research Council Canada - National Science Library

    Maria Gooyandeh Hagh

    2015-01-01

    This paper presents an empirical investigation to determine the key criteria that viral marketing practitioners believe should be implemented to measure about the success of viral marketing campaigns...

  7. Expression and characterization of highly antigenic domains of chicken anemia virus viral VP2 and VP3 subunit proteins in a recombinant E. coli for sero-diagnostic applications.

    Science.gov (United States)

    Lai, Guan-Hua; Lin, Ming-Kuem; Lien, Yi-Yang; Fu, Jiun-Hau; Chen, Hsi-Jien; Huang, Chi-Hung; Tzen, Jason T C; Lee, Meng-Shiou

    2013-08-13

    Chicken anemia virus (CAV) is an important viral pathogen that causes anemia and severe immunodeficiency syndrome in chickens worldwide. Generally, CAV infection occurs via vertical transmission in young chicks that are less than two weeks old, which are very susceptible to the disease. Therefore, epidemiological investigations of CAV infection and/or the evaluation of the immunization status of chickens is necessary for disease control. Up to the present, systematically assessing viral protein antigenicity and/or determining the immunorelevant domain(s) of viral proteins during serological testing for CAV infection has never been performed. The expression, production and antigenic characterization of CAV viral proteins such as VP1, VP2 and VP3, and their use in the development of diagnostic kit would be useful for CAV infection prevention. Three CAV viral proteins VP1, VP2 and VP3 was separately cloned and expressed in recombinant E. coli. The purified recombinant CAV VP1, VP2 and VP3 proteins were then used as antigens in order to evaluate their reactivity against chicken sera using indirect ELISA. The results indicated that VP2 and VP3 show good immunoreactivity with CAV-positive chicken sera, whereas VP1 was found to show less immunoreactivity than VP2 and VP3. To carry out the further antigenic characterization of the immunorelevant domains of the VP2 and VP3 proteins, five recombinant VP2 subunit proteins (VP2-435N, VP2-396N, VP2-345N, VP2-171C and VP2-318C) and three recombinant VP3 subunit proteins (VP3-123N, VP3-246M, VP3-366C), spanning the defined regions of VP2 and VP3 were separately produced by an E. coli expression system. These peptides were then used as antigens in indirect ELISAs against chicken sera. The results of these ELISAs using truncated recombinant VP2 and VP3 subunit proteins as coating antigen showed that VP2-345N, VP2-396N and VP3-246M gave good immunoreactivity with CAV-positive chicken sera compared to the other subunit proteins

  8. Human hepatitis B viral e antigen and its precursor P20 inhibit T lymphocyte proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Purvina, Maija; Hoste, Astrid; Rossignol, Jean-Michel [Universite de Versailles-Saint-Quentin-en-Yvelines, Laboratoire de Genetique et Biologie Cellulaire, EA 4589, 45 avenue des Etats-Unis, 78035 Versailles (France); Lagaudriere-Gesbert, Cecile, E-mail: cecile.lagaudriere-gesbert@u-psud.fr [Universite de Versailles-Saint-Quentin-en-Yvelines, Laboratoire de Genetique et Biologie Cellulaire, EA 4589, 45 avenue des Etats-Unis, 78035 Versailles (France)

    2012-01-27

    Highlights: Black-Right-Pointing-Pointer P20, precursor of the HBeAg, interacts with the cellular protein gC1qR. Black-Right-Pointing-Pointer HBeAg and P20 bind to T cell surface and inhibit mitogen-induced T cell division. Black-Right-Pointing-Pointer HBeAg and P20 inhibition of T cell proliferation is gC1qR and IL-1RAcP-independent. -- Abstract: The hepatitis B virus (HBV) Precore protein is processed through the secretory pathway directly as HBeAg or with the generation of an intermediate (P20). Precore gene has been shown to be implicated in viral persistence, but the functions of HBeAg and its precursors have not been fully elucidated. We show that the secreted proteins HBeAg and P20 interact with T cell surface and alter Kit-225 and primary T cells proliferation, a process which may facilitate the establishment of HBV persistence. Our data indicate that the N-terminal end of Precore is important for these inhibitory effects and exclude that they are dependent on the association of HBeAg and P20 with two characterized cell surface ligands, the Interleukin-1 Receptor Accessory Protein and gC1qR (present study).

  9. [Evaluation of Mascia Brunelli rapid antigen test in the diagnosis of group A streptococcal pharyngitis].

    Science.gov (United States)

    Barış, Ayşe; Anlıaçık, Nur; Bulut, Mehmet Emin; Deniz, Rıdvan; Yücel, Elif; Aktaş, Elif

    2017-01-01

    Pharyngitis in most cases is due to viral microorganisms however drug therapy without the detection of etiological agent leads to unnecessary use of antibiotics. On the other hand, when the etiologic agent is group A beta-hemolytic streptococci (GAS) it is important to identify the etiologic agent rapidly which will guide the treatment with appropriate antibiotics. The use of highly sensitive rapid tests will contribute significantly to early diagnosis and appropriate therapy. The aim of this study is to evaluate the efficacy of Mascia Brunelli rapid antigen test for the detection of GAS in throat swab samples. A total of 833 throat swab samples submitted to our laboratory with pre-diagnosis of pharyngitis were assessed between June 2016 and August 2016. The samples were simultaneously cultured and tested by rapid Mascia Brunelli Strep-A Card (Mascia Brunelli S.p.a, Italy). For identification, bacitracin sensitivity, PYR test and latex agglutination test in addition to Bruker MALDI-TOF MS (Daltonics, Germany) system were used. The density of GAS growth in the culture was noted. The samples that were false negative with Mascia Brunelli test were re-tested with QuickVue + Strep A Test (Quidel Corporation, San Diego, USA) rapid antigen test. A total of 833 patients, 376 (45.2%) female and 457 (54.8%) male were included in the study. The age range was between 0-94 years with a mean value of 7.86 ± 6.72. 125 (15%) and 94 (11.28%) of the samples were positive with culture and rapid antigen test, respectively. Mascia Brunelli antigen test gave negative results for 31 culture positive samples. Of these 31 samples, 28 were found positive by QuickVue + Strep A antigen test. As a result, the sensitivity of the test was found to be independent of the inoculum effect. The culture positivity rate in patients between 5-15 years was 18.4%. The sensitivity, specificity, positive predictive value, negative predictive value and the accuracy of Mascia Brunelli antigen test, with

  10. Adaptive memory of human NK-like CD8+ T-cells to ageing, and viral and tumor antigens

    Directory of Open Access Journals (Sweden)

    MARIA LUISA PITA-LOPEZ

    2016-12-01

    Full Text Available Human NK-like CD8+ T-cells are singular T-cells that express both T and NK cell markers such as CD56; their frequencies depend on their differentiation and activation during their lifetime. There is evidence of the presence of these innate CD8+ T-cells in the human umbilical cord, highlighting the necessity of investigating whether the NK-like CD8+ T-cells arise in the early stages of life (gestation. Based on the presence of cell surface markers, these cells have also been referred to as CD8+KIR+ T-cells, innate CD8+ T-cells, CD8+CD28−KIR+ T-cells or NKT-like CD8+CD56+ cells. However, the functional and co-signaling significance of these NK cell receptors on NK-like CD8+ T-cells is less clear. Also, the diverse array of co-stimulatory and co-inhibitory receptors are spatially and temporally regulated and may have distinct overlapping functions on NK-like CD8+ T-cell priming, activation, differentiation, and memory responses associated with different cell phenotypes. Currently, there is no consensus regarding the functional properties and phenotypic characterization of human NK-like CD8+ T-cells. Environmental factors, such as aging, autoimmunity, inflammation, viral antigen re-exposure or the presence of persistent tumor antigens have been shown to allow differentiation (adaptation of the NK-like CD8+ T-cells; the elucidation of this differentiation process and a greater understanding of the characteristics of these cells could be important for their eventual in potential therapeutic applications aimed at improving protective immunity. This review will attempt to elucidate a understanding of the characteristics of these cells with the goal towards their eventual use in potential therapeutic applications aimed at improving protective immunity.

  11. Cost-effectiveness of testing hepatitis B-positive pregnant women for hepatitis B e antigen or viral load.

    Science.gov (United States)

    Fan, Lin; Owusu-Edusei, Kwame; Schillie, Sarah F; Murphy, Trudy V

    2014-05-01

    To estimate the cost-effectiveness of testing pregnant women with hepatitis B (hepatitis B surface antigen [HBsAg]-positive) for hepatitis B e antigen (HBeAg) or hepatitis B virus (HBV) DNA, and administering maternal antiviral prophylaxis if indicated, to decrease breakthrough perinatal HBV transmission from the U.S. health care perspective. A Markov decision model was constructed for a 2010 birth cohort of 4 million neonates to estimate the cost-effectiveness of two strategies: testing HBsAg-positive pregnant women for 1) HBeAg or 2) HBV load. Maternal antiviral prophylaxis is given from 28 weeks of gestation through 4 weeks postpartum when HBeAg is positive or HBV load is high (10 copies/mL or greater). These strategies were compared with the current recommendation. All neonates born to HBsAg-positive women received recommended active-passive immunoprophylaxis. Effects were measured in quality-adjusted life-years (QALYs) and all costs were in 2010 U.S. dollars. The HBeAg testing strategy saved $3.3 million and 3,080 QALYs and prevented 486 chronic HBV infections compared with the current recommendation. The HBV load testing strategy cost $3 million more than current recommendation, saved 2,080 QALYs, and prevented 324 chronic infections with an incremental cost-effectiveness ratio of $1,583 per QALY saved compared with the current recommendations. The results remained robust over a wide range of assumptions. Testing HBsAg-positive pregnant women for HBeAg or HBV load followed by maternal antiviral prophylaxis if HBeAg-positive or high viral load to reduce perinatal hepatitis B transmission in the United States is cost-effective.

  12. Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors.

    Directory of Open Access Journals (Sweden)

    Hannah Karlsson

    Full Text Available CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs.

  13. ARCHITECT® HIV Ag/Ab Combo assay: correlation of HIV-1 p24 antigen sensitivity and RNA viral load using genetically diverse virus isolates.

    Science.gov (United States)

    Brennan, Catherine A; Yamaguchi, Julie; Vallari, Ana; Swanson, Priscilla; Hackett, John R

    2013-06-01

    HIV antigen/antibody (Ag/Ab) combination assays represent a significant advancement in assays used for diagnosing HIV infection based on their ability to detect acute and chronic infections. During acute HIV infection (AHI), detection depends on assay sensitivity for p24 Ag. To directly compare the Ag sensitivity of the ARCHITECT(®) HIV Ag/Ab Combo assay to RNA viral load using cell culture supernatants of virus isolates. HIV-1 isolates allow correlation in the total absence of an antibody response to infection and across genetically diverse HIV-1 group M strains. Thirty-five HIV-1 isolates comprising subtypes A-D, F and G, CRF01_AE, CRF02_AG, and unique recombinant forms were evaluated. Cell-free culture supernatant for each isolate was diluted to four levels and tested in the HIV Combo assay to determine a signal to cutoff ratio and the RealTime(®) HIV-1 assay to quantify RNA. The RNA copies/mL at the HIV Combo assay cutoff was determined. The median RNA copies/mL at the HIV Combo assay cutoff was 57,900 for individual virus isolates (range 26,440-102,400). A single plot of all the data gave a value of 58,500RNA copies/mL. An analysis of data published for acute HIV infection in human subjects gave a similar result; HIV Combo detected 97% of AHIs with RNA copies/mL > 30,700. Based on analysis of virus isolates, the ARCHITECT HIV Combo assay can detect p24 Ag when RNA is above approximately 58,000copies/mL. The correlation of viral load and Ag sensitivity was consistent across genetically diverse HIV-1 group M strains. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. A viral vectored prime-boost immunization regime targeting the malaria Pfs25 antigen induces transmission-blocking activity.

    Directory of Open Access Journals (Sweden)

    Anna L Goodman

    Full Text Available The ookinete surface protein Pfs25 is a macrogamete-to-ookinete/ookinete stage antigen of Plasmodium falciparum, capable of exerting high-level anti-malarial transmission-blocking activity following immunization with recombinant protein-in-adjuvant formulations. Here, this antigen was expressed in recombinant chimpanzee adenovirus 63 (ChAd63, human adenovirus serotype 5 (AdHu5 and modified vaccinia virus Ankara (MVA viral vectored vaccines. Two immunizations were administered to mice in a heterologous prime-boost regime. Immunization of mice with AdHu5 Pfs25 at week 0 and MVA Pfs25 at week 10 (Ad-MVA Pfs25 resulted in high anti-Pfs25 IgG titers, consisting of predominantly isotypes IgG1 and IgG2a. A single priming immunization with ChAd63 Pfs25 was as effective as AdHu5 Pfs25 with respect to ELISA titers at 8 weeks post-immunization. Sera from Ad-MVA Pfs25 immunized mice inhibited the transmission of P. falciparum to the mosquito both ex vivo and in vivo. In a standard membrane-feeding assay using NF54 strain P. falciparum, oocyst intensity in Anopheles stephensi mosquitoes was significantly reduced in an IgG concentration-dependent manner when compared to control feeds (96% reduction of intensity, 78% reduction in prevalence at a 1 in 5 dilution of sera. In addition, an in vivo transmission-blocking effect was also demonstrated by direct feeding of immunized mice infected with Pfs25DR3, a chimeric P. berghei line expressing Pfs25 in place of endogenous Pbs25. In this assay the density of Pfs25DR3 oocysts was significantly reduced when mosquitoes were fed on vaccinated as compared to control mice (67% reduction of intensity, 28% reduction in prevalence and specific IgG titer correlated with efficacy. These data confirm the utility of the adenovirus-MVA vaccine platform for the induction of antibodies with transmission-blocking activity, and support the continued development of this alternative approach to transmission-blocking malaria subunit

  15. Viral phylodynamics.

    Directory of Open Access Journals (Sweden)

    Erik M Volz

    Full Text Available Viral phylodynamics is defined as the study of how epidemiological, immunological, and evolutionary processes act and potentially interact to shape viralphylogenies. Since the coining of the term in 2004, research on viral phylodynamics has focused on transmission dynamics in an effort to shed light on how these dynamics impact viral genetic variation. Transmission dynamics can be considered at the level of cells within an infected host, individual hosts within a population, or entire populations of hosts. Many viruses, especially RNA viruses, rapidly accumulate genetic variation because of short generation times and high mutation rates. Patterns of viral genetic variation are therefore heavily influenced by how quickly transmission occurs and by which entities transmit to one another. Patterns of viral genetic variation will also be affected by selection acting on viral phenotypes. Although viruses can differ with respect to many phenotypes, phylodynamic studies have to date tended to focus on a limited number of viral phenotypes. These include virulence phenotypes, phenotypes associated with viral transmissibility, cell or tissue tropism phenotypes, and antigenic phenotypes that can facilitate escape from host immunity. Due to the impact that transmission dynamics and selection can have on viral genetic variation, viral phylogenies can therefore be used to investigate important epidemiological, immunological, and evolutionary processes, such as epidemic spread[2], spatio-temporal dynamics including metapopulation dynamics[3], zoonotic transmission, tissue tropism[4], and antigenic drift[5]. The quantitative investigation of these processes through the consideration of viral phylogenies is the central aim of viral phylodynamics.

  16. Development of an enhanced bovine viral diarrhea virus subunit vaccine based on E2 glycoprotein fused to a single chain antibody which targets to antigen-presenting cells

    Directory of Open Access Journals (Sweden)

    Andrea Pecora

    2015-03-01

    Full Text Available Bovine viral diarrhea virus (BVDV is an important cause of economic losses worldwide. E2 is an immunodominant protein and a promising candidate to develop subunit vaccines. To improve its immunogenicity, a truncated E2 (tE2 was fused to a single chain antibody named APCH, which targets to antigen-presenting cells. APCH-tE2 and tE2 proteins were expressed in the baculovirus system and their immunogenicity was firstly compared in guinea pigs. APCH-tE2 vaccine was the best one to evoke a humoral response, and for this reason, it was selected for a cattle vaccination experiment. All the bovines immunized with 1.5 µg of APCH-tE2 developed high levels of neutralizing antibodies against BVDV up to a year post-immunization, demonstrating its significant potential as a subunit vaccine. This novel vaccine is undergoing scale-up and was transferred to the private sector. Nowadays, it is being evaluated for registration as the first Argentinean subunit vaccine for cattle.

  17. Development of an enhanced bovine viral diarrhea virus subunit vaccine based on E2 glycoprotein fused to a single chain antibody which targets to antigen-presenting cells.

    Science.gov (United States)

    Pecora, Andrea; Malacari, Darío A; Pérez Aguirreburualde, María S; Bellido, Demian; Escribano, José M; Dus Santos, María J; Wigdorovitz, Andrés

    2015-01-01

    Bovine viral diarrhea virus (BVDV) is an important cause of economic losses worldwide. E2 is an immunodominant protein and a promising candidate to develop subunit vaccines. To improve its immunogenicity, a truncated E2 (tE2) was fused to a single chain antibody named APCH, which targets to antigen-presenting cells. APCH-tE2 and tE2 proteins were expressed in the baculovirus system and their immunogenicity was firstly compared in guinea pigs. APCH-tE2 vaccine was the best one to evoke a humoral response, and for this reason, it was selected for a cattle vaccination experiment. All the bovines immunized with 1.5 μg of APCH-tE2 developed high levels of neutralizing antibodies against BVDV up to a year post-immunization, demonstrating its significant potential as a subunit vaccine. This novel vaccine is undergoing scale-up and was transferred to the private sector. Nowadays, it is being evaluated for registration as the first Argentinean subunit vaccine for cattle. Copyright © 2014 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

  18. Epidemiological Characteristics of Carrying Viral Hepatitis B Antigen in the Population of the Nišava District

    Directory of Open Access Journals (Sweden)

    Kostić Marina

    2014-06-01

    Full Text Available The aim of this paper was to determine the trend of diseases and epidemiological characteristics of viral antigen carrying of hepatitis B for better implementation of prevention and control of the disease activity. The annual reports, reports of diseases - deaths from infectious diseases, epidemiological survey of the Public Health Institute (IPH Niš were used as the material. The period from 2002 to 2011 in the Nišava District was considered. A descriptive method was used. HBsAg carrying shows an upward trend (y=15+3.27 x. Most carriers are males (57.27%, live in urban areas (98.16/ 100.000 population, average age 41.92 years old ±SD 18.59, pensioners (22.42%. 54.05% are nephrology patients (almost all retirees under the age of 60 years old. Only 15.76% were hospitalized. The data on the vaccination status are insufficient. In 5.45%, co-infection with hepatitis C virus was found. 63.33% belong to the group of patients for whom there were no data on the mode of transmission. Hemodialysis patients make 16.67%, blood donors 9.39%, 6.36% pregnant women and injecting drug users 1.21%. The upward trend of carrying, the presence of all known risk groups in the population of carrying in the Nišava District points to the need for improved epidemiological surveillance, strict application of protective measures, conducting of statutory vaccination of all categories of people exposed to particular risk of infection as well as continuing education on preventive measures of both population and health care providers.

  19. Self-Amplifying mRNA Vaccines Expressing Multiple Conserved Influenza Antigens Confer Protection against Homologous and Heterosubtypic Viral Challenge

    Science.gov (United States)

    Magini, Diletta; Giovani, Cinzia; Mangiavacchi, Simona; Maccari, Silvia; Cecchi, Raffaella; Ulmer, Jeffrey B.; De Gregorio, Ennio; Geall, Andrew J.; Brazzoli, Michela; Bertholet, Sylvie

    2016-01-01

    Current hemagglutinin (HA)-based seasonal influenza vaccines induce vaccine strain-specific neutralizing antibodies that usually fail to provide protection against mismatched circulating viruses. Inclusion in the vaccine of highly conserved internal proteins such as the nucleoprotein (NP) and the matrix protein 1 (M1) was shown previously to increase vaccine efficacy by eliciting cross-reactive T-cells. However, appropriate delivery systems are required for efficient priming of T-cell responses. In this study, we demonstrated that administration of novel self-amplifying mRNA (SAM®) vectors expressing influenza NP (SAM(NP)), M1 (SAM(M1)), and NP and M1 (SAM(M1-NP)) delivered with lipid nanoparticles (LNP) induced robust polyfunctional CD4 T helper 1 cells, while NP-containing SAM also induced cytotoxic CD8 T cells. Robust expansions of central memory (TCM) and effector memory (TEM) CD4 and CD8 T cells were also measured. An enhanced recruitment of NP-specific cytotoxic CD8 T cells was observed in the lungs of SAM(NP)-immunized mice after influenza infection that paralleled with reduced lung viral titers and pathology, and increased survival after homologous and heterosubtypic influenza challenge. Finally, we demonstrated for the first time that the co-administration of RNA (SAM(M1-NP)) and protein (monovalent inactivated influenza vaccine (MIIV)) was feasible, induced simultaneously NP-, M1- and HA-specific T cells and HA-specific neutralizing antibodies, and enhanced MIIV efficacy against a heterologous challenge. In conclusion, systemic administration of SAM vectors expressing conserved internal influenza antigens induced protective immune responses in mice, supporting the SAM® platform as another promising strategy for the development of broad-spectrum universal influenza vaccines. PMID:27525409

  20. Self-Amplifying mRNA Vaccines Expressing Multiple Conserved Influenza Antigens Confer Protection against Homologous and Heterosubtypic Viral Challenge.

    Directory of Open Access Journals (Sweden)

    Diletta Magini

    Full Text Available Current hemagglutinin (HA-based seasonal influenza vaccines induce vaccine strain-specific neutralizing antibodies that usually fail to provide protection against mismatched circulating viruses. Inclusion in the vaccine of highly conserved internal proteins such as the nucleoprotein (NP and the matrix protein 1 (M1 was shown previously to increase vaccine efficacy by eliciting cross-reactive T-cells. However, appropriate delivery systems are required for efficient priming of T-cell responses. In this study, we demonstrated that administration of novel self-amplifying mRNA (SAM® vectors expressing influenza NP (SAM(NP, M1 (SAM(M1, and NP and M1 (SAM(M1-NP delivered with lipid nanoparticles (LNP induced robust polyfunctional CD4 T helper 1 cells, while NP-containing SAM also induced cytotoxic CD8 T cells. Robust expansions of central memory (TCM and effector memory (TEM CD4 and CD8 T cells were also measured. An enhanced recruitment of NP-specific cytotoxic CD8 T cells was observed in the lungs of SAM(NP-immunized mice after influenza infection that paralleled with reduced lung viral titers and pathology, and increased survival after homologous and heterosubtypic influenza challenge. Finally, we demonstrated for the first time that the co-administration of RNA (SAM(M1-NP and protein (monovalent inactivated influenza vaccine (MIIV was feasible, induced simultaneously NP-, M1- and HA-specific T cells and HA-specific neutralizing antibodies, and enhanced MIIV efficacy against a heterologous challenge. In conclusion, systemic administration of SAM vectors expressing conserved internal influenza antigens induced protective immune responses in mice, supporting the SAM® platform as another promising strategy for the development of broad-spectrum universal influenza vaccines.

  1. A multicenter evaluation of the Biotest legionella urinary antigen EIA

    DEFF Research Database (Denmark)

    Harrison, Timothy; Uldum, Søren; Alexiou-Daniel, Stella

    1998-01-01

    OBJECTIVES: To undertake a multicenter study to evaluate the Biotest legionella urinary antigen enzyme immunoassay (EIA) performance against those EIAs already in use in 14 European laboratories. METHODS: Each laboratory examined urine specimens from appropriate patients using both their current...... in the laboratories' current EIAs, and in 94.6% of those specimens which were positive for Legionella pneumophila serogroup 1. CONCLUSION: The Biotest EIA is simple to use and specific and the results obtained in different laboratories show excellent agreement. The assay compares well existing EIAs, at least for L...

  2. Seasonal Evaluation of Antigenic Bacterial Infections Among Working Class in the Inner City of Houston

    Directory of Open Access Journals (Sweden)

    Ebere C. Anyanwu

    2004-01-01

    Full Text Available This paper evaluates the monthly, quarterly, and seasonal variation of antigenic bacterial infections among the working class in the inner city of Houston using the Wellcogen Rapid Test methods. One of the aims was to demonstrate how this method could be used effectively in screening patients at risk and preventing the spread of antigenic bacteria such as Streptococcus pneumoniae, Haemophilus influenzae b, Streptococcus (Strep b, and Neisseria meningitidis (mainly group c and b. A total of 2,837 patients were screened for bacterial infections; 908 (32% were male and 1,929 (68% were female. The age range was between 2 and 70 years. Of the total group, 356 (12.5% patients were positive; 203 (57% were female while 153 (43% were male (male/female ratio of 1:1.3. Medically underserved and immune suppressed populations are the most affected by these bacterial infections. Blacks are the most affected (48% compared to Native Americans (1%, but children under 10 years of age have the highest incidence. This research showed, in addition, that the Wellcogen Rapid Tests are effective (356 cases identified for a rapid screening of infectious bacteria. Explanation for these results was probably due to poor living conditions, poor hygiene, and viral immune suppression in adults and immature immune systems in neonates and children under 10 years of age.

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

    Science.gov (United States)

    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

  4. CD8 T Cells in Innate Immune Responses: Using STAT4-Dependent but Antigen-Independent Pathways to Gamma Interferon during Viral Infection

    Science.gov (United States)

    Suarez-Ramirez, Jenny E.; Tarrio, Margarite L.; Kim, Kwangsin; Demers, Delia A.

    2014-01-01

    ABSTRACT The cytokine gamma interferon (IFN-γ), with antimicrobial and immunoregulatory functions, can be produced by T cells following stimulation through their T cell receptors (TCRs) for antigen. The innate cytokines type 1 IFNs and interleukin-12 (IL-12) can also stimulate IFN-γ production by natural killer (NK) but not naive T cells. High basal expression of signal transducer and activator of transcription 4 (STAT4), used by type 1 IFN and IL-12 to induce IFN-γ as well as CD25, contributes to the NK cell responses. During acute viral infections, antigen-specific CD8 T cells are stimulated to express elevated STAT4 and respond to the innate factors with IFN-γ production. Little is known about the requirements for cytokine compared to TCR stimulation. Primary infections of mice with lymphocytic choriomeningitis virus (LCMV) demonstrated that although the elicited antigen-specific CD8 T cells acquired STAT4-dependent innate cytokine responsiveness for IFN-γ and CD25 induction ex vivo, TCR stimulation induced these through STAT4-independent pathways. During secondary infections, LCMV-immune CD8 T cells had STAT4-dependent IFN-γ expression at times of innate cytokine induction but subsequently expanded through STAT4-independent pathways. At times of innate cytokine responses during infection with the antigen-distinct murine cytomegalovirus virus (MCMV), NK and LCMV-immune CD8 T cells both had activation of pSTAT4 and IFN-γ. The T cell IFN-γ response was STAT4 and IL-12 dependent, but antigen-dependent expansion was absent. By dissecting requirements for STAT4 and antigen, this work provides novel insights into the endogenous regulation of cytokine and proliferative responses and demonstrates conditioning of innate immunity by experience. PMID:25336459

  5. Development and evaluation of two subunit vaccine candidates containing antigens of hepatitis E virus, rotavirus, and astrovirus.

    Science.gov (United States)

    Xia, Ming; Wei, Chao; Wang, Leyi; Cao, Dianjun; Meng, Xiang-Jin; Jiang, Xi; Tan, Ming

    2016-05-19

    Hepatitis E virus (HEV), rotavirus (RV), and astrovirus (AstV) are important pathogens that transmit through a common fecal-oral route, causing hepatitis (HEV) and gastroenteritis (RV and AstV) respectively in humans. In this study, we developed and evaluated two subunit vaccine candidates that consisted of the same protruding or spike protein antigens of the three viruses in two formats, a fusion of the three antigens into one molecule (fused vaccine) vs. a mixture of the three free antigens together (mixed vaccine). Both vaccines were easily made via E. coli expression system. Mouse immunization experiments showed that the fused vaccine elicited significantly higher antibody responses against the three viral antigens than those induced by the mixed vaccine. In addition, the mouse post-immune antisera of the fused vaccine revealed significantly higher neutralizing titers against HEV infection in cell culture, as well as significantly higher 50% blocking titers (BT50) against RV VP8-HBGA receptor interactions than those of the post-immune antisera after immunization of the mixed vaccine. Thus, the fused vaccine is a promising trivalent vaccine candidate against HEV, RV, and AstV, which is worth for further development.

  6. Immunological evaluation of some antigens of Lucilia sericata larvae.

    Science.gov (United States)

    Mohamed, Hoda S; Fahmy, Magdy M; Attia, Marwa M; El Khateeb, Rabab M; Shalaby, Hatem A; Mohamed, Mai A

    2017-12-01

    The present study aimed to select an antigen of Lucilia sericata larvae showing both high antigenicity and cross-reactive binding abilities with other related antigens of L. sericata larvae for obtaining a promising candidate vaccine antigen. The ELISA results primary concluded that among the excretory secretory (ES) and midgut (MG) antigens of the different larval instars of L. sericata, MGL2 could be characterized as antigen which was able to reflect the highest level of antigenicity and cross-reactivity with the other tested L. sericata antigens. The results were extended to spot the light on the relation between different protein bands in MGL2 and rabbit hyper- immune sera (HIS) raised against the other tested antigens using SDS-PAGE and Western blot technique. Analysis by SDS-PAGE of ES and MG antigens of the different larval instars of L. sericata revealed common protein bands at molecular weights of about 10, 12, 16, 20, 28, 33 and 46 kDa. Western blotting of MGL2 antigen transferred to nitrocellulose sheet revealed reaction by MGL2 HIS to five polypeptide bands; 20, 28, 33, 46 and 63 kDa. Three bands of 28, 33 and 63 kDa were the most prominent bands detected whereas; there was a weak reaction with bands of 20 and 46 kDa. But what was apparent in Western blot was a strong reaction of all tested HIS with a polypeptide band of 63 kDa. This band might be considered to be the main cause of cross reactive binding ability of MGL2 antigen that had been recorded previously in ELISA technique.

  7. Evaluation of Gastrothylax crumenifer antigenic preparation in serodiagnosis of paramphistomiasis in sheep.

    Science.gov (United States)

    Ahmad, Tariq; Reshi, Mohammad Latif; Cheshti, M Z; Tanveer, Syed; Shah, Zaffar Amin; Fomada, Bashir Ahmad; Raina, O K

    2014-04-01

    An evaluation of Gastrothylax crumenifer crude antigen preparation viz., Somatic Antigen (SAg), Excretory Secretory Antigen (ESAg) and Egg Antigen (EAg) in serodiagnosis of disease was undertaken. Test sera samples were obtained from 30 Paramphistomiasis Positive and 30 Gastrothylax free sheep slaughtered at Hazratbal Kashmir. The referral antigenic preparation were evaluated against Paramphistomiasis positive sera, via., control negative sera, using double immunodiffusion test (DID), (IEP) Immunoelectrophoretic assay and ELISA. The performance of referral antigens, as assessed from percent sensitivity and specificity, revealed an increasing trend from DID (Double immunodiffusion-An immunological technique used in the detection, identification and quantification of antibodies and antigens) to IEP (immunoelectrophoresis-A general name for a number of biochemical methods for separation and characterization of proteins based on electrophoresis and reaction with antibodies), followed by ELISA, detecting higher number of sheep positive for paramphistomiasis. In ELISA the ESAg and SAg were evaluated as most reactive antigens with no significant difference and EAg was the least antigenic. In IEP, EAg had the higher sensitivity (60%) and analogous specificity of SAg and ESAg. The formation of the preceptin lines in the proximity to EAg containing wells (cathode end) in IEP was suggestive of higher molecular weight of G. crumenifer specific protein molecules with slower rate of migration. Purification and characterization of G. crumenifer and identification of specific antigenic molecules, particularly in EAg has been suggested for qualitative improvement of diagnostic value of the antigens in the tests used here in.

  8. Evaluation of the Korean Isolate-1 Tachyzoite Antigen for Serodiagnosis of Toxoplasmosis

    OpenAIRE

    Shin, Eun-Hee; Kim, Dong-Hee; Lin, Aifen; Lee, Jo-Woon-Yi; Kim, Hyo-Jin; Ahn, Myoung-Hee; Chai, Jong-Yil

    2008-01-01

    To evaluate the usefulness of the Korean Isolate-1 (KI-1) antigen for serodiagnosis of toxoplasmosis, antigen profiles of KI-1 tachyzoites were analyzed in comparison with RH tachyzoites by SDS-PAGE and immunoblotting. ELISA was performed on latex agglutination (LA)-positive and negative serum samples using KI-1 and RH antigens. Immunoblotting of the KI-1 antigen showed multiple antigen bands with molecular sizes of 22-105 kDa. Among them, 1 and 6 common bands were noted against a KI-1-infect...

  9. Local CD4 and CD8 T-cell reactivity to HSV-1 antigens documents broad viral protein expression and immune competence in latently infected human trigeminal ganglia.

    Directory of Open Access Journals (Sweden)

    Monique van Velzen

    2013-08-01

    Full Text Available Herpes simplex virus type 1 (HSV-1 infection results in lifelong chronic infection of trigeminal ganglion (TG neurons, also referred to as neuronal HSV-1 latency, with periodic reactivation leading to recrudescent herpetic disease in some persons. HSV-1 proteins are expressed in a temporally coordinated fashion during lytic infection, but their expression pattern during latent infection is largely unknown. Selective retention of HSV-1 reactive T-cells in human TG suggests their role in controlling reactivation by recognizing locally expressed HSV-1 proteins. We characterized the HSV-1 proteins recognized by virus-specific CD4 and CD8 T-cells recovered from human HSV-1-infected TG. T-cell clusters, consisting of both CD4 and CD8 T-cells, surrounded neurons and expressed mRNAs and proteins consistent with in situ antigen recognition and antiviral function. HSV-1 proteome-wide scans revealed that intra-TG T-cell responses included both CD4 and CD8 T-cells directed to one to three HSV-1 proteins per person. HSV-1 protein ICP6 was targeted by CD8 T-cells in 4 of 8 HLA-discordant donors. In situ tetramer staining demonstrated HSV-1-specific CD8 T-cells juxtaposed to TG neurons. Intra-TG retention of virus-specific CD4 T-cells, validated to the HSV-1 peptide level, implies trafficking of viral proteins from neurons to HLA class II-expressing non-neuronal cells for antigen presentation. The diversity of viral proteins targeted by TG T-cells across all kinetic and functional classes of viral proteins suggests broad HSV-1 protein expression, and viral antigen processing and presentation, in latently infected human TG. Collectively, the human TG represents an immunocompetent environment for both CD4 and CD8 T-cell recognition of HSV-1 proteins expressed during latent infection. HSV-1 proteins recognized by TG-resident T-cells, particularly ICP6 and VP16, are potential HSV-1 vaccine candidates.

  10. Evaluation of ascitic soluble human leukocyte antigen-G for distinguishing malignant ascites from benign ascites.

    Science.gov (United States)

    Sun, Juan; Chang, Yan-Xiang; Niu, Chun-Yan

    2017-11-01

    The overexpression of soluble human leukocyte antigen-G is associated with malignant tumours. The purpose of our study was to detect soluble human leukocyte antigen-G concentrations in ascites and to evaluate the value of ascitic soluble human leukocyte antigen-G for the diagnosis of malignant ascites. Enzyme-linked immunosorbent assay was used to detect soluble human leukocyte antigen-G levels in 64 patients with malignant ascites and 30 patients with benign ascites. Receiver operating characteristic curves were used to evaluate the diagnostic efficacy of ascitic soluble human leukocyte antigen-G for the detection of malignant ascites. Ascitic soluble human leukocyte antigen-G levels were significantly higher in the malignant ascites group than in the benign ascites group (20.718 ± 3.215 versus 12.467 ± 3.678 µg/L, t = 7.425, p human leukocyte antigen-G was 0.957 (95% confidence interval, 0.872-0.992). At a cut-off value of 19.60 µg/L, the sensitivity and specificity of ascitic soluble human leukocyte antigen-G were 87.5% (95% confidence interval, 71.0%-96.5%) and 100% (95% confidence interval, 88.4%-100%), respectively. With respect to area under the receiver operating characteristic curve, sensitivity and specificity, ascitic carcinoembryonic antigen (0.810, 68.75% and 83.33%, respectively) and carbohydrate antigen 19-9 (0.710, 65.63% and 70%, respectively) significantly differed (all p human leukocyte antigen-G was 75%, which was higher than the corresponding rates for ascitic carcinoembryonic antigen (31.25%) and carbohydrate antigen 19-9 (6.25%; both p human leukocyte antigen-G exhibited good performance for diagnosing malignant ascites, and particularly those that were cytology-negative and biopsy-positive.

  11. [Evaluation of Abbott Fourth Generation HIV Antigen and Antibody Assays.].

    Science.gov (United States)

    Kang, Hee Jung; Yoo, Kyeong Ha; Kim, Han Sung; Cho, Hyoun Chan

    2006-02-01

    In order to reduce the diagnostic window period between the time of human immunodeficiency virus (HIV) infection and serological diagnosis, new fourth generation screening assays which detect HIV p24 antigen and specific antibody simultaneously have been developed. In this study, we evaluated the performance of a new fourth generation assay. We compared a new fourth generation assay, Architect HIV Ag/Ab combo, with another fourth generation assay AxSYM HIV Ag/Ab combo and a third generation assay, AxSYM HIV 1/2 gO for their performance. The assays were evaluated using 3 HIV seroconversion panels, 305 sera of healthy subjects and 100 sera of patients with HBsAg or anti-HCV antibodies. Within-run and total coefficient variations of the three screening assays were analyzed for the evaluation of precision. Architect HIV Ag/Ab combo shortened the window period by 8.7+/-2.1 days relative to AxSYM HIV 1/2 gO and 2.0+/-2.0 days relative to AxSYM HIV Ag/Ab combo in seroconversion panels. Architect HIV Ag/Ab combo presented the best performance in precision among the three reagents; total CV for positive control was 3.6%, 9.6% and 4.6% for Architect HIV Ag/Ab combo, AxSYM HIV Ag/Ab combo and AxSYM HIV 1/2 gO, respectively. Specificities of three assays were not different in this study. HIV Ag/Ab combined assays reduced the diagnostic window as compared to the third generation screening assays, enabling an earlier diagnosis of HIV infection. A new fourth generation assay, Architect HIV Ag/Ab combo presents a better performance than AxSYM HIV Ag/Ab combo, showing improved seroconversion sensitivity and precision.

  12. Evaluation of the siemens HIV antigen-antibody immunoassay.

    Science.gov (United States)

    Vallefuoco, Luca; Aden Abdi, Fatima; Sorrentino, Rosanna; Spalletti-Cernia, Daniela; Mazzarella, Claudia; Barbato, Sara; Perna, Enzo; Buffolano, Wilma; Di Nicuolo, Giuseppe; Portella, Giuseppe

    2014-01-01

    Fourth-generation assays for the simultaneous detection of human immunodeficiency virus (HIV) antigen and antibodies are available on the international market and are currently used for blood donor screening and for HIV diagnosis. In this study we evaluated the performance of the novel automated fourth-generation ADVIA Centaur® HIV Ag/Ab Combo assay. The assay detected seroconversion at the same bleed or at least one bleed earlier in panels with respect to other assays and showed a detection efficacy equal to those of other assays in a low-titer panel. Samples obtained from blood donors (n = 2,778) or from HIV-positive patients (HIV-1 B subtype, n = 82; non-B subtype, n = 71) were also tested, showing a good correlation with other fourth-generation assays. We assessed the performance of 3 fourth-generation assays for detecting in utero transmitted anti-HIV antibodies and found a more specific detection efficiency with the ADVIA Centaur HIV Ag/Ab Combo assay compared to the other fourth-generation assays.

  13. Evaluating viral marketing: isolating the key criteria in insurance industry

    Directory of Open Access Journals (Sweden)

    Maria Gooyandeh Hagh

    2015-06-01

    Full Text Available This paper presents an empirical investigation to determine the key criteria that viral marketing practitioners believe should be implemented to measure about the success of viral marketing campaigns in insurance industry. The study designs a questionnaire in Likert scale where the effects of four independent variables, personal, message, media and tools characteristics are measured on an Iranian insurance firm’s reputation as well as service expansion. Cronbach alphas were measured for all components of the survey and they were all well above the minimum acceptable level. Using regression analysis, the study has determined positive and meaningful relationships between insurance firm’s reputation as well as service expansion and four independent variables.

  14. [Evaluation of serum specific IgM detection in diagnosis of respiratory viral infections in children].

    Science.gov (United States)

    Song, Qin-wei; Zhu, Ru-nan; Deng, Jie; Wang, Fang; Zhao, Lin-qing; Sun, Yu; Ding, Ya-xin; Qian, Yuan

    2012-06-01

    The present study was designed to explore the practical application of the rapid etiological diagnosis by detecting specific IgM antibody against common respiratory viruses in children with acute lower respiratory infections (ALRI). Clinical specimens including nasopharyngeal aspirates and serum of acute phase from hospitalized children were collected from 207 infants and children with acute lower respiratory infections from March 2009 to September 2010. Seven common respiratory virus antigens were identified from the collected nasopharyngeal aspirates by direct immunofluorescence assay (DFA). ELISA was used to detect specific IgM antibody against RSV, ADV, IFVA, IFVB and PIV, while indirect immunofluorescence assay (IFA) was used to detect specific IgM antibody against RSV, ADV, IFVA, IFVB, PIV1, PIV2 and PIV3 in collected acute phase serum. The overall positive rates to detect viral antigen by using DFA, ELISA and IFA was 67.6%, 57.5% and 39.6%, respectively. The consistent rate of ELISA and IFA versus accepted DFA were 21.7% and 31.4%, respectively. The average days from onset of the symptoms to blood sample collection for those with the consistent results by ELISA and DFA were 12.0 d for ADV, 9.6 d for PIV2, 9.5 d for IFV, and 5.3 d for RSV, respectively, and by IFA and DFA were 15.0 d for PIV3, 9.2 d for ADV, and 7.4 d for RSV, respectively. Among all age groups, the consistent rate of serum viral IgM and antigen detections was highest in children younger than 3 years old. Although there were differences between serum IgM antibody and viral antigen detections, specific IgM antibody detection was of value in early and rapid etiological diagnosis of pediatric ALRI, especially for young children. It could provide serologic evidence of respiratory virus infection. The diagnostic rate of pathogen could be improved if it was used in combination with viral antigen diagnostic methods.

  15. An evaluation of a recombinant multiepitope based antigen for detection of Toxoplasma gondii specific antibodies.

    Science.gov (United States)

    Hajissa, Khalid; Zakaria, Robaiza; Suppian, Rapeah; Mohamed, Zeehaida

    2017-12-29

    The inefficiency of the current tachyzoite antigen-based serological assays for the serodiagnosis of Toxoplasma gondii infection mandates the need for acquirement of reliable and standard diagnostic reagents. Recently, epitope-based antigens have emerged as an alternative diagnostic marker for the achievement of highly sensitive and specific capture antigens. In this study, the diagnostic utility of a recombinant multiepitope antigen (USM.TOXO1) for the serodiagnosis of human toxoplasmosis was evaluated. An indirect enzyme-linked immunosorbent assay (ELISA) was developed to evaluate the usefulness of USM.TOXO1 antigen for the detection of IgG antibodies against Toxoplasma gondii in human sera. Whereas the reactivity of the developed antigen against IgM antibody was evaluated by western blot and Dot enzyme immunoassay (dot-EIA) analysis. The diagnostic performance of the new antigens in IgG ELISA was achieved at the maximum values of 85.43% and 81.25% for diagnostic sensitivity and specificity respectively. The USM.TOXO1 was also proven to be reactive with anti- T. gondii IgM antibody. This finding makes the USM.TOXO1 antigen an attractive candidate for improving the toxoplasmosis serodiagnosis and demonstrates that multiepitope antigens could be a potential and promising diagnostic marker for the development of high sensitive and accurate assays.

  16. Early Viral Kinetics with Telbivudine, Tenofovir or Combination of Both in Immunotolerant Patients with Hepatitis B e Antigen-Positive Chronic Hepatitis B.

    Science.gov (United States)

    Leung, Nancy W Y; Herrmann, Eva; Lau, George K K; Chan, Henry L Y; So, Tokutei M K; Zeuzem, Stefan; Dong, Yu; Trylesinski, Aldo; Naoumov, Nikolai V

    2014-12-01

    Viral kinetics has proved useful in understanding antiviral potency, determining antiviral profiles and optimizing treatment strategy. This was a randomized, open-label study comparing the viral kinetics in 46 hepatitis B e antigen-positive patients during 12-week treatment with telbivudine monotherapy, tenofovir monotherapy or the combination of telbivudine plus tenofovir. A standard biphasic mathematical model was used to compare hepatitis B virus (HBV) DNA decay parameters. Forty-six patients received telbivudine (n = 16), tenofovir (n = 14) or telbivudine plus tenofovir (n = 16). From baseline to Week 12, the mean (SD) reduction in HBV DNA levels was not significantly different between treatment groups: -3.9 (0.9) log10 copies/mL in telbivudine group, -4.2 (0.7) log10 copies/mL in tenofovir group, and -4.4 (1.0) log10 copies/mL in combination group. No significant difference was observed among the three groups for viral clearance rate per day (0.97, 1.02, and 0.88, respectively) or for infected cell loss rate per day (0.04, 0.05, and 0.05, respectively). Antiviral efficiency in blocking viral production was similar in the monotherapy groups (median; 99.7% in telbivudine group and 99.4% in tenofovir group), but was slightly better and more homogeneous in the combination treatment group than in the monotherapy groups: mean (SD), 99.1% (0.8%) and 98.8% (1.6%), respectively (Wald-Wolfowitz test; P = 0.038). All treatments were well tolerated and no serious adverse event was reported during the study. Of the 46 patients in the safety population, 23 experienced adverse events. Most of the adverse events were not suspected to be related to the study drug by the investigators. Monotherapy with telbivudine or tenofovir showed similar antiviral effectiveness in HBV DNA reduction and viral kinetics of HBV DNA decay. Efficiency in blocking viral production was slightly improved in the combination treatment group compared to the monotherapy groups.

  17. Establishing China's national standards of antigen content and neutralizing antibody responses for evaluation of enterovirus 71 (EV71) vaccines.

    Science.gov (United States)

    Liang, Zhenglun; Mao, Qunying; Gao, Qiang; Li, Xiuling; Dong, Chenghong; Yu, Xiang; Yao, Xin; Li, Fengxiang; Yin, Weidong; Li, Qihan; Shen, Xinliang; Wang, Junzhi

    2011-12-06

    Enterovirus 71 (EV71) is a highly infectious agent that causes hand-foot-mouth disease (HFMD) in humans. Effective vaccination against EV71 infection is critically important, given the recent outbreak of HFMD in the Asia-Pacific region, where it has shown significant mortality and morbidity. There is currently no approved anti-viral therapy available to treat the disease. While several vaccine manufacturers are actively developing EV71 vaccines, there are no international reference standards available to conduct quality control on EV71 vaccines or to assess the effectiveness of EV71 vaccines in immunized populations. In the current report, antigen reference standard based on the C4 subtype of the EV71 vaccine strain was developed. In addition, neutralizing antibody (NTAb) reference panels were analyzed and standards with various neutralizing titers were selected. These reference antigens were used to calibrate vaccine samples from several producers and found that five EV71 antigens and the national reference standards showed good linearity and parallelism. Moreover, mice immunized with various vaccines at doses standardized by these national references showed comparable NTAb responses. Finally, the national NTAb reference panels were found to effectively reduce assay discrepancy between different labs. Taken together, these national reference standards are highly valuable for the standardization and evaluation of EV71 vaccines. Copyright © 2011 Elsevier Ltd. All rights reserved.

  18. Interplay among viral antigens, cellular pathways and tumor microenvironment in the pathogenesis of EBV-driven lymphomas.

    Science.gov (United States)

    Dolcetti, Riccardo; Dal Col, Jessica; Martorelli, Debora; Carbone, Antonino; Klein, Eva

    2013-12-01

    Epstein-Barr virus (EBV) is a ubiquitous human γ-herpes virus that has established an elegant strategy to persist as a life-long asymptomatic infection in memory B lymphocytes. EBV has potent transforming properties for B lymphocytes and it is pathogenically associated with a variety of lymphomas of B or NK/T cell origin. The viral latency programs expressed can hijack or deregulate cellular pathways critical for cell proliferation and survival, while impairing anti-viral immune responses. Similar effects may also be induced by EBV-encoded micro-RNAs, which may have a pathogenic role particularly in lymphomas showing a restricted expression of viral proteins. Of note, recent data have challenged the view that only the EBV latency is relevant for lymphomagenesis, suggesting that lytic EBV replication may also contribute to the development of EBV-associated lymphoproliferations. The recent advances in the elucidation of the mechanisms underlying EBV-induced cell transformation and immune evasion are providing the rationale for innovative and tailored treatment approaches for EBV-driven lymphomas. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. Human polyoma JC virus minor capsid proteins, VP2 and VP3, enhance large T antigen binding to the origin of viral DNA replication: evidence for their involvement in regulation of the viral DNA replication.

    Science.gov (United States)

    Saribas, A Sami; Mun, Sarah; Johnson, Jaslyn; El-Hajmoussa, Mohammad; White, Martyn K; Safak, Mahmut

    2014-01-20

    JC virus (JCV) lytically infects the oligodendrocytes in the central nervous system in a subset of immunocompromized patients and causes the demyelinating disease, progressive multifocal leukoencephalopathy. JCV replicates and assembles into infectious virions in the nucleus. However, understanding the molecular mechanisms of its virion biogenesis remains elusive. In this report, we have attempted to shed more light on this process by investigating molecular interactions between large T antigen (LT-Ag), Hsp70 and minor capsid proteins, VP2/VP3. We demonstrated that Hsp70 interacts with VP2/VP3 and LT-Ag; and accumulates heavily in the nucleus of the infected cells. We also showed that VP2/VP3 associates with LT-Ag through their DNA binding domains resulting in enhancement in LT-Ag DNA binding to Ori and induction in viral DNA replication. Altogether, our results suggest that VP2/VP3 and Hsp70 actively participate in JCV DNA replication and may play critical roles in coupling of viral DNA replication to virion encapsidation. © 2013 Published by Elsevier Inc.

  20. INTEGRAL EVALUATION OF THE CYTOKINE SYSTEM IN VIRAL MYOCARDITIS

    Directory of Open Access Journals (Sweden)

    Peremot S. D.

    2017-10-01

    Full Text Available The need for an individual approach in the choice of means for the prevention of complications in inflammatory processes in cardiomyocytes, the course of which unfolds against a persistent viral infection, dictates the need to determine the general mechanisms for maintaining and progressing of the pathological process and an objective evaluation of immunological changes. The aim of the study was to determine changes in the system of inflammatory mediators in patients with subacute and chronic herpesviral infectious myocarditis on the basis of an integral assessment of the levels of opposing groups of cytokines. Materials & methods. To achieve this goal, we conducted a determination and analysis of changes in the cytokine profile in 87 patients with subacute (from 2 to 6 months and chronic (more than 6 months myocarditis due to an integral assessment of the mediator levels of inflammation of opposing groups in patients with herpesviral myocarditis on treatment in medical institutions of the Kharkov city. The average age of the patients was (27 ± 7.4 years. The control group was attracted to 40 people without clinical manifestations of cardiovascular diseases and in whose anamnesis there were no data on the transferred inflammatory diseases of the myocardium. Both groups of subjects were comparable in age and gender. The main group of subjects was divided into two subgroups. The first was 44 patients with subacute flow, the second - 43 patients with chronic infectious myocarditis. The diagnosis was established in accordance with the recommendations of the Association of Cardiologists of Ukraine and experts of the European Society of Cardiology, according to the formation of definitions of diseases in the International Classification of Diseases (ICD-10 of the tenth revision. The removal of material from patients was carried out according to the rules for the collection of infectious material. The concentration of cytokines: IL-2, IL-4, IL-6, IL

  1. Virus-induced asthma attack: The importance of allergic inflammation in response to viral antigen in an animal model of asthma.

    Science.gov (United States)

    Skappak, Christopher; Ilarraza, Ramses; Wu, Ying-Qi; Drake, Matthew G; Adamko, Darryl J

    2017-01-01

    Asthma exacerbation can be a life-threatening condition, and is most often triggered by common respiratory viruses. Poor asthma control and worsening of respiratory function is associated with increased airway inflammation, including eosinophilia. Prevention of asthma exacerbation relies on treatment with corticosteroids, which preferentially inhibit allergic inflammation like eosinophils. Human studies demonstrate that inactivated virus can trigger eosinophil activation in vitro through antigen presentation and memory CD4+ lymphocytes. We hypothesized that animals with immunologic memory to a respiratory virus would also develop airway hyperresponsiveness in response to a UV-inactivated form of the virus if they have pre-existing allergic airway inflammation. Guinea pigs were ovalbumin-sensitized, infected with live parainfluenza virus (PIV), aerosol-challenged with ovalbumin, and then re-inoculated 60 days later with live or UV-inactivated PIV. Some animals were either treated with dexamethasone prior to the second viral exposure. Lymphocytes were isolated from parabronchial lymph nodes to confirm immunologic memory to the virus. Airway reactivity was measured and inflammation was assessed using bronchoalveolar lavage and lung histology. The induction of viral immunologic memory was confirmed in infected animals. Allergen sensitized and challenged animals developed airway hyperreactivity with eosinophilic airway inflammation when re-exposed to UV-inactivated PIV, while non-sensitized animals did not. Airway hyperreactivity in the sensitized animals was inhibited by pre-treatment with dexamethasone. We suggest that the response of allergic inflammation to virus antigen is a significant factor causing asthma exacerbation. We propose that this is one mechanism explaining how corticosteroids prevent virus-induced asthma attack.

  2. Quantitative removal of hepatitis B viral antigens from serum by a monoclonal IgM coupled to a biocompatible solid-phase support.

    Science.gov (United States)

    Marciniak, R A; Wands, J R; Bruns, R R; Malchesky, P S; Nose, Y; Haber, E

    1983-06-01

    We covalently linked to regenerated cellulose filters a high-affinity monoclonal IgM produced against epitopes that reside on hepatitis B viral surface antigen (HBsAg). Conditions were established whereby as much as 250 micrograms of anti-HBsAg IgM could be linked to 2-4 mg of regenerated cellulose acetate by using cyanogen bromide and trichloro-s-triazine coupling agents. The immunoreactivity of the monoclonal anti-HBsAg IgM was preserved, and quantitative binding studies with HBsAg suggests that more than one functional binding site on the IgM molecule was operative. The specificity of the monoclonal anti-HBsAg IgM was established by demonstrating that a nonspecific monoclonal IgM (against influenza hemagglutinin), when coupled to the filters under identical conditions, had no effect on removal of HBsAg from serum. Most importantly, the monoclonal anti-HBsAg IgM-coupled filters quantitatively removed low levels of HBsAg from serum; after the third pass through the filter, HBsAg was undetectable in the perfusate. Further, the stability of the covalent bond between the anti-HBsAg IgM and regenerated cellulose acetate was shown by the lack of detectable murine monoclonal anti-HBsAg IgM in filtered serum despite 50 passages through the filter. Thus, we have demonstrated that monoclonal IgM antibodies with predefined specificity, when coupled to a biocompatible solid-phase support, may serve as a high-affinity and specific immunoabsorbant for quantitative removal and recovery of viral antigens from human serum. By using this approach, specific removal and recovery of many other substances from serum or plasma would seem possible.

  3. Evaluating the use of dedicated swab for rapid antigen detection ...

    African Journals Online (AJOL)

    Background: Group A streptococcus (GAS) is the most common and fearful bacterial cause in pediatric acute pharyngitis due to its serious complications. Several generations of rapid antigen detection tests (RADTs) have been developed to facilitate rapid detection of GAS pharyngitis. We assessed the value of using a ...

  4. Influence of hepatitis B virus genotypes on the intra- and extracellular expression of viral DNA and antigens.

    Science.gov (United States)

    Sugiyama, Masaya; Tanaka, Yasuhito; Kato, Takanobu; Orito, Etsuro; Ito, Kiyoaki; Acharya, Subrat K; Gish, Robert G; Kramvis, Anna; Shimada, Takashi; Izumi, Namiki; Kaito, Masahiko; Miyakawa, Yuzo; Mizokami, Masashi

    2006-10-01

    Various genotypes of the hepatitis B virus (HBV) induce liver disease of distinct severity, but the underlying virological differences are not well defined. Huh7 cells were transfected with plasmids carrying 1.24-fold the HBV genome of different genotypes/subgenotypes (2 strains each for Aa/A1, Ae/A2, Ba/B2 and D; 3 each for Bj/B1 and C). HBV DNA levels in cell lysates, determined by Southern hybridization, were the highest for C followed by Bj/Ba and D/Ae (P B e antigen (HBeAg) was excreted in a trend similar to that of HBV DNA with smaller differences. Secretion of hepatitis B surface antigen (HBsAg) was most abundant for Ae followed by Aa, Ba, Bj/C and remotely by D, which was consistent with mRNA levels. Cellular stress determined by the reporter assay for Grp78 promoter was higher for C and Ba than the other genotypes/subgenotypes (P type plasminogen activator (uPA/SCID), with the liver replaced for human hepatocytes, were inoculated with virions passed in mouse and recovered from culture supernatants. HBV DNA levels in their sera were higher for C than Ae by 2 logs during 4-7 weeks after inoculation. In conclusion, virological differences among HBV genotypes were demonstrated both in vitro and in vivo. These differences may influence HBV infections with distinct genotypes in clinical and epidemiological settings.

  5. The Oncogenic Small Tumor Antigen of Merkel Cell Polyomavirus Is an Iron-Sulfur Cluster Protein That Enhances Viral DNA Replication.

    Science.gov (United States)

    Tsang, Sabrina H; Wang, Ranran; Nakamaru-Ogiso, Eiko; Knight, Simon A B; Buck, Christopher B; You, Jianxin

    2015-11-25

    Merkel cell polyomavirus (MCPyV) plays an important role in Merkel cell carcinoma (MCC). MCPyV small T (sT) antigen has emerged as the key oncogenic driver in MCC carcinogenesis. It has also been shown to promote MCPyV LT-mediated replication by stabilizing LT. The importance of MCPyV sT led us to investigate sT functions and to identify potential ways to target this protein. We discovered that MCPyV sT purified from bacteria contains iron-sulfur (Fe/S) clusters. Electron paramagnetic resonance analysis showed that MCPyV sT coordinates a [2Fe-2S] and a [4Fe-4S] cluster. We also observed phenotypic conservation of Fe/S coordination in the sTs of other polyomaviruses. Since Fe/S clusters are critical cofactors in many nucleic acid processing enzymes involved in DNA unwinding and polymerization, our results suggested the hypothesis that MCPyV sT might be directly involved in viral replication. Indeed, we demonstrated that MCPyV sT enhances LT-mediated replication in a manner that is independent of its previously reported ability to stabilize LT. MCPyV sT translocates to nuclear foci containing actively replicating viral DNA, supporting a direct role for sT in promoting viral replication. Mutations of Fe/S cluster-coordinating cysteines in MCPyV sT abolish its ability to stimulate viral replication. Moreover, treatment with cidofovir, a potent antiviral agent, robustly inhibits the sT-mediated enhancement of MCPyV replication but has little effect on the basal viral replication driven by LT alone. This finding further indicates that MCPyV sT plays a direct role in stimulating viral DNA replication and introduces cidofovir as a possible drug for controlling MCPyV infection. MCPyV is associated with a highly aggressive form of skin cancer in humans. Epidemiological surveys for MCPyV seropositivity and sequencing analyses of healthy human skin suggest that MCPyV may represent a common component of the human skin microbial flora. However, much of the biology of the virus

  6. Bovine viral diarrhoea virus antigen in foetal calf serum batches and consequences of such contamination for vaccine production.

    Science.gov (United States)

    Makoschey, B; van Gelder, P T J A; Keijsers, V; Goovaerts, D

    2003-09-01

    A protocol to test foetal calf serum (FCS) for contamination with bovine viral diarrhoea virus (BVDV) is described. Following this protocol, which combines cell culture methods and detection of pestivirus RNA, seven batches of FCS were tested. Infectious BVDV was detected in four of those batches. One of the remaining batches contained a relatively high number of non-infectious BVDV particles. A sample of this batch was formulated with aluminium hydroxide and aluminium phosphate as adjuvant into an experimental vaccine preparation. This product was injected twice into BVDV seronegative cattle with a 4 week interval. Blood samples taken 4 weeks after the second application were negative for BVDV specific antibodies. Our data stress that detection of BVDV RNA is not sufficient for a complete risk assessment on FCS. Discrimination between infectious and non-infectious BVDV is essential. This can only be achieved by cell culture methods.

  7. Piecewise mixed-effects models with skew distributions for evaluating viral load changes: A Bayesian approach.

    Science.gov (United States)

    Huang, Yangxin; Dagne, Getachew A; Zhou, Shumin; Wang, Zhongjun

    2015-12-01

    Studies of human immunodeficiency virus dynamics in acquired immuno deficiency syndrome (AIDS) research are very important in evaluating the effectiveness of antiretroviral (ARV) therapies. The potency of ARV agents in AIDS clinical trials can be assessed on the basis of a viral response such as viral decay rate or viral load change in plasma. Following ARV treatment, the profile of each subject's viral load tends to follow a 'broken stick'-like dynamic trajectory, indicating multiple phases of decline and increase in viral loads. Such multiple-phases (change-points) can be described by a random change-point model with random subject-specific parameters. One usually assumes a normal distribution for model error. However, this assumption may be unrealistic, obscuring important features of within- and among-subject variations. In this article, we propose piecewise linear mixed-effects models with skew-elliptical distributions to describe the time trend of a response variable under a Bayesian framework. This methodology can be widely applied to real problems for longitudinal studies. A real data analysis, using viral load data from an AIDS study, is carried out to illustrate the proposed method by comparing various candidate models. Biologically important findings are reported, and these findings also suggest that it is very important to assume a model with skew distribution in order to achieve reliable results, in particular, when the data exhibit skewness. © The Author(s) 2011.

  8. Analysis of DNA-vaccinated fish reveals viral antigen in muscle, kidney, and thymus, and transient histopathologic changes

    Science.gov (United States)

    Garver, K.A.; Conway, C.M.; Elliott, D.G.; Kurath, G.

    2005-01-01

    A highly efficacious DNA vaccine against a fish rhabdovirus, infectious hematopoietic necrosis virus (IHNV), was used in a systematic study to analyze vaccine tissue distribution, persistence, expression patterns, and histopathologic effects. Vaccine plasmid pIHNw-G, containing the gene for the viral glycoprotein, was detected immediately after intramuscular injection in all tissues analyzed, including blood, but at later time points was found primarily in muscle tissue, where it persisted to 90 days. Glycoprotein expression was detected in muscle, kidney, and thymus tissues, with levels peaking at 14 days and becoming undetectable by 28 days. Histologic examination revealed no vaccine-specific pathologic changes at the standard effective dose of 0.1 ??g DNA per fish, but at a high dose of 50 ??g an increased inflammatory response was evident. Transient damage associated with needle injection was localized in muscle tissue, but by 90 days after vaccination no damage was detected in any tissue, indicating the vaccine to be safe and well tolerated. ?? Springer Science+Business Media, Inc. 2005.

  9. Impact of acute malaria on pre-existing antibodies to viral and vaccine antigens in mice and humans.

    Directory of Open Access Journals (Sweden)

    Simran Banga

    Full Text Available Vaccine-induced immunity depends on long-lived plasma cells (LLPCs that maintain antibody levels. A recent mouse study showed that Plasmodium chaubaudi infection reduced pre-existing influenza-specific antibodies--raising concerns that malaria may compromise pre-existing vaccine responses. We extended these findings to P. yoelii infection, observing decreases in antibodies to model antigens in inbred mice and to influenza in outbred mice, associated with LLPC depletion and increased susceptibility to influenza rechallenge. We investigated the implications of these findings in Malian children by measuring vaccine-specific IgG (tetanus, measles, hepatitis B before and after the malaria-free 6-month dry season, 10 days after the first malaria episode of the malaria season, and after the subsequent dry season. On average, vaccine-specific IgG did not decrease following acute malaria. However, in some children malaria was associated with an accelerated decline in vaccine-specific IgG, underscoring the need to further investigate the impact of malaria on pre-existing vaccine-specific antibodies.

  10. Targeting 4-1BB (CD137 to enhance CD8 T cell responses with poxviruses and viral antigens

    Directory of Open Access Journals (Sweden)

    Yuan eZhao

    2012-11-01

    Full Text Available Attenuated vaccinia virus (VACV vectors are considered prime vaccine candidates for use in immunotherapy of infectious disease. In spite of this, recent data show that the level of attenuation may hamper the efficient generation of protective CD8 T cells. This suggests that additional adjuvant-like activities may need to be combined with attenuated VACV for optimal vaccination. Stimulatory reagents to the TNFR family molecule 4-1BB (CD137 may represent such an adjuvant for vaccination. Previous murine studies have found that 4-1BB can participate in optimal priming of effector and memory CD8 T cells in response to several virus infections, and concordantly direct stimulation of 4-1BB with agonist reagents effectively boosts the CD8 T cell response against those viruses. In contrast, we recently reported that 4-1BB plays no role in the response to a virulent strain of VACV, questioning whether agonists of 4-1BB will be useful adjuvants for vaccination with VACV vectors. Here we show that agonist anti-4-1BB strongly enhanced the primary viral-specific effector CD8 T cell response during infection with live virulent VACV and attenuated VACV, and during immunization with VACV peptides given in IFA. However, accumulation of memory CD8 T cells was enhanced only following infection with virulent VACV or with peptide vaccination, but not with attenuated VACV, correlating in part with more transient expression of 4-1BB on CD8 T cells with attenuated virus. Our data therefore suggest that 4-1BB may be a promising candidate for targeting as an adjuvant for short-term enhancement of CD8 T cell responses with VACV vaccine strategies, but additional receptors may need to be engaged with 4-1BB to allow long-term CD8 T cell immunity with attenuated VACV vectors.

  11. Multicenter Evaluation of the Cepheid Xpert Hepatitis C Virus Viral Load Assay.

    Science.gov (United States)

    McHugh, M P; Wu, A H B; Chevaliez, S; Pawlotsky, J M; Hallin, M; Templeton, K E

    2017-05-01

    Viral load monitoring for hepatitis C virus (HCV) is necessary to diagnose infection and monitor response to therapy, but the tests involved are currently confined to specialist institutions. There is a need for a fast, accurate assay with limited operator input to enhance the access to viral load monitoring. We evaluated the quantification of HCV RNA in serum and plasma by the Cepheid Xpert HCV Viral Load assay in comparison to the Abbott RealTi m e HCV assay. Serum and plasma samples were gathered from HCV-infected individuals at four international sites. These were tested with the Xpert HCV Viral Load assay, and results were compared to quantification by the Abbott RealTi m e HCV assay. An external quality assessment panel of eight samples was also tested. In total, 614 samples were analyzed in the study, and the qualitative results agreed on the two platforms for 588 (95.8%) samples. Further analysis of 396 samples quantified by both tests showed strong correlation (correlation coefficient r = 0.99) across the quantifiable range, with Bland-Altman plot data showing a mean difference (±1.96 standard deviation) of 0.03 ± 0.44 log 10 IU/ml. In the external quality assessment panel, the Xpert HCV Viral Load assay results (quantified in log 10 IU per milliliter) were within 1 standard deviation of the target value for all but one sample, which was also similarly misquantified by the Abbott RealTime HCV assay. The Xpert HCV Viral Load assay performs well compared to a market-leading HCV viral load test and should be considered for instances where rapid near-to-patient testing is required. Copyright © 2017 American Society for Microbiology.

  12. [Evaluation of the protective performance of a positive pressure bio-protective clothing against viral aerosol].

    Science.gov (United States)

    Li, Na; Wen, Zhan-bo; Yang, Wen-hui; Wang, Jie; Li, Jin-song; Hu, Ling-fei; Dong, Xiao-kai; Liu, Ke-yang; Cao, Jie

    2012-01-01

    To evaluate the protective performance of a positive pressure bio-protective clothing against viral aerosol. The suspension of indicating virus phage Phi-X174 was made for viral aerosol generating in a hermetic cabin. The diameter of viral aerosol particles were measured with a aerodynamics size analyzer. By adjusting the inner humidity of the cabin, the protective efficiency of the positive pressure bio-protective clothing against viral aerosol in high and low windshield conditions was determined with Andersen six-stage air sampler sampling and plage forming unit (PFU) counting, respectively. The mass median diameter of Phage Phi-X174 aerosol particles was about 0.922 µm and the background concentration is beyond 2 × 10⁴ particles/m³. The protective efficiency of the clothing against phage Phi-X174 aerosol particles was above 99.9% under different test conditions with the range of viral aerosol concentration between 0 - 23 PFU/m³. Airflow (P = 0.84), environment humidity conditions (P = 0.33) and sampling time (P = 0.07) did not affect the protective efficiency statistically. The positive pressure bio-protective clothing provided a relatively high efficiency against phage Phi-X174 aerosol regardless of airflow rate, environment humidity and sampling time.

  13. Design evaluation and optimization for models of hepatitis C viral dynamics.

    Science.gov (United States)

    Guedj, Jeremie; Bazzoli, Caroline; Neumann, Avidan U; Mentré, France

    2011-05-10

    Mathematical modeling of hepatitis C viral (HCV) kinetics is widely used for understanding viral pathogenesis and predicting treatment outcome. The standard model is based on a system of five non-linear ordinary differential equations (ODE) that describe both viral kinetics and changes in drug concentration after treatment initiation. In such complex models parameter estimation is challenging and requires frequent sampling measurements on each individual. By borrowing information between study subjects, non-linear mixed effect models can deal with sparser sampling from each individual. However, the search for optimal designs in this context has been limited by the numerical difficulty of evaluating the Fisher information matrix (FIM). Using the software PFIM, we show that a linearization of the statistical model avoids most of the computational burden, while providing a good approximation to the FIM. We then compare the precision of the parameters that can be expected using five study designs from the literature. We illustrate the usefulness of rationalizing data sampling by showing that, for a given level of precision, optimal design could reduce the total number of measurements by up 50 per cent. Our approach can be used by a statistician or a clinician aiming at designing an HCV viral kinetics study. Copyright © 2011 John Wiley & Sons, Ltd.

  14. Viral Marketing Past Present Future

    OpenAIRE

    Nessipbekova, Zarina

    2010-01-01

    The work studies the viral marketing. These are past viral campaigns, viral campaigns today, and evaluates their actuality. The work tries to predict the development of viral marketing on the basis of the research done by the author.

  15. Evaluation of a new in-clinic method for the detection of canine heartworm antigen.

    Science.gov (United States)

    Lee, Alice C Y; Bowman, Dwight D; Lucio-Forster, Araceli; Beall, Melissa J; Liotta, Janice L; Dillon, Ray

    2011-05-11

    Canine heartworm is endemic in many parts of the world, and veterinarians rely on rapid in-clinic antigen tests to screen for this infection. Recently, an in-clinic, instrument-based rotor employing a colloidal gold agglutination immunoassay was launched in the marketplace (VetScan VS2(®) Canine Heartworm (HW) Antigen Test Kit; Abaxis, Inc.). Because of the widespread use of heartworm prevention and possible false negative test results in dogs with low heartworm burdens, the performance of the VetScan VS2(®) HW test and a commercially available in-clinic, membrane-based ELISA test (SNAP(®) Heartworm RT Test; IDEXX Laboratories) was compared using samples from dogs with low heartworm burdens and/or low levels of circulating antigen. Ninety serum samples were evaluated using the two methods. Testing was performed according to the manufacturer's product insert by personnel blinded to sample status. The samples were derived from two populations: dogs with necropsy-confirmed heartworm status (40 with 1-4 female worms, 30 with no worms), and field dogs (20) confirmed positive for antigen by microtiter plate ELISA (PetChek(®) Heartworm PF Antigen Test; IDEXX Laboratories). All 40 dogs with heartworms on necropsy were also confirmed to have circulating antigen by the PetChek HW ELISA. In necropsy-negative dogs (n=30), neither the VetScan VS2 HW nor SNAP HW tests detected heartworm antigen. Of the samples testing positive for antigen by PetChek HW (n=60), the VetScan VS2 HW and SNAP HW tests detected antigen in 15 and 56 samples, respectively. Percent agreement (plus 95% confidence interval) for each test relative to the PetChek HW qualitative result was 50% (40-60%) for VetScan VS2 HW and 96% (89-98%) for SNAP HW. Relative to the presence or absence of female worms at necropsy, agreement was 61% (50-72%) for VetScan VS2 HW and 99% (92-99.6%) for SNAP HW tests. It is clinically important that dogs with low heartworm burdens and/or low levels of circulating heartworm

  16. An evaluation of purified Salmonella Typhi protein antigens for the serological diagnosis of acute typhoid fever.

    Science.gov (United States)

    Tran Vu Thieu, Nga; Trinh Van, Tan; Tran Tuan, Anh; Klemm, Elizabeth J; Nguyen Ngoc Minh, Chau; Voong Vinh, Phat; Pham Thanh, Duy; Ho Ngoc Dan, Thanh; Pham Duc, Trung; Langat, Pinky; Martin, Laura B; Galan, Jorge; Liang, Li; Felgner, Philip L; Davies, D Huw; de Jong, Hanna K; Maude, Rapeephan R; Fukushima, Masako; Wijedoru, Lalith; Ghose, Aniruddha; Samad, Rasheda; Dondorp, Arjen M; Faiz, Abul; Darton, Thomas C; Pollard, Andrew J; Thwaites, Guy E; Dougan, Gordon; Parry, Christopher M; Baker, Stephen

    2017-08-01

    The diagnosis of typhoid fever is a challenge. Aiming to develop a typhoid diagnostic we measured antibody responses against Salmonella Typhi (S. Typhi) protein antigens and the Vi polysaccharide in a cohort of Bangladeshi febrile patients. IgM against 12 purified antigens and the Vi polysaccharide was measured by ELISA in plasma from patients with confirmed typhoid fever (n = 32), other confirmed infections (n = 17), and healthy controls (n = 40). ELISAs with the most specific antigens were performed on plasma from 243 patients with undiagnosed febrile disease. IgM against the S. Typhi protein antigens correlated with each other (rho > 0.8), but not against Vi (rho 0.85, respectively. Applying a dynamic cut-off to patients with undiagnosed febrile disease suggested that 34-58% had an IgM response indicative of typhoid. We evaluated the diagnostic potential of several S. Typhi antigens; our assays give good sensitivity and specificity, but require further assessment in differing patient populations. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  17. Mechanisms of Antigen Adsorption Onto an Aluminum-Hydroxide Adjuvant Evaluated by High-Throughput Screening.

    Science.gov (United States)

    Jully, Vanessa; Mathot, Frédéric; Moniotte, Nicolas; Préat, Véronique; Lemoine, Dominique

    2016-06-01

    The adsorption mechanism of antigen on aluminum adjuvant can affect antigen elution at the injection site and hence the immune response. Our aim was to evaluate adsorption onto aluminum hydroxide (AH) by ligand exchange and electrostatic interactions of model proteins and antigens, bovine serum albumin (BSA), β-casein, ovalbumin (OVA), hepatitis B surface antigen, and tetanus toxin (TT). A high-throughput screening platform was developed to measure adsorption isotherms in the presence of electrolytes and ligand exchange by a fluorescence-spectroscopy method that detects the catalysis of 6,8-difluoro-4-methylumbelliferyl phosphate by free hydroxyl groups on AH. BSA adsorption depended on predominant electrostatic interactions. Ligand exchange contributes to the adsorption of β-casein, OVA, hepatitis B surface antigen, and TT onto AH. Based on relative surface phosphophilicity and adsorption isotherms in the presence of phosphate and fluoride, the capacities of the proteins to interact with AH by ligand exchange followed the trend: OVA aluminum-containing adjuvant. Copyright © 2016 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

  18. Viral and bacterial contamination in a sedimentary aquifer in Uruguay: evaluation of coliforms as regional indicators of viral contamination.

    Science.gov (United States)

    Gamazo, Pablo; Colina, Rodney; Victoria, Matias; Alvareda, Elena; Burutatran, Luciana; Ramos, Julian; Olivera, María; Soler, Joan

    2015-04-01

    In many areas of Uruguay groundwater is the only source of water for human consumption and for industrial-agricultural economic activities. Traditionally considered as a safe source, groundwater is commonly used without any treatment. The Uruguayan law requires bacteriological (fecal) analysis for most water uses, but virological analyses are not mentioned in the legislation. In the Salto district, where groundwater is used for human consumption and for agricultural activities, bacterial contamination has been detected in several wells but no viruses analysis have been performed. The Republic University (UDELAR), with the support of the National Agency for Research and Innovation (ANII), is studying the incidence of virus and fecal bacteria in groundwater on an intensive agriculture area of the Salto district. An initial screening campaign of 44 wells was performed in which, besides total and fecal coliforms, rotavirus and adenovirus were detected. A subgroup of the screening wells (15) where selected for bimonthly sampling during a year. In accordance with literature results, single well data analysis shows that coliform and viral contamination can be considered as independent variables. However, when spatial data is integrated, coliform and viral contamination show linear correlation. In this work we present the survey results, we analyse the temporal incidence of variables like precipitation, temperature and chemical composition in well contamination and we discuss the value of coliforms as global indicator of viral contamination for the Salto aquifer.

  19. Evaluation and molecular characterization of human adenovirus in drinking water supplies: viral integrity and viability assays.

    Science.gov (United States)

    Fongaro, Gislaine; Nascimento, Mariana A do; Rigotto, Caroline; Ritterbusch, Giseli; da Silva, Alessandra D' A; Esteves, Paulo A; Barardi, Célia R M

    2013-05-28

    Human adenoviruses (HAdVs) are the second-leading cause of childhood gastroenteritis worldwide. This virus is commonly found in environmental waters and is very resistant to water disinfection and environmental stressors, especially UV light inactivation. Molecular techniques, such as PCR-based methods (Polymerase Chain Reaction), are commonly used to detect and identify viral contamination in water, although PCR alone does not allow the discrimination between infectious and non-infectious viral particles. A combination of cell culture and PCR has allowed detection of infectious viruses that grow slowly or fail to produce cytopathic effects (CPE) in cell culture. This study aimed to assess the integrity and viability of human adenovirus (HAdV) in environmental water and evaluate circulating strains by molecular characterization in three sites of the water supply in Florianópolis, Santa Catarina Island, Brazil: Peri Lagoon water, spring source water, and water from the public water supply system. Water samples were collected, concentrated and HAdV quantified by real-time PCR. Viral integrity was evaluated by enzymatic assay (DNase I) and infectivity by plaque assay (PA) and integrated cell culture using transcribed mRNA (ICC-RT-qPCR). Samples containing particles of infectious HAdV were selected for sequencing and molecular characterization. The analyzed sites contained 83, 66 and 58% undamaged HAdV particles (defined as those in which the genetic material is protected by the viral capsid) at Peri Lagoon, spring source water and public supply system water, respectively. Of these, 66% of the particles (by PA) and 75% (by ICC-RT-qPCR) HAdV were shown to be infectious, due to being undamaged in Peri Lagoon, 33% (by PA) and 58% (by ICC-RT-qPCR) in spring source water and 8% (by PA) and 25% (by ICC-RT-qPCR) in the public water supply system. ICC-RT-qPCR, a very sensitive and rapid technique, was able to detect as low as 1 × 102 HAdV genome copies per milliliter of

  20. In vitro production of Trypanosoma equiperdum antigen and its evaluation for use in serodiagnosis of dourine.

    Science.gov (United States)

    Bassarak, Björn; Moser, Irmgard; Menge, Christian

    2016-06-15

    A modified Baltz's in vitro cultivation system for the propagation of Trypanosoma equiperdum strain OVI was established to develop a replacement for the conventional production procedure of dourine diagnostic antigen in rats. To increase trypanosome yields we designed an optimized culture medium by addition of supplemental compounds. Trypanosomes were adapted to this medium by two succeeding cultivation steps which led to a substantial proliferation rate and an increased cell density tolerance, respectively. As a result, adapted parasites could be propagated to maximum cell densities of >2×10(6) cells/ml, facilitating in vitro antigen production in preparative quantities comparable to the conventional method. A panel of 180 horse field sera, previously sent for testing to the German National Reference Laboratory for Dourine, was tested by complement fixation test using culture-derived as well as conventionally produced dourine antigen. Cohen's kappa values for results obtained with two batches of culture-derived antigen as compared to conventional antigen were 0.91 (95% confidence interval [CI]: 82.2-99.7) and 0.83 (95% CI: 70.3-95.3), respectively. Performance of antigens for diagnostic purposes was characterized in an inter-laboratory comparative study deploying 14 sera from horses with defined dourine statuses. Complement fixation test results from 15 participating European laboratories showed a diagnostic sensitivity of 94.1% (95% CI: 89.4-98.7) and a diagnostic specificity of 96.2% (95% CI: 92.5-99.9) for conventional antigen and a slightly higher diagnostic sensitivity of 96.0% (95% CI: 92.2-99.8) and a diagnostic specificity of 97.1% (95% CI: 94.0-100) for culture-derived antigen. We conclude that our novel approach for dourine antigen production from in vitro-grown trypanosomes described and evaluated herein meets the requirements for the prospective purpose in quantitative and qualitative terms and should be considered by the competent authorities as an

  1. In vitro evaluation of marine-microorganism extracts for anti-viral activity

    Directory of Open Access Journals (Sweden)

    Yasuhara-Bell Jarred

    2010-08-01

    Full Text Available Abstract Viral-induced infectious diseases represent a major health threat and their control remains an unachieved goal, due in part to the limited availability of effective anti-viral drugs and measures. The use of natural products in drug manufacturing is an ancient and well-established practice. Marine organisms are known producers of pharmacological and anti-viral agents. In this study, a total of 20 extracts from marine microorganisms were evaluated for their antiviral activity. These extracts were tested against two mammalian viruses, herpes simplex virus (HSV-1 and vesicular stomatitis virus (VSV, using Vero cells as the cell culture system, and two marine virus counterparts, channel catfish virus (CCV and snakehead rhabdovirus (SHRV, in their respective cell cultures (CCO and EPC. Evaluation of these extracts demonstrated that some possess antiviral potential. In sum, extracts 162M(4, 258M(1, 298M(4, 313(2, 331M(2, 367M(1 and 397(1 appear to be effective broad-spectrum antivirals with potential uses as prophylactic agents to prevent infection, as evident by their highly inhibitive effects against both virus types. Extract 313(2 shows the most potential in that it showed significantly high inhibition across all tested viruses. The samples tested in this study were crude extracts; therefore the development of antiviral application of the few potential extracts is dependent on future studies focused on the isolation of the active elements contained in these extracts.

  2. Technical evaluation of the Beckman Coulter OV-Monitor (CA 125 antigen) immunoassay.

    Science.gov (United States)

    Yagmur, Eray; Driesch, Reinhard; Gressner, Axel M; Kiefer, Paul

    2006-01-01

    The CA 125 antigen is a large (200-1000 kDa) glycoprotein, present within normal and benign ovarian tissue. We evaluated the analytical performance of the newly available Beckman Coulter OV-Monitor (CA 125 antigen) immunoassay on the Beckman Coulter UniCel DxI 800 analyzer. The evaluation was performed according to NCCLS recommendations. The lowest level of CA 125 antigen detectable was 0.374 U/mL. Serial dilution of two pooled CA 125 antigen-rich samples provided a linear response (pUniCel DxI 800 showed linear regression values of r=0.961 vs. the Bayer ADVIA Centaur system and r=0.981 vs. the Abbott AxSYM system. Considering the limited number of serum samples analyzed, our data indicate that the Beckman Coulter OV-Monitor immunoassay has excellent analytical performance and shows satisfactory correlation with automated immunoassays on the Abbott AxSYM and Bayer ADVIA Centaur systems. It is easy to perform, accurate and suitable for measurements in routine clinical laboratories.

  3. Evaluation of a Recombinant Trypanosoma cruzi Mucin-Like Antigen for Serodiagnosis of Chagas' Disease ▿

    Science.gov (United States)

    De Marchi, Claudia R.; Di Noia, Javier M.; Frasch, Alberto C. C.; Amato Neto, Vicente; Almeida, Igor C.; Buscaglia, Carlos A.

    2011-01-01

    Chagas' disease is caused by the protozoan parasite Trypanosoma cruzi and is one of the most important endemic problems in Latin America. Lately, it has also become a health concern in the United States and Europe. Currently, a diagnosis of Chagas' disease and the screening of blood supplies for antiparasite antibodies are achieved by conventional serological tests that show substantial variation in the reproducibility and reliability of their results. In addition, the specificity of these assays is curtailed by antigenic cross-reactivity with sera from patients affected by other endemic diseases, such as leishmaniasis. Here we used a highly sensitive chemiluminescent enzyme-linked immunosorbent assay (CL-ELISA) to evaluate a recombinant protein core of a mucin-like molecule (termed trypomastigote small surface antigen [TSSA]) for the detection of specific serum antibodies in a broad panel of human sera. The same samples were evaluated by CL-ELISA using as the antigen either a mixture of native T. cruzi trypomastigote mucins or an epimastigote extract and, for further comparison, by conventional serologic tests, such as an indirect hemagglutination assay and indirect immunofluorescence assay. TSSA showed ∼87% sensitivity among the seropositive Chagasic panel, a value which was increased up to >98% when only parasitologically positive samples were considered. More importantly, TSSA showed a significant increase in specificity (97.4%) compared to those of currently used assays, which averaged 80 to 90%. Overall, our data demonstrate that recombinant TSSA may be a useful antigen for the immunodiagnosis of Chagas' disease. PMID:21880857

  4. Evaluation of a recombinant Trypanosoma cruzi mucin-like antigen for serodiagnosis of Chagas' disease.

    Science.gov (United States)

    De Marchi, Claudia R; Di Noia, Javier M; Frasch, Alberto C C; Amato Neto, Vicente; Almeida, Igor C; Buscaglia, Carlos A

    2011-11-01

    Chagas' disease is caused by the protozoan parasite Trypanosoma cruzi and is one of the most important endemic problems in Latin America. Lately, it has also become a health concern in the United States and Europe. Currently, a diagnosis of Chagas' disease and the screening of blood supplies for antiparasite antibodies are achieved by conventional serological tests that show substantial variation in the reproducibility and reliability of their results. In addition, the specificity of these assays is curtailed by antigenic cross-reactivity with sera from patients affected by other endemic diseases, such as leishmaniasis. Here we used a highly sensitive chemiluminescent enzyme-linked immunosorbent assay (CL-ELISA) to evaluate a recombinant protein core of a mucin-like molecule (termed trypomastigote small surface antigen [TSSA]) for the detection of specific serum antibodies in a broad panel of human sera. The same samples were evaluated by CL-ELISA using as the antigen either a mixture of native T. cruzi trypomastigote mucins or an epimastigote extract and, for further comparison, by conventional serologic tests, such as an indirect hemagglutination assay and indirect immunofluorescence assay. TSSA showed ∼87% sensitivity among the seropositive Chagasic panel, a value which was increased up to >98% when only parasitologically positive samples were considered. More importantly, TSSA showed a significant increase in specificity (97.4%) compared to those of currently used assays, which averaged 80 to 90%. Overall, our data demonstrate that recombinant TSSA may be a useful antigen for the immunodiagnosis of Chagas' disease.

  5. Evaluation of hepatitis B virus transmission and antiviral therapy among hepatitis B surface antigen-positive pregnant women.

    Science.gov (United States)

    Tekin Koruk, Suda; Batirel, Ayse; Kose, Sukran; Cetin Akhan, Sila; Aygen, Bilgehan; Tulek, Necla; Hatipoglu, Çigdem; Bulut, Cemal; Yıldız, Orhan; Sacligil, Cahide; Sirmatel, Fatma; Altunok, Elif

    2015-12-01

    The aim of the present study was to assess the potential risk of hepatitis B virus (HBV) vertical transmission among Turkish parturient women and to evaluate the efficacy and safety of antiviral agents. Data were collected retrospectively from 114 HBV-infected pregnant women and their infants in eight health institutions in Turkey. The baseline characteristics of the women were: mean age, 28.3 ± 5.2 years; alanine aminotransferase, 57.4 ± 139.0 U/L; aspartate aminotransferase, 56.6 ± 150.0 U/L; and HBV DNA, 8.3 × 10(7) ± 2.6 × 10(8) copies/mL. Family history of HBV infection was detected in 53.5% (n = 61). In total, 60 (52.6%) pregnant women received tenofovir (60.0%), lamivudine (33.3%) or telbivudine (6.7%) therapy at the median gestational age of 22.2 ± 8.5 (1-36) weeks. All infants were vaccinated and hepatitis B immune globulin was administered, with 81 of them (71.1%) available for follow-up. After completion of HBV vaccination course, 71 (87.7%) infants had protective anti-HBs levels, three (3.7%) were hepatitis B surface antigen-positive, and seven (8.6%) were hepatitis B surface antigen-negative with nonprotective anti-HBs levels. Five of the infants had low gestational birthweight but no other birth defects were observed. According to our results, viral load may not be the only effecting factor for transmission of HBV to children of infected mothers. Pregnant women with high viral load should be followed-up closely during pregnancy. They should begin to take tenofovir or telbivudine, which are category B drugs for pregnancy, at the beginning of the third trimester at the latest. We need new treatment strategies; and close follow-up of mothers and children is another important issue. © 2015 Japan Society of Obstetrics and Gynecology.

  6. Evaluation of host and viral factors associated with severe dengue based on the 2009 WHO classification.

    Science.gov (United States)

    Pozo-Aguilar, Jorge O; Monroy-Martínez, Verónica; Díaz, Daniel; Barrios-Palacios, Jacqueline; Ramos, Celso; Ulloa-García, Armando; García-Pillado, Janet; Ruiz-Ordaz, Blanca H

    2014-12-11

    Dengue fever (DF) is the most prevalent arthropod-borne viral disease affecting humans. The World Health Organization (WHO) proposed a revised classification in 2009 to enable the more effective identification of cases of severe dengue (SD). This was designed primarily as a clinical tool, but it also enables cases of SD to be differentiated into three specific subcategories (severe vascular leakage, severe bleeding, and severe organ dysfunction). However, no study has addressed whether this classification has advantage in estimating factors associated with the progression of disease severity or dengue pathogenesis. We evaluate in a dengue outbreak associated risk factors that could contribute to the development of SD according to the 2009 WHO classification. A prospective cross-sectional study was performed during an epidemic of dengue in 2009 in Chiapas, Mexico. Data were analyzed for host and viral factors associated with dengue cases, using the 1997 and 2009 WHO classifications. The cost-benefit ratio (CBR) was also estimated. The sensitivity in the 1997 WHO classification for determining SD was 75%, and the specificity was 97.7%. For the 2009 scheme, these were 100% and 81.1%, respectively. The 2009 classification showed a higher benefit (537%) with a lower cost (10.2%) than the 1997 WHO scheme. A secondary antibody response was strongly associated with SD. Early viral load was higher in cases of SD than in those with DF. Logistic regression analysis identified predictive SD factors (secondary infection, disease phase, viral load) within the 2009 classification. However, within the 1997 scheme it was not possible to differentiate risk factors between DF and dengue hemorrhagic fever or dengue shock syndrome. The critical clinical stage for determining SD progression was the transition from fever to defervescence in which plasma leakage can occur. The clinical phenotype of SD is influenced by the host (secondary response) and viral factors (viral load). The 2009

  7. Strategic evaluation of vaccine candidate antigens for the prevention of Visceral Leishmaniasis.

    Science.gov (United States)

    Duthie, Malcolm S; Favila, Michelle; Hofmeyer, Kimberley A; Tutterrow, Yeung L; Reed, Steven J; Laurance, John D; Picone, Alessandro; Guderian, Jeffrey; Bailor, H Remy; Vallur, Aarthy C; Liang, Hong; Mohamath, Raodoh; Vergara, Julie; Howard, Randall F; Coler, Rhea N; Reed, Steven G

    2016-05-27

    Infection with Leishmania parasites results in a range of clinical manifestations and outcomes, the most severe of which is visceral leishmaniasis (VL). Vaccination will likely provide the most effective long-term control strategy, as the large number of vectors and potential infectious reservoirs renders sustained interruption of Leishmania parasite transmission extremely difficult. Selection of the best vaccine is complicated because, although several vaccine antigen candidates have been proposed, they have emerged following production in different platforms. To consolidate the information that has been generated into a single vaccine platform, we expressed seven candidates as recombinant proteins in E. coli. After verifying that each recombinant protein could be recognized by VL patients, we evaluated their protective efficacy against experimental L. donovani infection of mice. Administration in formulation with the Th1-potentiating adjuvant GLA-SE indicated that each antigen could elicit antigen-specific Th1 responses that were protective. Considering the ability to reduce parasite burden along with additional factors such as sequence identity across Leishmania species, we then generated a chimeric fusion protein comprising a combination of the 8E, p21 and SMT proteins. This E. coli -expressed fusion protein was also demonstrated to protect against L. donovani infection. These data indicate a novel recombinant vaccine antigen with the potential for use in VL control programs. Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.

  8. Antigenic and immunogenic evaluation of Helicobacter pylori FlaA epitopes

    Directory of Open Access Journals (Sweden)

    Mansoor Zarei

    2017-08-01

    Full Text Available Objective(s: Helicobacter pyloriare among most common human pathogens affecting at least half of the world’s population. Mobility is one of the important primary factors in bacterial colonization and invasion. The purpose of this research is cloning, expression, and purification of FlaA protein specific epitopes in order to evaluate their antigenicity and immunogenicity. Materials and Methods: The antigenic region of the flaA gene was bioinformatically predicted using Epitope mapping software’s and the predicted epitopes were expressed in a prokaryotic expression vector. The antigen was injected into the animal model (mice BALB/c and some indicators including IgG1, IgG2a, IgA, IFN-γ, and IL 5 were measured. Results: The immunogenicity studies in animal models by measuring serum antibodies (IgG1, IgG2a, and IgA and cytokines (IFN-γ and IL5 revealed that the rFlaA induces a proper immune response in animal models. Conclusion: The recombinant FlaA protein is antigenic and immunogenic. Therefore, it might be used in order to design of specific diagnostic kits and recombinant vaccines against H. pylori.

  9. Evaluation the Surface Antigen of the Salmonella typhimurium ATCC 14028 Ghosts Prepared by “SLRP”

    Directory of Open Access Journals (Sweden)

    Amara A. Amro

    2014-01-01

    Full Text Available Recently, bacterial ghosts (BGs were prepared using a protocol based on critical chemical concentrations. It has been given the name “sponge like” (SL protocol and used in its reduced form “sponge like reduced protocol” (SLRP. While specific antibody for Salmonella is available on the market under the commercial names (of some kits such as Febrile Antigen Kit (N.S. BIO-TEC, we used the described Kit to investigate the validity of the SLRP. In this study, using SLRP we succeeded to prepare STGs with correct surface antigens could interact with their specific antibodies. Additionally the study has included oral vaccination with STGs with challenge test. The rats serums have been evaluated against both of the O and H antigens. The antigen-antibody interaction (agglutination results of both the SLRP and the animal experiments prove that we have correct STGs able to immunize the rats against viable Salmonella. STGs could be used as vaccine and as adjuvant and in the antibodies and in the diagnostic kits production. This study is an additional step for the establishment of correct BGs for immunological purposes.

  10. Phenotype and functional evaluation of ex vivo generated antigen-specific immune effector cells with potential for therapeutic applications

    Science.gov (United States)

    Han, Shuhong; Huang, Yuju; Liang, Yin; Ho, Yuchin; Wang, Yichen; Chang, Lung-Ji

    2009-01-01

    Ex vivo activation and expansion of lymphocytes for adoptive cell therapy has demonstrated great success. To improve safety and therapeutic efficacy, increased antigen specificity and reduced non-specific response of the ex vivo generated immune cells are necessary. Here, using a complete protein-spanning pool of pentadecapeptides of the latent membrane protein 2A (LMP2A) of Epstein-Barr virus (EBV), a weak viral antigen which is associated with EBV lymphoproliferative diseases, we investigated the phenotype and function of immune effector cells generated based on IFN-γ or CD137 activation marker selection and dendritic cell (DC) activation. These ex vivo prepared immune cells exhibited a donor- and antigen-dependent T cell response; the IFN-γ-selected immune cells displayed a donor-related CD4- or CD8-dominant T cell phenotype; however, the CD137-enriched cells showed an increased ratio of CD4 T cells. Importantly, the pentadecapeptide antigens accessed both class II and class I MHC antigen processing machineries and effectively activated EBV-specific CD4 and CD8 T cells. Phenotype and kinetic analyses revealed that the IFN-γ and the CD137 selections enriched more central memory T (Tcm) cells than did the DC-activation approach, and after expansion, the IFN-γ-selected effector cells showed the highest level of antigen-specificity and effector activities. While all three approaches generated immune cells with comparable antigen-specific activities, the IFN-γ selection followed by ex vivo expansion produced high quality and quantity of antigen-specific effector cells. Our studies presented the optimal approach for generating therapeutic immune cells with potential for emergency and routine clinical applications. PMID:19660111

  11. Evaluating Recombinant Antigen ROP1 Efficacy in Diagnosis of Toxoplasma Gondii Infection

    Directory of Open Access Journals (Sweden)

    F Keshavarzi

    2015-07-01

    Full Text Available Introduction:Toxoplasma gondii is a ubiquitous obligate intracellular parasite with a relatively broad host range infecting both mammals and birds. Toxoplasma proteins are strong antigens that can begin strong immune reactions, among which Rhoptry protein 1 (ROP1 can be named discharging from rhoptry cell-organ. ROP1 is regarded as a competitor for recombinant vaccines against toxoplasmosis. Therefore, the main objective of the current study was to evaluate the cloning and expression of ROP1 Toxoplasma gondii in a cloning vector as well as to create this recombinant antigen in order to be applied for later uses. Methods:Genomic DNA of Toxoplasma gondii was removed and reproduced by PCR, then the PCR product was cloned into the EcoR1 and BamH1 sites of cloning vector, pUET1, and transformed into Escherichia coli BL21 plysS strain. Moreover, pcROP1 was sub-cloned into the HindIII and EcoRI sites of the pcDNA3 in order to produce recombining eukaryotic declaration vector. The cloned ROP1 was verified by PCR, limitation enzymes (HindIII and BglΙ digestion and nucleotide sequencing. Then, this recombinant antigen was covered applying IgM and ELISAIgG. Results:The study results demonstrated that a fragment of 757 bp was separated. In addition, nucleotide sequence analysis of the ROP1 cloned in pUET1vector revealed high homology (96% with RH strain Gene Bank Accession (No. M71274. Conclusion:The recombinant ROP1 antigen in an IgM Rec-ELISA test can be replaced with the tachyzoite antigen in IgG and IgM serologic tests.

  12. [Evaluation of four antigens for the detection of anti-Mycobacterium bovis antibodies by enzyme immunoassay].

    Science.gov (United States)

    Ritacco, V; Lopez, B; Barrera, L; Torrea, G; Nader, A; de Kantor, I N; Fliess, E

    1988-01-01

    An enzyme-linked immunosorbent assay (ELISA) for the diagnosis of bovine tuberculosis through the detection of specific seric antibodies has recently been developed in our laboratory. In order to assess its reproducibility and select the most adequate antigen, four bovine PPDs from different sources were evaluated in parallel: PPD M. bovis strain AN5, CEPANZO standard (CPZ), PPD M. bovis strain AN5, European Economic Community standard (EEC), PPD M. bovis strain AN5, prepared from non heated bacilli, killed by phenol (P) and PPD. M. bovis BCG strain prepared at the Pasteur Institute, Paris (BCG). Sera from 22 healthy cattle from tuberculosis free area and 20 bacteriologically confirmed tuberculous animals were employed in simultaneous assays. Antibody mean and standard deviations from healthy cattle expressed as optical density (OD) values were 45 +/- 22 when CPZ was used as antigen, 24 +/- 10 with EEC, 103 +/- 56 with P and 56 +/- 20 with BCG. Mean O.D. from tuberculous cattle were 588 +/- 158, 510 +/- 234, 782 +/- 138 and 441 +/- 189 with antigens CPZ, EEC, P and BCG respectively. A close correlation was observed when results obtained with EEC and P were compared with that of CPZ (r: 0.97 and 0.94 respectively). A lower specificity was achieved when BCG was used as antigen being also lower its correlation with the results obtained with CPZ (r: 0.87). It is concluded that our ELISA would achieve similar sensitivity and specificity if CPZ, EEC and P were used as antigens. On the other hand, BCG would not be suitable for this assay.

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

    Science.gov (United States)

    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.

  14. Evaluate the efficiency of Antigen 60 (A60 protein from BCG strain of Mycobacterium bovis as a diagnostic antigen

    Directory of Open Access Journals (Sweden)

    Nafiseh Shakibamehr

    2016-01-01

    Conclusion: Results of reactions of the injected A60 and standard human tuberculin shows the effectiveness of this antigen in comparison with standard human tuberculin. Detection of antibody in the serum of patients is a rapid and repeatable method. A60 with 89% sensitivity and 94% specificity could be an appropriate matter for the diagnosis of tuberculosis. Because this method can be performed without radioactive materials or advanced and expensive equipment, it will provide results quickly.

  15. Multicenter evaluation of a new rapid automated human immunodeficiency virus antigen detection assay.

    Science.gov (United States)

    Weber, B; Mühlbacher, A; Michl, U; Paggi, G; Bossi, V; Sargento, C; Camacho, R; Fall, E H; Berger, A; Schmitt, U; Melchior, W

    1999-03-01

    Although human immunodeficiency virus (HIV) antigen assays are of limited value for monitoring antiretroviral therapy, they play an important role for confirmatory testing of fourth generation HIV screening enzyme immunoassay (EIA) reactive samples. In a multicenter study, a new automated rapid p24 antigen assay, Elecsys HIV Ag (Roche Diagnostics Boehringer Mannheim GmbH, Penzberg, Germany), was compared to FDA licensed tests (Abbott HIV-1 Ag monoclonal and Coulter HIV-1 p24 antigen assay). In the evaluation 27 seroconversion panels were included, sera from the acute phase of infection, single and follow-up samples from HIV antibody positive patients, dilution series of HIV antigen positive standards, sera and cell culture supernatants infected with different HIV-1 subtypes (A-H, and O) HIV-2 and recombinant HIV-1 (gag/env) isolates. To challenge the specificity of the new assay, 2565 unselected blood donors, sera from pregnant women, dialysis and hospitalized patients and 407 potentially cross-reactive samples were investigated. Acute HIV infection was detected in three to eight seroconversion panels earlier with Elecsys HIV Ag than with the alternative assays. Higher numbers of serum samples from HIV infected patients tested positive by Elecsys HIV Ag than with the comparative assays. All HIV-1 subtypes and HIV-2 isolates were recognized with Elecsys HIV Ag. Abbott HIV-1 Ag monoclonal and Coulter HIV-1 p24 antigen assay showed a variable sensitivity for the different HIV-1 subtypes. The specificity of Elecsys HIV Ag and Coulter HIV-1 p24 antigen assay were 99.8 and 99.93%, respectively. All the eight sera that were false reactive by Elecsys HIV Ag were tested negative with the Elecsys HIV Ag Neutralization Test. In conclusion, Elecsys HIV Ag was more sensitive than the alternative assays and showed a high specificity in combination with the neutralization assay. The very short incubation time of 18 min and the fully automated procedure of Elecsys HIV Ag which

  16. Evaluating of Clinical and Laboratory Findings of Patients with Acute Viral Hepatitis A

    Directory of Open Access Journals (Sweden)

    Mustafa Taşkesen

    2008-09-01

    Full Text Available Acute viral hepatitis A is usually, which asymptomatic and anicteric, has often been seen in children. In this study, the files of 42 patients with acute viral hepatitis A infection who were admitted at the Department of Pediatrics, Medical Faculty, Dicle University, between January 2006 and February 2007, were evaluated retrospectively. Male/female rate was determined to be 1.4, and the mean ages of patients were 6.9±3.5 years (18months to14 years. The most common symptoms on admission were jaundice (73.8%, vomiting (52.3%, fever (42.8% and abdominal pain (38%. The seasons on admission were winter (47.6%, autumn (38.1% and spring (14.3%. In laboratory findings mean of aspartate aminotransferase (AST 1875U/L (82-6340U/L, alanine aminotransferase (ALT 1697U/L (144-4876 U/L, total bilirubin 12.1mg/dl (0.9-47mg/dl, and direct bilirubin 8.9mg/dl (0.4-27.3 mg/dl were determined. The most frequent complications were anemia, hepatic encephalopathy, thrombocytopenia, and the mortality rate was determined as 2.4%.

  17. A neonatal mouse model of coxsackievirus A10 infection for anti-viral evaluation.

    Science.gov (United States)

    Li, Shuxuan; Zhao, Huan; Yang, Lisheng; Hou, Wangheng; Xu, Longfa; Wu, Yangtao; Wang, Wei; Chen, Chunye; Wan, Junkai; Ye, Xiangzhong; Liang, Zhenglun; Mao, Qunying; Cheng, Tong; Xia, Ningshao

    2017-08-01

    Epidemiological data indicate that coxsackievirus A10 (CVA10) has become one of the main causative agents of hand, foot and mouth disease (HFMD) and in recent years has often been found to co-circulate with other enteroviruses, which poses a challenge for the prevention and control of HFMD. Although most CVA10-associated HFMD cases present mild symptoms, severe manifestations and even death can also occur. However, the study of the pathogenesis and the development of drugs and vaccines for CVA10 infection are still far from complete. In this study, we established a neonatal mouse model for anti-viral evaluation and characterized the pathology of CVA10 infection. To develop the mouse model, both inbred and outbred mouse strains were used to compare their sensitivity to CVA10 infection; then, one-day-old BALB/c mice were selected and inoculated intraperitoneally with a CVA10 clinical strain, CVA10-FJ-01. Clinical symptoms, such as wasting, hind-limb paralysis and even death were observed in the CVA10-infected mice. Moreover, pathological examination and immunohistochemistry staining showed that severe myonecrosis with inflammatory infiltration was observed in CVA10-infected mice, indicating that CVA10 exhibited strong tropism to muscle tissue. Using real-time PCR, we also found that the viral load in the blood and muscle was higher than that in other organs/tissues at different time points post-infection, suggesting that CVA10 had a strong tropism to mice muscle and that viremic spread may also contribute to the death of the CVA10-infected mice. Additionally, to evaluate the neonatal mouse model of CVA10 infection, female mice were immunized with formalin-inactivated CVA10 and then allowed to mate after the third immunization. The results showed that maternal antibodies could protect mice against CVA10 infection. In summary, the results demonstrated that the neonatal mice model was a useful tool for evaluating the protective effects of CVA10 vaccines and anti-viral

  18. Valuable Virality

    NARCIS (Netherlands)

    Akpinar, E.; Berger, Jonah

    2017-01-01

    Given recent interest in social media, many brands now create content that they hope consumers will view and share with peers. While some campaigns indeed go “viral,” their value to the brand is limited if they do not boost brand evaluation or increase purchase. Consequently, a key question is how

  19. Evaluation of serum squamous cell carcinoma antigen as a novel biomarker for diagnosis of hepatocellular carcinoma in Egyptian patients.

    Science.gov (United States)

    Hussein, M M; Ibrahim, A A; Abdella, H M; Montasser, I F; Hassan, M I

    2008-01-01

    Hepatocellular carcinoma (HCC) is the fifth most common malignancy in the world. In Egypt, HCC was reported to account for about 4.7% of chronic liver disease (CLD) patients. Squamous cell carcinoma antigen (SCCA) has been reported to be strongly expressed in HCC tissue hampering its extensive use in clinical practice. To evaluate the clinical usefulness of serum SCCA levels as a serological marker for early detection of HCC among high-risk patients compared to AFP. The study comprised of three groups. Group A included 30 patients with CLD diagnosed based on clinical, laboratory, and ultrasonographical investigations; group B included 49 patients with HCC diagnostically confirmed by spiral CT, elevated alfafetoprotein (AFP), and/or liver biopsy; and group C, the control group, included 15 healthy subjects matched for age and sex. All groups were subjected to thorough history taking, full clinical examination, and laboratory investigations including liver functions, viral markers, and AFP and SCCA estimation using ELISA technique. This study revealed a highly significant difference between patients with HCC, CLD, and controls regarding serum SCCA levels (5.138 +/- 7.689, 1.133 +/- 0.516, and 0.787 +/- 0.432 ng/ml, respectively). SCCA level was persistently elevated in patients with HCC with normal AFP levels representing its useful role in early detection and follow-up of patients treated for HCC. The area under the curve (AUC) of SCCA was 0.869 (95% CI 0.783-0.929), the cut-off value was established at 1.5 ng/ml with sensitivity of 77.6% and specificity of 84.4%). The difference between AUC of SCCA and that of AFP was 0.09 which mounted statistical significance. SCCA could represent a useful tool as a marker for detection of HCC.

  20. REPRODUCIBILITY OF ALKALINE ANTIGENS OF Leishmania major-LIKE AND Leishmania (V. braziliensis EVALUATED BY IgG-ELISA. COMPARISON OF ANTIGENS ADDED OF A PROTEIN INHIBITOR (PMSF OR NOT

    Directory of Open Access Journals (Sweden)

    Celeste Beatriz Julieta

    1998-01-01

    Full Text Available This paper deals with the analysis of 10 batches of L.major-like and L.(V. braziliensis antigens added or not of a proteases inhibitor evaluated by means of an IgG-ELISA on three consecutive days using positive standard sera from patients with diagnosis of American Leishmaniasis previously tested for the presence of IgG antibodies by means of ELISA. The statistical analysis showed that for L. (V. braziliensis the PMSF-containing antigen did not show any difference among batches or days of testing; the L.(V. braziliensis antigen without PMSF showed statistical significance for differences among batches and a two-way ANOVA showed significant differences between antigens. L.major-like antigen prepared with or without PMSF showed differences among batches; all 3 days of testing displayed differences for the PMSF antigen but only for days 1 and 2 for the antigen without inhibitor. A two-way ANOVA showed differences among batches of the antigens but not for antigens with and without the protein inhibitor. According to the statistical analysis the L.major-like antigen added or not of PMSF has shown that it is the choice antigen for mucocutaneous leishmaniasis serology.

  1. Understanding MHC class I presentation of viral antigens by human dendritic cells as a basis for rational design of therapeutic vaccines

    NARCIS (Netherlands)

    N. van Montfoort (Nadine); E. van der Aa (Evelyn); A.M. Woltman (Andrea)

    2014-01-01

    textabstractEffective 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

  2. INDUCTION OF A SECRETORY IGA RESPONSE IN THE MURINE FEMALE UROGENITAL TRACT BY IMMUNIZATION OF THE LUNGS WITH LIPOSOME-SUPPLEMENTED VIRAL SUBUNIT ANTIGEN

    NARCIS (Netherlands)

    DEHAAN, A; RENEGAR, KB; SMALL, PA; WILSCHUT, J

    This study demonstrates that liposomes administered to the lower respiratory tract of mice have the capacity to stimulate secretory IgA (s-IgA) antibody production in the female urogenital system. Total respiratory tract immunization of mice with influenza virus subunit antigen simply mixed with

  3. EARLY SERODIAGNOSIS OF ACUTE HUMAN CYTOMEGALOVIRUS-INFECTION BY ENZYME-LINKED-IMMUNOSORBENT-ASSAY USING RECOMBINANT ANTIGENS

    NARCIS (Netherlands)

    VORNHAGEN, R; PLACHTER, B; HINDERER, W; THE, TH; VANZANTEN, J; MATTER, L; SCHMIDT, CA; SONNEBORN, HH; JAHN, G

    DNA fragments from eight different reading frames of human cytomegalovirus (HCMV) were generated by PCR and subsequently cloned and expressed in Escherichia coli in fusion with glutathione S-transferase. The recombinant viral antigens were evaluated in immunoblot analyses. The most reactive antigens

  4. Benchmarking viromics: an in silico evaluation of metagenome-enabled estimates of viral community composition and diversity

    Directory of Open Access Journals (Sweden)

    Simon Roux

    2017-09-01

    Full Text Available Background Viral metagenomics (viromics is increasingly used to obtain uncultivated viral genomes, evaluate community diversity, and assess ecological hypotheses. While viromic experimental methods are relatively mature and widely accepted by the research community, robust bioinformatics standards remain to be established. Here we used in silico mock viral communities to evaluate the viromic sequence-to-ecological-inference pipeline, including (i read pre-processing and metagenome assembly, (ii thresholds applied to estimate viral relative abundances based on read mapping to assembled contigs, and (iii normalization methods applied to the matrix of viral relative abundances for alpha and beta diversity estimates. Results Tools specifically designed for metagenomes, specifically metaSPAdes, MEGAHIT, and IDBA-UD, were the most effective at assembling viromes. Read pre-processing, such as partitioning, had virtually no impact on assembly output, but may be useful when hardware is limited. Viral populations with 2–5 × coverage typically assembled well, whereas lesser coverage led to fragmented assembly. Strain heterogeneity within populations hampered assembly, especially when strains were closely related (average nucleotide identity, or ANI ≥97% and when the most abundant strain represented <50% of the population. Viral community composition assessments based on read recruitment were generally accurate when the following thresholds for detection were applied: (i ≥10 kb contig lengths to define populations, (ii coverage defined from reads mapping at ≥90% identity, and (iii ≥75% of contig length with ≥1 × coverage. Finally, although data are limited to the most abundant viruses in a community, alpha and beta diversity patterns were robustly estimated (±10% when comparing samples of similar sequencing depth, but more divergent (up to 80% when sequencing depth was uneven across the dataset. In the latter cases, the use of normalization

  5. Development and evaluation of two truncated recombinant NP antigen-based indirect ELISAs for detection of bovine parainfluenza virus type 3 antibodies in cattle.

    Science.gov (United States)

    Yang, Yong; Wang, Feng-Xue; Sun, Na; Cao, Li; Zhang, Shu-Qin; Zhu, Hong-Wei; Guo, Li; Cheng, Shi-Peng; Wen, Yong-Jun

    2015-09-15

    Bovine parainfluenza virus type 3 (BPIV3) is one of the most important viral respiratory pathogens in both young and adult cattle. Nucleocapsid protein (NP) is the most abundant viral protein and the main regulator of virus replication and transcription. In this study, amino acid sequence data of BPIV3 NP was used to identify potential linear epitopic regions, which were subsequently used to design truncated recombinant NP antigens. The amino-terminal region (aa 9-157, NP-N) and the carboxy-terminal region (aa 391-500, NP-C) were selected, and these two truncated recombinant BPIV3 NP proteins were expressed in Escherichia coli based on the results of prediction studies. Furthermore, Enzyme-Linked Immunosorbent Assays (ELISAs) were established using the truncated recombinant BPIV3-N proteins as antigens, and 154 clinical samples were used to evaluate the newly established ELISA systems in comparison with a virus neutralisation test (VNT) as a reference. The results showed that a high coincidence rate was observed for the data that were obtained by the two methods. The sensitivity of NP-N ELISA and NP-C ELISA were 98.4% and 94.6%, respectively, and the specificity of both ELISAs was 100% with reference to the VNTs. Our data indicated that both ends of NP have high immunogenicity during BPIV3 infection and that they were good targets for serodiagnosis. The ELISAs based on the two truncated proteins were especially suitable for use in large-scale epidemiological investigations. Copyright © 2015 Elsevier B.V. All rights reserved.

  6. Recombinant hepatitis B surface antigen production in Aspergillus niger: evaluating the strategy of gene fusion to native glucoamylase

    CSIR Research Space (South Africa)

    James, ER

    2012-10-01

    Full Text Available Microbiology and Biotechnology October 2012/ Vol. 96, No.2 Recombinant hepatitis B surface antigen production in Aspergillus niger: evaluating the strategy of gene fusion to native glucoamylase ER James a,c & WH van Zyl b & PJ van Zyl c & JF Görgens..., Pretoria 0001, South Africa Abstract This study demonstrates the potential of Aspergillus niger as a candidate expression system for virus- like particle production using gene fusion. Hepatitis B surface antigen (HBsAg) production, targeted...

  7. A murine model of coxsackievirus A16 infection for anti-viral evaluation.

    Science.gov (United States)

    Liu, Qingwei; Shi, Jinping; Huang, Xulin; Liu, Fei; Cai, Yicun; Lan, Ke; Huang, Zhong

    2014-05-01

    Coxsackievirus A16 (CA16) is one of the main causative agents of hand, foot and mouth disease (HFMD), which is a common infectious disease in children. CA16 infection may lead to severe nervous system damage and even death in humans. However, study of the pathogenesis of CA16 infection and development of vaccines and anti-viral agents are hindered partly by the lack of an appropriate small animal model. In the present study, we developed and characterized a murine model of CA16 infection. We show that neonatal mice are susceptible to CA16 infection via intraperitoneal inoculation. One-day-old mice infected with 2×10(6)TCID50 of CA16/SZ05 strain consistently exhibited clinical signs, including reduced mobility, and limb weakness and paralysis. About 57% of the mice died within 14days after infection. Significant damage in the brainstem, limb muscles and intestines of the infected mice in the moribund state was observed by histological examination, and the presence of CA16 in neurons of the brainstem was demonstrated by immunohistochemical staining with a CA16-specific polyclonal antibody, strongly suggesting the involvement of the central nervous system in CA16 infection. Analysis of virus titers in various organs/tissues collected at 3, 6 and 9days post-infection, showed that skeletal muscle was the major site of virus replication at the early stage of infection, while the virus mainly accumulated in the brain at the late stage. In addition, susceptibility of mice to CA16 infection was found to be age dependent. Moreover, different CA16 strains could exhibit varied virulence in vivo. Importantly, we demonstrated that post-exposure treatment with an anti-CA16 monoclonal antibody fully protected mice against lethal CA16 infection. Collectively, these results indicate the successful development of a CA16 infection mouse model for anti-viral evaluation. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. Evaluation of reproductive protection against bovine viral diarrhea virus and bovine herpesvirus-1 afforded by annual revaccination with modified-live viral or combination modified-live/killed viral vaccines after primary vaccination with modified-live viral vaccine.

    Science.gov (United States)

    Walz, Paul H; Givens, M Daniel; Rodning, Soren P; Riddell, Kay P; Brodersen, Bruce W; Scruggs, Daniel; Short, Thomas; Grotelueschen, Dale

    2017-02-15

    The objective of this study was to compare reproductive protection in cattle against bovine viral diarrhea virus (BVDV) and bovine herpesvirus 1 (BoHV-1) provided by annual revaccination with multivalent modified-live viral (MLV) vaccine or multivalent combination viral (CV) vaccine containing temperature-sensitive modified-live BoHV-1 and killed BVDV when MLV vaccines were given pre-breeding to nulliparous heifers. Seventy-five beef heifers were allocated into treatment groups A (n=30; two MLV doses pre-breeding, annual revaccination with MLV vaccine), B (n=30; two MLV doses pre-breeding, annual revaccination with CV vaccine) and C (n=15; saline in lieu of vaccine). Heifers were administered treatments on days 0 (weaning), 183 (pre-breeding), 366 (first gestation), and 738 (second gestation). After first calving, primiparous cows were bred, with pregnancy assessment on day 715. At that time, 24 group A heifers (23 pregnancies), 23 group B heifers (22 pregnancies), and 15 group C heifers (15 pregnancies) were commingled with six persistently infected (PI) cattle for 16days. Ninety-nine days after PI removal, cows were intravenously inoculated with BoHV-1. All fetuses and live offspring were assessed for BVDV and BoHV-1. Abortions occurred in 3/23 group A cows, 1/22 group B cows, and 11/15 group C cows. Fetal infection with BVDV or BoHV-1 occurred in 4/23 group A offspring, 0/22 group B offspring, and 15/15 group C offspring. This research demonstrates efficacy of administering two pre-breeding doses of MLV vaccine with annual revaccination using CV vaccine to prevent fetal loss due to exposure to BVDV and BoHV-1. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  9. Effect of cell culture system on the production of human viral antigens Efeito do sistema de cultura celular na produção de antígenos virais humanos

    Directory of Open Access Journals (Sweden)

    Ronaldo Zucatelli Mendonça

    2004-06-01

    Full Text Available A comparative study was performed in the production of different viral antigens by using microcarrier systems and traditional systems. Vero, BHK and MA 104 cells were cultivated in microcarriers (2mg/ml using a bioreactor with a working capacity of 3.7 liters, in parallel with conventional Roux bottles. After four days (BHK cells, and seven days of culture (Vero and MA-104 cells, the cells were infected with 0.1 MOI (multiplicity of infection of rabies virus, measles virus, poliovirus and rotavirus. The yields of the cells and virus in microcarriers and in the conventional system were determined. It was observed that in the microcarrier system, an average increase of twenty-fold more cells/ml was obtained in relation to the conventional monolayer culture, using Roux bottle. On the other hand, cells grown in Roux bottles presented 1.3 to 6.7 more viruses/ml culture than those in the microcarrier systems. However, the overall data showed that yieldings, in terms of viruses per batch, were statistically similar for both systems (p > 0.05. The amount of viral antigen production seems to depend not only on cell concentration, but also on other culture factors such as the characteristic of the cell-growth surface. Thus, the present findings provide a baseline for further improvements and strategies to be established for a scaling-up virus production since depending on the type of virus the optimal conditions found for a small-scale virus production seem unsuitable for large-scale production, requiring new standardization and evaluation.Foi realizado estudo comparativo na produção de diferentes antígenos virais usando sistema de microcarregador e sistema tradicional. Células Vero, BHK e MA-104 foram cultivadas em microcarregadores (2mg/ml utilizando-se biorreatores com capacidade de 3,7 litros e, em paralelo, no sistema convencional com garrafas Roux. Após quatro dias de cultura para as células BHK e sete dias para as células Vero e MA-104, as c

  10. Evaluation of the humoral immune response to human leukocyte antigens in Brazilian renal transplant candidates.

    Directory of Open Access Journals (Sweden)

    Patricia Keiko Saito

    Full Text Available Pre-transplant sensitization to human leukocyte antigens (HLA is a risk factor for graft failure. Studies of the immunological profile related to anti-HLA antibodies in Brazilian renal transplant candidates are few. In this study, we evaluated the humoral immune response to HLA antigens in 269 renal transplant candidates, in Paraná State, Brazil. The HLA typing was performed by the polymerase chain reaction sequence-specific oligonucleotide method (PCR-SSO combined with Luminex technology, using an SSO-LABType commercial kit (One Lambda, Inc., Canoga Park, CA, USA. The percentages of panel-reactive antibodies (PRA and the specificity of anti-HLA antibodies were determined using the LS1PRA and LS2PRA commercial kits (One Lambda, Inc.. The PRA-positive group consisted of 182 (67.7% patients, and the PRA-negative group of 87 (32.3% patients. The two groups differed significantly only with respect to gender. Females were the most sensitized. Among the 182 patients with PRA- positive, 62 (34.1% were positive for class I and negative for class II, 39 (21.4% were negative for class I and positive for class II, and 81 (44.5% were positive for both classes I and II. The HLA-A*02, A*24, A*01, B*44, B*35, B*15, DRB1*11, DRB1*04 and DRB1*03 allele groups were the most frequent. The specificities of anti-HLA antibodies were more frequent: A34, B57, Cw15, Cw16, DR51, DQ8 and DP14. This study documented the profile of anti-HLA antibodies in patients with chronic renal failure who were on waiting lists for an organ in Paraná, and found high sensitization to HLA antigens in the samples.

  11. Development and Evaluation of a Rapid Antigen Detection and Serotyping Lateral Flow Antigen Detection System for Foot-and-Mouth Disease Virus.

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    Kazuki Morioka

    Full Text Available We developed a lateral flow strip using monoclonal antibodies (MAbs which allows for rapid antigen detection and serotyping of foot-and-mouth disease virus (FMDV. This FMDV serotyping strip was able to detect all 7 serotypes and distinguish serotypes O, A, C and Asia1. Its sensitivities ranged from 10(3 to 10(4 of a 50% tissue culture infectious dose of each FMDV stain; this is equal to those of the commercial product Svanodip (Boehringer Ingelheim Svanova, Uppsala, Sweden, which can detect all seven serotypes of FMDV, but does not distinguish them. Our evaluation of the FMDV serotyping strip using a total of 118 clinical samples (vesicular fluids, vesicular epithelial emulsions and oral and/or nasal swabs showed highly sensitive antigen detection and accuracy in serotyping in accordance with ELISA or RT-PCR. To the best of our knowledge, this is the first report on any FMDV serotyping strip that provides both rapid antigen detection and serotyping of FMDV at the same time on one strip without extra devices. This method will be useful in both FMD-free countries and FMD-infected countries, especially where laboratory diagnosis cannot be carried out.

  12. Detection of PMTV Using Polyclonal Antibodies Raised Against a Capsid-Specific Peptide Antigen / Detección de PMTV Utilizando Anticuerpos Policlonales Contra un Péptido Antigénico Derivado de la Cápside Viral

    Directory of Open Access Journals (Sweden)

    Yuliana Gallo García

    2013-12-01

    Full Text Available Potato mop-top virus (PMTV; genus Pomovirus;family Virgaviridae is the causing agent of the spraing disease in potato (Solanum tuberosum. PMTV is transmitted by Spongospora subterranea f. sp. subterranea (Sss. This disease has a widespread distribution in potato growing regions around the world. The possibility of obtaining strain specific antibodies at low cost can greatly increase the sensitivity and use of serological tests in seed certification programs, plant breeding and quarantine regulations to avoid dissemination of this injurious virus. This work presents an alternative procedure for the production of PMTV specific antibodies useful in serological test such as ELISAand lateral flow. In contrast to standard methods requiring theisolation of viral particles or expression of recombinant capsid, this method uses peptides mimicking the N-terminal region of PMTV capsid protein as antigen for the production of specific polyclonal antibodies. The antibodies were tested against bait plants grown in soil infested with viruliferous Sss, as well as potato plants obtained from naturally Sss infested fields in Colombia. PMTV was detected in 9/14 and 24/28 foliage samples of N. benthamiana and S. phureja, respectively. In the case of field plants, the virus wasdetected in eight out of 12 root tissues evaluated. The minimumpeptide concentration detected by ELISA was of the order of 0.1 nM. / Potato mop-top virus (PMTV; género Pomovirus; familia Virgaviridae es transmitido por Spongospora subterranea f. sp. subterranea (Sss, agente causal de la sarna polvosa de la papa. Esta enfermedad tiene una amplia distribución en las regiones cultivadoras de papa alrededor del mundo. La posibilidad de obtener anticuerpos específicos contra cepas de este virus, puede incrementar la sensibilidad y la utilización de pruebas serológicas en programas de certificación de semilla, mejoramiento genético y regulaciones cuarentenarias que eviten su diseminaci

  13. Evaluation of extraction methods from paraffin wax embedded tissues for PCR amplification of human and viral DNA

    OpenAIRE

    Chan, P; Chan, D.; To, K; Yu, M.; Cheung, J.; Cheng, A

    2001-01-01

    Aim—To evaluate the efficiency of phenol/chloroform, microwave, and Qiagen spin column based DNA extractions from paraffin wax embedded tissue for use in the polymerase chain reaction (PCR). In addition, to assess the reliability of amplifying a housekeeping gene to indicate successful viral DNA extraction.

  14. Evaluation of temporal surveillance system sensitivity and freedom from bovine viral diarrhea in Danish dairy herds using scenario tree modelling

    DEFF Research Database (Denmark)

    Foddai, Alessandro; Stockmarr, Anders; Boklund, Anette

    2016-01-01

    The temporal sensitivity of the surveillance system (TemSSe) for Bovine Viral Diarrhea (BVD) in Danish dairy herds was evaluated. Currently, the Danish antibody blocking ELISA is used to test quarterly bulk tank milk (BTM). To optimize the surveillance system as an early warning system, we...

  15. Evaluation of a rapid antigen test for detection of Streptococcus pneumoniae in cerebrospinal fluid.

    Science.gov (United States)

    Boulos, Angel; Fairley, Derek; McKenna, James; Coyle, Peter

    2017-05-01

    Detection of Streptococcus pneumoniae antigen in cerebrospinal fluid (CSF) using lateral flow immunochromatography tests (ICTs) is an effective, rapid and low-cost method to diagnose pneumococcal meningitis. This study evaluated the diagnostic accuracy of the Uni-Gold ICT to detect pneumococcal antigen in CSF specimens, compared with gold standard bacteriology and quantitative real-time PCR (qPCR) testing. CSF specimens (n=69) from patients with suspected bacterial meningitis were included in the study. 13/69 (19%) were positive and 56/69 (81%) were negative for pneumococcus by the gold standard tests. The ICT had sensitivity of 85% (55%-98%), specificity of 96% (88%-100%), positive likelihood ratio of 23.7 (6-94) and negative likelihood ratio of 0.16 (0.04-0.57). Overall, a strong correlation between the ICT and qPCR results was seen (κ=0.81). In contrast, CSF microscopy and culture were exceptionally insensitive. The ICT method is sufficiently robust and accurate for use in algorithms to diagnose bacterial meningitis. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  16. Preclinical evaluation of TriMix and antigen mRNA-based antitumor therapy.

    Science.gov (United States)

    Van Lint, Sandra; Goyvaerts, Cleo; Maenhout, Sarah; Goethals, Lode; Disy, Aurélie; Benteyn, Daphné; Pen, Joeri; Bonehill, Aude; Heirman, Carlo; Breckpot, Karine; Thielemans, Kris

    2012-04-01

    The use of tumor-associated antigen (TAA) mRNA for therapeutic purposes is under active investigation. To be effective, mRNA vaccines need to deliver activation stimuli in addition to TAAs to dendritic cells (DC). In this study, we evaluated whether intranodal delivery of TAA mRNA together with TriMix, a mix of mRNA encoding CD40 ligand, constitutive active Toll-like receptor 4 and CD70, results in the in situ modification and maturation of DCs, hence, priming of TAA-specific T cells. We showed selective uptake and translation of mRNA in vivo by lymph node resident CD11c(+) cells. This process was hampered by codelivery of classical maturation stimuli but not by TriMix mRNA. Importantly, TriMix mRNA induced a T-cell-attracting and stimulatory environment, including recruitment of antigen-specific CD4(+) and CD8(+) T cells and CTLs against various TAAs. In several mouse tumor models, mRNA vaccination was as efficient in CTL induction and therapy response as vaccination with mRNA-electroporated DCs. Together, our findings suggest that intranodal administration of TAA mRNA together with mRNA encoding immunomodulating molecules is a promising vaccination strategy. ©2012 AACR.

  17. Viral hepatitis B- an overview

    African Journals Online (AJOL)

    1994-08-08

    Aug 8, 1994 ... Hepatitis B e antigen. (HBeAg) is a soluble non-structural, enigmatic antigen which is often detected in the blood of patients infected with replicating HBV which results in massive viral load in the blood. Both HBe and HBc are derived from the same section of HBV DNA but the HBe transcript contains an.

  18. Viral marketing on the Internet

    OpenAIRE

    Štverák, Martin

    2008-01-01

    Thesis provides an overview of viral marketing. It describes the process by which you can be inspired to implement viral campaign. The thesis includes analysis of specific viral Web project. The aim of this thesis is to create a breakdown of the various components of viral marketing, to establish conditions that should be satisfied for the viral marketing to success, suggesting how to use viral marketing on social network Facebook and evaluate the various components of this service for the pr...

  19. Influence of plasminogen activator inhibitor-1 (SERPINE1) 4G/5G polymorphism on circulating SERPINE-1 antigen expression in HCC associated with viral infection.

    Science.gov (United States)

    Divella, Rosa; Mazzocca, Antonio; Gadaleta, Cosimo; Simone, Giovanni; Paradiso, Angelo; Quaranta, Michele; Daniele, Antonella

    2012-01-01

    Hepatocarcinogenesis is heavily influenced by chronic hepatitis B (HBV) and C (HCV) infection. Elevated levels of plasminogen activator inhibitor-1 (SERPINE1/PAI-1) have been reported in patients with hepatocellular carcinoma (HCC) associated with viral infection. The gene encoding SERPINE1 is highly polymorphic and the frequently associated 4/5 guanosine (4G/5G) polymorphism in the gene promoter may influence its expression. Here, we investigated the distribution of genotypes and the frequency of alleles of the 4G/5G polymorphism in patients with HCC, the influence of the 4G/5G polymorphism on plasma SERPINE1 levels and its association with viral infection. A total of 75 patients with HCC were enrolled: 32 (42.6%) were HBV(+)/HCV(+), 11 (14.6%) were only HCV(+), and 32 (42.6%) were negative for both viruses. A control group of healthy donors was also enrolled (n=50). SERPINE1 plasma concentrations were determined by ELISA and the detection of the promoter 4G/5G polymorphism was performed by an allele-specific PCR analysis. We found that the frequency of both the 4G/4G genotype (p=0.02) and the 4G allele (p=0.006) were significantly higher in patients with HCC compared to the control group, and particularly higher in patients with HCC co-infected with HBV(+)/HCV(+) than in those with no viral infection. We also found that patients with the 4G/4G genotype had significantly higher plasma SERPINE1 protein levels when compared with patients with the 4G/5G or 5G/5G genotype (p5G SERPINE1 polymorphism with a higher level of SERPINE1 protein in patients with HCC with HBV(+)/HCV(+) than those without infection, suggest the presence of two distinct pathogenic mechanisms in hepatocarcinogenesis, depending on the etiology.

  20. Comparison of serum, ear notches, and nasal and saliva swabs for Bovine viral diarrhea virus antigen detection in colostrum-fed persistently infected (PI) calves and non-PI calves.

    Science.gov (United States)

    Lanyon, Sasha R; Sims, Sarah K; Cockcroft, Peter D; Reichel, Michael P

    2014-11-01

    The diagnosis of neonatal and young calves persistently infected (PI) with Bovine viral diarrhea virus (BVDV) by antigen-capture enzyme-linked immunosorbent assay (ACE) may be complicated by interference from colostrum-derived specific antibodies. Ten calves, with 3 calves identified as PI and 7 as non-PI were used in the current study. All non-PI calves were shown to be seropositive for BVDV-specific antibodies by antibody enzyme-linked immunosorbent assay (Ab-ELISA) on serum. Serum samples, ear notch samples, and nasal and saliva swabs were collected from each calf from birth until 12 weeks of age and tested by ELISA for BVDV-specific antigen and antibodies. Following colostrum ingestion, Ab-ELISA sample-to-positive (S/P) ratios rose by a mean of 0.95 (95% confidence interval [CI] = 0.64-1.25) and 1.72 (95% CI = 1.55-1.89) in seropositive, non-PI calves and in PI calves, respectively. The mean S/P ratios then declined to approximately 1.1 in non-PI calves and 0.5 in PI calves at between 60 and 80 days of age. In PI calves, testing for antigen in serum and nasal and saliva swabs was subject to interference by colostrum-derived antibodies in calves up to 3 weeks of age. Nasal swabs were less affected than serum and saliva swabs. Ear notches maintained positive ACE corrected optical densities at all sample times, despite a drop in the signal following the ingestion of colostrum. © 2014 The Author(s).

  1. Viral Hepatitis

    Science.gov (United States)

    ... Home A-Z Health Topics Viral hepatitis Viral hepatitis > A-Z Health Topics Viral hepatitis (PDF, 90 ... liver. Source: National Cancer Institute Learn more about hepatitis Watch a video. Learn who is at risk ...

  2. Evaluation of three recombinant Leishmania infantum antigens in human and canine visceral leishmaniasis diagnosis.

    Science.gov (United States)

    Fonseca, Aliani Moura; Faria, Angélica Rosa; Rodrigues, Fernandes Tenório Gomes; Nagem, Ronaldo Alves Pinto; Magalhães, Rubens Daniel Miserani; Cunha, João Luís Reis; Bartholomeu, Daniella Castanheira; de Andrade, Hélida Monteiro

    2014-09-01

    Visceral leishmaniasis (VL) is a neglected disease and is fatal if untreated. Dogs serve as reservoirs for Leishmania infantum (syn. L. chagasi) due to their susceptibility to infection and high skin parasitism. Therefore, VL control in Brazil involves the elimination of seropositive dogs, among other actions. However, the most frequently used serological tests have limitations regarding sensitivity and specificity. In this study, we have selected three Leishmania antigens (C1, C8 and C9) and have produced them as recombinant proteins using pET-28a-TEV vector and Escherichia coli BL-21 as expression system. When tested in ELISA with human samples, the C9 antigen was the one showing the most promising results, with 68% sensitivity and 78% specificity. When testing canine samples, the C1, C8 and C9 antigens showed a sensitivity range from 70% to 80% and specificity range from 60% to 90%. The C1 antigen presented higher sensitivity (80%) and the C8 antigen presented higher specificity (90%). Due to it, we decided to mix and test C1 and C8 antigens together, resulting in the C18 antigen. The mix also yielded high percentages of detected symptomatic and asymptomatic dogs however it did not improve the performance of the diagnostic. Comparison of our tests with the tests recommended by the Brazilian Ministry of Health revealed that our antigens' sensitivities and the percentage of detected asymptomatic dogs were much higher. Our results suggest that the C1, C8, C18 and C9 recombinant proteins are good antigens to diagnose canine visceral leishmaniasis and could potentially be used in screening tests. To diagnose human visceral leishmaniasis, the C9 antigen presented reasonable results, but more optimization must be performed for this antigen to provide better performance. Copyright © 2014 Elsevier B.V. All rights reserved.

  3. An evaluation of purified Salmonella Typhi protein antigens for the serological diagnosis of acute typhoid fever

    NARCIS (Netherlands)

    Tran Vu Thieu, Nga; Trinh van, Tan; Tran Tuan, Anh; Klemm, Elizabeth J.; Nguyen Ngoc Minh, Chau; Voong Vinh, Phat; Pham Thanh, Duy; Ho Ngoc Dan, Thanh; Pham Duc, Trung; Langat, Pinky; Martin, Laura B.; Galan, Jorge; Liang, Li; Felgner, Philip L.; Davies, D. Huw; de Jong, Hanna K.; Maude, Rapeephan R.; Fukushima, Masako; Wijedoru, Lalith; Ghose, Aniruddha; Samad, Rasheda; Dondorp, Arjen M.; Faiz, Abul; Darton, Thomas C.; Pollard, Andrew J.; Thwaites, Guy E.; Dougan, Gordon; Parry, Christopher M.; Baker, Stephen

    2017-01-01

    The diagnosis of typhoid fever is a challenge. Aiming to develop a typhoid diagnostic we measured antibody responses against Salmonella Typhi (S. Typhi) protein antigens and the Vi polysaccharide in a cohort of Bangladeshi febrile patients. IgM against 12 purified antigens and the Vi polysaccharide

  4. Antigens evaluation of Fasciola hepática for three inmulogic methods

    Directory of Open Access Journals (Sweden)

    Erasmo Colona

    2014-06-01

    Full Text Available Excretion–secretion antigens (ESA of Fasciola hepatica were evaluated in bovine feces samples from endemic area (Andrés Avelino Cáceres, Central Peruvian Andean. We employed ELISA (Fascidig, Indirect hemagglutination (IHA and Latex agglutination (LA; the results were correlationated with Dennis coproparasitologic method (DC. A number of 106 bovine feces samples were obtained from diferent places of the Region between October and November 1995 (rain time. AES was not detected for LA, the specificity for IHA was 63,6% and the sensibility 35% in relation to DC; the specificity for Elisa (Fascidig was 68,2% and the sensibility was 60% with regard to DC; and in relation to IHA the specificity was 61,7% and the sensibility 50%. The correlation between ELISA (Fascidig and DC was significative, it indicates that ELISA is the best method for the fasciolosis diagnostic in the prepatent and patent phases, while DC only is recommended for the patent phase diagnostic. Diagnostic of infection or prevalence for IHA, ELISA and DC was 35,8%, 42,5% and 37,7% respectively. The results indicate that in the investigated area the prevalence of Fasciola hepatica is high. Thanks to this investigation it was possible to evaluate three immunologic methods and to determine the performance of ELISA-Fascidig for the diagnostic of fasciolosis both prepatent and patent infection phases by means of the demonstration of coproantigens.

  5. Evaluation of three Brucella soluble antigens used in an indirect Elisa to discriminate S19 vaccinated from naturally infected cattle.

    Science.gov (United States)

    Abalos, P; Daffner, J; Pinochet, L

    2000-01-01

    An O-polysaccharide (O-chain) and a hot-water extracted polysaccharide (PS), both obtained from Brucella abortus 1119-3, and a B. melitensis 16M native hapten (NH) were evaluated by indirect enzyme linked immunosorbent assay (ELISA) on three groups of cattle sera. The sera tested were: (a) 75 sera from cows naturally infected with B. abortus; (b) 130 sera from non-infected and non-vaccinated cattle; and (c) 61 sera from non-infected heifers recently vaccinated with B. abortus Strain 19 (S19). Sensitivity (Se), specificity (Sp) and the capability to discriminate vaccinated cattle (ADV) were determined. Using PS antigen, Se was 100% and the Sp was 97.7%, while the highest Sp was obtained by using the O-chain (99.2% ). For the NH antigen, Se was 94.7% and the Sp was 90.0%. The ADV of the three antigens was approximately 85%. Statistical analysis showed significant differences between O-chain/PS and O-chain/NH antigens. The agreement among antigens determined by kappa coefficient was 0.899 for O-chain/PS, 0.845 for O-chain/NH and 0.795 for PS/NH.

  6. Diagnóstico rápido de citomegalovirus (CMV en pacientes inmunocomprometidos mediante anticuerpos monoclonales que reconocen proteinas precoces virales Rapid diagnosis of cytomegalovirus infection in immunocompromised patients by using monoclonal antibodies against early viral antigens

    Directory of Open Access Journals (Sweden)

    Maritza Alvarez

    1989-06-01

    Full Text Available Se aplicó la técnica de detección de antigenos precoces fluorescentes (DAPF usando el anticuerpo monoclonal E-13 McAb, mediante el cual se lograron detectar 15 casos positivos a CMV de 75 muestras de orina o sangre ("buffy coat" tomadas de 52 pacientes inmunocomprometidos ingresados en el Instituto de Nefrología de ciudad Habana. Aplicando las técnicas clásicas de aislamiento en fibroblastos humanos diploides (MRC-5, se lograron aislar 12 cepas de CMV de casos previamente positivos por DAPF; lográndose además un aislamiento en una muestra reportada negativa por fluorescencia. Se observó una coincidencia de un 80% entre ambas técnicas. Se detectó la presencia de anticuerpos IgG contra CMV en todos los casos estudiados, utilizando para ello la técnica ELISA.A technique was applied to detect early fluorescent antigens (DEFA of cytomegalovirus (CMV using the E13 monoclonal antibodies in 52 immunocompromised patients hospitalized in the Nephrology Institute of Havana. Of the 75 urine or blood (buffy coat samples taken, 15 were found positive to CMV. Using classical diploide human fibroblast isolation technique, 12 CMV strains were isloation of previously detected positive samples by DEFA. In addition, CMV was isolated from one sample reported to be negative by DEFA. A coincidence of 80% was found between both techniques. With the ELISA test, all the sample studied have IgG antibodies to CMV.

  7. Hepatitis B surface antigen titer is a good indicator of durable viral response after entecavir off-treatment for chronic hepatitis B

    Directory of Open Access Journals (Sweden)

    Han Ah Lee

    2016-09-01

    Full Text Available Background/Aims Clear indicators for stopping antiviral therapy in chronic hepatitis B (CHB patients are not yet available. Since the level of hepatitis B surface antigen (HBsAg is correlated with covalently closed circular DNA, the HBsAg titer might be a good indicator of the off-treatment response. This study aimed to determine the relationship between the HBsAg titer and the entecavir (ETV off-treatment response. Methods This study analyzed 44 consecutive CHB patients (age, 44.6±11.4 years, mean±SD; men, 63.6%; positive hepatitis B envelope antigen (HBeAg at baseline, 56.8%; HBV DNA level, 6.8±1.3 log10 IU/mL treated with ETV for a sufficient duration and in whom treatment was discontinued after HBsAg levels were measured. A virological relapse was defined as an increase in serum HBV DNA level of >2000 IU/mL, and a clinical relapse was defined as a virological relapse with a biochemical flare, defined as an increase in the serum alanine aminotransferase level of >2 × upper limit of normal. Results After stopping ETV, virological relapse and clinical relapse were observed in 32 and 24 patients, respectively, during 20.8±19.9 months of follow-up. The cumulative incidence rates of virological relapse were 36.2% and 66.2%, respectively, at 6 and 12 months, and those of clinical relapse were 14.3% and 42.3%. The off-treatment HBsAg level was an independent factor associated with clinical relapse (hazard ratio, 2.251; 95% confidence interval, 1.076–4.706; P=0.031. When patients were grouped according to off-treatment HBsAg levels, clinical relapse did not occur in patients with an off-treatment HBsAg level of ≤2 log10 IU/mL (n=5, while the incidence rates of clinical relapse at 12 months after off-treatment were 28.4% and 55.7% in patients with off-treatment HBsAg levels of >2 and ≤3 log10 IU/mL (n=11 and >3 log10 IU/mL (n=28, respectively. Conclusion The off-treatment HBsAg level is closely related to clinical relapse after treatment

  8. Improved vaccine protection against retrovirus infection after co-administration of adenoviral vectors encoding viral antigens and type I interferon subtypes

    Directory of Open Access Journals (Sweden)

    Groitl Peter

    2011-09-01

    Full Text Available Abstract Background Type I interferons (IFNs exhibit direct antiviral effects, but also distinct immunomodulatory properties. In this study, we analyzed type I IFN subtypes for their effect on prophylactic adenovirus-based anti-retroviral vaccination of mice against Friend retrovirus (FV or HIV. Results Mice were vaccinated with adenoviral vectors encoding FV Env and Gag proteins alone or in combination with vectors encoding IFNα1, IFNα2, IFNα4, IFNα5, IFNα6, IFNα9 or IFNβ. Only the co-administration of adenoviral vectors encoding IFNα2, IFNα4, IFNα6 and IFNα9 resulted in strongly improved immune protection of vaccinated mice from subsequent FV challenge infection with high control over FV-induced splenomegaly and reduced viral loads. The level of protection correlated with augmented virus-specific CD4+ T cell responses and enhanced antibody titers. Similar results were obtained when mice were vaccinated against HIV with adenoviral vectors encoding HIV Env and Gag-Pol in combination with various type I IFN encoding vectors. Here mainly CD4+ T cell responses were enhanced by IFNα subtypes. Conclusions Our results indicate that certain IFNα subtypes have the potential to improve the protective effect of adenovirus-based vaccines against retroviruses. This correlated with augmented virus-specific CD4+ T cell and antibody responses. Thus, co-expression of select type I IFNs may be a valuable tool for the development of anti-retroviral vaccines.

  9. Epstein–Barr virus nuclear antigen 3C interact with p73: Interplay between a viral oncoprotein and cellular tumor suppressor

    Energy Technology Data Exchange (ETDEWEB)

    Sahu, Sushil Kumar; Mohanty, Suchitra; Kumar, Amit [Division of Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Chandrasekharpur, Bhubaneswar 751023 (India); Kundu, Chanakya N. [School of Biotechnology, KIIT University, Bhubaneswar (India); Verma, Subhash C. [Department of Microbiology and Immunology, University of Nevada, School of Medicine, Reno, NV 89557 (United States); Choudhuri, Tathagata, E-mail: tatha@ils.res.in [Division of Infectious Disease Biology, Institute of Life Sciences, Nalco Square, Chandrasekharpur, Bhubaneswar 751023 (India); Department of Biotechnology, Siksha Bhavana, Visva Bharati, Santiniketan, Bolpur (India)

    2014-01-05

    The p73 protein has structural and functional homology with the tumor suppressor p53, which plays an important role in cell cycle regulation, apoptosis, and DNA repair. The p73 locus encodes both a tumor suppressor (TAp73) and a putative oncogene (ΔNp73). p73 May play a significant role in p53-deficient lymphomas infected with Epstein–Barr virus (EBV). EBV produces an asymptomatic infection in the majority of the global population, but it is associated with several human B-cell malignancies. The EBV-encoded Epstein–Barr virus nuclear antigen 3C (EBNA3C) is thought to disrupt the cell cycle checkpoint by interacting directly with p53 family proteins. Doxorubicin, a commonly used chemotherapeutic agent, induces apoptosis through p53 and p73 signaling such that the lowΔNp73 level promotes the p73-mediated intrinsic pathway of apoptosis. In this report, we investigated the mechanism by which EBV infection counters p73α-induced apoptosis through EBNA3C. - Highlights: • EBV-encoded EBNA3C suppresses doxorubicin-induced apoptosis in B-cell lymphomas. • EBNA3C binds to p73 to suppress its apoptotic effect. • EBNA3C maintains latency by regulating downstream mitochondrial pathways.

  10. Association of cytotoxic T-lymphocyte antigen 4 genetic polymorphism, hepatitis C viral infection and B-cell non-Hodgkin lymphoma: an Egyptian study.

    Science.gov (United States)

    Khorshied, Mervat Mamdooh; Gouda, Heba Mahmoud; Khorshid, Ola M Reda

    2014-05-01

    Abstract Genetic and environmental factors are involved in the pathogenesis of non-Hodgkin lymphoma (NHL). The present study aimed to investigate the association between cytotoxic T-lymphocyte antigen 4 (CTLA-4) genetic polymorphism, hepatitis C virus (HCV) infection and B-cell NHL risk in Egypt. Genotyping of CTLA-4 single nucleotide polymorphisms (SNPs) was performed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay for 181 adult patients with B-NHL and 200 controls. Our study revealed that CTLA-4 + 49 A/G polymorphism conferred increased risk of B-NHL (odds ratio [OR] = 1.7, 95% confidence interval [CI] = 1.36-2.565). The prevalence of HCV infection in individuals harboring the mutant genotype + 49 A/G and - 318 C/T SNPs was higher in patients with B-NHL and was associated with increased risk of B-NHL (OR = 2.79, 95% CI = 1.24-6.93 for + 49 A/G and OR = 3.9, 95% CI = 1.01-15.98 for - 318 C/T). In conclusion, some SNPs of CTLA-4 are genetic risk factors for B-NHL. Moreover, this study identified an association of CTLA-4 + 49 A/G and - 318 C/T promoter polymorphisms with HCV infection.

  11. Comparative Evaluation of Native Antigens for the Development of Brucellosis Antibody Detection System

    Directory of Open Access Journals (Sweden)

    Yasmin Bano

    2015-09-01

    Full Text Available Brucellosis is a highly infectious zoonotic disease and an economically important infection of humans and livestock with a worldwide distribution. The main mode of transmission of this disease to humans is through the consumption of infected milk, milk products, and uncooked or raw meat. The present study was designed to prepare few native antigens, that is, sonicated antigen (SA, cell envelope (CE antigen, and freeze and thaw (FT antigen from Brucella abortus S99 culture and to test them in a highly sensitive and specific indirect enzyme-linked immunosorbent assay (I-ELISA in both a microtiter plate and a dot-blot format for the development of field-based diagnosis. All 50 suspected bovine samples were tested by plate as well as in dot ELISA formats for all the three antigens prepared. The CE antigen was found to be more suitable as it had the maximum agreement with the Rose Bengal plate agglutination test results followed by the SA and the least agreement was found with that of the FT antigen. This detection system in microtiter plates and a dot-blot format will be useful for the rapid screening of samples for the disease surveillance and routine diagnosis.

  12. Evaluation of seven viral isolates as potential biocontrol agents against Pseudoplusia includens (Lepidoptera: Noctuidae) caterpillars.

    Science.gov (United States)

    Alexandre, Talita M; Ribeiro, Zilda Maria A; Craveiro, Saluana R; Cunha, Fabiane; Fonseca, Ines Cristina B; Moscardi, Flávio; Castro, Maria Elita B

    2010-09-01

    The caterpillar Pseudoplusia includens (Walker, 1857) (Lepidoptera, Noctuidae), known as soybean looper, is a pest that has recently assumed greater importance in soybean in Brazil. Isolates of nucleopolyhedroviruses (NPVs) of this pest have been identified from cotton in Guatemala and soybean farms in Brazil, providing an interesting perspective of potential use of viral insecticide against the insect in lieu to chemical insecticides. With the objective to contribute to the characterization studies of this virus, morphological and molecular analyses and biological activity were carried out with seven P. includens viral isolates (I-A to I-G). Electron microscopy of viral samples, purified from macerated infected larvae, showed particles with typical morphology of the Baculoviridae family, genus Alphabaculovirus (Nucleopolyhedrovirus - NPV) presenting virions with only a single nucleocapsid per envelope (SNPV) occluded in a protein matrix, forming occlusion bodies (OB). This virus was then classified as P. includens single nucleopolyhedrovirus (PsinSNPV). OB particles analyzed in SDS-polyacrylamide gel showed an intense band corresponding in size to NPV polyhedrin protein. DNA restriction profiles of the PsinSNPV isolates showed differences in the fragment size and number suggesting the existence of genotypic variants, except between I-E and I-F profiles that were similar. Among the isolates tested for infectivity against P. includens, I-A, I-E and I-F were the most virulent. Survival times (ST(50)) varied according to viral concentration, with significant differences among isolates for the three higher concentrations. Copyright 2010 Elsevier Inc. All rights reserved.

  13. Antibodies against the mono-methylated arginine-glycine repeat (MMA-RG) of the Epstein-Barr virus nuclear antigen 2 (EBNA2) identify potential cellular proteins targeted in viral transformation.

    Science.gov (United States)

    Ayoubian, Hiresh; Fröhlich, Thomas; Pogodski, Dagmar; Flatley, Andrew; Kremmer, Elisabeth; Schepers, Aloys; Feederle, Regina; Arnold, Georg J; Grässer, Friedrich A

    2017-08-01

    The Epstein-Barr virus is a human herpes virus with oncogenic potential. The virus-encoded nuclear antigen 2 (EBNA2) is a key mediator of viral tumorigenesis. EBNA2 features an arginine-glycine (RG) repeat at amino acids (aa)339-354 that is essential for the transformation of lymphocytes and contains symmetrically (SDMA) and asymmetrically (ADMA) di-methylated arginine residues. The SDMA-modified EBNA2 binds the survival motor neuron protein (SMN), thus mimicking SMD3, a cellular SDMA-containing protein that interacts with SMN. Accordingly, a monoclonal antibody (mAb) specific for the SDMA-modified RG repeat of EBNA2 also binds to SMD3. With the novel mAb 19D4 we now show that EBNA2 contains mono-methylated arginine (MMA) residues within the RG repeat. Using 19D4, we immune-precipitated and analysed by mass spectrometry cellular proteins in EBV-transformed B-cells that feature MMA motifs that are similar to the one in EBNA2. Among the cellular proteins identified, we confirmed by immunoprecipitation and/or Western blot analyses Aly/REF, Coilin, DDX5, FXR1, HNRNPK, LSM4, MRE11, NRIP, nucleolin, PRPF8, RBM26, SMD1 (SNRDP1) and THRAP3 proteins that are either known to contain MMA residues or feature RG repeat sequences that probably serve as methylation substrates. The identified proteins are involved in splicing, tumorigenesis, transcriptional activation, DNA stability and RNA processing or export. Furthermore, we found that several proteins involved in energy metabolism are associated with MMA-modified proteins. Interestingly, the viral EBNA1 protein that features methylated RG repeat motifs also reacted with the antibodies. Our results indicate that the region between aa 34-52 of EBNA1 contains ADMA or SDMA residues, while the region between aa 328-377 mainly contains MMA residues.

  14. Vitamin A deficient mice exhibit increased viral antigens and enhanced cytokine/chemokine production in nasal tissues following respiratory virus infection despite the presence of FoxP3+ T cells.

    Science.gov (United States)

    Penkert, Rhiannon R; Surman, Sherri L; Jones, Bart G; Sealy, Robert E; Vogel, Peter; Neale, Geoffrey; Hurwitz, Julia L

    2016-03-01

    The World Health Organization (WHO) estimates that 250 million children under the age of five suffer from vitamin A deficiencies (VAD). Individuals with VAD experience higher rates of mortality and increased morbidity during enteric and respiratory infections compared with those who are vitamin A sufficient. Previously, our laboratory has demonstrated that VAD mice have significantly impaired virus-specific IgA and CD8(+) T-cell responses in the airways. Here, we demonstrate that VAD mice experience enhanced cytokine/chemokine gene expression and release in the respiratory tract 10 days following virus infection compared with control vitamin A sufficient animals. Cytokines/chemokines that are reproducibly up-regulated at the gene expression and protein levels include IFNγ and IL-6. Despite previous indications that cytokine dysregulation in VAD animals might reflect low forkhead box P3 (FoxP3)-positive regulatory T-cell frequencies, we found no reduction in FoxP3(+) T cells in VAD respiratory tissues. As an alternative explanation for the high cytokine levels, we found that the extent of virus infection and the persistence of viral antigens were increased on day 10 post-infection in VAD animals compared with controls, and consequently that respiratory tract tissues had an increased potential to activate virus-specific T cells. Results encourage cautious management of viral infections in patients with VAD, as efforts to enhance FoxP3(+) T cell frequencies and quell immune effectors could potentially exacerbate disease if the virus has not been cleared. © The Japanese Society for Immunology. 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  15. Species-specific and cross-reactive IgG1 antibody binding to viral capsid protein 1 (VP1 antigens of human rhinovirus species A, B and C.

    Directory of Open Access Journals (Sweden)

    Jua Iwasaki

    Full Text Available BACKGROUND: Human rhinoviruses (HRV are associated with upper and lower respiratory illnesses, including severe infections causing hospitalization in both children and adults. Although the clinical significance of HRV infections is now well established, no detailed investigation of the immune response against HRV has been performed. The purpose of this study was to assess the IgG1 antibody response to the three known HRV species, HRV-A, -B and -C in healthy subjects. METHODS: Recombinant polypeptides of viral capsid protein 1 (VP1 from two genotypes of HRV-A, -B and -C were expressed as glutathione S-transferase (GST fusion proteins and purified by affinity and then size exclusion chromatography. The presence of secondary structures similar to the natural antigens was verified by circular dichroism analysis. Total and species-specific IgG1 measurements were quantitated by immunoassays and immunoabsorption using sera from 63 healthy adults. RESULTS: Most adult sera reacted with the HRV VP1 antigens, at high titres. As expected, strong cross-reactivity between HRV genotypes of the same species was found. A high degree of cross-reactivity between different HRV species was also evident, particularly between HRV-A and HRV-C. Immunoabsorption studies revealed HRV-C specific titres were markedly and significantly lower than the HRV-A and HRV-B specific titres (P<0.0001. A truncated construct of HRV-C VP1 showed greater specificity in detecting anti-HRV-C antibodies. CONCLUSIONS: High titres of IgG1 antibody were bound by the VP1 capsid proteins of HRV-A, -B and -C, but for the majority of people, a large proportion of the antibody to HRV-C was cross-reactive, especially to HRV-A. The improved specificity found for the truncated HRV-C VP1 indicates species-specific and cross-reactive regions could be defined.

  16. Evaluation by ELISA of Anisakis simplex Larval Antigen Purified by Affinity Chromatography

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    Rodero M

    2002-01-01

    Full Text Available In order to improve the specificity and sensitivity of the techniques for the human anisakidosis diagnosis, a method of affinity chromatography for the purification of species-specific antigens from Anisakis simplex third-stage larvae (L3 has been developed. New Zealand rabbits were immunized with A. simplex or Ascaris suum antigens or inoculated with Toxocara canis embryonated eggs. The IgG specific antibodies were isolated by means of protein A-Sepharose CL-4B beads columns. IgG anti-A. simplex and -A. suum were coupled to CNBr-activated Sepharose 4B. For the purification of the larval A. simplex antigens, these were loaded into the anti-A. simplex column and bound antigens eluted. For the elimination of the epitopes responsible for the cross-reactions, the A. simplex specific proteins were loaded into the anti-A. suum column. To prove the specificity of the isolated proteins, immunochemical analyses by polyacrylamide gel electrophoresis were carried out. Further, we studied the different responses by ELISA to the different antigenic preparations of A. simplex used, observing their capability of discriminating among the different antisera raised in rabbits (anti-A. simplex, anti-A. suum, anti-T. canis. The discriminatory capability with the anti-T. canis antisera was good using the larval A. simplex crude extract (CE antigen. When larval A. simplex CE antigen was loaded into a CNBr-activated Sepharose 4B coupled to IgG from rabbits immunized with A. simplex CE antigen, its capability for discriminate between A. simplex and A. suum was improved, increasing in the case of T. canis. The best results were obtained using larval A. simplex CE antigen loaded into a CNBr-activated Sepharose 4B coupled to IgG from rabbits immunized with adult A. suum CE antigen. When we compared the different serum dilution and antigenic concentration, we selected the working serum dilution of 1/400 and 1 mg/ml of antigenic concentration.

  17. Evaluation of dengue NS1 antigen rapid tests and ELISA kits using clinical samples.

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    Subhamoy Pal

    Full Text Available Early diagnosis of dengue virus (DENV infection can improve clinical outcomes by ensuring close follow-up, initiating appropriate supportive therapies and raising awareness to the potential of hemorrhage or shock. Non-structural glycoprotein-1 (NS1 has proven to be a useful biomarker for early diagnosis of dengue. A number of rapid diagnostic tests (RDTs and enzyme-linked immunosorbent assays (ELISAs targeting NS1 antigen (Ag are now commercially available. Here we evaluated these tests using a well-characterized panel of clinical samples to determine their effectiveness for early diagnosis.Retrospective samples from South America were used to evaluate the following tests: (i "Dengue NS1 Ag STRIP" and (ii "Platelia Dengue NS1 Ag ELISA" (Bio-Rad, France, (iii "Dengue NS1 Detect Rapid Test (1st Generation" and (iv "DENV Detect NS1 ELISA" (InBios International, United States, (v "Panbio Dengue Early Rapid (1st generation" (vi "Panbio Dengue Early ELISA (2nd generation" and (vii "SD Bioline Dengue NS1 Ag Rapid Test" (Alere, United States. Overall, the sensitivity of the RDTs ranged from 71.9%-79.1% while the sensitivity of the ELISAs varied between 85.6-95.9%, using virus isolation as the reference method. Most tests had lower sensitivity for DENV-4 relative to the other three serotypes, were less sensitive in detecting secondary infections, and appeared to be most sensitive on Day 3-4 post symptom onset. The specificity of all evaluated tests ranged from 95%-100%.ELISAs had greater overall sensitivity than RDTs. In conjunction with other parameters, the performance data can help determine which dengue diagnostics should be used during the first few days of illness, when the patients are most likely to present to a clinic seeking care.

  18. Binding of the heterogeneous ribonucleoprotein K (hnRNP K to the Epstein-Barr virus nuclear antigen 2 (EBNA2 enhances viral LMP2A expression.

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    Henrik Gross

    Full Text Available The Epstein-Barr Virus (EBV -encoded EBNA2 protein, which is essential for the in vitro transformation of B-lymphocytes, interferes with cellular processes by binding to proteins via conserved sequence motifs. Its Arginine-Glycine (RG repeat element contains either symmetrically or asymmetrically di-methylated arginine residues (SDMA and ADMA, respectively. EBNA2 binds via its SDMA-modified RG-repeat to the survival motor neurons protein (SMN and via the ADMA-RG-repeat to the NP9 protein of the human endogenous retrovirus K (HERV-K (HML-2 Type 1. The hypothesis of this work was that the methylated RG-repeat mimics an epitope shared with cellular proteins that is used for interaction with target structures. With monoclonal antibodies against the modified RG-repeat, we indeed identified cellular homologues that apparently have the same surface structure as methylated EBNA2. With the SDMA-specific antibodies, we precipitated the Sm protein D3 (SmD3 which, like EBNA2, binds via its SDMA-modified RG-repeat to SMN. With the ADMA-specific antibodies, we precipitated the heterogeneous ribonucleoprotein K (hnRNP K. Specific binding of the ADMA- antibody to hnRNP K was demonstrated using E. coli expressed/ADMA-methylated hnRNP K. In addition, we show that EBNA2 and hnRNP K form a complex in EBV- infected B-cells. Finally, hnRNP K, when co-expressed with EBNA2, strongly enhances viral latent membrane protein 2A (LMP2A expression by an unknown mechanism as we did not detect a direct association of hnRNP K with DNA-bound EBNA2 in gel shift experiments. Our data support the notion that the methylated surface of EBNA2 mimics the surface structure of cellular proteins to interfere with or co-opt their functional properties.

  19. Luteolin-7-O-Glucoside Present in Lettuce Extracts Inhibits Hepatitis B Surface Antigen Production and Viral Replication by Human Hepatoma Cells in Vitro

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    Xiao-Xian Cui

    2017-12-01

    Full Text Available Hepatitis B virus (HBV infection is endemic in Asia and chronic hepatitis B (CHB is a major public health issue worldwide. Current treatment strategies for CHB are not satisfactory as they induce a low rate of hepatitis B surface antigen (HBsAg loss. Extracts were prepared from lettuce hydroponically cultivated in solutions containing glycine or nitrate as nitrogen sources. The lettuce extracts exerted potent anti-HBV effects in HepG2 cell lines in vitro, including significant HBsAg inhibition, HBV replication and transcription inhibition, without exerting cytotoxic effects. When used in combination interferon-alpha 2b (IFNα-2b or lamivudine (3TC, the lettuce extracts synergistically inhibited HBsAg expression and HBV replication. By using differential metabolomics analysis, Luteolin-7-O-glucoside was identified and confirmed as a functional component of the lettuce extracts and exhibited similar anti-HBV activity as the lettuce extracts in vitro. The inhibition rate on HBsAg was up to 77.4%. Moreover, both the lettuce extracts and luteolin-7-O-glucoside functioned as organic antioxidants and, significantly attenuated HBV-induced intracellular reactive oxygen species (ROS accumulation. Luteolin-7-O-glucoside also normalized ROS-induced mitochondrial membrane potential damage, which suggests luteolin-7-O-glucoside inhibits HBsAg and HBV replication via a mechanism involving the mitochondria. Our findings suggest luteolin-7-O-glucoside may have potential value for clinical application in CHB and may enhance HBsAg and HBV clearance when used as a combination therapy.

  20. [Evaluation of new fourth-generation human immunodeficiency virus antigen and antibody detection assay with enzyme-linked fluorescent immunoassay].

    Science.gov (United States)

    Shima, Takako; Sudo, Koji; Kondo, Makiko; Kurai, Hanako; Sagara, Hiroko; Imai, Mitsunobu

    2007-09-01

    We evaluated the fourth-generation HIV screening assay VIDAS HIV DUOII (DUOII) based on ELFA for simultaneous detection of anti-HIV-1 and anti-HIV-2 antibodies and HIV-1 p24 antigen through comparison with other HIV antigen-antibody detection assays. Materials were 1228 HIV-negative specimens, 95 HIV-antibody-positive specimens, and HIV commercial panels. The specificity of DUOII was 99.8% and sensitivity 100%, detecting all of HIV-1 group M subtype A, B, B', C, D, A/E, F, G, B/D, HIV-1 group O, and HIV-2. The sensitivity test to HIV-1 p24 antigen was 5pg/ mL, higher than other assays. DUOII was equivalent to or superior in detecting results earlier than other assays in an evaluation using 10 commercial HIV-1 seroconversion panels of primary infection. DUOII detects anti-HIV IgM antibody, so no negative sample was found in the second window between p24 antigen disappearance and raised anti-HIV IgG antibody. DUOII has sufficient specificity and sensitivity for HIV screening, and detects primary infection sooner than other assays. These results indicate that DUOII is useful and reliable in HIV screening.

  1. Evaluation of the cross‐reactivity of antigens in Glupearl 19S and other hydrolysed wheat proteins in cosmetics

    Science.gov (United States)

    Nakamura, Masashi; Yagami, Akiko; Hara, Kazuhiro; Sano‐Nagai, Akiyo; Kobayashi, Tsukane

    2016-01-01

    Summary Background In Japan, over 2000 users of a facial soap containing Glupearl 19S (GP19S), a hydrolysed wheat protein (HWP), developed immediate‐type systemic wheat allergy (HWP‐IWA), and ∼70% of them developed associated contact urticaria. Objectives We investigated whether HWP‐IWA patients cross‐react with other HWPs, and analysed HWP antigenic characteristics. Methods We used 10 types of HWP that are commercially available as cosmetic ingredients, and 16 subjects with HWP‐IWA. We performed an enzyme‐linked immunosorbent assay (ELISA) to evaluate the reactivity to each HWP, and western blotting to evaluate the characteristics of the antigens by using HWP‐IWA patients' serum IgE antibodies. We also performed prick tests with the HWPs. Results The patients reacted to four other HWPs in addition to GP19S, according to ELISA, and this was confirmed by strong reactions in the prick tests to the same four types of HWP. Smears of antigens with molecular weights ranging from the high range to the low range were seen on western blotting with the four HWPs that showed strong reactions in the ELISA and prick tests. Conclusions HWP‐IWA patients cross‐react with other HWPs. The antigens that they cross‐reacted to had a molecular weight distribution similar to that of GP19S present in the HWPs. PMID:27027256

  2. Evaluation of Diagnos Malaria Stix test (antigen detection assay) for diagnosis of malaria.

    Science.gov (United States)

    Khan, Haris M; Shujatullah, Fatima; Shahid, M; Raza, Adil; Malik, Ritu

    2010-06-01

    Malaria is one of the most common parasitic infection in India. The diagnosis largely depends on peripheral blood smear examination. Newer diagnostic methods like various antigen detection assays are now in use for prompt diagnosis and treatment. This study was done to determine the effectiveness of Diagnos Malaria Stix (antigen detection) assay in diagnosis of malaria. This involves detection of PfHRP-2 antigen and P.V. specific pLDH antigen. 162 patients with signs and symptoms of malaria included in the study. Leishman stained blood smear examination was done for all patients. Commercially available Diagnos Malaria Stix assay was used. Diagnos Malaria Stix showed sensitivity, specificity positive and negative predictive values of 100% each while Sensitivity, specificity, positive and negative predictive values of Leishman stained blood smear examination were 45.45%, 100%, 100% and 92% respectively.

  3. Evaluation of the antigen-experienced B-cell receptor repertoire in healthy children and adults

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    Hanna IJspeert

    2016-10-01

    Full Text Available Upon antigen recognition via their B cell receptor (BR B cells migrate to the germinal center where they undergo somatic hypermutation (SHM to increase their affinity for the antigen, and class switch recombination (CSR to change the effector function of the secreted antibodies. These steps are essential to create an antigen-experienced BR repertoire that efficiently protects the body against pathogens. At the same time, the BR repertoire should be selected to protect against responses to self-antigen or harmless antigens. Insights into the processes of SHM, selection and CSR can be obtained by studying the antigen experienced BR repertoire. Currently, a large reference data set of healthy children and adults, which ranges from neonates to the elderly, is not available. In this study, we analyzed the antigen-experienced repertoire of 38 healthy donors (HD, ranging from cord blood to 74 years old, by sequencing IGA and IGG transcripts using next generation sequencing. This resulted in a large, freely available reference data set containing 412,890 IGA and IGG transcripts. We used this data set to study mutation levels, SHM patterns, antigenic selection and CSR from birth to elderly HD. Only small differences were observed in SHM patterns, while the mutation levels increase in early childhood, and stabilize at six years of age at around 7%. Furthermore, comparison of the antigen-experienced repertoire with sequences from the naïve immune repertoire showed that features associated with auto-immunity such as long CDR3 length and IGHV4-34 usage are reduced in the antigen experienced repertoire. Moreover, IGA2 and IGG2 usage was increased in HD in higher age categories, while IGG1 usage was decreased. In addition, we studied clonal relationship in the different samples. Clonally related sequences were found with different subclasses. Interestingly, we found transcripts with the same CDR1-CDR3 sequence, but different subclasses. Together these data

  4. Viral encephalitis

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    Marcus Tulius T Silva

    2013-09-01

    Full Text Available While systemic viral infections are exceptionally common, symptomatic viral infections of the brain parenchyma itself are very rare, but a serious neurologic condition. It is estimated that viral encephalitis occurs at a rate of 1.4 cases per 100.000 inhabitants. Geography is a major determinant of encephalitis caused by vector-borne pathogens. A diagnosis of viral encephalitis could be a challenge to the clinician, since almost 70% of viral encephalitis cases are left without an etiologic agent identified. In this review, the most common viral encephalitis will be discussed, with focus on ecology, diagnosis, and clinical management.

  5. Comparative evaluation of novel African swine fever virus (ASF) antibody detection techniques derived from specific ASF viral genotypes with the OIE internationally prescribed serological tests.

    Science.gov (United States)

    Gallardo, C; Soler, A; Nieto, R; Carrascosa, A L; De Mia, G M; Bishop, R P; Martins, C; Fasina, F O; Couacy-Hymman, E; Heath, L; Pelayo, V; Martín, E; Simón, A; Martín, R; Okurut, A R; Lekolol, I; Okoth, E; Arias, M

    2013-02-22

    The presence of antibodies against African swine fever (ASF), a complex fatal notifiable OIE disease of swine, is always indicative of previous infection, since there is no vaccine that is currently used in the field. The early appearance and subsequent long-term persistence of antibodies combined with cost-effectiveness make antibody detection techniques essential in control programmes. Recent reports appear to indicate that the serological tests recommended by the OIE for ASF monitoring are much less effective in East and Southern Africa where viral genetic and antigenic diversity is the greatest. We report herein an extensive analysis including more than 1000 field and experimental infection sera, in which the OIE recommended tests are compared with antigen-specific ELISAs and immuno-peroxidase staining of cells (IPT). The antibody detection results generated using new antigen-specific tests, developed in this study, which are based on production of antigen fractions generated by infection and virus purification from COS-1 cells, showed strong concordance with the OIE tests. We therefore conclude that the lack of success is not attributable to antigenic polymorphism and may be related to the specific characteristics of the local breeds African pigs. Copyright © 2012 Elsevier B.V. All rights reserved.

  6. X-ray diffraction analysis of crystals from the human major histocompatibility antigen HLA-B*2706 in complex with a viral peptide and with a self-peptide

    Energy Technology Data Exchange (ETDEWEB)

    Zawacka, Anna [Institut für Immungenetik, Charité - Universitätsmedizin Berlin, Campus Virchow-Klinikum, Humboldt-Universität zu Berlin, Spandauer Damm 130, 14050 Berlin (Germany); Loll, Bernhard; Biesiadka, Jacek; Saenger, Wolfram [Institut für Chemie und Biochemie/Kristallographie, Freie Universität Berlin, Takustrasse 6, 14195 Berlin (Germany); Uchanska-Ziegler, Barbara; Ziegler, Andreas, E-mail: andreas.ziegler@charite.de [Institut für Immungenetik, Charité - Universitätsmedizin Berlin, Campus Virchow-Klinikum, Humboldt-Universität zu Berlin, Spandauer Damm 130, 14050 Berlin (Germany)

    2005-12-01

    The crystallization of HLA-B*2706 in complex with two peptides is reported. The human leukocyte antigen (HLA) alleles HLA-B*2704 and HLA-B*2706 show an ethnically restricted distribution and are differentially associated with ankylosing spondylitis, with HLA-B*2706 lacking association with this autoimmune disease. However, the products of the two alleles differ by only two amino acids, at heavy-chain residues 114 (His in HLA-B*2704; Asp in HLA-B*2706) and 116 (Asp in HLA-B*2704; Tyr in HLA-B*2706). Both residues could be involved in contacting amino acids of a bound peptide, suggesting that peptides presented by these subtypes play a role in disease pathogenesis. Two HLA-B*2706–peptide complexes were crystallized using the hanging-drop vapour-diffusion method with PEG as precipitant. Data sets were collected to resolutions of 2.70 Å (viral peptide pLMP2, RRRWRRLTV; space group P2{sub 1}2{sub 1}2{sub 1}) and 1.83 Å (self-peptide pVIPR, RRKWRRWHL; space group P2{sub 1}). Using HLA-B*2705 complexed with the pGR peptide (RRRWHRWRL) as a search model, unambiguous molecular-replacement solutions were found for both HLA-B*2706 complexes.

  7. Evaluation of Innate Immune Biomarkers in Saliva for Diagnostic Potential of Bacterial and Viral Respiratory Infection

    Science.gov (United States)

    2014-02-03

    up-regulated in bacterial and viral infections, such as Staphylococcus aureus and Influenza virus, and plays a key role in cell-mediated immunity... Streptococcus sore throat compact pneumoniae 2 58 male white Saliva/serum Cough, fatigue, fever, Vitek 2 Streptococcus sore throat compact pneumoniae 3 52...fever, fatigue, Vitek 2 Streptococcus sore throat compact pneumoniae 8 44 female white Saliva/serum Cough, fever, muscle Vitek 2 Klebsiella

  8. Evaluation of vaccinal effectiveness of preparations containing membrane antigens of Leishmania (L.) amazonensis in experimental cutaneous leishmaniasis model.

    Science.gov (United States)

    Ribeiro, João G; Ferreira, Amália S; Macedo, Sharon R A; Rossi, Norton R D L P; da Silva, Mayara C P; Guerra, Rosane N M; de Barros, Neuza B; Nicolete, Roberto

    2017-06-01

    American tegumentary leishmaniasis (ATL) is considered a neglected disease, for which an effective vaccine or an efficient diagnosis is not yet available and whose chemotherapeutic arsenal is threatened by the emergence of resistance by etiological agents such as Leishmania amazonensis. ATL is endemic in poor countries and has a high incidence in Brazil. Vaccines developed from native parasite fractions have led to the identification of defined antigenic subunits and the development of vaccine adjuvant technology. The purpose of the present study was to develop and compare preparations based on membrane antigens from L. amazonensis, as a biotechnological prototype for the immunoprophylaxis of the disease in a murine experimental model. For this purpose, batches of biodegradable polymeric micro/nanoparticles were produced, characterized and compared with other parasite's antigens in solution. All preparations containing membrane antigens presented low toxicity on murine macrophages. The in vivo evaluation of immunization efficacy was performed against a challenge with L. amazonensis, along with an evaluation of the immune response profile generated in BALB/C mice. The animals were followed for sample processing and quantification of serum-specific cytokines, nitrites and antibodies. The sera of animals immunized with the non-encapsulated antigen formulations showed higher intensities of nitrites and total IgGs. This approach evidenced the importance of the biological studies involving the immune response of the host against the parasite being interconnected and related to the subfractionation of its proteins in the search for more effective vaccine candidates. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. [Evaluation of the Recombinant Protein Tp0965 of Treponema Pallidum as Perspective Antigen for the Improved Serological Diagnosis of Syphilis].

    Science.gov (United States)

    Runina, A V; Starovoitova, A S; Deryabin, D G; Kubanov, A A

    2016-01-01

    BACKGRAUND. Treponemal tests based on the detection of antibodies against the Treponema pallidum antigens are the most specific methods for serological diagnosis of syphilis. Due to the inability to cultivate this bacterium in vitro, the most promising sources of antigens for diagnostics are recombinant proteins of T. pallidum. Evaluation of the analytical value of certain T. pallidum proteins is the approach to improve sensitivity, specificity, and reproducibility of syphilis serological tests, including possibilities of differential diagnosis of various forms of the disease. The aim of the research was to evaluate the analytical values (sensitivity and specificity) of recombinant protein Tp0965 of T. pallidum as a candidate antigen for serological diagnosis of syphilis. tp0965 gene was cloned into the expression vector pET28a and the construct was used for the transformation of E. coli BL-21 (DE3) cells and further expression and purification of the recombinant protein. The collected protein was used as T. pallidum antigen for serum analysis (ELISA) of groups of patients with various forms of syphilis (n=84) and the group of healthy donors (n = 25). High frequency of positive ELISA results was shown with serum of patients with syphilis, compared to the group of healthy donors. The sensitivity of serological reactions using recombinant protein Tp0965 was 98.8%, specificity--87.5%. The highest sensitivity (100%) was detected in the groups of patients with primary, secondary and early latent syphilis while in the group of patients with late latent syphilis it decreased to 95.2%. We concluded that due to its specificity T. pallidum recombinant protein Tp0965 can be used as a novel perspective antigen for development of syphilis serological diagnostic assays (for primary and early latent forms).

  10. Evaluation of extraction methods from paraffin wax embedded tissues for PCR amplification of human and viral DNA.

    Science.gov (United States)

    Chan, P K; Chan, D P; To, K F; Yu, M Y; Cheung, J L; Cheng, A F

    2001-05-01

    To evaluate the efficiency of phenol/chloroform, microwave, and Qiagen spin column based DNA extractions from paraffin wax embedded tissue for use in the polymerase chain reaction (PCR). In addition, to assess the reliability of amplifying a housekeeping gene to indicate successful viral DNA extraction. DNA samples extracted from 20 blocks of cervical carcinoma tissues using the three methods were subjected to PCRs targeting 509 bp and 355 bp of the beta globin gene, and 450 bp and 150 bp of human papillomavirus (HPV) DNA. Microwave extraction showed the highest positive rate for beta globin PCR, whereas the spin column method was the most efficient for HPV DNA extraction. When the 509 bp beta globin and 450 bp HPV PCR results were correlated, two of 10, eight of 12, and nine of 10 beta globin positive extractions prepared by means of the phenol/chloroform, microwave, and spin column methods, respectively, yielded HPV DNA of the expected size. For the beta globin negative samples, HPV was detected in three of 10, two of eight, and four of 10 samples. HPV DNA extraction was most efficient using the Qiagen spin column and had the highest positive predictive value when a housekeeping gene was used as an indicator of successful viral DNA extraction; the phenol/chloroform method was the least efficient. The potential drawbacks of some extraction methods when using a human housekeeping gene to assess the quality of viral DNA extraction need to be considered.

  11. Treatment of viral encephalitis.

    Science.gov (United States)

    Domingues, Renan Barros

    2009-03-01

    Several viruses may cause central nervous system diseases with a broad range of clinical manifestations. The time course of the viral encephalitis can be acute, subacute, or chronic. Pathologically there are encephalitis with direct viral entry into the CNS in which brain parenchyma exhibits neuronal damaging and viral antigens and there are postinfectious autoimmune encephalitis associated with systemic viral infections with brain tissue presenting perivascular aggregation of immune cells and myelin damaging. Some virus affect previously healthy individuals while others produce encephalitis among imunocompromised ones. Factors such evolving lifestyles and ecological changes have had a considerable impact on the epidemiology of some viral encephalitis [e.g. West-Nile virus, and Japanese B virus]. Citomegalovirus and JC virus are examples of infections of the brain that have been seen more frequently because they occur in immunocompromised patients. In the other hand many scientific achievements in neuroimaging, molecular diagnosis, antiviral therapy, immunomodulatory treatments, and neurointensive care have allowed more precise and earlier diagnoses and more efficient treatments, resulting in improved outcomes. In this article, we will present the current drug options in the management of the main acute and chronic viral infection of the central nervous system of immunocompetent and immunocompromised adults, focusing on drugs mechanisms of action, efficacy, and side effects. The early diagnosis and correct management of such diseases can reduce mortality and neurological sequelae; however, even with recent treatment advances, potentially devastating outcomes are still possible.

  12. Evaluation of PCR based assays for the improvement of proportion estimation of bacterial and viral pathogens in diarrheal surveillance

    Directory of Open Access Journals (Sweden)

    Hongxia eGuan

    2016-03-01

    Full Text Available AbstractDiarrhea can be caused by a variety of bacterial, viral and parasitic organisms. Laboratory diagnosis is essential in the pathogen-specific burden assessment. In the pathogen spectrum monitoring in the diarrheal surveillance, culture methods are commonly used for the bacterial pathogens’ detection whereas nucleic acid based amplification, the non-cultural methods are used for the viral pathogens. Different methodology may cause the inaccurate pathogen spectrum for the bacterial pathogens because of their different culture abilities with the different media, and for the comparison of bacterial vs. viral pathogens. The application of nucleic acid-based methods in the detection of viral and bacterial pathogens will likely increase the number of confirmed positive diagnoses, and will be comparable since all pathogens will be detected based on the same nucleic acid extracts from the same sample. In this study, bacterial pathogens, including diarrheagenic Escherichia coli (DEC, Salmonella spp., Shigella spp., Vibrio parahaemolyticus and V. cholerae, were detected in 334 diarrheal samples by PCR-based methods using nucleic acid extracted from stool samples and associated enrichment cultures. A protocol was established to facilitate the consistent identification of bacterial pathogens in diarrheal patients. Five common enteric viruses were also detected by RT-PCR, including rotavirus, sapovirus, norovirus (I and II, human astrovirus, and enteric adenovirus. Higher positive rates were found for the bacterial pathogens, showing the lower proportion estimation if only using culture methods. This application will improve the quality of bacterial diarrheagenic pathogen survey, providing more accurate information pertaining to the pathogen spectrum associated with finding of food safety problems and disease burden evaluation.

  13. Viral marketing

    OpenAIRE

    Bláhová, Adéla

    2012-01-01

    The aim of my thesis is to provide a comprehensive overview of the viral marketing and to analyze selected viral campaigns. There is a description of advantages and disadvantages of this marketing tool. In the end I suggest for which companies viral marketing is an appropriate form of the promotion.

  14. Evaluation of immune response elicited by inulin as an adjuvant with filarial antigens in mice model.

    Science.gov (United States)

    Mahalakshmi, N; Aparnaa, R; Kaliraj, P

    2014-10-01

    Filariasis caused by infectious parasitic nematodes has been identified as the second leading source of permanent and long-term disability in Sub-Saharan Africa, Asia and Latin America. Several vaccine candidates were identified from infective third-stage larvae (L3) which involves in the critical transition from arthropod to human. Hitherto studies of these antigens in combination with alum adjuvant have shown to elicit its characteristic Th2 responses. Inulin is a safe, non-toxic adjuvant that principally stimulates the innate immune response through the alternative complement pathway. In the present study, the immune response elicited by inulin and alum as adjuvants were compared with filarial antigens from different aetiological agents: secreted larval acidic protein 1 (SLAP1) from Onchocerca volvulus and venom allergen homologue (VAH) from Brugia malayi as single or as cocktail vaccines in mice model. The study revealed that inulin can induce better humoral response against these antigens than alum adjuvant. Antibody isotyping disclosed inulin's ability to elevate the levels of IgG2a and IgG3 antibodies which mediates in complement-dependent cytotoxicity and antibody-dependent cell-mediated cytotoxicity (ADCC), respectively, in mice. Splenocyte analysis showed that T cells prestimulated with inulin have higher stimulation index (P inulin formulation had induced higher cytotoxicity with filarial antigens (as single P inulin to deplete the levels of Treg and brought a balance in Th1/Th2 arms against filarial antigens in mice. © 2014 John Wiley & Sons Ltd.

  15. Development of a luciferase based viral inhibition assay to evaluate vaccine induced CD8 T-cell responses.

    Science.gov (United States)

    Naarding, Marloes A; Fernandez, Natalia; Kappes, John C; Hayes, Peter; Ahmed, Tina; Icyuz, Mert; Edmonds, Tara G; Bergin, Philip; Anzala, Omu; Hanke, Tomas; Clark, Lorna; Cox, Josephine H; Cormier, Emmanuel; Ochsenbauer, Christina; Gilmour, Jill

    2014-07-01

    Emergence of SIV and HIV specific CD8 T cells has been shown to correlate with control of in vivo replication. Poor correlation between IFN-γ ELISPOT responses and in vivo control of the virus has triggered the development of more relevant assays to assess functional HIV-1 specific CD8 T-cell responses for the evaluation and prioritization of new HIV-1 vaccine candidates. We previously established a viral inhibition assay (VIA) that measures the ability of vaccine-induced CD8 T-cell responses to inhibit viral replication in autologous CD4 T cells. In this assay, viral replication is determined by measuring p24 in the culture supernatant. Here we describe the development of a novel VIA, referred to as IMC LucR VIA that exploits replication-competent HIV-1 infectious molecular clones (IMCs) in which the complete proviral genome is strain-specific and which express the Renilla luciferase (LucR) gene to determine viral growth and inhibition. The introduction of the luciferase readout does provide significant improvement of the read out time. In addition to switching to the LucR read out, changes made to the overall protocol resulted in the miniaturization of the assay from a 48 to a 96-well plate format, which preserved sample and allowed for the introduction of replicates. The overall assay time was reduced from 13 to 8 days. The assay has a high degree of specificity, and the previously observed non-specific background inhibition in cells from HIV-1 negative volunteers has been reduced dramatically. Importantly, we observed an increase in positive responses, indicating an improvement in sensitivity compared to the original VIA. Currently, only a limited number of "whole-genome" IMC-LucR viruses are available and our efforts will focus on expanding the panel to better evaluate anti-viral breadth. Overall, we believe the IMC LucR VIA provides a platform to assess functional CD8 T-cell responses in large-scale clinical trial testing, which will enhance the ability to

  16. [Evaluation of antigenic properties of Campylobacter jejuni and Campylobacter coli proteins in a western-immunoblot].

    Science.gov (United States)

    Rokosz, Natalia; Waldemar, Rastawicki; Jagielski, Marek

    2008-01-01

    Campylobacter jejuni and Campylobacter coli are the most common bacterial cause for acute diarrheal illnesses in developed countries. The aim of this study was to evaluate the antigenic properties of Campylobacterjejuni and Campylobacter coli proteins in western-blot assay. Whole-cell components of Campulobacter jejuni and Campylobacter coli were separated by sodium dodecyl sulfate-polyacrylamide gel electroforesis. Using this method we detected in all seven C. jejuni strains 21 peptides migrating between 180-29 kDa. All three Ccoli strains had a 17 bands migrating with the same molecular weight range. Proteins were transferred electrophoretically to nitrocellulose paper for immunoblotting experiments. The 74 kDa protein reacted strongly in all classes ofimmmunoglobulin with all tested human serum samples. We observed that this protein reacted also with human immunoglobulins for Salmonella and Yersinia sp. This cross-reaction observed for this protein could give false positive results in routine diagnosis of C. jejuni infections. The proteins with molecular weight of: 92, 62, 56, 52, 45-43, 29 kDa were most recognized in the 20 human serum samples. The other proteins of Cljejuni and C. coli, particularly in the 68-50 kDa and 45-31 kDa regions, were recognized occasionally and the response to these in reconvalescent sera was usually weak. The result of this study showed that the proteins with molecular weight: 92, 62, 56, 52, 45-43 and 29 kDa can be use in routine serological diagnostic of campylobacteriosis.

  17. Long-term outcome of hepatitis B e antigen-positive patients with compensated cirrhosis treated with interferon alfa. European Concerted Action on Viral Hepatitis (EUROHEP)

    DEFF Research Database (Denmark)

    Fattovich, G; Giustina, G; Realdi, G

    1997-01-01

    The aim of this study was to evaluate whether interferon alfa (IFN-alpha) treatment-associated virological and biochemical remission improves survival in a cohort of 90 white patients with compensated cirrhosis caused by hepatitis B (Child A) followed for a mean period of 7 years. Inclusion...

  18. Avaliação hematológica e histopatológica de camundongos BALB/c e C57BL/6 expostos aos antígenos recombinantes Cytoplasmic Repetitive Antigen e Flagellar Repetitive Antigen de Trypanosoma cruzi Hematological and histopathological evaluation of BALB/c and C57BL/6 mice exposed to Cytoplasmic Repetitive Antigen and Flagellar Repetitive Antigen recombinant antigens of Trypanosoma cruzi

    Directory of Open Access Journals (Sweden)

    Valéria Rêgo Alves Pereira

    2003-12-01

    Full Text Available Os antígenos recombinantes Cytoplasmic Repetitive Antigen e Flagellar Repetitive Antigen de Trypanosoma cruzi foram inoculados em camundongos BALB/c e C57BL/6 e o seu efeito avaliado a nível hematológico e histopatológico. Os resultados mostraram que o padrão histológico normal dos órgãos e o perfil hematológico dos camundongos não foram modificados sugerindo que esses antígenos não parecem causar dano ao animal.The Cytoplasmic Repetitive Antigen and Flagellar Repetitive Antigen recombinant antigens of Trypanosoma cruzi were inoculated into BALB/c and C57BL/6 mice and its effects evaluated at hematological and histopathological levels. The results showed that the histological pattern of the organs and the hematological profile of mice were not modified suggesting that these antigens are not harmful for the animal.

  19. Controlled attenuation parameter for evaluating liver steatosis in chronic viral hepatitis.

    Science.gov (United States)

    Ferraioli, Giovanna; Tinelli, Carmine; Lissandrin, Raffaella; Zicchetti, Mabel; Dal Bello, Barbara; Filice, Gaetano; Filice, Carlo

    2014-06-07

    To assess the performance of controlled attenuation parameter (CAP) in patients with chronic viral hepatitis. CAP is a new technique that measures the attenuation in the liver of an ultrasound beam, which is directly related to lipid accumulation. Consecutive patients undergoing liver biopsy for chronic viral hepatitis were studied using the M probe of FibroScan device (Echosens, Paris, France). The device estimates liver steatosis in decibel per meter (dB/m). An expert operator performed all measurements. Steatosis was graded according to Kleiner's classification. Pearson or Spearman rank coefficient was used to test correlation between two study variables. Linear regression was used for multivariate model to assess the association between CAP and other variables. Receiver operating characteristic curve analysis was performed to calculate area under the curve (AUROC) for S0 vs S1-S3 and S0-S1 vs S2-S3. 115 subjects (85 males and 30 females) were prospectively studied. The mean values of CAP were 227.1 ± 43.1 for S0; 254.6 ± 38.9 for S1; 297.8 ± 49.4 dB/m for S2-S3. In univariate analysis CAP showed a significant correlation with age, body mass index (BMI), degree of steatosis, and cholesterol. Multivariate regression analysis confirmed the correlation with the degree of steatosis [coefficient, 1.2 (0.60-1.83); P Controlled attenuation parameter could be a useful tool in the clinical management of patients with chronic viral hepatitis for detecting liver steatosis.

  20. Evaluation of Dirofilaria immitis antigen detection comparing heated and unheated serum in dogs with experimental heartworm infections.

    Science.gov (United States)

    Carmichael, James; McCall, Scott; DiCosty, Utami; Mansour, Abdelmoneim; Roycroft, Linda

    2017-11-09

    To evaluate whether heated serum allows for earlier detection of Dirofilaria immitis antigen, dogs with experimental D. immitis infections underwent weekly blood sampling to compare antigen results using both heated and unheated serum. One of two isolates (JYD-34 or Big Head™) were used to infect naïve laboratory beagle dogs. Serum was collected from dogs weekly and divided into two aliquots, heated and unheated. The samples designated as heated were placed in a heat block at 104 °C for 10 min then centrifuged with collection of the resulting supernatant. Two commercial ELISAs, DiroCHEK® (Synbiotics Corporation, Zoetis) and PetChek® (IDEXX Laboratories, Inc.), were used to conduct D. immitis antigen testing on all serum samples. There was no statistical difference in the mean number of days from infection to positive D. immitis antigen status between the two commercial testing kits (DiroCHEK® versus PetChek®) with either heated or unheated serum. When unheated serum was utilized, very strong agreement between the two assays was demonstrated using Lin's concordance correlation coefficient (R c  = 0.98). However, when heated serum was compared, Lin's concordance correlation coefficient was only R c  = 0.64, showing a lesser agreement. There was a statistical difference in the mean number of days from infection to a positive test result for unheated serum when compared to mean days to positive status with heated serum. For DiroCHEK® the heated serum yielded a positive result 126.9 ± 18.9 days postinfection while the unheated serum yielded a positive result 162.6 ± 23.0 days postinfection; this was a significant 35.7 ± 32.2 days longer, on average, compared with heated serum. With PetChek® the heated serum yielded a positive result 131.5 ± 11.7 days postinfection while the unheated serum yielded a positive result 162.8 ± 23.8 days postinfection; this was a significant 31.3 ± 25.5 days longer, on average, compared with heated

  1. Dengue viruses cluster antigenically but not as discrete serotypes

    NARCIS (Netherlands)

    L. Katzelnick (Leah); J.M. Fonville (Judith); G.D. Gromowski (Gregory D.); J.B. Arriaga (Jose Bustos); A. Green (Angela); S.L. James (Sarah ); L. Lau (Louis); M. Montoya (Magelda); C. Wang (Chunling); L.A. Van Blargan (Laura A.); C.A. Russell (Colin); H.M. Thu (Hlaing Myat); T.C. Pierson (Theodore C.); P. Buchy (Philippe); J.G. Aaskov (John G.); J.L. Muñoz-Jordán (Jorge L.); N. Vasilakis (Nikos); R.V. Gibbons (Robert V.); R.B. Tesh (Robert B.); A.D.M.E. Osterhaus (Albert); R.A.M. Fouchier (Ron); A. Durbin (Anna); C.P. Simmons (Cameron P.); E.C. Holmes (Edward C.); E. Harris (Eva); S.S. Whitehead (Stephen S.); D.J. Smith (Derek James)

    2015-01-01

    textabstractThe four genetically divergent dengue virus (DENV) types are traditionally classified as serotypes. Antigenic and genetic differences among the DENV types influence disease outcome, vaccine-induced protection, epidemic magnitude, and viral evolution.We scharacterized antigenic diversity

  2. Evaluation of Salmonella enterica Type III Secretion System Effector Proteins as Carriers for Heterologous Vaccine Antigens

    Science.gov (United States)

    Hegazy, Wael Abdel Halim; Xu, Xin; Metelitsa, Leonid

    2012-01-01

    Live attenuated strains of Salmonella enterica have a high potential as carriers of recombinant vaccines. The type III secretion system (T3SS)-dependent translocation of S. enterica can be deployed for delivery of heterologous antigens to antigen-presenting cells. Here we investigated the efficacy of various effector proteins of the Salmonella pathogenicity island (SPI2)-encoded T3SS for the translocation of model antigens and elicitation of immune responses. The SPI2 T3SS effector proteins SifA, SteC, SseL, SseJ, and SseF share an endosomal membrane-associated subcellular localization after translocation. We observed that all effector proteins could be used to translocate fusion proteins with the model antigens ovalbumin and listeriolysin into the cytosol of host cells. Under in vitro conditions, fusion proteins with SseJ and SteC stimulated T-cell responses that were superior to those triggered by fusion proteins with SseF. However, in mice vaccinated with Salmonella carrier strains, only fusion proteins based on SseJ or SifA elicited potent T-cell responses. These data demonstrate that the selection of an optimal SPI2 effector protein for T3SS-mediated translocation is a critical parameter for the rational design of effective Salmonella-based recombinant vaccines. PMID:22252866

  3. Antigenic variation: Molecular and genetic mechanisms of relapsing disease

    Energy Technology Data Exchange (ETDEWEB)

    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.

  4. Preparation and evaluation of Salmonella Enteritidis antigen conjugated with nanogold for screening of poultry flocks

    Directory of Open Access Journals (Sweden)

    Hazem Mohammed Ibrahim

    2017-08-01

    Full Text Available Aim: The present work aimed to develop lateral flow immunochromatographic strip (ICS test for detection of Salmonella Enteritidis (SE specific antibodies in chicken sera. Materials and Methods: A rapid lateral flow immunochromatographic test (LFIT has been developed, in which SE Group D antigen labeled with the gold chloride molecules laid on the conjugate pad. Staphylococcus aureus protein A was used as capture antibody at the test line (T of a nitrocellulose (NC membrane and anti-SE antigen-specific rabbit antibodies were used as capture antibody at the control line (C of the NC strip in the lateral flow layout device. Results: Using the developed LFIT, the minimal amount of SE-specific antibodies that can be detected in chicken serum sample was 1427 enzyme-linked immunosorbent assay (ELISA unit/100 μl that was equal to 0.1 μg (Ab/100 μl sample. 100 suspected serum samples collected from a poultry flock were tested with the prepared SE-LFIT kits and the locally prepared stained Salmonella antigen, and the results were compared with those obtained from examination of these samples with Salmonella Group D antibody ELISA kit as the gold standard test. The sensitivity, specificity, and accuracy of the prepared SE-LFIT antigen kits were 94.4%, 90%, and 94%, respectively, while those obtained with stained Salmonella antigen were 88.8%, 90%, and 89%, respectively. Conclusion: The developed test is a simple field rapid test of high sensitivity, specificity, and accuracy that can improve and facilitates rapid field surveillance of salmonellosis among chickens.

  5. Cloning, expression and evaluation of the efficacy of a recombinant Baylisascaris schroederi Bs-Ag3 antigen in mice.

    Science.gov (United States)

    Wang, Tao; He, Guangzhi; Yang, Guangyou; Fei, Yuxiang; Zhang, Zhihe; Wang, Chengdong; Yang, Zhi; Lan, Jingchao; Luo, Li; Liu, Li

    2008-12-09

    The gene Bs-Ag3 enconding a antigen of 37kDa from Baylisascaris schroederi (giant panda isolates), as well as the recombinant Bs-Ag3, obtained by cloning and expression of the Bs-Ag3 gene in heterologous host Escherichia coli BL-21 (DE3), were used to evaluate their ability to induce immune protective responses in BALB/c mice against L3-challenge infection in a mouse-B. schroederi model. There was a significant reduction (62.91%) of mice vaccinated with rBs-Ag3 coupled with Freund's complete adjuvant (FCA) in recovery of challenged B. schroederi L3 compared with either controls or mice vaccinated with FCA alone. Our data indicate recombinant Bs-Ag3 may be a potential target as a vaccine antigen for giant panda ascariasis.

  6. Evaluation of envelope domain III-based single chimeric tetravalent antigen and monovalent antigen mixtures for the detection of anti-dengue antibodies in human sera.

    Science.gov (United States)

    Batra, Gaurav; Nemani, Satish K; Tyagi, Poornima; Swaminathan, Sathyamangalam; Khanna, Navin

    2011-03-15

    Flavivirus cross-reactive antibodies in human sera interfere with the definitive identification of dengue virus (DENV) infections especially in areas with multiple co-circulating flaviviruses. Use of DENV envelope domain-III (EDIII) can partially resolve the problem. This study has examined the effect of (i) incorporating the EDIIIs of four DENV serotypes into a single chimeric antigen, and (ii) immobilizing the antigen through specific interaction on the sensitivity and specificity of anti-DENV antibody detection. A sera panel (n = 164) was assembled and characterized using commercial kits for infection by DENV and a host of other pathogens. Anti-DENV antibodies of both IgM and IgG classes in this panel were detected in indirect ELISAs using a mixture of monovalent EDIIIs, a chimeric EDIII-based tetravalent antigen, EDIII-T, and a biotinylated version of the latter as coating antigens. The sensitivity and specificity of these assays were compared to those obtained using the PanBio Dengue IgG/IgM ELISAs. The performance of dengue IgG and IgM indirect ELISAs, using either a physical mixture of four EDIIIs or the single chimeric EDIII-T antigen, were comparable. Coating of a biotinylated version of the tetravalent antigen on streptavidin plates enhanced sensitivity without compromising specificity. The incorporation of the EDIIIs of the four DENV serotypes into a single chimeric antigen did not adversely affect assay outcome in indirect ELISAs. Oriented, rather than random, immobilization of the tetravalent antigen enhanced sensitivity of detection of anti-DENV antibodies with retention of 100% specificity.

  7. EVALUATION OF HUMAN LEUKOCYTE ANTIGEN CLASS I AND II ANTIGENS IN HELICOBACTER PYLORI-POSITIVE PEDIATRIC PATIENTS WITH ACTIVE GASTRITIS AND DUODENAL ULCER

    Directory of Open Access Journals (Sweden)

    Sevim GÖNEN

    2017-10-01

    Full Text Available ABSTRACT BACKGROUND: As being the first bacteria determined to be carcinogenic, Helicobacter pylori (H. pylori is a pathogen localized in the stomach in more than half of the world population. Some earlier studies have found a relation between tissue histocompatibility antigens and gastric cancers depending on the regions. OBJECTIVE: The present study aimed to determine the distribution of human leukocyte antigen (HLA class I and class II antigens in H. pylori-positive pediatric patients with active gastritis and duodenal ulcer, excluding cancer cases, in our center. METHODS: The study included 40 patients diagnosed with H. pylori-positive active gastritis and duodenal ulcer and 100 controls consisting of healthy donor candidates. The HLA class I and class II antigens were studied in the isolated DNA samples using the polymerase chain reaction sequence-specific oligonucleotide probes. RESULTS: The frequency of HLA-B*51 antigen was significantly higher in the patient group than in the control group (40% vs 17%; P=0.003. There was no difference between the two groups in terms of the frequencies of HLA-A, HLA-C, HLA-DR, and HLA-DQ antigens. CONCLUSION: It was determined that HLA-B*51 plays a critical role in H. pylori infection.

  8. Immunohistochemical evaluation of epithelial antigen Ber-Ep4 and CD10: new markers for endometriosis?

    Science.gov (United States)

    Capobianco, G; Wenger, J M; Marras, V; Cosmi, E; Ambrosini, G; Dessole, M; Cherchi, P L

    2013-01-01

    Early and certain diagnoses of endometriosis are mandatory to begin the correct treatment and to exclude the risk of endometriosis-associated ovarian carcinoma (EOC) and endometrial stromal sarcoma (ESS). To assess the immunohistochemical expression of Ber-Ep4, an epithelial antigen, and CD10 in endometriosis. Forty-eight women underwent laparoscopic surgery for endometriosis and endometriotic samples were recovered for histology. In all surgical specimens Ber-Ep4 and CD10 were searched by an immnohistochemical method. The authors evaluated the correlations among the immunohistochemical positivity and the location of endometriosis. Most cases (40/48 83.34%) were represented by ovarian endometriotic cyst. Among the eight remaining cases, three (3/48, 6.25%) were pelvic endometriotic lesions, two (2/48, 4.17%) peritoneum of vesico-uterine pouch, one vaginal lesion (2.08%), one salpinx lesion (2.08%), and one inguinal location (2.08%). Ber-Ep4 and CD10 were expressed in 90% and in 100% of the ovarian lesions, respectively. In pelvic lesions Ber-Ep4 and CD10 showed both 66.67% of positivity and had the same pattern in peritoneal, salpinx, vaginal, and inguinal lesions (50%, 100%, 100%, 100%, respectively). Ber-Ep4 was negative in 6/48 (12.5%) cases whereas CDO10 was negative in 2/48 (4.17%) cases of endometriosis. The sensitivity of Ber-Ep4 and CD10 for endometriosis diagnosis were 87.50% and 95.83%, respectively. Immunohistochemistry for Ber-Ep4 showed positivity in all cases of endometriosis with typical cubic epithelium, whereas CD10 was positive in 1/2 (50%) atypical case. Immunohistochemical expression of Ber-Ep4 and CD10 was positive in most cases of endometriosis and was useful in differential diagnosis with mesothelial cysts. Ber-Ep4 was negative in cases of hyperplastic epithelium or cytological atypia; these cases are not well-differentiated and could be optimally treated by surgery and not by hormonal therapy because of the risk of cancer degeneration.

  9. Expression, purification and preliminary X-ray crystallographic analysis of the human major histocompatibility antigen HLA-B*1402 in complex with a viral peptide and with a self-peptide

    Energy Technology Data Exchange (ETDEWEB)

    Kumar, Pravin [Institut für Immungenetik, Charité - Universitätsmedizin Berlin, Humboldt-Universität zu Berlin, Thielallee 73, 14195 Berlin (Germany); Vahedi-Faridi, Ardeschir [Institut für Chemie und Biochemie/Kristallographie, Freie Universität Berlin, Takustrasse 6, 14195 Berlin (Germany); Merino, Elena; López de Castro, José A. [Centro de Biología Molecular Severo Ochoa, Consejo Superior de Investigaciones Cientificas, Universidad Autónoma de Madrid, Facultad de Ciencias, Universidad Autónoma, 28049 Madrid (Spain); Volz, Armin; Ziegler, Andreas [Institut für Immungenetik, Charité - Universitätsmedizin Berlin, Humboldt-Universität zu Berlin, Thielallee 73, 14195 Berlin (Germany); Saenger, Wolfram, E-mail: saenger@chemie.fu-berlin.de [Institut für Chemie und Biochemie/Kristallographie, Freie Universität Berlin, Takustrasse 6, 14195 Berlin (Germany); Uchanska-Ziegler, Barbara, E-mail: saenger@chemie.fu-berlin.de [Institut für Immungenetik, Charité - Universitätsmedizin Berlin, Humboldt-Universität zu Berlin, Thielallee 73, 14195 Berlin (Germany)

    2007-07-01

    The crystallization of HLA-B*1402 in complex with two peptides is reported. The product of the human major histocompatibility (HLA) class I allele HLA-B*1402 only differs from that of allele HLA-B*1403 at amino-acid position 156 of the heavy chain (Leu in HLA-B*1402 and Arg in HLA-B*1403). However, both subtypes are known to be differentially associated with the inflammatory rheumatic disease ankylosing spondylitis (AS) in black populations in Cameroon and Togo. HLA-B*1402 is not associated with AS, in contrast to HLA-B*1403, which is associated with this disease in the Togolese population. The products of these alleles can present peptides with Arg at position 2, a feature shared by a small group of other HLA-B antigens, including HLA-B*2705, the prototypical AS-associated subtype. Complexes of HLA-B*1402 with a viral peptide (RRRWRRLTV, termed pLMP2) and a self-peptide (IRAAPPPLF, termed pCatA) were prepared and were crystallized using polyethylene glycol as precipitant. The complexes crystallized in space groups P2{sub 1} (pLMP2) and P2{sub 1}2{sub 1}2{sub 1} (pCatA) and diffracted synchrotron radiation to 2.55 and 1.86 Å resolution, respectively. Unambiguous solutions for both data sets were obtained by molecular replacement using a peptide-complexed HLA-B*2705 molecule (PDB code) as a search model.

  10. Evaluation of Mycobacterium leprae antigens in the serological monitoring of a clofazimine-based chemotherapeutic study of dapsone resistant lepromatous leprosy patients in Cebu, Philippines

    NARCIS (Netherlands)

    Douglas, J. T.; Hirsch, D. S.; Fajardo, T. T.; Cellona, R. V.; Abalos, R. M.; de la Cruz, E. C.; Madarang, M. G.; de Wit, M. Y.; Klatser, P. R.

    1989-01-01

    Thirty-one dapsone resistant lepromatous leprosy patients receiving clofazimine based therapy were serologically monitored throughout their 5-year period of treatment. Sequentially collected sera were used to examine 4 Mycobacterium leprae antigens to evaluate their usefulness in ELISA's for

  11. EXPERIMENTAL LIPOSOMAL VIRAL VACCINE SAFETY

    Directory of Open Access Journals (Sweden)

    Romanova OA

    2016-12-01

    Full Text Available Introduction. With the transport links development there is rather important issue respiratory viral infections spread, especially influenza. The only method controlling influenza is vaccination. Search and development effective and safe vaccines is important. Material and methods. In base SO "Mechnikov Institute Microbiology and Immunology National Ukrainian Academy Medical Sciences" in the scientific theme "Developing new approaches to creating viral vaccines and study specific activity depending of type and degree component`s modification" was created several experimental influenza vaccine with subsequent component`s modification for selecting the most optimal pattern of safety and immunogenicity. In assessing the influenza vaccine safety is using a few criteria, including, reactivity, as measured by the frequency of local and systemic adverse (negative effects, which due to its introduction, and for lipid content drugs, ability to influence oxidation processes. At present study phase was determined: a systemic reaction and local reaction of delayed-type hypersensitivity (foot pad swelling assay;b lipids and proteins peroxidation processes after administration officinal and experimental vaccines (content protein’s carbonyl groups, lipid’s hydroperoxides, activity of glutathione-peroxidase.Study objects were trivalent seasonal influenza vaccine, "Vaxigrip" (Sanofi Pasteur, S.A., France, "Inflexal V" (Biotech Ltd. Berne, Switzerland and experimental vaccine samples. Highest immunogenicity vaccines had undergone improvements and modifications using adjuvant systems and acylation influenza proteins. Liposomes 2 – the experimental influenza vaccine with a liposome negative charge and antigenic composition like split vaccines "Vaksihryp". Liposomes 2.1 - the adjuvantexperimental influenza vaccine with modifications liposomal components (etoniy and chlorophyllipt molecules embedded in liposomal membrane. Liposomes 2.2 - the adjuvant

  12. Evaluation of a non-viral vaccine in smallpox-vaccinated individuals and immunized HLA-transgenic mice.

    Science.gov (United States)

    Taylor, Alice P; Makabi-Panzu, Boby; Chen, Xiaochuan; Gold, David V; Goldenberg, David M

    2012-06-01

    The current poxvirus vaccine is associated with rare, but serious adverse events. Therefore, we investigated a non-replicating approach to vaccine design. Peptides encoding potential HLA-binding motifs were derived from the orthopoxvirus genes, D8L, A27L, and C12L (the IL-18-binding protein [vIL18BP105]), all of which are preserved among poxviruses that infect humans, and which may be a target of host immunity. The peptides were tested with poxvirus-vaccinated human PBMC and serum for eliciting memory responses, as well as with splenocytes and serum from peptide-immunized, human HLA-DR04 transgenic (HLA tg) mice. vIL18BP105 induced 5-fold proliferation of vaccinated-donor PBMC over non-vaccinated (P<0.001), including IL-2-producing CD8+ cells. Serum IgG recognizing vIL18BP105 was detected (P<0.002 vs non-vaccinated) by ELISA. Viral peptides were conjugated to the HLA-targeting mAb, L243, for immunization of HLA tg mice. Splenocytes from vIL18BP105-L243-immunized mice proliferated upon exposure to vIL18BP105 (P<0.001). Proliferating splenocytes were interferon-γ-producing CD4(+)CD45RA(neg). vIL18BP105-L243-immunized mice generated IgG more rapidly than free-peptide-immunized mice. Peptide-specific antibody was also detected when different L243-peptide conjugates were combined. vIL18BP, by eliciting human memory responses, is a viable antigen for inclusion in a virus-free vaccine. The immunogenicity of peptides was boosted by conjugation to L243, whether administered alone or combined. Copyright © 2012 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

  13. Immunohistochemical evaluation of a new epithelial antigen, BER-EP4 in ovarian cancer: a propos of 62 cases.

    Science.gov (United States)

    Capobianco, G; Marras, V; Battista Meloni, G; Dessole, S; Ashqar, N; Cherchi, C; Dessole, F; Cherchi, P L

    2012-01-01

    To assess the immunohistochemical expression of BerEP4, a new epithelial antigen in ovarian cancer. We studied 62 cases of ovarian cancer in which BerEP4, CEA and CA-125 were investigated by an immunohistochemical method. We evaluated the correlations among immunohistochemical positivity and the grading, histotype and stage of disease. BerEP4 was positive in 45 out of 62 cases (72.58%), CA-125 in 36 out of 62 cases (58.06%) and CEA in ten out of 62 cases (16.13%). BerEP4 was present both in serous and in mucinous tumors (80.96% vs. 80.77%). CA-125 was mainly expressed in serous vs mucinous tumors (66.67% vs. 57.69%). CEA was more prevalent in mucinous vs. serous tumors. Ber-EP4 was mainly expressed in G1 (75%) and G2 (77.27%). CA-125 was more present in G1 and G3 (both 62.50%) than G2 (50%), whereas CEA showed positivity in G1: 12.50%, G2: 22.73% and G3: 12.50%. There were no differences among the three antigens studied with regard to clinical stage. In our study Ber-EP4 was positive in 45 out of 62 cases (72.58%) of primary epithelial ovarian cancers. The presence of this antigen seemed to be related to the histotype and grading but not to clinical stage.

  14. Evaluation of Helicobacter pylori antigen positivity in stool samples of patients with dyspeptic complaints in a tertiary care hospital

    Directory of Open Access Journals (Sweden)

    Mehmet Burak Selek

    2013-12-01

    Full Text Available Objective: Helicobacter pylori is a microorganism associatedwith gastritis, peptic ulcer disease and gastriccancer. We aimed to figure out the positivity rate in stoolsamples of outpatients with dyspeptic complaints visitinggastroenterology department and to evaluate its relationwith age, gender and seasonal changes.Methods: Between January 01, 2012 and December 31,2012, stool samples of 330 adult outpatients admitted togastroenterology department are investigated with an immunochromatographictest kit using monoclonal antibodiesfor detection of H. pylori antigen.Results: Among 330 patients’ stool samples tested, 67(20.3% were positive. 18.6% of men and 22.2% of womenwere detected as positive. According to age groups,17.1% patients were positive for 15-35 age groups,27.1% patients were positive for 36-55 age groups and18.2% patients were positive for above 56. Seasonal differenceof H. pylori antigen positivity in stool samples wasstatistically significant (p=0.001. Highest positivity rate29.7% was detected for winter months (December-January-February. According to logistic regression analysis,winter is found as a risk factor with statistically significant2.295 times greater risk [p=0001, Exp (B = 2.925, 95.0%C.I. for EXP (B = 1.668-5.129].Conclusion: H. pylori antigen positivity rate of our study islower than other previously conducted studies in Turkey.But, positivity rates are higher among women comparedto men, concordant with other studies. Even more, detectionof high positivity rates in winter shows primary infectionand/or relapse can be affected by seasonal changes.Key words: Helicobacter pylori, gastroenterology, stool antigen test

  15. Pemasaran ViralViral Marketing

    OpenAIRE

    Situmorang, James Rianto

    2010-01-01

    Viral marketing is an extremely powerful and effective form of internet marketing. Itis a new form of word-of-mouth through internet. In viral marketing, someone passeson a marketing message to someone else and so on. Viral marketing proposes thatmessages can be rapidly disseminated from consumer to consumer leading to largescale market acceptance. The analogy of a virus is used to described the exponentialdiffusion of information in an electronic environment and should not be confusedwith th...

  16. Evaluation of disease and viral biomarkers as triggers for therapeutic intervention in respiratory mousepox - an animal model of smallpox.

    Science.gov (United States)

    Parker, Scott; Chen, Nanhai G; Foster, Scott; Hartzler, Hollyce; Hembrador, Ed; Hruby, Dennis; Jordan, Robert; Lanier, Randall; Painter, George; Painter, Wesley; Sagartz, John E; Schriewer, Jill; Mark Buller, R

    2012-04-01

    The human population is currently faced with the potential use of natural or recombinant variola and monkeypox viruses as biological weapons. Furthermore, the emergence of human monkeypox in Africa and its expanding environs poses a significant natural threat. Such occurrences would require therapeutic and prophylactic intervention with antivirals to minimize morbidity and mortality of exposed populations. Two orally-bioavailable antivirals are currently in clinical trials; namely CMX001, an ether-lipid analog of cidofovir with activity at the DNA replication stage and ST-246, a novel viral egress inhibitor. Both of these drugs have previously been evaluated in the ectromelia/mousepox system; however, the trigger for intervention was not linked to a disease biomarker or a specific marker of virus replication. In this study we used lethal, intranasal, ectromelia virus infections of C57BL/6 and hairless SKH1 mice to model human disease and evaluate exanthematous rash (rash) as an indicator to initiate antiviral treatment. We show that significant protection can be provided to C57BL/6 mice by CMX001 or ST-246 when therapy is initiated on day 6 post infection or earlier. We also show that significant protection can be provided to SKH1 mice treated with CMX001 at day 3 post infection or earlier, but this is four or more days before detection of rash (ST-246 not tested). Although in this model rash could not be used as a treatment trigger, viral DNA was detected in blood by day 4 post infection and in the oropharyngeal secretions (saliva) by day 2-3 post infection - thus providing robust and specific markers of virus replication for therapy initiation. These findings are discussed in the context of current respiratory challenge animal models in use for the evaluation of poxvirus antivirals. Copyright © 2012 Elsevier B.V. All rights reserved.

  17. Identification of sVSG117 as an immunodiagnostic antigen and evaluation of a dual-antigen lateral flow test for the diagnosis of human African trypanosomiasis.

    Directory of Open Access Journals (Sweden)

    Lauren Sullivan

    2014-07-01

    Full Text Available The diagnosis of human African trypanosomiasis (HAT caused by Trypanosoma brucei gambiense relies mainly on the Card Agglutination Test for Trypanosomiasis (CATT. There is no immunodiagnostic for HAT caused by T. b. rhodesiense. Our principle aim was to develop a prototype lateral flow test that might be an improvement on CATT.Pools of infection and control sera were screened against four different soluble form variant surface glycoproteins (sVSGs by ELISA and one, sVSG117, showed particularly strong immunoreactivity to pooled infection sera. Using individual sera, sVSG117 was shown to be able to discriminate between T. b. gambiense infection and control sera by both ELISA and lateral flow test. The sVSG117 antigen was subsequently used with a previously described recombinant diagnostic antigen, rISG65, to create a dual-antigen lateral flow test prototype. The latter was used blind in a virtual field trial of 431 randomized infection and control sera from the WHO HAT Specimen Biobank.In the virtual field trial, using two positive antigen bands as the criterion for infection, the sVSG117 and rISG65 dual-antigen lateral flow test prototype showed a sensitivity of 97.3% (95% CI: 93.3 to 99.2 and a specificity of 83.3% (95% CI: 76.4 to 88.9 for the detection of T. b. gambiense infections. The device was not as good for detecting T. b. rhodesiense infections using two positive antigen bands as the criterion for infection, with a sensitivity of 58.9% (95% CI: 44.9 to 71.9 and specificity of 97.3% (95% CI: 90.7 to 99.7. However, using one or both positive antigen band(s as the criterion for T. b. rhodesiense infection improved the sensitivity to 83.9% (95% CI: 71.7 to 92.4 with a specificity of 85.3% (95% CI: 75.3 to 92.4. These results encourage further development of the dual-antigen device for clinical use.

  18. [Field evaluation for diagnostic accuracy of the rapid test SD Bioline Malaria Antigen Pf/Pv® in Colombia].

    Science.gov (United States)

    Mendoza, Nohora Marcela; Cucunubá, Zulma Milena; Aponte, Samanda; González, Nohora Elizabeth; Bernal, Sindy Durley

    2013-01-01

    Rapid diagnostic tests (RDT) have been postulated as a way to ensure access to malaria diagnosis in remote areas. Despite its widespread use, there are no field studies to evaluate the accuracy of the SD Bioline Malaria Antigen Pf/Pv in Colombia RDT. To evaluate the diagnostic accuracy of the SD Bioline Malaria Antigen Pf/Pv® RDT in two departments endemic for malaria, comparing diagnosis with thick film corrected with PCR. A retrospective study was carried out to evaluate sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), concordance and sensitivity limits according to parasitemia ranges for the SD Bioline Malaria Antigen Pf/Pv ® test in Cordoba and Choco. The results were compared with microscopy corrected by PCR. A total of 383 samples processed, 121 were positive (75 for P. vivax , 42 for P. falciparum and 4 for mixed infection) and 262 negative samples. P. vivax: sensitivity 92.0% (95% CI: 83.6-96.3), specificity 98.7% ( 95% CI: 96.7-99.5), PPV 94.5% (95% CI: 86.7-97.9), NPV 98.1% (95% CI: 95.8-99.1), Cohen's kappa coefficient was 0.90 (0.80-1.00). P. falciparum: sensitivity 88.1% (95% CI: 75.0-94.8), specificity 97.9% (95% CI: 95.8-99.0), PPV 84.1% (95% CI: 70.6-92.1), NPV 98.5% (95% IC: 96.6-99.4), Cohen's kappa coefficient 0.80 (95% CI: 0.70-0.90). The test performed well, being better for P. vivax as compared to P. falciparum. There are still difficulties of RDT to detect low parasitemias. The non amplification of Pfhrp2 and Pfhrp3 genes in two samples diagnosed as mixed infection, suggest a possible deletion of these two genes together.

  19. [Use of infusoria Paramecium caudatum for the evaluation of the toxicity of antigens of different causative quarantine agents].

    Science.gov (United States)

    Zhukova, S I; Adel'shin, F K; Khrapova, N P; Piven', N N; Proshina, O B; Zasiadkina, A V; Plekhanova, N G; Avrorova, I V

    2005-01-01

    The results of the evaluation of the toxicity of bacterial antigens obtained from the causative agents of plaque, glanders, melioidosis, cholera on infusoria of the species P. caudatum, as well as on cell lines L-929, CHO K-1 and peritoneal macrophages of BALB/c mice, are presented. As revealed in this study, the method of toxicity determination on infusoria is similar in its sensitivity to the methods of testing on. CHO K-1 and L-929 cells, but the former is simpler, more available and permits the determination of toxic doses producing disturbances in the vital activity of the infusoria, but not leading to their death.

  20. Evaluation of the fully automated Cobas Core enzyme immunoassay for the quantitation of antibodies against hepatitis B virus surface antigen.

    Science.gov (United States)

    Doche, C; Thomé, M; Dimet, I; Bienvenu, J

    1996-04-01

    The Roche Cobas Core automated enzyme immunoassay of antibodies against hepatitis B virus surface antigen (anti-HBs) was evaluated using a protocol complying with the Société Française de Biologie Clinique guidelines. Results showed good precision (CV below 6 and 9% for intra- and inter-assay precision) and accuracy (according to the Paul-Ehrlich-Institute standard); the detection limit of the test was 2 IU/l. The manufacturer's specifications were confirmed and a satisfactory correlation (r = 0.901) was found with an automated method (IMx, Abbott) used for comparison.

  1. Performance evaluation of a new fourth-generation HIV combination antigen-antibody assay.

    Science.gov (United States)

    Mühlbacher, A; Schennach, H; van Helden, J; Hebell, T; Pantaleo, G; Bürgisser, P; Cellerai, C; Permpikul, P; Rodriguez, M I; Eiras, A; Alborino, F; Cunningham, P; Axelsson, M; Andersson, S; Wetlitzky, O; Kaiser, C; Möller, P; de Sousa, G

    2013-02-01

    Education and diagnostic tests capable of early detection represent our most effective means of preventing transmission of human immunodeficiency virus (HIV). The importance of early detection is underlined by studies demonstrating increased life expectancy following early initiation of antiviral treatment. The Elecsys(®) HIV combi PT assay is a fourth-generation antigen-antibody combination assay developed to allow earlier detection of seroconversion, and to have increased sensitivity and improved specificity. We aimed to determine how early the assay could detect infection compared with existing assays; whether all HIV variants could be detected; and the assay's specificity using samples from blood donors, routine specimens, and patients with potential cross-reacting factors. Samples were identified as positive by the Elecsys(®) assay 4.9 days after a positive polymerase chain reaction result (as determined by the panel supplier), which was earlier than the 5.3-7.1 days observed with comparators. The analytical sensitivity of the Elecsys(®) HIV combi PT assay for the HIV-1 p24 antigen was 1.05 IU/mL, which compares favorably with the comparator assays. In addition, the Elecsys(®) assay identified all screened HIV subtypes and displayed greater sensitivity to HIV-2 homologous antigen and antibodies to HIV-1 E and O and HIV-2 than the other assays. Overall, the specificity of the Elecsys(®) assay was 99.88 % using samples from blood donors and 99.81 % when analyzing unselected samples. Potential cross-reacting factors did not interfere with assay performance. The Elecsys(®) HIV combi PT assay is a sensitive and specific assay that has been granted the CE mark according to Directive 2009/886/EC.

  2. Evaluation of an immunoblot methodology for the detection of relevant Entamoeba histolytica antigens by antibodies induced in human amebiasis.

    Science.gov (United States)

    Argüello-García, R; Sánchez-Guillén, M C; Garduño, G; Valadez-Salazar, A; Martínez-García, M C; Muñoz, O; Ortega-Pierres, M G

    1990-01-01

    The complexity of the clinical spectrum in human amebiasis and the high variability in laboratory methods used to detect Entamoeba histolytica infections have impeded the collection and evaluation of reliable epidemiological data. Thus, more sensitive, specific and standardized methods are needed in order to accurately identify infections with this parasite. An important step in the development of serological diagnostic methods is the identification and isolation of specific parasite antigens which are immunogenic in the host. In this work, we have standardized an electroimmunotransfer blot technique to characterize E. histolytica antigens recognized by antibodies present during human amebic infections. An important aspect was an investigation of technical variations in the preparation of cell lysates including the use of different protease inhibitors and solubilizing agents. The highest yield of protein was achieved by homogenization of trophozoites in the presence of 10 mM p-hydroxymercuribenzoate (pHMB) as a protease inhibitor and by lysis using Triton X-100 and a mixture of protease inhibitors. Recovery of degraded vs non-degraded proteins in the cell extracts was evaluated by gradient polyacrylamide gel electrophoresis. Both quantitative and qualitative differences were noted between the different methods of preparing soluble cell extracts. A more complete set of antigenic components was obtained by homogenization and use of pHMB. Thus parasite extracts prepared by this method were selected for protein transfer. In this, the optimal protein concentration was of 120 micrograms of protein per cm of gel width and efficient transfer of proteins to nitrocellulose sheets was achieved at 100 V for 2 hrs and at 4 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)

  3. Evaluation of performance of human immunodeficiency virus antigen/antibody combination assays in Taiwan.

    Science.gov (United States)

    Chang, Chun-Kai; Kao, Cheng-Feng; Lin, Pi-Han; Huang, Hui-Lin; Ho, Shu-Yuan; Wong, Kuo-Chen; Lin, Bo-Chang; Yeh, Chang-Ching; Lee, Chia-Yeh; Kao, Chuan-Liang; Lee, Chun-Nan; Chang, Sui-Yuan; Yang, Jyh-Yuan

    2017-08-01

    The fourth-generation human immunodeficiency virus (HIV) combination assay, which can simultaneously detect the presence of anti-HIV antibody and HIV antigen, has been shown to shorten the window period in HIV diagnosis compared with the third-generation HIV antibody immunoassay. This study was aimed to determine the performance of HIV combination assays in Taiwan, where the HIV-1 seroprevalence is 0.007% and HIV-2 infection has never been reported. Performance of three fourth-generation HIV Ag/Ab combination assays (Dia.Pro, Wantai, and Bio-Rad) and one third-generation HIV Ab immunoassay (AxSYM HIV 1/2 gO) was assessed. A total of 152 specimens, including 86 confirmed HIV-seropositive and 66 HIV-seronegative samples, were used in the study. The sensitivity of four assays varied from 98.8% to 100%, and specificity varied from 98.5% to 100%. Performance of the 75 equivocal samples, the HIV status of which was confirmed later, in terms of negative prediction varied from 81.8% to 87.5%. The Bio-Rad and Dia.Pro assays exhibited higher sensitivity for the detection of p24 antigen among the three fourth-generation HIV combination assays. The three fourth-generation HIV Ag/Ab combination assays exhibited better sensitivity, specificity, and negative prediction than the third-generation HIV Ab immunoassay. Copyright © 2015. Published by Elsevier B.V.

  4. EVALUATION OF AN O-ANTIGEN ELISA FOR SCREENING CATTLE HERDS FOR SALMONELLA-TYPHIMURIUM

    DEFF Research Database (Denmark)

    Hoorfar, Jeffrey; Bitsch, V.

    1995-01-01

    A total of 2585 serum samples from 62 dairy herds located in four different regions of Denmark were tested in an O-antigen (0:1,4,5,12)-based ELISA for the detection of antibodies against Salmonella typhimurium. Ten closed herds from an island with no reported occurrence of salmonellosis...... the salmonellosis-free island were ELISA-negative, and all but one of the 12 S typhimurium-infected herds were ELISA-positive, which resulted in a herd test sensitivity of 0.92 and herd test specificity of 1.0. Eleven of the 12 S typhimurium-infected herds were negative in a blocking ELISA based on a monoclonal...... antibody to the 0:9 antigen of the serogroup D salmonellas, indicating the possibility of rapid serogroup-specific screening of herds by means of these two tests. Ten other randomly selected herds with clinical outbreaks of S dublin were all, to a large extent, positive in the 0:1,4,5,12-ELISA, whereas a S...

  5. The use of an in vitro microneutralization assay to evaluate the potential of recombinant VP5 protein as an antigen for vaccinating against Grass carp reovirus

    Directory of Open Access Journals (Sweden)

    Xu Dan

    2011-03-01

    Full Text Available Abstract Background Grass carp reovirus (GCRV is the causative pathogen of grass carp hemorrhagic disease, one of the major diseases damaging grass carp Ctenopharyngon idellus breeding industry in China. Prevention and control of the disease is impeded largely due to the lack of research in economic subunit vaccine development. This study aimed to evaluate the potential of viral outer shell protein VP5 as subunit vaccine. Methods The vp5 gene was isolated from the viral genome through RT-PCR and genetically engineered to express the recombinant VP5 protein in E coli. The viral origin of the recombinant protein was confirmed by Western blot analysis with a monoclonal antibody against viral VP5 protein. Polyclonal antibody against the recombinant VP5 protein was prepared from mice. A microneutralization assay was developed to test its neutralizing ability against GCRV infection in cell culture. Results The GST-VP5 fusion protein (rVP5 was produced from E. Coli with expected molecular weight of 90 kDa. The protein was purified and employed to prepare anti-VP5 polyclonal antibody from mice. The anti-VP5 antibody was found to neutralize GCRV through in vitro microneutralization assay and viral progeny quantification analysis. Conclusions The present study showed that the viral VP5 protein was involved in viral infection and bacterially-expressed VP5 could be suitable for developing subunit vaccine for the control of GCRV infection.

  6. Viral vector-based influenza vaccines

    Science.gov (United States)

    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

  7. Evaluation of Serum Specific Antibody against Recombinant ESAT-6 Antigen in Patients with Tuberculosis and Comparing to Normal Controls

    Directory of Open Access Journals (Sweden)

    Homeira Izadi

    2017-02-01

    Full Text Available Background & Objective: Tuberculosis (TB is a zoonotic disease which is caused by Mycobacterium tuberculosis. Because of common structural and secretory antigens between pathogen and nonpathogenic mycobacterium, the specific diagnosis of TB is difficult. Therefore, it is very important to find a new method with high specificity and sensitivity for accurate and rapid diagnosis of tuberculosis. In this study, the serodiagnostic potential of Mycobacterium tuberculosis recombinant ESAT-6 in TB infected patients was evaluated by Enzyme Linked Immunosorbent Assay (ELISA. Materials & Methods: 55 TB patients with active disease and 28 healthy controls have been collected and evaluated in different dilutions in ELISA methods for the presence of specific anti-ESAT-6 antibody. The specificity and the sensitivity of this method was compared with the culture test. Results: TB patients have high levels of specific antibody against ESAT-6 antigens. The specificity and the sensitivity of this method was calculated as 80.90% and 85.45%, respectively. Conclusion: These findings provide useful information on the importance of ESAT-6 protein and suggested this serologic test as a good alternative method for rapid and prefect diagnosis of tuberculosis.

  8. Deteccion de citomegalovirus mediante la tecnica de inmunoperoxidasa y aislamiento viral Cytomegalovirus detection by Immunoperoxidase assay and viral isolation

    Directory of Open Access Journals (Sweden)

    Maritza Alvarez

    1991-06-01

    Full Text Available En el presente estudio se comparó la técnica de inmunoperoxidasa para la detección de citomegalovirus (IPCMV utilizando anticuerpos monoclonales que reconocen proteínas precoces virales con el método convencional de aislamiento viral en fibroblastos humanos. Un total de 150 muestras de orina fueron examinadas encontrando una sensibilidad de un 89.8% y una especificidad de 91.3% de la técnica de IPCMV comparada con el aislamiento viral. Una de las ventajas que presentó la IPCMV fue la rapidez con que fueron obtenidos los resultados (48 horas mientras que el aislamiento viral fue como promedio 14 días.An Immunoperoxidase assay was applied to detect early antigens of Cytomegalovirus (CMV in 150 urine samples from immunocompromised patients, using the commercial available monoclonal antibody against CMV El3. The detection of early antigen by IP (IPCMV is compared to the conventional cell culture isolation regarding specificity and sensitivity in order to evaluate is usefulness in the diagnostic of CMV infections. The IPCMV showed a sensitivity of 89.8% and a specificity of 91.3% when compared to the isolation method. The great advantage of the IPCMV is based on the shorter time results are achieved, since 48-72 Hs can be enough to provide evidence of CMV infection, while in the isolation technique cytopatho-genic effect was present around 14 days after sample inoculation.

  9. Viral Hepatitis

    Science.gov (United States)

    ... Us FAQs Ask a Question Toll Free Numbers Homeless Veterans Chat VA » Health Care » Viral Hepatitis » Veterans and ... Vet Centers) War Related Illness & Injury Study Center Homeless Veterans Returning Service Members Rural Veterans Seniors & Aging Veterans ...

  10. An evaluation of circulating bovine viral diarrhea virus type 2 maternal antibody level and response to vaccination in Angus calves.

    Science.gov (United States)

    Downey, E D; Tait, R G; Mayes, M S; Park, C A; Ridpath, J F; Garrick, D J; Reecy, J M

    2013-09-01

    Vaccination against viruses has been shown to help prevent bovine respiratory disease in cattle. However, both passively acquired maternal antibody concentration and calf age have been shown to impact the ability of the immune system of a calf to respond to vaccination. The objectives of this study were to identify and evaluate environmental and management factors that affect 1) passively acquired bovine viral diarrhea virus (BVDV) type 2 antibody level, 2) decay rate of passively acquired BVDV type 2 antibody level, and 3) responses to BVDV type 2 vaccinations. A 2-shot modified live vaccine was administered to 1,004 Angus calves that were weaned at either the initial vaccination (n = 508) or the booster vaccination (n = 496). Calves weaned at the initial vaccination averaged 139 d whereas calves weaned at booster vaccination averaged 128 d of age. Bovine viral diarrhea virus type 2 antibodies were measured in 3 approximately 21-d intervals, serially collected serum samples to quantify antibody levels at initiation and end of vaccination protocol in addition to responses to initial, booster, and overall vaccination protocol. Amount of passively transferred antibody in the calf increased as dam age increased from 2 to 6 yr (P 0.05). Calf age nested within birth year-season and dam age affected both initial and final antibody level, initial response, booster response, and overall antibody response to vaccination. The level of circulating, passively acquired maternal antibodies present at the time of vaccination had a significant (P calf to mount an overall antibody response to vaccination, maternal antibodies in circulation need to be less than 3.12 titers. However, the age at which a calf reached this antibody threshold was dependent on dam age. This information will help cattle managers and consultants design vaccination protocols to successfully mount an antibody response to vaccination.

  11. Evaluation of Nicotiana tabacum plants transformed for the expression of verocytotoxic Escherichia coli antigens.

    Directory of Open Access Journals (Sweden)

    Angela Lombardi

    2015-07-01

    Full Text Available Two transgenic Nicotiana tabacum plants, carrying respectively the F18 adhesive fimbriae and the B subunit of verocytotoxin genes from O138 Verocytotoxic E.coli serotype were developed as a model of edible vaccine. Tobacco plants were transformed by agroinfection according to Rossi et al. (2013 stably.  The F18 adhesive fimbriae and VT2e B-subunit were placed under control of the GLOB promoter for the seed-specific protein expression. Agrobacterium tumefaciens binary vector system is an efficient tool to transform plant cells; however, the exogenous gene integrates at semi-random into the nuclear chromosome. PCR products, using specific oligonucleotides putatively encoding the B-subunit of VT2e-B and F18 fimbriae were identified on agarose gel (1.5% - 0.9% as bands with a length of 270 and 519 base pairs, respectively. We showed that the foreign VT2e-B and F18 fimbriae genes were stably integrated into the tobacco genome. Northern blot and Western blot analyses carried out respectively on total mRNA and total soluble protein extract obtained from seeds. For each line, the obtained amount of antigens is sufficient for subsequent oral immunization trials. Three lines of tobacco seeds (F18, VT2e-B, and WT were seeded in homogeneous conditions and were harvested simultaneously. Tobacco plants were analysed also by optical and electronic microscope in different phases of growth. Germination of transgenic seeds were delayed of three/five days compared to WT in two replicated experiments, suggesting that genetic manipulation may influenced mechanisms leading to germination. In conclusion the genes coding for VT2e-B and the F18 are stably maintained in the seeds and obtained tobacco seeds represent a valid strategy to ferry antigenic proteins to the gut and a promising non-invasive method of vaccination in pig industry.

  12. Evaluation of antigens stability of tobacco seeds as edible vaccine against VTEC strains

    Directory of Open Access Journals (Sweden)

    Luciana Rossi

    2015-11-01

    Full Text Available Plants have represent a promising alternative for biopharmaceutical proteins (Ma et al., 2003; Rossi et al., 2014. Many plant based edible vaccines have been shown to be effective in inducing local immune responses (Rossi et al., 2013. Edible vaccines can activate both mucosal and systemic immunity, as they come in contact with the digestive tract lining. This dual effect would provide first-line defense against pathogens invading through the mucosa. The antigens are released in the intestines are taken up by M cells that are present over the Payer’s patches (in the ileum and the gut associated lymphoid tissue (GALT. Edible vaccines represent an important worldwide goal for the prevention of the enteric diseases, also in livestock. In particular, the enteric infections are a significant clinical problem in pigs. Verocytotoxic Escherichia (E. coli strains are responsible for serious enterotoxaemia that causes important economic losses in the pig industry. The production of a vaccine for oral administration of transgenic seeds could be a practical and efficient system to prevent the infection and to reduce the antibiotic use. This study was focused on tobacco plants, previously transformed by agroinfection for the seed-specific expression of antigenic proteins (F18 adhesive fimbriae and the B subunit of the Vt2e toxin as model of edible vaccines against verocytotoxic E. coli strains. The dietary administration of transgenic tobacco seeds promotes a significant increase in the number of mucosal IgA-producing cells of the tunica propria in both small and large intestine in mice (Rossi et al., 2013. A protective effect of oral administration of transgenic tobacco seeds was also observed against verocytotoxic Escherichia coli infection in piglets (Rossi et al., 2014. The aim of this study was to assess the seed-expression stability, that is a important requirement in the vaccine production, of F 18 and Vt2e-B heterologous genes into the progeny of

  13. Quantitative evaluation of viral fitness due to a single nucleotide polymorphism in the Marek's disease virus UL41 gene via an in vitro competition assay.

    Science.gov (United States)

    Mao, Weifeng; Niikura, Masahiro; Silva, Robert F; Cheng, Hans H

    2008-03-01

    Marek's disease, a T cell lymphoma, is an economically important disease of poultry caused by the Marek's disease virus (MDV), a highly cell-associated alphaherpesvirus. A greater understanding of viral gene function and the contribution of sequence variation to virulence should facilitate efforts to control Marek's disease in chickens. To characterize a naturally occurring single nucleotide polymorphism (SNP; AY510475:g.108,206C>T) in the MDV UL41 gene that results in a missense mutation (AAS01683:p.Arg377Cys), bacterial artificial chromosome (BAC)-derived MDVs that differed only in the UL41 SNP were evaluated using a head-to-head competition assay in vitro. Monitoring the frequency of each SNP by pyrosequencing during virus passage determined the ratio of each viral genome in a single monolayer, which is a very sensitive method to monitor viral fitness. MDV with the UL41*Cys allele showed enhanced fitness in vitro. To evaluate the mechanism of altered viral fitness caused by this SNP, the virion-associated host shutoff (vhs) activity of both UL41 alleles was determined. The UL41*Cys allele had no vhs activity, which suggests that enhanced fitness in vitro for MDV with inactive vhs was due to reduced degradation of viral transcripts. The in vitro competition assay should be applicable to other MDV genes and mutations.

  14. Economic and clinical effects of evaluating rapid viral response to peginterferon alfa-2b plus ribavirin for the initial treatment of chronic hepatitis C.

    Science.gov (United States)

    Wong, John B; Davis, Gary L; McHutchison, John G; Manns, Michael P; Albrecht, Janice K

    2003-11-01

    Evaluation of 12-wk viral response to initial antiviral therapy for chronic hepatitis C has been recommended to minimize antiviral-associated morbidity and costs. The aim of this study was to examine the economic and clinical effects of evaluating rapid viral response during antiviral therapy for treatment naive chronic hepatitis C patients. We applied viral response and drug dosage from an international randomized clinical trial of ribavirin plus peginterferon alfa-2b or ribavirin plus interferon alfa-2b to a previously published computer cohort simulation to project lifelong clinical and economic outcomes. Natural history and economic estimates were based on published literature, expert panel estimates, and actual variable and reimbursement cost data. The assessment of 12-wk rapid viral response reduced antiviral treatment duration by 40-44% and antiviral costs by 44-45% (savings of $15,116-16,268 for peginterferon plus ribavirin and $8300 for interferon plus ribavirin) compared to full 48-wk dosing. With the 12-wk evaluation, the marginal cost-effectiveness of peginterferon plus ribavirin versus interferon plus ribavirin was $13,600-22,800 compared with $14,600-25,000 per discounted quality adjusted life-year gained with the 24-wk evaluation. For genotype 1, hepatitis C infected patients, 12-wk testing for peginterferon plus ribavirin remaining preferred and cost-effective compared with interferon plus ribavirin. For genotype 2 or 3, hepatitis C infected patients, 12-wk testing yielded similar results to those of 24-wk treatment. Assessment of 12-wk viral response in genotype 1, hepatitis C infected patients should reduce peginterferon plus ribavirin morbidity and costs and improve its cost-effectiveness; however, for genotype 2 and 3, hepatitis C infected patients, 12-wk testing and 24-wk treatment have similar outcomes. Decisions regarding continuation of antiviral treatment should also consider the variability in the accuracy of quantitative viral assays as

  15. A simple, rapid, and sensitive system for the evaluation of anti-viral drugs in rats

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xiaoguang [Tohoku University Graduate School of Medicine, Department of Internal Medicine/Division of Emerging Infectious Diseases, Sendai 980-8575 (Japan); Department of Medical Microbiology, Harbin Medical University, Harbin 150086 (China); Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811 (Japan); Qian, Hua [Tohoku University Graduate School of Medicine, Department of Internal Medicine/Division of Emerging Infectious Diseases, Sendai 980-8575 (Japan); Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811 (Japan); Miyamoto, Fusako [Tohoku University Graduate School of Medicine, Department of Internal Medicine/Division of Emerging Infectious Diseases, Sendai 980-8575 (Japan); Naito, Takeshi [Laboratory of Virus Control, Institute for Virus Research, Kyoto University, 53 Kawaramachi, Shogoin, Sakyo-ku, Kyoto 606-8507 (Japan); Kawaji, Kumi [Tohoku University Graduate School of Medicine, Department of Internal Medicine/Division of Emerging Infectious Diseases, Sendai 980-8575 (Japan); Kajiwara, Kazumi [Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto 606-8501 (Japan); JST Innovation Plaza Kyoto, Japan Science and Technology Agency, Nishigyo-ku, Kyoto 615-8245 (Japan); Hattori, Toshio [Tohoku University Graduate School of Medicine, Department of Internal Medicine/Division of Emerging Infectious Diseases, Sendai 980-8575 (Japan); Matsuoka, Masao [Laboratory of Virus Control, Institute for Virus Research, Kyoto University, 53 Kawaramachi, Shogoin, Sakyo-ku, Kyoto 606-8507 (Japan); Watanabe, Kentaro; Oishi, Shinya; Fujii, Nobutaka [Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto 606-8501 (Japan); and others

    2012-07-27

    Highlights: Black-Right-Pointing-Pointer We established a novel, simple and rapid in vivo system for evaluation of anti-HIV-1 drugs with rats. Black-Right-Pointing-Pointer The system may be applicable for other antiviral drugs, and/or useful for initial screening in vivo. Black-Right-Pointing-Pointer In this system, TRI-1144 displayed the most potent anti-HIV-1 activity in vivo. -- Abstract: The lack of small animal models for the evaluation of anti-human immunodeficiency virus type 1 (HIV-1) agents hampers drug development. Here, we describe the establishment of a simple and rapid evaluation system in a rat model without animal infection facilities. After intraperitoneal administration of test drugs to rats, antiviral activity in the sera was examined by the MAGI assay. Recently developed inhibitors for HIV-1 entry, two CXCR4 antagonists, TF14016 and FC131, and four fusion inhibitors, T-20, T-20EK, SC29EK, and TRI-1144, were evaluated using HIV-1{sub IIIB} and HIV-1{sub BaL} as representative CXCR4- and CCR5-tropic HIV-1 strains, respectively. CXCR4 antagonists were shown to only possess anti-HIV-1{sub IIIB} activity, whereas fusion inhibitors showed both anti-HIV-1{sub IIIB} and anti-HIV-1{sub BaL} activities in rat sera. These results indicate that test drugs were successfully processed into the rat sera and could be detected by the MAGI assay. In this system, TRI-1144 showed the most potent and sustained antiviral activity. Sera from animals not administered drugs showed substantial anti-HIV-1 activity, indicating that relatively high dose or activity of the test drugs might be needed. In conclusion, the novel rat system established here, 'phenotypic drug evaluation', may be applicable for the evaluation of various antiviral drugs in vivo.

  16. Evaluation of Diagnostic Value in Using a Panel of Multiple Tumor-Associated Antigens for Immunodiagnosis of Cancer

    Directory of Open Access Journals (Sweden)

    Peng Wang

    2014-01-01

    Full Text Available To determine whether a panel of multiple tumor-associated antigens (TAAs would enhance antibody detection, the diagnostic value of autoantibodies to a panel of multiple TAAs in cancer has been evaluated. The TAAs used in this study was composed of eight TAAs including Imp1, p62, Koc, p53, C-myc, Cyclin B1, Survivin, and p16 full-length recombinant proteins. Enzyme-linked immunosorbent assay and immunoblotting were used to detect antibodies in 304 cancer sera and also 58 sera from normal individuals. The antibody frequency to any individual TAA in cancer was variable but rarely exceeded 20%. With the successive addition of TAAs to a final combination of total of eight antigens, there was a stepwise increase of positive antibody reactions reaching a sensitivity of 63.5% and a specificity of 86.2% in the combined cancer group. In different types of cancer, the ranges of positive and negative likelihood ratio were 4.07–4.76 and 0.39–0.51, respectively, and the ranges of positive and negative predictive values were 74.2–88.7% and 58.8–75.8%, respectively. Agreement rate and Kappa value were 67.1% and 0.51, respectively. These results further support our previous hypothesis that detection of anti-TAAs autoantibodies for diagnosis of certain type of cancer can be enhanced by using a miniarray of several TAAs.

  17. Evaluation of the BioFire FilmArray respiratory panel and the GenMark eSensor respiratory viral panel on lower respiratory tract specimens.

    Science.gov (United States)

    Ruggiero, Phyllis; McMillen, Tracy; Tang, Yi-Wei; Babady, N Esther

    2014-01-01

    We evaluated the performance characteristics of the FilmArray respiratory panel and the eSensor respiratory viral panel on clinical and spiked lower respiratory tract specimens (LRTS). The overall agreement between the two methods was 89.5% (51/57). The lower limit of detection of both assays for all targets in LRTS was comparable to that for nasopharyngeal swab specimens.

  18. Preclinical evaluation of multi antigenic HCV DNA vaccine for the prevention of Hepatitis C virus infection.

    Science.gov (United States)

    Lee, Hyojin; Jeong, Moonsup; Oh, Jooyeon; Cho, Youngran; Shen, Xuefei; Stone, John; Yan, Jian; Rothkopf, Zachary; Khan, Amir S; Cho, Byung Mun; Park, Young K; Weiner, David B; Son, Woo-Chan; Maslow, Joel N

    2017-03-07

    Direct-acting antiviral treatment for hepatitis C virus (HCV) infection is costly and does not protect from re-infection. For human and chimpanzees, recovery from acute HCV infection correlates with host CD4+ and CD8+ T cell responses. DNA plasmids targeting the HCV non-structural antigens NS3, NS4, and NS5, were previously reported to induce robust and sustained T cell responses in mice and primates. These plasmids were combined with a plasmid encoding cytokine IL-28B, together named as VGX-6150. The dose-dependent T cell response and safety of VGX-6150 administered intramuscularly and followed by electroporation was assessed in mice. Immune responses plateaued at 20 μg/dose with IL-28B demonstrating significant immunoadjuvant activity. Mice administered VGX-6150 at 40, 400, and 800 μg given either as a single injection or as 14 injections given bi-weekly over 26 weeks showed no vaccine related changes in any clinical parameter compared to placebo recipients. There was no evidence of VGX-6150 accumulation at the injection site or in any organ 1 month following the 14th vaccination. Based on these studies, the approximate lethal dose (ALD) exceeds 800 μg/dose and the NOAEL was 800 μg/dose in mouse. In conclusion, VGX-6150 appears safe and a promising preventive vaccine candidate for HCV infection.

  19. Evaluating Quantum Dot Performance in Homogeneous FRET Immunoassays for Prostate Specific Antigen.

    Science.gov (United States)

    Bhuckory, Shashi; Lefebvre, Olivier; Qiu, Xue; Wegner, Karl David; Hildebrandt, Niko

    2016-02-04

    The integration of semiconductor quantum dots (QDs) into homogeneous Förster resonance energy transfer (FRET) immunoassay kits for clinical diagnostics can provide significant advantages concerning multiplexing and sensitivity. Here we present a facile and functional QD-antibody conjugation method using three commercially available QDs with different photoluminescence (PL) maxima (605 nm, 655 nm, and 705 nm). The QD-antibody conjugates were successfully applied for FRET immunoassays against prostate specific antigen (PSA) in 50 µL serum samples using Lumi4-Tb (Tb) antibody conjugates as FRET donors and time-gated PL detection on a KRYPTOR clinical plate reader. Förster distance and Tb donor background PL were directly related to the analytical sensitivity for PSA, which resulted in the lowest limits of detection for Tb-QD705 (2 nM), followed by Tb-QD655 (4 nM), and Tb-QD605 (23 nM). Duplexed PSA detection using the Tb-QD655 and Tb-QD705 FRET-pairs demonstrated the multiplexing ability of our immunoassays. Our results show that FRET based on QD acceptors is suitable for multiplexed and sensitive biomarker detection in clinical diagnostics.

  20. Evaluation of the matrix effect of thermophilic anaerobic digestion on inactivation of infectious laryngotracheitis virus using real-time PCR and viral cell culture.

    Science.gov (United States)

    Gao, Tiejun; Bowlby, Evelyn; Tong, Yupin; Wu, John T Y; Wong, Lester; Tower, Robert J; Pang, Xiaoli; Li, Xiaomei

    2012-04-01

    The matrix effect of the thermophilic anaerobic digestion (TAD) process on inactivation of infectious laryngotracheitis virus (ILTV) was evaluated. Viral cell culture and real-time PCR were used for assessing removal of the viral infectivity and degradation of viral DNA, respectively. Results showed that the TAD-derived matrix alone can inactivate the virus and destroy the nucleic acid helix core of ILTV in a time-and- dose-dependent manner. No cytopathogenic effect (CPE) was observed in the cells exposed to ILTV pre-treated with TAD matrix for 1.5h in experiment 1 and for 16h in experiment 2. There was a significant statistical difference between TAD matrix treated and non-treated cultures (p<0.001, Chi-test). Amplifiable ILT viral DNA was reduced 2.27 log by 1.5h-treatment and was not present by 16h-treatment with TAD matrix, indicating complete viral DNA fragmentation. The TAD process is an environmentally friendly way for disposing of poultry biowaste and carcasses. Copyright © 2012 Elsevier Ltd. All rights reserved.

  1. Evaluation of neutralization patterns of the five unique Argentine equine arteritis virus field strains reported Evaluación de los patrones de neutralización de las únicas cinco cepas argentinas descritas de arteritis viral equina

    Directory of Open Access Journals (Sweden)

    M. G. Echeverría

    2010-02-01

    Full Text Available Equine viral arteritis (EVA is a contagious viral disease that frequently causes mild or subclinical infections in adult horses. Only one EAV serotype has been described. However, there are differences in antigenicity, pathogenicity and neutralization characteristics of virus field strains. The interaction of two viral proteins, GP5 and M, is critical for infectivity and amino acid changes in the GP5 sequences have an effect on the neutralizing phenotype, regardless the effects of other viral proteins. The objective of the present study was to evaluate the neutralization phenotypes of the 5 unique Argentine EAV strains reported and to compare them with the neutralization phenotypes of the EAV-UCD reference strain, with special emphasis on the analysis of M and GP5 proteins. The strains had a similar neutralization phenotype pattern when anti-EAV serum, derived from EAV seropositive horses, was used in the analysis. Meanwhile, low titers were observed when equine polyclonal anti-EAV reference sera were used in the assay. Argentine strains have almost the same amino acid substitutions, with the exception of LP01 strain, that mainly involves the first variable region V1, especially in neutralization sites B and C. However, they are fairly different from the EAV-UCD strain. Nevertheless, the nucleotide and amino acid differences observed among the Argentine strains LP02/R, LP02/C, LP02/P and LP-LT-ARG did not show any variations in the neutralization phenotype.La arteritis viral equina (AVE ocasiona infecciones, en su mayoría subclínicas, pero puede causar abortos y enfermedad respiratoria. Si bien se ha descrito un solo serotipo de AVE, existen diferencias en cuanto a la antigenicidad, patogenicidad y patrones de neutralización en las cepas de campo. Los ORF5 y ORF6 del virus codifican las proteínas de envoltura GP5 y M; la interacción entre estas proteínas es crítica para la infectividad. Los cambios en las secuencias de aminoácidos en la

  2. Evaluation of the efficacy of a recombinant Entamoeba histolytica cysteine proteinase gene (EhCP5) antigen in Minipig.

    Science.gov (United States)

    He, Guang-Zhi; Deng, Shu-Xuan; Tian, Wei-Yi; Feng, Yong

    2012-03-01

    Entamoeba histolytica cysteine proteinase gene 5(EhCP5) is one of the major proteinase genes of all EhCP-transcripts. The amebiasis cysteine proteinase gene encoding an antigen from E. histolytica, as well as the recombinant EhCP5, obtained by cloning and expression of the EhCP5 gene in heterologous host Escherichia coli BL-21 (DE3), were used to evaluate their ability to induce immune protective responses in Minipig against challenge infection in a minipig-E. histolytica model. There was a 52.27% reduction (P<0.001) in the group of recovery of challenged E. histolytica compared with that in the control group. Specific anti-EhCP5 antibodies from immune protected minipig had significantly higher levels of immunoglobulin G (IgG) (P<0.0001). Our data will help to know the mechanism of vaccinal protection of E. histolytica. Copyright © 2011 Elsevier Inc. All rights reserved.

  3. Effect of HCV Core Antigen and RNA Clearance during Therapy with Direct Acting Antivirals on Hepatic Stiffness Measured with Shear Wave Elastography in Patients with Chronic Viral Hepatitis C

    Directory of Open Access Journals (Sweden)

    Mariusz Łucejko

    2018-01-01

    Full Text Available To assess a combination of novel measures of therapeutic success in the treatment of chronic hepatitis C (CHC infection, we evaluated liver stiffness (LS with shear wave elastography and hepatitis C virus core antigen (HCVcAg concentrations. We followed 34 patients during and after treatment with direct acting antivirals. All patients achieved a sustained virologic and serologic response and a significant increase of albumin levels. Decreases of alanine aminotransferase (ALT activity and alpha-fetoprotein (AFP level were observed during the treatment and follow-up period. A significant decrease in LS was observed between baseline, end of treatment (EOT, and at 24- and 96-week post-treatment follow-up. LS decline between EOT and 96-week follow-up (FU96 was observed in 79% of patients. Significant LS changes were seen in patients with advanced fibrosis, particularly in cirrhotics and in patients with ALT exceeding 100 IU/mL. There was a positive correlation between ALT activity and LS changes at the baseline versus FU96. A negative correlation was demonstrated between individual HCVcAg baseline concentrations and reduction of LS at the baseline versus FU96. In conclusion, we observed that LS significantly declined during and after antiviral treatment. It was accompanied by improvement in some liver function measures, and disappearance of both HCVcAg and HCV ribonucleic acid (HCV RNA.

  4. Evaluation of whole blood IFNgamma test using PPD and recombinant antigen challenge for diagnosis of pulmonary and extra-pulmonary tuberculosis.

    Science.gov (United States)

    Kalantri, Yatiraj; Hemvani, Nanda; Chitnis, D S

    2009-06-01

    Quantiferon TB gold (QFT-G) with recombinant antigen cocktail is well evaluated for diagnosis of pulmonary tuberculosis (PTB). However, diagnosis of extra-pulmonary tuberculosis (EPTB) is more difficult due to limitations of conventional techniques. This study compares recombinant antigens based QFT-G and low cost PPD based interferon test for the diagnosis of PTB and EPTB. IFNgamma release, with recombinant antigens and PPD, was assayed by ELISA from 140 cases of EPTB, 100 cases of PTB along with acid fast bacillus (AFB) detection, AFB culture on LJ and MGIT BACTEC. Sensitivity and specificity for QFT-G recombinant antigens was 84.29% and 96%, while for PPD based interferon was 70% and 84% for EPTB group. The sensitivity was far superior to AFB smear and culture for both the antigens. Nine samples were identified as non-tubercular mycobacteria (NTM) in the EPTB group and all were negative for QFT-G, but six of them were positive for PPD based test. Results of the study show that QFT-G using recombinant antigen is sensitive and specific for both PTB and EPTB diagnosis. The PPD based test is economic and offers comparable performance for PTB and EPTB diagnosis and also useful for diagnosis of NTM.

  5. Comparative evaluation of hepatitis B surface antigen-loaded elastic liposomes and ethosomes for human dendritic cell uptake and immune response.

    Science.gov (United States)

    Mishra, Dinesh; Mishra, Pradyumna Kumar; Dabadghao, Sunil; Dubey, Vaibhav; Nahar, Manoj; Jain, Narendra K

    2010-02-01

    The aim of the present study was to evaluate two vesicular carrier systems, ethosomes and elastic liposomes loaded with hepatitis B surface antigen, for in vitro qualitative and quantitative uptake by human dendritic cells (DCs) and ability to stimulate T lymphocytes. Quantitative uptake of antigen-loaded carriers was documented by flow cytometry, and internalization of the systems by the DCs was studied using spectral bioimaging. Ability of antigen-pulsed DCs to stimulate autologous peripheral blood lymphocytes and levels of TH1/TH2 cytokines were also examined using flow cytometry. Both vesicular carrier systems as antigen delivery modules and DCs as antigen-presenting cells were able to generate a protective immune response. However, ethosomes were found to have higher internalizing ability and immunogenicity in comparison with elastic liposomes. These properties of ethosomes coupled with their skin-navigating potential, make it an attractive vehicle for development of a transcutaneous vaccine against hepatitis B in preference to elastic liposomes. Two carrier systems for more potent vaccine administration - ethosomes and elastic liposomes loaded with hepatitis B surface antigen - are compared. Ethosomes demonstrated higher internalizing ability and immunogenicity. Due to their known skin-navigating potential, ethosomes may represent an attractive vehicle for development of a transcutaneous vaccine against hepatitis B. Copyright 2010 Elsevier Inc. All rights reserved.

  6. Experimental infection with the Paderborn isolate of classical swine fever virus in 10-week-old pigs: determination of viral replication kinetics by quantitative RT-PCR, virus isolation and antigen ELISA

    DEFF Research Database (Denmark)

    Uttenthal, Åse; Storgaard, Torben; Oleksiewicz, M.B.

    2003-01-01

    We performed experimental infection in 10-week-old pigs with the Paderborn isolate of classical swine fever virus (CSFV). Despite being epidemiologically linked to the major CSFV outbreak in The Netherlands in 1997, the in vivo replication kinetics of this isolate have to our knowledge not been...... described in detail previously. We found that oronasal infection with 10(4.7) TCID50 produced mortality in three out of five pigs after 29-31 days, and severe clinical symptoms in one out of five pigs, while one out of five pigs exhibited no clinical symptoms. At this infection dose, pigs had viral RNA...... viral RNA in serum for more than 30 days, and exhibited only mild clinical symptoms. We observed an excellent correlation between clinical symptoms and viral RNA loads in serum, while serum antibody levels were low. Clinically affected pigs had up to 1000-fold higher serum viral RNA loads than did pigs...

  7. Evaluation of homeopathy in broiler chickens exposed to live viral vaccines and administered Calendula officinalis extract.

    Science.gov (United States)

    Barbour, Elie K; Sagherian, Vatché; Talhouk, Salma; Talhouk, Rabih; Farran, Mohamad T; Sleiman, Fawwak T; Harakeh, Steve

    2004-08-01

    In this study it was determined that a Calendula officinalis water extract can reduce the immune response to three different viruses in broiler chickens, associated with improvement in body weights. The experiment was conducted on broiler chickens divided into two groups of 105 birds each. The first group received a Calendula officinalis water extract orally, while the second group received drinking water only. All birds in the two groups were similarly exposed to three different live vaccine viruses. Quantitative assessment of humoral immunity to each of the 3 viruses and records of bursal and thymus weight indices were taken. Performance, as observed in weight records at 21 and 41 days of age, feed conversion, and% mortality up to market age, was also evaluated. There was a reduction in immune response to IB virus at 42 days of age, to ND virus at 29 and 42 days of age, and to IBD virus at 14, 29, and 42 days of age in the Calendula officinals-treated birds in comparison with controls. This immune reduction in Calendula officinalis-treated birds was associated with insignificant reduction in the bursal weight index at 42 days of age and an improvement in mean weights at 21 and 41 days of age; the feed conversion and mortality rates were similar in the two groups (P>0.05). Calendula officinalis had an immunomodulation effect against three different live viruses in broiler chickens.

  8. Evaluation of the Protective Effect of Deoxyribonucleic Acid Vaccines Encoding Granule Antigen 2 and 5 Against Acute Toxoplasmosis in BALB/c Mice.

    Science.gov (United States)

    Ching, Xiao Teng; Fong, Mun Yik; Lau, Yee Ling

    2017-06-01

    AbstractToxoplasma gondii infects a broad range of warm-blooded hosts, including humans. Important clinical manifestations include encephalitis in immunocompromised patients as well as miscarriage and fetal damage during early pregnancy. Toxoplasma gondii dense granule antigen 2 and 5 (GRA2 and GRA5) are essential for parasitophorous vacuole development of the parasite. To evaluate the potential of GRA2 and GRA5 as recombinant DNA vaccine candidates, these antigens were cloned into eukaryotic expression vector (pcDNA 3.1C) and evaluated in vaccination experiments. Recombinant DNA vaccines constructed with genes encoding GRAs were validated in Chinese hamster ovary cells before evaluation using lethal challenge of the virulent T. gondii RH strain in BALB/c mice. The DNA vaccines of pcGRA2 and pcGRA5 elicited cellular-mediated immune response with significantly higher levels of interferon-gamma, interleukin-2 (IL-2), IL-4, and IL-10 (P < 0.05) compared with controls. A mixed T-helper cell 1 (Th1)/Th2 response was associated with slightly prolonged survival. These findings provide evidence that DNA vaccination with GRA2 and GRA5 is associated with Th1-like cell-mediated immune responses. It will be worthwhile to construct recombinant multiantigen combining full-length GRA2 or/and GRA5 with various antigenic proteins such as the surface antigens and rhoptry antigens to improve vaccination efficacy.

  9. Viral Gastroenteritis

    Science.gov (United States)

    ... help relieve the symptoms of viral gastroenteritis in adults: drinking plenty of liquids such as fruit juices, sports ... as the child is hungry giving infants breast milk or full strength ... solutions Older adults and adults with weak immune systems should also ...

  10. [Viral superantigens].

    Science.gov (United States)

    Us, Dürdal

    2016-07-01

    Superantigens (SAgs) are microbial proteins produced by various microorganisms that elicit excessive and strong stimulation of T cells via an unconventional mechanism. They cause polyclonal activation of T cells in a non-specific manner, by binding to a particular variable-beta (Vβ) chain of T-cell receptor (TCR) and MHC class II molecule, in unprocessed form and outside of peptide-binding cleft, forming a bridge between the antigen presenting cell and the T cell. SAgs are classified into three groups, namely 1) exogenous (soluble proteins and exotoxins secreted by microorganisms), 2) endogenous (transmembrane proteins encoded by viruses which are integrated into the genome) and 3) B-cell SAgs (proteins which stimulate predominantly B cells). The best characterized and mostly studied SAgs are staphylococcal and streptococcal exotoxins, however it is well-known that many other microorganisms also possess SAg activities. Despite the presence of several viruses that cause severe infections in humans, the number of viruses that have proteins identified with SAg property in their pathogenesis, is relatively low. To date, the defined viruses that encoded SAgs are as follows; mouse mammary tumor virus (MMTV) (Marrack, et al. 1991), rabies virus (Lafon, et al. 1992), Epstein-Barr virus (EBV) (Sutkowski, et al. 1996), human endogenous retrovirus (HERV) (Conrad, et al. 1997), human immunodeficiency virus (HIV) (Posnett, et al. 1995; Torres, et al. 1996; Townsley-Fuchs, et al. 1997) and Ebola virus (Leroy, et al. 2011). SAgs were first described in the MMTV, a polymorphic B-type retrovirus that is either contained in the genome as an endogenous provirus (germline transmission) or exogenous infectious virus that transmits vertically via breast milk. Both MMTV forms encode SAgs. The SAg-mediated massive T cell activation is required for the spread of exogenous MMTV from intestines to mammary glands, facilitating the transmission of infectious virus. On the other hand

  11. Evaluation of recombinant porin (rOmp2a) protein as a potential antigen candidate for serodiagnosis of Human Brucellosis.

    Science.gov (United States)

    Pathak, Prachi; Kumar, Ashu; Thavaselvam, Duraipandian

    2017-07-11

    Brucellosis is an important zoonotic disease caused by different Brucella species and human brucellosis is commonly prevalent in different states of India. Among various Brucella species, B. melitensis is most pathogenic to human and included as category B biothreat which can cause infection through aerosol, cut, wounds in skin and contact with infected animals. The diagnosis of human brucellosis is very important for proper treatment and management of disease as there is no vaccine available for human use. The present study was designed to clone, express and purify immunodominant recombinant omp2a (rOmp2a) porin protein of B. melitensis and to evaluate this new antigen candidate for specific serodiagnosis of human brucellosis by highly sensitive iELISA (indirect enzyme linked immunosorbent assay). Omp2a gene of B. melitensis 16 M strain was cloned and expressed in pET-SUMO expression system. The recombinant protein was purified under denaturing conditions using 8 M urea. The purified recombinant protein was confirmed by western blotting by reacting with anti-HIS antibody. The sero-reactivity of the recombinant protein was also checked by reacting with antisera of experimentally infected mice with B. melitensis 16 M at different time points. Serodiagnostic potential of recombinant porin antigen was tested against 185 clinical serum samples collected from regions endemic to brucellosis in southern part of India by iELISA. The samples were grouped into five groups. Group 1 contained cultured confirmed positive serum samples of brucellosis (n = 15), group 2 contained sera samples from positive cases of brucellosis previously tested by conventional methods of RBPT (n = 28) and STAT (n = 26), group 3 contained sera samples negative by RBPT(n = 36) and STAT (n = 32), group 4 contained sera samples of other febrile illness and PUO case (n = 35) and group 5 contained confirmed negative sera samples from healthy donors (n = 23). The rOmp2a was found to be

  12. Viral hepatitis

    DEFF Research Database (Denmark)

    Gottwein, Judith M; Bukh, Jens

    2013-01-01

    With millions of humans infected yearly with HCV, leading to cirrhosis and cancer, a vaccine is urgently needed. Cultured virus particles constitute the antigen in most antiviral vaccines. A study in mice demonstrated induction of neutralizing antibodies by immunization with cell-culture-derived ...

  13. Viral pathogens.

    Science.gov (United States)

    Ragni, M V; Sherman, K E; Jordan, J A

    2010-07-01

    Despite continuous improvement in safety and purity of blood products for individuals with haemophilia, transmissible agents continue to affect individuals with haemophilia. This chapter addresses three viral pathogens with significant clinical impact: HIV, hepatitis C and parvovirus B19. Hepatitis C is the leading cause of chronic hepatitis and the major co-morbid complication of haemophilia treatment. Clinically, asymptomatic intermittent alanine aminotransferase elevation is typical, with biopsy evidence of advanced fibrosis currently in 25%. Current treatment is effective in up to 70%, and many new agents are in development. For those progressing to end-stage liver disease, liver transplantation outcomes are similar to those in non-haemophilia subjects, although pretransplant mortality is higher. HIV infection, the second leading co-morbid condition in haemophilia, is managed as a chronic infection with highly active antiretroviral therapy (HAART). HAART also slows hepatitis C virus (HCV) progression in those with HIV/HCV co-infection. Viral inactivation and recombinant technologies have effectively prevented transfusion-transmitted viral pathogens in haemophilia. Human parvovirus B19 infection, typically associated with anaemia or, rarely severe aplastic crisis, is a non-lipid enveloped virus, for which standard inactivation techniques are ineffective. Thus, nucleic acid testing (NAT) to screen the blood supply for B19 DNA is currently under consideration by the Food and Drug Administration. To the extent, viral inactivation, recombinant, and NAT technologies are available worldwide, and the lifespan for those with haemophilia is approaching that of the normal population. The purpose of this chapter is to provide an update on three clinically significant transfusion-transmitted viral pathogens.

  14. Antigenic drift of viruses within a host: a finite site model with demographic stochasticity.

    Science.gov (United States)

    Sasaki, A; Haraguchi, Y

    2000-09-01

    We theoretically study the antigenic drift of viruses within an infected host, as observed in human immunodeficiency virus (HIV) and equine infectious anemia virus (EIAV) infections, assuming that a finite number of antigen-determining sites at the viral envelop gene are responsible for the specific immune response. The pattern of antigen evolution becomes more complex than that predicted from the previous one-dimensional antigen space models. If the viral growth rate is sufficiently large, the demographic stochasticity for the fate of a new antigen mutant can be neglected. The high dimensionality in the way a virus escapes the immune defense in genotype space could then causes a rapid increase in the antigenic diversity and the total viral density, until finally the whole antigen genotypes are used up. The viral population is then driven to extinction in a host by the enhanced immune response to all genotypes. In contrast, if the viral growth rate is moderate or small so that only a small fraction of new antigen mutants can survive during the initial endangered period of random extinction, the viral antigenic diversity and the total density remain bounded, thereby enabling them to persist for a prolonged period by shifting the dominant antigen types. The phylogenetic pattern of antigen divergence is well characterized by the mean number of surviving antigen mutants from an antigen genotype. The substitution rate at antigen-determining sites increases as the efficiency of host immune response increases.

  15. APLASTIC ANEMIA AND VIRAL HEPATITIS

    Directory of Open Access Journals (Sweden)

    Laura Cudillo

    2009-11-01

    Liver histology is characterized by T cell infiltrating the parenchyma as reported in acute hepatitis. Recently in HAA it has been demonstrated intrahepatic  and blood lymphocytes with  T cell repertoire similar to that of confirmed viral acute hepatitis. The expanded T cell clones return to a normal distribution after response to immunosuppressive treatment, suggesting the antigen or T cell clearance. Therapeutic options are the same as acquired aplastic anemia.

  16. Comparative evaluation of Schistosoma mansoni, Schistosoma intercalatum, and Schistosoma haematobium alkaline phosphatase antigenicity by the alkaline phosphatase immunoassay (APIA).

    Science.gov (United States)

    Cesari, I M; Ballén, D E; Mendoza, L; Ferrer, A; Pointier, J-P; Kombila, M; Richard-Lenoble, D; Théron, A

    2014-04-01

    To know if alkaline phosphatase (AP) from schistosomes other than Schistosoma mansoni can be used as diagnostic marker for schistosomiasis in alkaline phosphatase immunocapture assay (APIA), we comparatively tested n-butanol extracts of adult worm membranes from a Venezuelan (JL) strain of S. mansoni (Ven/AWBE/Sm); a Cameroonian (EDEN) strain of Schistosoma intercalatum (Cam/AWBE/Si) and a Yemeni strain of Schistosoma haematobium (Yem/AWBE/Sh). APIA was evaluated with sera of patients from Venezuela, Senegal, and Gabon infected with S. mansoni, from Gabon infected with S. intercalatum or S. haematobium, from Chine infected with Schistosoma japonicum and from Cambodian patients infected with Schistosoma mekongi. Results indicate that 92.5% (37/40) of Venezuela sera, 75% (15/20) of Senegal sera, 39.5% (17/43) of S. haematobium sera, and 19.2% (5/26) S. intercalatum sera were APIA-positive with the Ven/AWBE/Sm preparation. APIA with the Cam/AWBE/Si preparation showed that 53.8% of S. intercalatum-positive sera had anti-AP antibodies, and 51.2% S. haematobium-positive sera cross-immunocapturing the S. intercalatum AP. APIA performed with Yem/AWBE/Sh showed that 55.8% S. haematobium sera were positive. Only two out of nine S. japonicum sera were APIA-positive with the Ven/AWBE/Sm and Cam/AWBE/Si, and no reaction was observed with Cambodian S. mekongi-positive sera. AP activity was shown to be present in all the schistosome species/strains studied. The use of APIA as a tool to explore the APs antigenicity and the presence of Schistosoma sp. infections through the detection of anti-Schistosoma sp. AP antibodies in a host, allowed us to demonstrate the antigenicity of APs of S. mansoni, S. intercalatum, and S. haematobium.

  17. Evaluation of a Direct Immunofluorescent Assay and/or Conjunctival Cytology for Detection of Canine Distemper Virus Antigen.

    Science.gov (United States)

    Athanasiou, Labrini V; Kantere, Maria C; Kyriakis, Constantinos S; Pardali, Dimitra; Adamama Moraitou, Katerina; Polizopoulou, Zoe S

    2017-11-29

    Canine distemper is a common and potentially lethal multisystemic disease caused by the Canine distemper virus (CDV). We evaluated the diagnostic performance of direct immunofluorescent assay (FA) and cytology to detect CDV antigen in conjunctival cells compared with an established polymerase chain reaction (PCR) detection assay used as a gold standard for CDV diagnosis. Samples were collected from 57 young dogs presenting with central nervous system signs compatible with distemper disease. Exfoliative epithelial cells were collected from the right and left conjunctiva of each animal using nylon-bristled cytobrushes for cytology and cotton swabs for FA and PCR. For the direct FA, samples were stained with anti-CDV polyclonal antiserum conjugated to fluorescein isothiocyanate and imaged using a fluorescent microscope. Out of 57 dogs tested, 19 were PCR positive (15 positive in direct FA and 4 positive in cytology, including one that was negative by PCR), whereas 37 dogs were negative in all methods. A good agreement was observed between the FA and PCR, with a κ-value of 0.833 (95% CI: 0.678-0.989). Meanwhile, there was poor agreement between cytology and PCR (κ-value of 0.164; 95% CI: -0.045 to 0.373) and a fair agreement between FA and cytology (κ-value of 0.231; 95% CI: -0.026 to 0.487). Our results indicated a poor performance of cytology for the detection of CDV antigen. In contrast, FA is a 100% specific and an adequately sensitive assay (sensitivity: 78.95%, negative likelihood ratio: 0.21, 95% CI: 0.09-0.50) for antemortem diagnosis of canine distemper.

  18. Towards improved diagnosis of von Willebrand disease: comparative evaluations of several automated von Willebrand factor antigen and activity assays.

    Science.gov (United States)

    Favaloro, Emmanuel J; Mohammed, Soma

    2014-12-01

    von Willebrand disease (VWD) is reportedly the most common bleeding disorder and arises from deficiency and/or defects of von Willebrand factor (VWF). Laboratory diagnosis and typing has important management implications and requires a wide range of tests, including VWF activity and antigen, and involves differential identification of qualitative vs quantitative defects. We have assessed several VWF antigen and activity assays (collagen binding [VWF:CB], ristocetin cofactor [VWF:RCo] and the new Siemens INNOVANCE assay [VWF:Ac], employing latex particles and gain of function recombinant glycoprotein Ib to facilitate VWF binding and agglutination without need for ristocetin) using different instrumentation, including the new Sysmex CS-5100, with a large sample test set (n=600). We included retrospective plus prospective study designs, and also evaluated desmopressin responsiveness plus differential sensitivity to high molecular weight VWF. VWF:Ag and VWF:RCo results from different methods were respectively largely comparable, although some notable differences were evident, including one high false normal VWF:Ag value (105 U/dL) on a type 3 VWD sample, possibly due to heterophile antibody interference in the latex-based CS-5100 methodology. VWF:Ac was largely comparable to VWF:RCo, but VWF:CB showed discrepant findings to both VWF:RCo and VWF:Ac with some patients, most notably patients with type 2M VWD. (a) VWF:Ag on different platforms are largely interchangeable, as are VWF:RCo on different platforms, except for occasional (some potentially important) differences, and manufacturer recommended methods may otherwise require some assay optimization; (b) VWF:RCo and VWF:Ac are largely interchangeable, except for occasional differences that may also relate to assay design (differing optimizations); (c) VWF:CB provides an additional activity to supplement VWF:RCo or VWF:Ac activity assays, and is not interchangeable with either. Crown Copyright © 2014. Published by

  19. Viral antibody titers are influenced by HLA-DR2 phenotype.

    Science.gov (United States)

    Lio, D; Caccamo, N; D'Anna, C; Cigna, D; Candore, G; Caruso, C

    1994-01-01

    Antibody serum levels against herpes simplex type 1 virus, cytomegalovirus, viral capsid antigens of Epstein-Barr virus, and rubella virus were evaluated in a sample of the Sicilian population. Results demonstrated that HLA-DR2-positive individuals showed a significant increase in antibody titers, when compared to HLA-DR2-negative individuals. These observations seem to be in contrast with the reported association of the HLA-DR2 phenotype with an ineffective immune response against several infectious pathogens. On the other hand, an increased humoral response to viral antigens need not be interpreted as a marker for effective control of virus infections. In fact, the response to virus infections is related to the T-cell-mediated immune response restricted by class-I- or class-II-encoded proteins. Thus, the above-mentioned HLA-DR2-related susceptibility to certain viral infections could be associated with a preferential induction of an increased (although ineffective) antibody synthesis against viral antigens.

  20. A viral metagenomic approach on a non-metagenomic experiment: Mining next generation sequencing datasets from pig DNA identified several porcine parvoviruses for a retrospective evaluation of viral infections.

    Directory of Open Access Journals (Sweden)

    Samuele Bovo

    Full Text Available Shot-gun next generation sequencing (NGS on whole DNA extracted from specimens collected from mammals often produces reads that are not mapped (i.e. unmapped reads on the host reference genome and that are usually discarded as by-products of the experiments. In this study, we mined Ion Torrent reads obtained by sequencing DNA isolated from archived blood samples collected from 100 performance tested Italian Large White pigs. Two reduced representation libraries were prepared from two DNA pools constructed each from 50 equimolar DNA samples. Bioinformatic analyses were carried out to mine unmapped reads on the reference pig genome that were obtained from the two NGS datasets. In silico analyses included read mapping and sequence assembly approaches for a viral metagenomic analysis using the NCBI Viral Genome Resource. Our approach identified sequences matching several viruses of the Parvoviridae family: porcine parvovirus 2 (PPV2, PPV4, PPV5 and PPV6 and porcine bocavirus 1-H18 isolate (PBoV1-H18. The presence of these viruses was confirmed by PCR and Sanger sequencing of individual DNA samples. PPV2, PPV4, PPV5, PPV6 and PBoV1-H18 were all identified in samples collected in 1998-2007, 1998-2000, 1997-2000, 1998-2004 and 2003, respectively. For most of these viruses (PPV4, PPV5, PPV6 and PBoV1-H18 previous studies reported their first occurrence much later (from 5 to more than 10 years than our identification period and in different geographic areas. Our study provided a retrospective evaluation of apparently asymptomatic parvovirus infected pigs providing information that could be important to define occurrence and prevalence of different parvoviruses in South Europe. This study demonstrated the potential of mining NGS datasets non-originally derived by metagenomics experiments for viral metagenomics analyses in a livestock species.

  1. A viral metagenomic approach on a non-metagenomic experiment: Mining next generation sequencing datasets from pig DNA identified several porcine parvoviruses for a retrospective evaluation of viral infections.

    Science.gov (United States)

    Bovo, Samuele; Mazzoni, Gianluca; Ribani, Anisa; Utzeri, Valerio Joe; Bertolini, Francesca; Schiavo, Giuseppina; Fontanesi, Luca

    2017-01-01

    Shot-gun next generation sequencing (NGS) on whole DNA extracted from specimens collected from mammals often produces reads that are not mapped (i.e. unmapped reads) on the host reference genome and that are usually discarded as by-products of the experiments. In this study, we mined Ion Torrent reads obtained by sequencing DNA isolated from archived blood samples collected from 100 performance tested Italian Large White pigs. Two reduced representation libraries were prepared from two DNA pools constructed each from 50 equimolar DNA samples. Bioinformatic analyses were carried out to mine unmapped reads on the reference pig genome that were obtained from the two NGS datasets. In silico analyses included read mapping and sequence assembly approaches for a viral metagenomic analysis using the NCBI Viral Genome Resource. Our approach identified sequences matching several viruses of the Parvoviridae family: porcine parvovirus 2 (PPV2), PPV4, PPV5 and PPV6 and porcine bocavirus 1-H18 isolate (PBoV1-H18). The presence of these viruses was confirmed by PCR and Sanger sequencing of individual DNA samples. PPV2, PPV4, PPV5, PPV6 and PBoV1-H18 were all identified in samples collected in 1998-2007, 1998-2000, 1997-2000, 1998-2004 and 2003, respectively. For most of these viruses (PPV4, PPV5, PPV6 and PBoV1-H18) previous studies reported their first occurrence much later (from 5 to more than 10 years) than our identification period and in different geographic areas. Our study provided a retrospective evaluation of apparently asymptomatic parvovirus infected pigs providing information that could be important to define occurrence and prevalence of different parvoviruses in South Europe. This study demonstrated the potential of mining NGS datasets non-originally derived by metagenomics experiments for viral metagenomics analyses in a livestock species.

  2. A Review of Intra- and Extracellular Antigen Delivery Systems for Virus Vaccines of Finfish

    Directory of Open Access Journals (Sweden)

    Hetron Mweemba Munang’andu

    2015-01-01

    Full Text Available 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.

  3. Comparative evaluation of commercially available point-of-care heartworm antigen tests using well-characterized canine plasma samples

    Directory of Open Access Journals (Sweden)

    Lindsay A. Starkey

    2017-11-01

    Full Text Available Abstract Background Dirofilaria immitis is a worldwide parasite that is endemic in many parts of the United States. There are many commercial assays available for the detection of D. immitis antigen, one of which was modified and has reentered the market. Our objective was to compare the recently reintroduced Witness® Heartworm (HW Antigen test Kit (Zoetis, Florham Park, NJ and the SNAP® Heartworm RT (IDEXX Laboratories, Inc., Westbrook, ME to the well-based ELISA DiroChek® Heartworm Antigen Test Kit (Zoetis, Florham Park, NJ. Methods Canine plasma samples were either received at the Auburn Diagnostic Parasitology Laboratory from veterinarians submitting samples for additional heartworm testing (n = 100 from 2008 to 2016 or purchased from purpose-bred beagles (n = 50, presumed negative in 2016. Samples were categorized as “positive,” “borderline” or “negative” using our established spectrophotometric cutoff value with the DiroChek® assay when a sample was initially received and processed. Three commercially available heartworm antigen tests (DiroChek®, Witness® HW, and SNAP® RT were utilized for simultaneous testing of the 150 samples in random order as per their package insert with the addition of spectrophotometric optical density (OD readings of the DiroChek® assay. Any samples yielding discordant test results between assays were further evaluated by heat treatment of plasma and retesting. Chi-square tests for the equality of proportions were utilized for statistical analyses. Results Concordant results occurred in 140/150 (93.3% samples. Discrepant results occurred in 10/150 samples tested (6.6%: 9/10 occurring in the borderline heartworm (HW category and 1/10 occurring in the negative HW category. The sensitivity and specificity of each test compared to the DiroChek® read by spectrophotometer was similar to what has been reported previously (Witness®: sensitivity 97.0% [94.1–99.4%], specificity 96.4% [95.5–100

  4. Evaluation of rapid one-step prostate specific antigen test against an ...

    African Journals Online (AJOL)

    Main Objective: To evaluate the analytical performance of the rapid one-step immunochromatographic PSA assay against an established ELISA method. Design: A comparative study was conducted in the Department of Chemical Pathology at the University of Zimbabwe, College Health Sciences, between June 2012 and ...

  5. Preclinical evaluation of TriMix and antigen mRNA-based antitumor therapy

    National Research Council Canada - National Science Library

    Van Lint, Sandra; Goyvaerts, Cleo; Maenhout, Sarah; Goethals, Lode; Disy, Aurélie; Benteyn, Daphné; Pen, Joeri; Bonehill, Aude; Heirman, Carlo; Breckpot, Karine; Thielemans, Kris

    2012-01-01

    ...). In this study, we evaluated whether intranodal delivery of TAA mRNA together with TriMix, a mix of mRNA encoding CD40 ligand, constitutive active Toll-like receptor 4 and CD70, results in the in...

  6. Evaluation of the effects of a new formulation of Leishmania major antigen in Balb/C and conventional white laboratory mice.

    Science.gov (United States)

    Latifynia, Afshineh; Hazrati, Saleh Mohaghegh

    2010-04-01

    Leishmaniasis is currently a threat in 80 countries around the world, and cutaneous Leishmaniasis accounts for more than 5% of new cases. It is a problematic disease in Iran, and preparing a protective vaccine has been a major goal of medical investigations. The objective of this study was to compare the protective effects of a cocktail vaccine candidate encoding various Leishmania major antigens in highly susceptible (Balb/C, or type I mice) and resistant (laboratory small white, or type II) mice. A new antigen formulation was evaluated in type I and II mice. Leishmania major promastigotes was cultured and harvested at different growth stages, and a cocktail made from the harvested organisms. The preparations were tested for sterility and contamination by endotoxin. Five different methods were utilized to produce a crude antigen preparation. The protein levels of the antigen preparations were measured using the Lowry method, and the antigens were intradermally injected using different protocols for type I and II mice. After 38 days, all mice were euthanized with diethyl ether, and spleens were removed. Histological sections were prepared, stained with hematoxylin and eosin and changes to the splenic white pulp (SWP) were studied microscopically. Compared with the control groups, there was a drastic change in white pulp structure. The size increases of the SWP were dependent on the injection group and mouse strain. There was a remarkable expansion of lymphoid follicles in the treated groups in both mice strains. The new antigen formulation was able to stimulate and expand the lymphoid constituents of spleen tissue. The SWP is where immune responses and antibodies are produced. Therefore, the effect of the antigen preparations on secondary immune responses, adaptive immunity, and antibody production is important in determining the susceptibility of mice to cutaneous Leishmaniasis and the induction of immunity. Copyright 2010 Taiwan Society of Microbiology. Published

  7. Evaluation of Humoral Immune Response against Somatic and Excretory-Secretory Antigens of Dicrocoelium Dendriticum in Infected Sheep by Western Blot

    Directory of Open Access Journals (Sweden)

    Mohammad Hosein Razi Jalali

    2013-12-01

    Full Text Available Introduction and objective: Dicrocoelium dendriticum is a worldwide spread parasite of liver, bile ducts and gallbladder of especially ruminants and humans as well. Identification of specific antigens is useful for early diagnosis of the infection. The goal of this study was the isolation and identification of excretory-secretory and somatic antigens from D. dendriticum by sodium dodecyl sulphate (SDS-PAGE and evaluation of humoral immune response against these antigens.   Methods: The parasites were collected and washed by phosphate buffered saline (PBS and supplemented by antibiotic for several times. For preparing somatic antigens, parasites were sonicated and centrifuged prior to collect supernatant. For preparing excretory-secretory antigens the viable parasites were transferred to the sterile medium. The samples were centrifuged and supernatants were collected. The sera of infected sheep with different infection degrees were collected too. Somatic and excretory-secretory proteins were isolated with SDS PAGE and stained with coomassie blue. Immunogenicity properties of the resulting proteins were determined using western blot analysis.   Results: The total extract of somatic antigens analyzed by SDS-PAGE revealed 21 proteins. In mild infection, bands of 130 KDa were immune dominant. In moderate infections 48, 80 and 130 KDa and in heavy infections 48, 60, 80, 130 KDa were detected as immune dominant bands. In excretory- secretory antigens seven bands of protein were detected. In mild infection 130 KDa, in moderate infection 100, 120 and 130 KDa and in heavy infection 45, 80, 85, 100, 120 and 130 KDa were immune dominant bands.   Conclusion: Probably the most immunogenic protein band during different degrees of infection was 130KDa that can be used for vaccination and inducing immunity.

  8. Prospective evaluation of a whole-blood test using Mycobacterium tuberculosis-specific antigens ESAT-6 and CFP-10 for diagnosis of active tuberculosis

    DEFF Research Database (Denmark)

    Ravn, Pernille; Munk, Martin E; Andersen, Ase B

    2005-01-01

    A new immunodiagnostic test based on the Mycobacterium tuberculosis-specific antigens CFP-10/ESAT-6(QFT-RD1) has been launched as an aid in the diagnosis of latent tuberculosis (TB) infection (LTBI). The aim of this study was to evaluate this test for the diagnosis of active TB. Eighty-two patients...

  9. Evaluation of Mycobacterium leprae antigens in the monitoring of a dapsone-based chemotherapy of previously untreated lepromatous patients in Cebu, Philippines

    NARCIS (Netherlands)

    Klatser, P. R.; de Wit, M. Y.; Fajardo, T. T.; Cellona, R. V.; Abalos, R. M.; de la Cruz, E. C.; Madarang, M. G.; Hirsch, D. S.; Douglas, J. T.

    1989-01-01

    Thirty-five previously untreated lepromatous patients receiving dapsone-based therapy were monitored throughout their 5-year period of treatment by serology and by pathology. Sequentially collected sera were used to evaluate the usefulness of four Mycobacterium leprae antigens as used in ELISA to

  10. Characterization of an F1 Deletion Mutant of Yersinia pestis CO92, Pathogenic Role of F1 Antigen in Bubonic and Pneumonic Plague, and Evaluation of Sensitivity and Specificity of F1 Antigen Capture-Based Dipsticks▿

    Science.gov (United States)

    Sha, Jian; Endsley, Janice J.; Kirtley, Michelle L.; Foltz, Sheri M.; Huante, Matthew B.; Erova, Tatiana E.; Kozlova, Elena V.; Popov, Vsevolod L.; Yeager, Linsey A.; Zudina, Irina V.; Motin, Vladimir L.; Peterson, Johnny W.; DeBord, Kristin L.; Chopra, Ashok K.

    2011-01-01

    We evaluated two commercial F1 antigen capture-based immunochromatographic dipsticks, Yersinia Pestis (F1) Smart II and Plague BioThreat Alert test strips, in detecting plague bacilli by using whole-blood samples from mice experimentally infected with Yersinia pestis CO92. To assess the specificities of these dipsticks, an in-frame F1-deficient mutant of CO92 (Δcaf) was generated by homologous recombination and used as a negative control. Based on genetic, antigenic/immunologic, and electron microscopic analyses, the Δcaf mutant was devoid of a capsule. The growth rate of the Δcaf mutant generally was similar to that of the wild-type (WT) bacterium at both 26 and 37°C, although the mutant's growth dropped slightly during the late phase at 37°C. The Δcaf mutant was as virulent as WT CO92 in the pneumonic plague mouse model; however, it was attenuated in developing bubonic plague. Both dipsticks had similar sensitivities, requiring a minimum of 0.5 μg/ml of purified F1 antigen or 1 × 105 to 5 × 105 CFU/ml of WT CO92 for positive results, while the blood samples were negative for up to 1 × 108 CFU/ml of the Δcaf mutant. Our studies demonstrated the diagnostic potential of two plague dipsticks in detecting capsular-positive strains of Y. pestis in bubonic and pneumonic plague. PMID:21367990

  11. Caution in evaluation of removal of virus by filtration: Misinterpretation due to detection of viral genome fragments by PCR.

    Science.gov (United States)

    Tsujikawa, Muneo; Ohkubo, Yuji; Masuda, Maki; Tanaka, Hiroyuki; Takahashi, Kadue; Sasaki, Yuko; Yunoki, Mikihiro; Ikuta, Kazuyoshi

    2011-12-01

    The testing of biological products at different stages of the manufacturing process currently involves quantitative polymerase chain reaction (Q-PCR)-based assays. Q-PCR techniques are able to detect not only the viral genome in viral particles but also fragments of degraded genome in samples. The ability of 15 and 19-nm filters to remove viruses was examined by conducting infectivity assays and Q-PCR assays using parvovirus B19 (B19), one of the smallest non-enveloped viruses. Although the filtered samples showed no infectivity, viral DNA was detected by Q-PCR. Interestingly, approximately 90% of the total viral genome in 15-nm filtrates had a detectable size of less than 0.5kb by the Q-PCR and as a consequence reduction factors were underestimated using Q-PCR. The reduction factors using Q-PCR might be underestimated due to the presence of a large amount of free B19 DNA which shows no infectivity in the tested filtrates. Therefore, the results of Q-PCR should be interpreted with caution. The careful design of primers is needed to eliminate amplification from fragments of viral DNA by Q-PCR. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Use of three-dimensional accelerometers to evaluate behavioral changes in cattle experimentally infected with bovine viral diarrhea virus.

    Science.gov (United States)

    Bayne, Jenna E; Walz, Paul H; Passler, Thomas; White, Brad J; Theurer, Miles E; van Santen, Edzard

    2016-06-01

    OBJECTIVE To assess the use of 3-D accelerometers to evaluate behavioral changes in cattle experimentally infected with a low-virulent strain of bovine viral diarrhea virus (BVDV). ANIMALS 20 beef steers (mean weight, 238 kg). PROCEDURES Calves were allocated to a BVDV (n = 10) or control (10) group. On day 0, calves in the BVDV group were inoculated with a low-virulent strain of BVDV (4 × 10(6) TCID50, intranasally), and calves in the control group were sham inoculated with BVDV-free medium (4 mL; intranasally). An accelerometer was affixed to the right hind limb of each calf on day -7 to record activity (lying, walking, and standing) continuously until 35 days after inoculation. Baseline was defined as days -7 to -1. Blood samples were collected at predetermined times for CBC, serum biochemical analysis, virus isolation, and determination of anti-BVDV antibody titers. RESULTS All calves in the BVDV group developed viremia and anti-BVDV antibodies but developed only subclinical or mild disease. Calves in the control group did not develop viremia or anti-BVDV antibodies. Mean time allocated to each activity did not differ significantly between the BVDV and control groups on any day except day 8, when calves in the BVDV group spent less time standing than the calves in the control group. Following inoculation, calves in both groups tended to spend more time lying and less time walking and standing than they did during baseline. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that behavioral data obtained by accelerometers could not distinguish calves subclinically infected with BVDV from healthy control calves. However, subtle changes in the behavior of the BVDV-infected calves were detected and warrant further investigation.

  13. In vitro evaluation of five rapid antigen detection tests for group A beta-haemolytic streptococcal sore throat infections.

    Science.gov (United States)

    Lasseter, Gemma M; McNulty, Cliodna A M; Richard Hobbs, F D; Mant, David; Little, Paul

    2009-12-01

    Using accurate and easy to use rapid antigen detection tests (RADTs) to identify group A beta-haemolytic Streptococci (GABHS) sore throat infections could reduce unnecessary antibiotic prescribing and antimicrobial resistance. Although there is no international consensus on the use of RADTs, these kits have been widely adopted in Finland, France and the USA. Yet in the UK, the Clinical Knowledge Summaries, that provide the main online guidance for GPs, discourage RADTs use, citing their poor sensitivity and inability to impact on prescribing decisions in acute sore throat infections. The purpose of this study was to evaluate the ease of use and in vitro accuracy (sensitivity and specificity) of the five most commonly used RADTs in Europe (OSOM Ultra, Quickvue Dipstick, Streptatest, Clearview Exact Strep A and IMI Test Pack). To ensure the RADTs were evaluated objectively, a standardized in vitro method using known concentrations of GABHS was used to remove the inherent biases associated with clinical studies. The IMI Test Pack was the easiest RADT to use overall. The ability to detect all positive GABHS (sensitivity) varied considerably between kits from 95% [95% confidence interval (CI): 88-98%], for the IMI Test Pack and OSOM, to 62% (95% CI: 51-72%) for Clearview, at the highest GABHS concentration. None of the RADTs gave any false-positive results with commensal flora-they were 100% specific. The IMI Test Pack is most suitable for use in primary care, as it had high sensitivity, high specificity and was easy to use.

  14. Evaluation of two Neospora caninum recombinant antigens for use in an enzyme-linked immunosorbent assay for the diagnosis of bovine neosporosis.

    Science.gov (United States)

    Lally, N C; Jenkins, M C; Dubey, J P

    1996-01-01

    Neospora caninum is a recently described apicomplexan parasite which causes paralysis and death in dogs. Neospora parasites also cause abortion and neonatal morbidity in cattle, sheep, goats, and horses, and neosporosis is emerging as an important cause of bovine abortion worldwide. The purpose of this study was to identify N. caninum cDNA clones encoding antigens that would be useful for the immunodiagnosis of bovine neosporosis. Two N. caninum tachyzoite cDNA clones expressing antigens that were recognized by serum from naturally and experimentally infected cattle were identified. The DNA sequences of these clones were determined, and the inserts were subcloned into the plasmid expression vector pTrcHisB. Both recombinant antigens, expressed as fusion proteins with a His6 tag, were purified on a nickel-chelating affinity column and evaluated in separate enzyme-linked immunosorbent assays (ELISAs). Both recombinant antigen ELISAs were capable of distinguishing between sera from Neospora-infected cows and sera from uninfected control cows. Furthermore, both assays were able to detect an antibody response in animals that were experimentally inoculated with N. caninum. Neither antigen showed evidence of cross-reactivity with serum from animals inoculated with the closely related parasites Toxoplasma gondii, Sarcocystis cruzi, Sarcocystis hominis, and Sarcocystis hirsuta. PMID:8705668

  15. Evaluation of a Salmonella vectored vaccine expressing Mycobacterium avium subsp. paratuberculosis antigens against challenge in a goat model.

    Directory of Open Access Journals (Sweden)

    Syed M Faisal

    Full Text Available Johnes disease (JD, caused by Mycobacterium avium subsp paratuberculosis (MAP, occurs worldwide as chronic granulomatous enteritis of domestic and wild ruminants. To develop a cost effective vaccine, in a previous study we constructed an attenuated Salmonella strain that expressed a fusion product made up of partial fragments of MAP antigens (Ag85A, Ag85B and SOD that imparted protection against challenge in a mouse model. In the current study we evaluated the differential immune response and protective efficacy of the Sal-Ag vaccine against challenge in a goat model as compared to the live attenuated vaccine MAP316F. PBMCs from goats vaccinated with Sal-Ag and challenged with MAP generated significantly lower levels of IFN-γ, following in vitro stimulation with either Antigen-mix or PPD jhonin, than PBMC from MAP316F vaccinated animals. Flow cytometric analysis showed the increase in IFN-γ correlated with a significantly higher level of proliferation of CD4, CD8 and γδT cells and an increased expression of CD25 and CD45R0 in MAP316F vaccinated animals as compared to control animals. Evaluation of a range of cytokines involved in Th1, Th2, Treg, and Th17 immune responses by quantitative PCR showed low levels of expression of Th1 (IFN-γ, IL-2, IL-12 and proinflammatory cytokines (IL-6, IL-8, IL-18, TNF-α in the Sal-Ag immunized group. Significant levels of Th2 and anti-inflammatory cytokines transcripts (IL-4, IL-10, IL-13, TGF-β were expressed but their level was low and with a pattern similar to the control group. Over all, Sal-Ag vaccine imparted partial protection that limited colonization in tissues of some animals upon challenge with wild type MAP but not to the level achieved with MAP316F. In conclusion, the data indicates that Sal-Ag vaccine induced only a low level of protective immunity that failed to limit the colonization of MAP in infected animals. Hence the Sal-Ag vaccine needs further refinement to increase its efficacy.

  16. [Vasculitis and viral infection].

    Science.gov (United States)

    Martínez Aguilar, N E; Guido Bayardo, R; Vargas Camaño, M E; Compañ González, D; Miranda Feria, A J

    1997-01-01

    Viruses have been implicated in vasculitis. To determine activity of viral infection associated with vasculitis. 17 patients with vasculitis had been in immunological and antiviral antibodies evaluation. Twenty five healthy controls sex and age matched with hematic biometry (BH) and AA. All subjects were negative to HIV and HBV. Viral activity was demonstrated in eight patients; vascular purpura (5), Takayasu disease (1), polyarteritis nodosa (1), erythema nodosum (1). None subject of control group had IgM activity. Antibodies response of IgG in patients were of lesser intensity than in control group. 14 abnormalities in BH were found in patients and 4 in control group. Immune response in patients, measured by lymphocyte subpopulations and circulating immune complexes was abnormal. In conclusion 47% showed viral activity, but the dominant feature was abnormal immune response in 82%.

  17. Evaluation of an ovine testis cell line (OA3.Ts) for propagation of capripoxvirus isolates and development of an immunostaining technique for viral plaque visualization.

    Science.gov (United States)

    Babiuk, Shawn; Parkyn, Geoff; Copps, John; Larence, June E; Sabara, Marta I; Bowden, Timothy R; Boyle, David B; Kitching, R Paul

    2007-09-01

    An ovine testis cell line (OA3.Ts) was evaluated and compared with primary lamb kidney (LK) cells for its utility in capripoxvirus propagation and titration. A comparison of OA3.Ts cell growth kinetics and morphology at low (Capripoxvirus infection of OA3.Ts and LK cells resulted in a similar cytopathic effect, which allowed for the detection of discrete viral plaques following immunostaining with capripoxvirus-specific antiserum.

  18. Evaluation of a New Device for Simplifying and Standardizing Stool Sample Preparation for Viral Molecular Testing with Limited Hands-On Time

    OpenAIRE

    Feghoul, Linda; Salmona, Maud; Cherot, Janine; Fahd, Mony; Dalle, Jean-Hugues; Vachon, Carole; Perrod, Aurélie; Bourgeois, Philippe; Scieux, Catherine; Baruchel, André; Simon, François; Legoff, Jérôme

    2016-01-01

    Sensitive molecular assays have greatly improved the diagnosis of viral gastroenteritis. However, the proper preparation of stool samples for clinical testing remains an issue. bioMérieux has developed a stool preprocessing device (SPD) that includes a spoon for calibrated sampling and a vial containing buffer, glass beads, and two filters. The resulting stool filtrate is used for nucleic acid extraction. The purpose of this study was to evaluate the performance of the SPD for the quantificat...

  19. Evaluation of the Xpert HCV Viral Load point-of-care assay from venepuncture-collected and finger-stick capillary whole-blood samples: a cohort study.

    Science.gov (United States)

    Grebely, Jason; Lamoury, Francois M J; Hajarizadeh, Behzad; Mowat, Yasmin; Marshall, Alison D; Bajis, Sahar; Marks, Philippa; Amin, Janaki; Smith, Julie; Edwards, Michael; Gorton, Carla; Ezard, Nadine; Persing, David; Kleman, Marika; Cunningham, Philip; Catlett, Beth; Dore, Gregory J; Applegate, Tanya L

    2017-07-01

    Point-of-care hepatitis C virus (HCV) RNA testing offers an advantage over antibody testing (which only indicates previous exposure), enabling diagnosis of active infection in a single visit. In this study, we evaluated the performance of the Xpert HCV Viral Load assay with venepuncture and finger-stick capillary whole-blood samples. Plasma and finger-stick capillary whole-blood samples were collected from participants in an observational cohort enrolled at five sites in Australia (three drug and alcohol clinics, one homelessness service, and one needle and syringe programme). We compared the sensitivity and specificity of the Xpert HCV Viral Load test for HCV RNA detection by venepuncture and finger-stick collection with the Abbott RealTime HCV Viral Load assay (gold standard). Of 210 participants enrolled between Feb 8, 2016, and July 27, 2016, 150 participants had viral load testing results for the three assays tested. HCV RNA was detected in 45 (30% [95% CI 23-38]) of 150 participants based on Abbott RealTime. Sensitivity of the Xpert HCV Viral Load assay for HCV RNA detection in plasma collected by venepuncture was 100·0% (95% CI 92·0-100·0) and specificity was 99·1% (95% CI 94·9-100·0). Sensitivity of the Xpert HCV Viral Load assay for HCV RNA detection in samples collected by finger-stick was 95·5% (95% CI 84·5-99·4) and specificity was 98·1% (95% CI 93·4-99·8). No adverse events caused by the index test or the reference standard were observed. The Xpert HCV Viral Load test can detect active infection from a finger-stick sample, which represents an advance over antibody-based tests that only indicate past or previous exposure. National Health and Medical Research Council (Australia), Cepheid, South Eastern Sydney Local Health District (Australia), and Merck Sharp & Dohme (Australia). Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Phase Ia clinical evaluation of the safety and immunogenicity of the Plasmodium falciparum blood-stage antigen AMA1 in ChAd63 and MVA vaccine vectors.

    Directory of Open Access Journals (Sweden)

    Susanne H Sheehy

    Full Text Available Traditionally, vaccine development against the blood-stage of Plasmodium falciparum infection has focused on recombinant protein-adjuvant formulations in order to induce high-titer growth-inhibitory antibody responses. However, to date no such vaccine encoding a blood-stage antigen(s alone has induced significant protective efficacy against erythrocytic-stage infection in a pre-specified primary endpoint of a Phase IIa/b clinical trial designed to assess vaccine efficacy. Cell-mediated responses, acting in conjunction with functional antibodies, may be necessary for immunity against blood-stage P. falciparum. The development of a vaccine that could induce both cell-mediated and humoral immune responses would enable important proof-of-concept efficacy studies to be undertaken to address this question.We conducted a Phase Ia, non-randomized clinical trial in 16 healthy, malaria-naïve adults of the chimpanzee adenovirus 63 (ChAd63 and modified vaccinia virus Ankara (MVA replication-deficient viral vectored vaccines encoding two alleles (3D7 and FVO of the P. falciparum blood-stage malaria antigen; apical membrane antigen 1 (AMA1. ChAd63-MVA AMA1 administered in a heterologous prime-boost regime was shown to be safe and immunogenic, inducing high-level T cell responses to both alleles 3D7 (median 2036 SFU/million PBMC and FVO (median 1539 SFU/million PBMC, with a mixed CD4(+/CD8(+ phenotype, as well as substantial AMA1-specific serum IgG responses (medians of 49 µg/mL and 41 µg/mL for 3D7 and FVO AMA1 respectively that demonstrated growth inhibitory activity in vitro.ChAd63-MVA is a safe and highly immunogenic delivery platform for both alleles of the AMA1 antigen in humans which warrants further efficacy testing. ChAd63-MVA is a promising heterologous prime-boost vaccine strategy that could be applied to numerous other diseases where strong cellular and humoral immune responses are required for protection.ClinicalTrials.gov NCT01095055.

  1. The Trypomastigote Small Surface Antigen from Trypanosoma cruzi improves treatment evaluation and diagnosis in pediatric Chagas disease.

    Science.gov (United States)

    Balouz, Virginia; Melli, Luciano J; Volcovich, Romina; Moscatelli, Guillermo; Moroni, Samanta; González, Nicolás; Ballering, Griselda; Bisio, Margarita; Ciocchini, Andrés; Buscaglia, Carlos A; Altcheh, Jaime

    2017-10-04

    Chagas disease is caused by the protozoan parasite Trypanosoma cruzi. Assessment of parasitological cure upon treatment with available drugs relies on achieving consistent negative results in conventional parasitological and serological tests, which may take years to assess. Here, we evaluated the use of a recombinant T. cruzi antigen termed TSSA as an early serological marker of drug efficacy in T. cruzi-infected children. A cohort of 78 pediatric patients born to T. cruzi-infected mothers was included in this study. Solely 39 of them were infected with T. cruzi, and were immediately treated with trypanocidal drugs. Serological responses against TSSA were evaluated in infected and non-infected populations during the follow-up period using an in-house ELISA test, and compared to conventional serological methods. Anti-TSSA antibody titers decreased significantly faster than anti-whole parasite antibodies detected by conventional serology in both T. cruzi-infected patients undergoing effective treatment and in those not infected. This differential kinetics allowed a significant reduction in the required follow-up periods to evaluate therapeutic responses or to rule out maternal-fetal transmission, respectively. Finally, we present the case of a congenitally-infected patient with atypical course, in which TSSA provided an early marker for T. cruzi infection. In conclusion, we showed that TSSA was efficacious both for rapid assessment of treatment efficiency and for early negative diagnosis in infants at risk of congenital T. cruzi infection. Based upon these findings we propose the inclusion of TSSA for refining the post-therapeutic cure criterion and other diagnostic needs in pediatric Chagas disease. Copyright © 2017 American Society for Microbiology.

  2. Identification of clone-9 antigenic protein of Theileria uilenbergi and evaluation of its application for serodiagnosis.

    Science.gov (United States)

    Abdo, Jassim; Liu, Zhijie; Yin, Hong; Kullmann, Birgit; Ahmed, Jabbar S; Seitzer, Ulrike

    2010-08-01

    The pathogenic protozoan parasite Theileria uilenbergi is one of the causative agents of theileriosis in small ruminants in China. The infection results in great economical losses in the northwest part of China. Efforts are underway to establish an enzyme-linked immunosorbent assay (ELISA) based on a T. uilenbergi immunodominant recombinantly expressed protein using different approaches in order to perform epidemiological studies in the area. In this study, we describe the possible use of the clone-9 protein for this purpose, which was identified as a potential immunogenic piroplasm protein by random sequencing of cDNA library clones followed by bioinformatic analyses. The clone-9 gene was partially recombinantly expressed and used for the development of an indirect ELISA for the detection of circulating antibodies in sera of T. uilenbergi-infected sheep. No cross-reactivity was observed in serum from animals infected with Theileria lestoquardi. The cut-off was calculated at 48.6% positivity using 25 serum samples from uninfected animals. A total of 101 field samples collected from an endemic area in China were used to evaluate the clone-9 ELISA for its use in the field.

  3. Prevalence and evaluation strategies for viral contamination in food products: Risk to human health-a review.

    Science.gov (United States)

    Shukla, Shruti; Cho, Hyunjeong; Kwon, O Jun; Chung, Soo Hyun; Kim, Myunghee

    2018-02-11

    Nowadays, viruses of foodborne origin such as norovirus and hepatitis A are considered major causes of foodborne gastrointestinal illness with widespread distribution worldwide. A number of foodborne outbreaks associated with food products of animal and non-animal origins, which often involve multiple cases of variety of food streams, have been reported. Although several viruses, including rotavirus, adenovirus, astrovirus, parvovirus, and other enteroviruses, significantly contribute to incidence of gastrointestinal diseases, systematic information on the role of food in transmitting such viruses is limited. Most of the outbreak cases caused by infected food handlers were the source of 53% of total outbreaks. Therefore, prevention and hygiene measures to reduce the frequency of foodborne virus outbreaks should focus on food workers and production site of food products. Pivotal strategies, such as proper investigation, surveillance, and reports on foodborne viral illnesses, are needed in order to develop more accurate measures to detect the presence and pathogenesis of viral infection with detailed descriptions. Moreover, molecular epidemiology and surveillance of food samples may help analysis of public health hazards associated with exposure to foodborne viruses. In this present review, we discuss different aspects of foodborne viral contamination and its impact on human health. This review also aims to improve understanding of foodborne viral infections as major causes of human illness as well as provide descriptions of their control and prevention strategies and rapid detection by advanced molecular techniques. Further, a brief description of methods available for the detection of viruses in food and related matrices is provided.

  4. First evaluation of the serum level of anti-hepatitis B surface antigen after vaccination in Libya.

    Science.gov (United States)

    Madour, A; Alkout, A; Vanin, S

    2013-12-01

    The hepatitis B virus (HBV) vaccination schedule in Libya follows international recommendations (1st dose at birth, 2nd after 1 month and 3rd after 6 months). This research aimed to evaluate the long-term protection of the HBV immunization programme in Tripoli and to determine the best age to administer booster doses. Serum levels of hepatitis B surface antigen were determined in 277 randomly selected children aged 1-12 years. The response to HBV vaccine in 1-3-year-olds was 93.2%, but this declined with age and at 7-9 years after initial vaccination only 53.1% of children had protective titres (> or = 10 mIU/mL). No significant differences between males and females in antibody persistence or response to vaccine were observed. We recommend continuing the HBV vaccination programme and that a booster dose be given to 6-year-old children to ensure maximum protection during the period of school entry and beyond.

  5. Viral epigenetics.

    Science.gov (United States)

    Milavetz, Barry I; Balakrishnan, Lata

    2015-01-01

    DNA tumor viruses including members of the polyomavirus, adenovirus, papillomavirus, and herpes virus families are presently the subject of intense interest with respect to the role that epigenetics plays in control of the virus life cycle and the transformation of a normal cell to a cancer cell. To date, these studies have primarily focused on the role of histone modification, nucleosome location, and DNA methylation in regulating the biological consequences of infection. Using a wide variety of strategies and techniques ranging from simple ChIP to ChIP-chip and ChIP-seq to identify histone modifications, nuclease digestion to genome wide next generation sequencing to identify nucleosome location, and bisulfite treatment to MeDIP to identify DNA methylation sites, the epigenetic regulation of these viruses is slowly becoming better understood. While the viruses may differ in significant ways from each other and cellular chromatin, the role of epigenetics appears to be relatively similar. Within the viral genome nucleosomes are organized for the expression of appropriate genes with relevant histone modifications particularly histone acetylation. DNA methylation occurs as part of the typical gene silencing during latent infection by herpesviruses. In the simple tumor viruses like the polyomaviruses, adenoviruses, and papillomaviruses, transformation of the cell occurs via integration of the virus genome such that the virus's normal regulation is disrupted. This results in the unregulated expression of critical viral genes capable of redirecting cellular gene expression. The redirected cellular expression is a consequence of either indirect epigenetic regulation where cellular signaling or transcriptional dysregulation occurs or direct epigenetic regulation where epigenetic cofactors such as histone deacetylases are targeted. In the more complex herpersviruses transformation is a consequence of the expression of the viral latency proteins and RNAs which again can

  6. In vitro evaluation of a soluble Leishmania promastigote surface antigen as a potential vaccine candidate against human leishmaniasis.

    Science.gov (United States)

    Chamakh-Ayari, Rym; Bras-Gonçalves, Rachel; Bahi-Jaber, Narges; Petitdidier, Elodie; Markikou-Ouni, Wafa; Aoun, Karim; Moreno, Javier; Carrillo, Eugenia; Salotra, Poonam; Kaushal, Himanshu; Negi, Narender Singh; Arevalo, Jorge; Falconi-Agapito, Francesca; Privat, Angela; Cruz, Maria; Pagniez, Julie; Papierok, Gérard-Marie; Rhouma, Faten Bel Haj; Torres, Pilar; Lemesre, Jean-Loup; Chenik, Mehdi; Meddeb-Garnaoui, Amel

    2014-01-01

    PSA (Promastigote Surface Antigen) belongs to a family of membrane-bound and secreted proteins present in several Leishmania (L.) species. PSA is recognized by human Th1 cells and provides a high degree of protection in vaccinated mice. We evaluated humoral and cellular immune responses induced by a L. amazonensis PSA protein (LaPSA-38S) produced in a L. tarentolae expression system. This was done in individuals cured of cutaneous leishmaniasis due to L. major (CCLm) or L. braziliensis (CCLb) or visceral leishmaniasis due to L. donovani (CVLd) and in healthy individuals. Healthy individuals were subdivided into immune (HHR-Lm and HHR-Li: Healthy High Responders living in an endemic area for L. major or L. infantum infection) or non immune/naive individuals (HLR: Healthy Low Responders), depending on whether they produce high or low levels of IFN-γ in response to Leishmania soluble antigen. Low levels of total IgG antibodies to LaPSA-38S were detected in sera from the studied groups. Interestingly, LaPSA-38S induced specific and significant levels of IFN-γ, granzyme B and IL-10 in CCLm, HHR-Lm and HHR-Li groups, with HHR-Li group producing TNF-α in more. No significant cytokine response was observed in individuals immune to L. braziliensis or L. donovani infection. Phenotypic analysis showed a significant increase in CD4+ T cells producing IFN-γ after LaPSA-38S stimulation, in CCLm. A high positive correlation was observed between the percentage of IFN-γ-producing CD4+ T cells and the released IFN-γ. We showed that the LaPSA-38S protein was able to induce a mixed Th1 and Th2/Treg cytokine response in individuals with immunity to L. major or L. infantum infection indicating that it may be exploited as a vaccine candidate. We also showed, to our knowledge for the first time, the capacity of Leishmania PSA protein to induce granzyme B production in humans with immunity to L. major and L. infantum infection.

  7. In Vitro Evaluation of a Soluble Leishmania Promastigote Surface Antigen as a Potential Vaccine Candidate against Human Leishmaniasis

    Science.gov (United States)

    Bahi-Jaber, Narges; Petitdidier, Elodie; Markikou-Ouni, Wafa; Aoun, Karim; Moreno, Javier; Carrillo, Eugenia; Salotra, Poonam; Kaushal, Himanshu; Negi, Narender Singh; Arevalo, Jorge; Falconi-Agapito, Francesca; Privat, Angela; Cruz, Maria; Pagniez, Julie; Papierok, Gérard-Marie; Rhouma, Faten Bel Haj; Torres, Pilar; Lemesre, Jean-Loup; Chenik, Mehdi; Meddeb-Garnaoui, Amel

    2014-01-01

    PSA (Promastigote Surface Antigen) belongs to a family of membrane-bound and secreted proteins present in several Leishmania (L.) species. PSA is recognized by human Th1 cells and provides a high degree of protection in vaccinated mice. We evaluated humoral and cellular immune responses induced by a L. amazonensis PSA protein (LaPSA-38S) produced in a L. tarentolae expression system. This was done in individuals cured of cutaneous leishmaniasis due to L. major (CCLm) or L. braziliensis (CCLb) or visceral leishmaniasis due to L. donovani (CVLd) and in healthy individuals. Healthy individuals were subdivided into immune (HHR-Lm and HHR-Li: Healthy High Responders living in an endemic area for L. major or L. infantum infection) or non immune/naive individuals (HLR: Healthy Low Responders), depending on whether they produce high or low levels of IFN-γ in response to Leishmania soluble antigen. Low levels of total IgG antibodies to LaPSA-38S were detected in sera from the studied groups. Interestingly, LaPSA-38S induced specific and significant levels of IFN-γ, granzyme B and IL-10 in CCLm, HHR-Lm and HHR-Li groups, with HHR-Li group producing TNF-α in more. No significant cytokine response was observed in individuals immune to L. braziliensis or L. donovani infection. Phenotypic analysis showed a significant increase in CD4+ T cells producing IFN-γ after LaPSA-38S stimulation, in CCLm. A high positive correlation was observed between the percentage of IFN-γ-producing CD4+ T cells and the released IFN-γ. We showed that the LaPSA-38S protein was able to induce a mixed Th1 and Th2/Treg cytokine response in individuals with immunity to L. major or L. infantum infection indicating that it may be exploited as a vaccine candidate. We also showed, to our knowledge for the first time, the capacity of Leishmania PSA protein to induce granzyme B production in humans with immunity to L. major and L. infantum infection. PMID:24786587

  8. In vitro evaluation of a soluble Leishmania promastigote surface antigen as a potential vaccine candidate against human leishmaniasis.

    Directory of Open Access Journals (Sweden)

    Rym Chamakh-Ayari

    Full Text Available PSA (Promastigote Surface Antigen belongs to a family of membrane-bound and secreted proteins present in several Leishmania (L. species. PSA is recognized by human Th1 cells and provides a high degree of protection in vaccinated mice. We evaluated humoral and cellular immune responses induced by a L. amazonensis PSA protein (LaPSA-38S produced in a L. tarentolae expression system. This was done in individuals cured of cutaneous leishmaniasis due to L. major (CCLm or L. braziliensis (CCLb or visceral leishmaniasis due to L. donovani (CVLd and in healthy individuals. Healthy individuals were subdivided into immune (HHR-Lm and HHR-Li: Healthy High Responders living in an endemic area for L. major or L. infantum infection or non immune/naive individuals (HLR: Healthy Low Responders, depending on whether they produce high or low levels of IFN-γ in response to Leishmania soluble antigen. Low levels of total IgG antibodies to LaPSA-38S were detected in sera from the studied groups. Interestingly, LaPSA-38S induced specific and significant levels of IFN-γ, granzyme B and IL-10 in CCLm, HHR-Lm and HHR-Li groups, with HHR-Li group producing TNF-α in more. No significant cytokine response was observed in individuals immune to L. braziliensis or L. donovani infection. Phenotypic analysis showed a significant increase in CD4+ T cells producing IFN-γ after LaPSA-38S stimulation, in CCLm. A high positive correlation was observed between the percentage of IFN-γ-producing CD4+ T cells and the released IFN-γ. We showed that the LaPSA-38S protein was able to induce a mixed Th1 and Th2/Treg cytokine response in individuals with immunity to L. major or L. infantum infection indicating that it may be exploited as a vaccine candidate. We also showed, to our knowledge for the first time, the capacity of Leishmania PSA protein to induce granzyme B production in humans with immunity to L. major and L. infantum infection.

  9. Preclinical evaluation of an mRNA HIV vaccine combining rationally selected antigenic sequences and adjuvant signals (HTI-TriMix).

    Science.gov (United States)

    Guardo, Alberto C; Joe, Patrick Tjok; Miralles, Laia; Bargalló, Manel E; Mothe, Beatriz; Krasniqi, Ahmet; Heirman, Carlo; García, Felipe; Thielemans, Kris; Brander, Christian; Aerts, Joeri L; Plana, Montserrat

    2017-01-28

    The development of a prophylactic vaccine against HIV-1 has so far not been successful. Therefore, attention has shifted more and more toward the development of novel therapeutic vaccines. Here, we evaluated a new mRNA-based therapeutic vaccine against HIV-1-encoding activation signals (TriMix: CD40L + CD70 + caTLR4) combined with rationally selected antigenic sequences [HIVACAT T-cell immunogen (HTI)] sequence: comprises 16 joined fragments from Gag, Pol, Vif, and Nef). For this purpose, peripheral blood mononuclear cells from HIV-1-infected individuals on cART, lymph node explants from noninfected humans, and splenocytes from immunized mice were collected and several immune functions were measured. Electroporation of immature monocyte-derived dendritic cells from HIV-infected patients with mRNA encoding HTI + TriMix potently activated dendritic cells which resulted in upregulation of maturation markers and cytokine production and T-cell stimulation, as evidenced by enhanced proliferation and cytokine secretion (IFN-γ). Responses were HIV specific and were predominantly targeted against the sequences included in HTI. These findings were confirmed in human lymph node explants exposed to HTI + TriMix mRNA. Intranodal immunizations with HTI mRNA in a mouse model increased antigen-specific cytotoxic T-lymphocyte responses. The addition of TriMix further enhanced cytotoxic responses. Our results suggest that uptake of mRNA, encoding strong activation signals and a potent HIV antigen, confers a T-cell stimulatory capacity to dendritic cells and enhances their ability to stimulate antigen-specific immunity. These findings may pave the way for therapeutic HIV vaccine strategies based on antigen-encoding RNA to specifically target antigen-presenting cells.

  10. [Evaluation of the efficacies of rapid antigen test, multiplex PCR, and real-time PCR for the detection of a novel influenza A (H1N1) virus].

    Science.gov (United States)

    Hwang, Yusun; Kim, Kyounghee; Lee, Miae

    2010-04-01

    In April 2009, a novel influenza A (H1N1) virus was detected in the US, and at the time of conducting this study, H1N1 infection had reached pandemic proportions. In Korea, rapid antigen tests and PCR assays have been developed to detect the H1N1 virus. We evaluated the efficacies of rapid antigen test, multiplex PCR, and real-time PCR for detecting the H1N1 virus. From August to September 2009, we tested 734 samples obtained from nasopharyngeal swab or nasal swab using rapid antigen test (SD Influenza Antigen, Standard Diagnostics, Inc., Korea) and multiplex PCR (Seeplex FluA ACE Subtyping, Seegene, Korea). We also tested 224 samples using the AdvanSure real-time PCR (LG Life Sciences, Korea) to compare the results obtained using real-time PCR with those obtained using multiplex PCR. Furthermore, 99 samples were tested using the AdvanSure real-time PCR and the AccuPower real-time PCR (Bioneer, Korea). In comparison with the results of multiplex PCR, the sensitivity and specificity of the rapid antigen test were 48.0% and 99.8%, respectively. The concordance rate for multiplex PCR and the AdvanSure real-time PCR was 99.6% (kappa=0.991, P=0.000), and that for the AdvanSure real-time PCR and the AccuPower real-time PCR was 97.0% (kappa=0.936, P=0.000). The rapid antigen test is significantly less sensitive than PCR assay; therefore, it is not useful for H1N1 detection; however multiplex PCR, the AdvanSure real-time PCR, and the Accu-Power real-time PCR can be useful for H1N1 detection.

  11. Viral bronchiolitis.

    Science.gov (United States)

    Florin, Todd A; Plint, Amy C; Zorc, Joseph J

    2017-01-14

    Viral bronchiolitis is a common clinical syndrome affecting infants and young children. Concern about its associated morbidity and cost has led to a large body of research that has been summarised in systematic reviews and integrated into clinical practice guidelines in several countries. The evidence and guideline recommendations consistently support a clinical diagnosis with the limited role for diagnostic testing for children presenting with the typical clinical syndrome of viral upper respiratory infection progressing to the lower respiratory tract. Management is largely supportive, focusing on maintaining oxygenation and hydration of the patient. Evidence suggests no benefit from bronchodilator or corticosteroid use in infants with a first episode of bronchiolitis. Evidence for other treatments such as hypertonic saline is evolving but not clearly defined yet. For infants with severe disease, the insufficient available data suggest a role for high-flow nasal cannula and continuous positive airway pressure use in a monitored setting to prevent respiratory failure. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Development of a stringent ELISA protocol to evaluate anti-viral hemorrhagic septicemia virus-specific antibodies in olive flounder Paralichthys olivaceus with improved specificity.

    Science.gov (United States)

    Kim, Hyoung Jun; Park, Jeong Su; Kwon, Se Ryun

    2015-07-01

    Olive flounder were vaccinated with polyinosinic:polycytidylic acid [Poly (I:C)] to prevent viral hemorrhagic septicemia (VHS). Vaccine efficacy was verified by detection of anti- VHS virus (VHSV) antibodies using enzyme-linked immunosorbent assay (ELISA). In the study, ELISA absorbance values of the negative control group [Poly (I:C)-MEM10] were saturated when an ELISA protocol, that includes pretreatment of the fish sera with 5% skim milk, was used. However, the saturated OD values in the negative control did not correlate with a specific immune response against VHSV, because the group showed low survival rate (only 10%) following the VHSV challenge. Also, OD values of Poly (I:C)- VHSV group were high, and the group showed high survival rate (97.5%) against VHSV challenge test. It was suggested that the high OD values were possibly due to the presence of anti-fetal bovine serum (FBS) cross-reactivity. To compensate this, we subtracted the absorbance of infectious hematopoietic necrosis (IHNV)-Ag plates from those of the VHSV-Ag plates. However, the average value for the Poly (I:C)-VHSV group (0.167) was lower than expected even though high survival rate. We used an advanced ELISA system to pre-treat fish sera with 5% skim milk and two novirhabdoviruses as capture antigens as well as 50% FBS. The corrected absorbance values for pre-treated fish sera from the negative control Poly (I:C)-MEM10 and experimental Poly (I:C)-VHSV groups averaged 0.033 and 0.579, respectively. The specific VHSV antibody response of the vaccinated group was assessed using fish sera pretreated with skim milk and FBS and by calculating the corrected absorbance values from ELISA with two novirhabdovirus capture antigens.

  13. Safety profile of the viral vectors of attenuated fowlpox strain FP9 and modified vaccinia virus Ankara recombinant for either of 2 preerythrocytic malaria antigens, ME-TRAP or the circumsporozoite protein, in children and adults in Kenya.

    Science.gov (United States)

    Bejon, Philip; Peshu, Norbert; Gilbert, Sarah C; Lowe, Brett S; Molyneux, Catherine S; Forsdyke, John; Lang, Trudie; Hill, Adrian V S; Marsh, Kevin

    2006-04-15

    We are developing a heterologous prime-boost vaccine strategy against malaria. This approach uses sequential immunization with different vectors to deliver a common preerythrocytic malaria antigen. Preliminary evidence of efficacy and safety has been previously documented in studies from an area where malaria is nonendemic. Additional safety data from an area where malaria is endemic are now required before larger-scale studies are undertaken to determine the efficacy of this vaccine strategy in the field. Other modified vaccinia virus Ankara (MVA) recombinants and prime-boost immunizations are being developed as vaccines against human immunodeficiency virus (HIV) infection, tuberculosis, and cancer, and MVA is a candidate attenuated smallpox vaccine. Candidate vaccines against malaria were intradermally administered to 73 adults (7 of whom were HIV positive) and 22 children in Kenya. These vaccines used the attenuated fowlpox strain FP9 and the MVA recombinant for either of 2 preerythrocytic malaria antigens, multiple preerythrocytic-stage epitopes joined with the preerythrocytic-stage antigen TRAP (ME-TRAP) and the circumsporozoite protein (CS). Adverse events were recorded. Reactogenicity was mild. MVA caused less frequent and less severe cutaneous reaction if given after FP9 priming. Half doses reduced the frequency and the severity of systemic reactogenicity, and particular vaccine lots were associated with different reactogenicities. Unexpectedly, prior immunity to the ME-TRAP antigen appeared to be protective against local reactions after immunization. Where the final intention is to use MVA after FP9 priming, previous testing of MVA alone overestimates reactogenicity. These recombinant vectors appear to be safe and suitable for use in larger-scale studies of children in Africa and of HIV-positive individuals.

  14. Equine IgE responses to non-viral vaccine components.

    Science.gov (United States)

    Gershwin, Laurel J; Netherwood, Kristina A; Norris, Meredith Somerville; Behrens, Nicole E; Shao, Matt X

    2012-12-14

    Vaccination of horses is performed annually or semi-annually with multiple viral antigens, either in a combination vaccine or as separate injections. While this practice undoubtedly prevents infection from such diseases as rabies, equine influenza, West Nile virus, and equine herpes virus, the procedure is not without repercussions. Hypersensitivity reactions, including fatal anaphylactic shock, after vaccination, although uncommon, have increased in incidence in recent years. Studies reported herein document the development of IgE antibodies against non-target antigen components of equine viral vaccines. We hypothesize that viral vaccines can induce an IgE response to non-target antigens, which could elicit an adverse response after vaccination with another viral vaccine containing the same component. In one study IgE responses to components of West Nile virus vaccine were evaluated by ELISA before and after vaccination in 30 horses. In a second five-year study 77 horses were similarly tested for IgE antibodies against bovine serum albumin (BSA), a component of most viral vaccines. Mast cell sensitization was evaluated in horses with high, moderate, and negative serum BSA specific IgE using an intradermal skin test with BSA. Over the five-year period high IgE responder horses showed gradually increasing BSA specific serum IgE levels and positive skin test reactivity, yet none had an adverse event. Sera from horses that had developed adverse vaccine reactions were also tested for IgE antibodies. Several of these horses had extremely high levels of BSA-specific IgE. These data suggest that non-essential protein components of vaccines may sensitize horses for future adverse responses to vaccination. Copyright © 2012 Elsevier Ltd. All rights reserved.

  15. Evaluation of a rapid diagnostic test, NanoSign® Influenza A/B Antigen, for detection of the 2009 pandemic influenza A/H1N1 viruses

    Science.gov (United States)

    2010-01-01

    Background This study evaluated the clinical accuracy and analytical sensitivity of the NanoSign® Influenza A/B antigen kit in detecting 2009 pandemic influenza A/H1N1 viruses. The kit is one of the most popular rapid diagnostic tests for detecting influenza in Republic of Korea. Results The NanoSign® Influenza A/B kit resulted in 79.4% sensitivity and 97.2% specificity compared to RT-PCR in the detection of the viruses from 1,023 specimens. In addition, the kit was able to detect two strains of novel influenza viruses, Influenza A/California/12/2009(H1N1) and clinically isolated wild-type novel influenza A/H1N1, both of which are spreading epidemically throughout the world. In addition, the correlation between NanoSign® Influenza A/B test and conventional RT-PCR was approximately 94%, indicating a high concordance rate. Analytical sensitivity of the kit was approximately 73 ± 3.65 ng/mL of the purified viral proteins and 1.13 ± 0.11 hemagglutination units for the cultured virus. Conclusions As the NanoSign® Influenza A/B kit showed relatively high sensitivity and specificity and the good correlation with RT-PCR, it will be very useful in the early control of influenza infection and in helping physicians in making early treatment decisions. PMID:20849665

  16. Characterization of an F1 deletion mutant of Yersinia pestis CO92, pathogenic role of F1 antigen in bubonic and pneumonic plague, and evaluation of sensitivity and specificity of F1 antigen capture-based dipsticks.

    Science.gov (United States)

    Sha, Jian; Endsley, Janice J; Kirtley, Michelle L; Foltz, Sheri M; Huante, Matthew B; Erova, Tatiana E; Kozlova, Elena V; Popov, Vsevolod L; Yeager, Linsey A; Zudina, Irina V; Motin, Vladimir L; Peterson, Johnny W; DeBord, Kristin L; Chopra, Ashok K

    2011-05-01

    We evaluated two commercial F1 antigen capture-based immunochromatographic dipsticks, Yersinia Pestis (F1) Smart II and Plague BioThreat Alert test strips, in detecting plague bacilli by using whole-blood samples from mice experimentally infected with Yersinia pestis CO92. To assess the specificities of these dipsticks, an in-frame F1-deficient mutant of CO92 (Δcaf) was generated by homologous recombination and used as a negative control. Based on genetic, antigenic/immunologic, and electron microscopic analyses, the Δcaf mutant was devoid of a capsule. The growth rate of the Δcaf mutant generally was similar to that of the wild-type (WT) bacterium at both 26 and 37 °C, although the mutant's growth dropped slightly during the late phase at 37 °C. The Δcaf mutant was as virulent as WT CO92 in the pneumonic plague mouse model; however, it was attenuated in developing bubonic plague. Both dipsticks had similar sensitivities, requiring a minimum of 0.5 μg/ml of purified F1 antigen or 1 × 10(5) to 5 × 10(5) CFU/ml of WT CO92 for positive results, while the blood samples were negative for up to 1 × 10(8) CFU/ml of the Δcaf mutant. Our studies demonstrated the diagnostic potential of two plague dipsticks in detecting capsular-positive strains of Y. pestis in bubonic and pneumonic plague.

  17. Variables that influence HIV-1 cerebrospinal fluid viral load in cryptococcal meningitis: a linear regression analysis

    Directory of Open Access Journals (Sweden)

    Cecchini Diego M

    2009-11-01

    Full Text Available Abstract Background The central nervous system is considered a sanctuary site for HIV-1 replication. Variables associated with HIV cerebrospinal fluid (CSF viral load in the context of opportunistic CNS infections are poorly understood. Our objective was to evaluate the relation between: (1 CSF HIV-1 viral load and CSF cytological and biochemical characteristics (leukocyte count, protein concentration, cryptococcal antigen titer; (2 CSF HIV-1 viral load and HIV-1 plasma viral load; and (3 CSF leukocyte count and the peripheral blood CD4+ T lymphocyte count. Methods Our approach was to use a prospective collection and analysis of pre-treatment, paired CSF and plasma samples from antiretroviral-naive HIV-positive patients with cryptococcal meningitis and assisted at the Francisco J Muñiz Hospital, Buenos Aires, Argentina (period: 2004 to 2006. We measured HIV CSF and plasma levels by polymerase chain reaction using the Cobas Amplicor HIV-1 Monitor Test version 1.5 (Roche. Data were processed with Statistix 7.0 software (linear regression analysis. Results Samples from 34 patients were analyzed. CSF leukocyte count showed statistically significant correlation with CSF HIV-1 viral load (r = 0.4, 95% CI = 0.13-0.63, p = 0.01. No correlation was found with the plasma viral load, CSF protein concentration and cryptococcal antigen titer. A positive correlation was found between peripheral blood CD4+ T lymphocyte count and the CSF leukocyte count (r = 0.44, 95% CI = 0.125-0.674, p = 0.0123. Conclusion Our study suggests that CSF leukocyte count influences CSF HIV-1 viral load in patients with meningitis caused by Cryptococcus neoformans.

  18. Single hemagglutinin mutations that alter both antigenicity and receptor binding avidity influence influenza virus antigenic clustering.

    Science.gov (United States)

    Li, Yang; Bostick, David L; Sullivan, Colleen B; Myers, Jaclyn L; Griesemer, Sara B; Stgeorge, Kirsten; Plotkin, Joshua B; Hensley, Scott E

    2013-09-01

    The hemagglutination inhibition (HAI) assay is the primary measurement used for identifying antigenically novel influenza virus strains. HAI assays measure the amount of reference sera required to prevent virus binding to red blood cells. Receptor binding avidities of viral strains are not usually taken into account when interpreting these assays. Here, we created antigenic maps of human H3N2 viruses that computationally account for variation in viral receptor binding avidities. These new antigenic maps differ qualitatively from conventional antigenic maps based on HAI measurements alone. We experimentally focused on an antigenic cluster associated with a single N145K hemagglutinin (HA) substitution that occurred between 1992 and 1995. Reverse-genetics experiments demonstrated that the N145K HA mutation increases viral receptor binding avidity. Enzyme-linked immunosorbent assays (ELISA) revealed that the N145K HA mutation does not prevent antibody binding; rather, viruses possessing this mutation escape antisera in HAI assays simply by attaching to cells more efficiently. Unexpectedly, we found an asymmetric antigenic effect of the N145K HA mutation. Once H3N2 viruses acquired K145, an epitope involving amino acid 145 became antigenically dominant. Antisera raised against an H3N2 strain possessing K145 had reduced reactivity to H3N2 strains possessing N145. Thus, individual mutations in HA can influence antigenic groupings of strains by altering receptor binding avidity and by changing the dominance of antibody responses. Our results indicate that it will be important to account for variation in viral receptor binding avidity when performing antigenic analyses in order to identify genuine antigenic differences among influenza virus variants.

  19. Recombinant viruses as vaccines against viral diseases

    Directory of Open Access Journals (Sweden)

    A.P.D. Souza

    2005-04-01

    Full Text Available Vaccine approaches to infectious diseases are widely applied and appreciated. Amongst them, vectors based on recombinant viruses have shown great promise and play an important role in the development of new vaccines. Many viruses have been investigated for their ability to express proteins from foreign pathogens and induce specific immunological responses against these antigens in vivo. Generally, gene-based vaccines can stimulate potent humoral and cellular immune responses and viral vectors might be an effective strategy for both the delivery of antigen-encoding genes and the facilitation and enhancement of antigen presentation. In order to be utilized as a vaccine carrier, the ideal viral vector should be safe and enable efficient presentation of required pathogen-specific antigens to the immune system. It should also exhibit low intrinsic immunogenicity to allow for its re-administration in order to boost relevant specific immune responses. Furthermore, the vector system must meet criteria that enable its production on a large-scale basis. Several viral vaccine vectors have thus emerged to date, all of them having relative advantages and limits depending on the proposed application, and thus far none of them have proven to be ideal vaccine carriers. In this review we describe the potential, as well as some of the foreseeable obstacles associated with viral vaccine vectors and their use in preventive medicine.

  20. Clinical Evaluation of Rapid Diagnostic Test Kit Using the Polysaccharide as a Genus-Specific Diagnostic Antigen for Leptospirosis in Korea, Bulgaria, and Argentina.

    Science.gov (United States)

    Lee, Jin-Woo; Park, Sungman; Kim, Seung Han; Christova, Iva; Jacob, Paulina; Vanasco, Norma B; Kang, Yeon-Mi; Woo, Ye-Ju; Kim, Min Soo; Kim, Young-Jin; Cho, Min-Kee; Kim, Yoon-Won

    2016-02-01

    Leptospirosis, a zoonotic disease that is caused by many serovars which are more than 200 in the world, is an emerging worldwide disease. Accurate and rapid diagnostic tests for leptospirosis are a critical step to diagnose the disease. There are some commercial kits available for diagnosis of leptospirosis, but the obscurity of a species- or genus-specific antigen of pathogenic Leptospira interrogans causes the reduced sensitivity and specificity. In this study, the polysaccharide derived from lipopolysaccharide (LPS) of nonpathogenic Leptospira biflexa serovar patoc was prepared, and the antigenicity was confirmed by immunoblot and enzyme linked immunosorbent assay (ELISA). The performance of the rapid diagnostic test (RDT) kit using the polysaccharide as a diagnostic antigen was evaluated in Korea, Bulgaria and Argentina. The sensitivity was 93.9%, 100%, and 81.0% and the specificity was 97.9%, 100%, and 95.4% in Korea (which is a rare region occurring with 2 serovars mostly), Bulgaria (epidemic region with 3 serovars chiefly) and Argentina (endemic region with 19 serovars mainly) respectively. These results indicate that this RDT is applicable for global diagnosis of leptospirosis. This rapid and effective diagnosis will be helpful for diagnosis and manage of leptospirosis to use and the polysaccharide of Leptospira may be called as genus specific antigen for diagnosis.

  1. Synthesis, antiviral evaluation and molecular docking studies of N4-aryl substituted/unsubstituted thiosemicarbazones derived from 1-indanones as potent anti-bovine viral diarrhea virus agents.

    Science.gov (United States)

    Soraires Santacruz, María C; Fabiani, Matías; Castro, Eliana F; Cavallaro, Lucía V; Finkielsztein, Liliana M

    2017-08-01

    A series of N4-arylsubstituted thiosemicarbazones derived from 1-indanones and a set of compounds lacking such substitution in the N4 position of the thiosemicarbazone moiety were synthesized and evaluated for their anti-bovine viral diarrhea virus (BVDV) activity. Among these, derivatives 2 and 15 displayed high activity (EC50=2.7±0.4 and 0.7±0.1µM, respectively) as inhibitors of BVDV replication. Novel key structural features related to the anti-BVDV activity were identified by structure-activity relationship (SAR) analysis. In a previous study, the thiosemicarbazone of 5,6-dimethoxy-1-indanone (5,6-TSC) was characterized as a non-nucleoside inhibitor (NNI) of the BVDV RNA-dependent RNA polymerase. In the present work, cross-resistance assays were performed with the most active compounds. Such studies were carried out on 5,6-TSC resistant BVDV (BVDV-TSCr T1) carrying mutations in the viral polymerase. This BVDV mutant was also resistant to compound 15. Molecular docking studies and MM/PBSA calculations were performed to assess the most active derivatives at the 5,6-TSC viral polymerase binding site. The differences in the interaction pattern and the binding affinity of derivative 15 either to the wild type or BVDV-TSCr T1 polymerase were key factors to define the mode of action of this compound. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Evaluation of a point-of-care immunoassay test kit 'StrongStep' for cryptococcal antigen detection.

    Directory of Open Access Journals (Sweden)

    Edward Mpoza

    Full Text Available HIV-associated cryptococcal meningitis is the leading cause of adult meningitis in Sub-Saharan Africa, accounting for 15%-20% of AIDS-attributable mortality. The development of point-of-care assays has greatly improved the screening and diagnosis of cryptococcal disease. We evaluated a point-of-care immunoassay, StrongStep (Liming Bio, Nanjing, Jiangsu, China lateral flow assay (LFA, for cryptococcal antigen (CrAg detection in cerebrospinal fluid (CSF and plasma.We retrospectively tested 143 CSF and 77 plasma samples collected from HIV-seropositive individuals with suspected meningitis from 2012-2016 in Uganda. We prospectively tested 90 plasma samples collected from HIV-seropositive individuals with CD4 cell count <100 cells/μL from 2016-2017 as part of a cryptococcal antigenemia screening program. The StrongStep CrAg was tested against a composite reference standard of positive Immy CrAg LFA (Immy, Norman, OK, USA or CSF culture with statistical comparison by McNemar's test.StrongStep CrAg had a 98% (54/55 sensitivity and 90% (101/112 specificity in plasma (P = 0.009, versus reference standard. In CSF, the StrongStep CrAg had 100% (101/101 sensitivity and 98% (41/42 specificity (P = 0.99. Adjusting for the cryptococcal antigenemia prevalence of 9% in Uganda and average cryptococcal meningitis prevalence of 37% in Sub-Saharan Africa, the positive predictive value of the StrongStep CrAg was 50% in plasma and 96% in CSF.We found the StrongStep CrAg LFA to be a sensitive assay, which unfortunately lacked specificity in plasma. In lower prevalence settings, a majority of positive results from blood would be expected to be false positives.

  3. A Latent Markov Modelling approach to the evaluation of Circulating Cathodic Antigen strips for Schistosomiasis diagnosis pre- and post-praziquantel treatment in Uganda

    DEFF Research Database (Denmark)

    Koukounari, Artemis; Donnelly, Christl A.; Moustaki, Irini

    2013-01-01

    Markov Models (LMMs). The aim was to evaluate the diagnostic accuracy of Circulating Cathodic Antigen (CCA) and of double Kato-Katz (KK) faecal slides over three consecutive days for Schistosoma mansoni infection simultaneously by age group at baseline and at two follow-up times post treatment...... as well as CCA for mapping surveys of S. mansoni infection, although additional diagnostic tools should be incorporated in schistosomiasis elimination programs....

  4. Evaluation of new monoclonal antibody-based latex agglutination test for detection of cryptococcal polysaccharide antigen in serum and cerebrospinal fluid.

    OpenAIRE

    Kiska, D L; Orkiszewski, D R; Howell, D; Gilligan, P H

    1994-01-01

    We evaluated the performance of CRYPTO-LEX (Trinity Laboratories, Inc., Raleigh, N. C.), a new mouse immunoglobulin M monoclonal antibody latex agglutination reagent which reacts with the capsular polysaccharide of the four serogroups of Cryptococcus neoformans. This test was compared with CALAS (Meridian Diagnostics, Cincinnati, Ohio) for the ability to detect cryptococcal antigen in serum and cerebrospinal fluid (CSF). A total of 580 clinical specimens (327 serum and 253 CSF samples), prima...

  5. Comparative evaluation of Strongyloides ratti and S. stercoralis larval antigen for diagnosis of strongyloidiasis in an endemic area of opisthorchiasis.

    Science.gov (United States)

    Eamudomkarn, Chatanun; Sithithaworn, Paiboon; Sithithaworn, Jiraporn; Kaewkes, Sasithorn; Sripa, Banchob; Itoh, Makoto

    2015-07-01

    The use of Strongyloides ratti as heterologous antigen for serodiagnosis of strongyloidiasis is preferable to Strongyloides from humans due to the ease and safety of antigen preparation. In Southeast Asia where Opisthorchis viverrini coexists with Strongyloides stercoralis, there has been no report in using S. ratti for serodiagnosis of S. stercoralis. In this study, performance of an enzyme-linked immunosorbent assay (ELISA) based on S. ratti was compared with that based on S. stercoralis for diagnosis of strongyloidiasis in areas where O. viverrini is co-endemic in Thailand. Of the 107 individuals, 50 (46.7 %) were positive for S. stercoralis by agar culture method and by ELISA; 82 (76.6 %) and 81 (75.7 %) were seropositive using S. ratti and S. stercoralis antigens, respectively. The levels of parasite-specific IgG to S. ratti and S. stercoralis antigen were significantly proportionally correlated (P < 0.001). Mixed infections with O. viverrini have little effect on diagnosis of strongyloidiasis. Of 42 subjects who were infected with other parasites, there were no cross-reaction with Angiostrongylus cantonensis, Taenia spp., hookworms, Paragonimus spp., Clonorchis sinensis, Ascaris lumbricoides except for Fasciola spp. (1 of 5), and Opisthorchis viverrini (5 of 20). In spite of cross-reactivities, the results suggest that the S. ratti antigen provides an useful option for diagnosis of strongyloidiasis in an endemic area of opisthorchiasis with high sensitivity comparable to the S. stercoralis antigen and provide a basis for effective control strategies for strongyloidiasis.

  6. Cloning, expression and characterization of SeM protein of Streptococcus equi subsp. equi and evaluation of its use as antigen in an indirect ELISA

    Directory of Open Access Journals (Sweden)

    C.M. Moraes

    2014-08-01

    Full Text Available Strangles is an economically important horse disease caused by Streptococcus equi subsp. equi. The diagnosis can be confirmed either directly by bacterial isolation and PCR or by ELISA, which is an indirect method based on the detection of serum antibodies. The aim of this study was to clone, express and characterize the SeM protein of Streptococcus equi subsp. equi, evaluate its use as antigen in indirect ELISA and determine its performance to distinguish sera of negative, vaccinated and positive animals. This was initially performed by cloning the gene encoding the SeM protein and its expression in Escherichia coli. Subsequently, the protein produced was characterized and used as antigen in ELISA. Serum samples for evaluation were taken from 40 negative foals, 46 horses vaccinated with a commercial vaccine against strangles and 46 horses diagnosed with the disease. The test showed high specificity and sensitivity, allowing discrimination between negative and positive, positive and vaccinated animals, and vaccinated animals and negative sera. Thus, it was concluded that the protein produced rSeM, which can be used as antigen for disease diagnosis, and the described ELISA might be helpful to evaluate the immune status of the herd.

  7. Differences in viral load among human respiratory syncytial virus genotypes in hospitalized children with severe acute respiratory infections in the Philippines.

    Science.gov (United States)

    Kadji, Francois Marie Ngako; Okamoto, Michiko; Furuse, Yuki; Tamaki, Raita; Suzuki, Akira; Lirio, Irene; Dapat, Clyde; Malasao, Rungnapa; Saito, Mariko; Pedrera-Rico, Gay Anne Granada; Tallo, Veronica; Lupisan, Socorro; Saito, Mayuko; Oshitani, Hitoshi

    2016-06-27

    Human respiratory syncytial virus (HRSV) is a leading viral etiologic agent of pediatric lower respiratory infections, including bronchiolitis and pneumonia. Two antigenic subgroups, HRSV-A and B, each contain several genotypes. While viral load may vary among HRSV genotypes and affect the clinical course of disease, data are scarce regarding the actual differences among genotypes. Therefore, this study estimated and compared viral load among NA1 and ON1 genotypes of HRSV-A and BA9 of HRSV-B. ON1 is a newly emerged genotype with a 72-nucleotide duplication in the G gene as observed previously with BA genotypes in HRSV-B. Children <5 years of age with an initial diagnosis of severe or very severe pneumonia at a hospital in the Philippines from September 2012 to December 2013 were enrolled. HRSV genotypes were determined and the viral load measured from nasopharyngeal swabs (NPS). The viral load of HRSV genotype NA1 were significantly higher than those of ON1 and BA9. Regression analysis showed that both genotype NA1 and younger age were significantly associated with high HRSV viral load. The viral load of NA1 was higher than that of ON1 and BA9 in NPS samples. HRSV genotypes may be associated with HRSV viral load. The reasons and clinical impacts of these differences in viral load among HRSV genotypes require further evaluation.

  8. [Functional analysis of hepatitis B virus immune escape mutants with insertion mutations in the surface antigen].

    Science.gov (United States)

    Yu, Shu-li; Yu, De-min; Zhang, Dong-hua; Jiang, Jie-hong; Chen, Jia; Deng, Lin; Zhang, Xin-xin

    2013-07-01

    To evaluate the influence of insertion mutations occurring in the hydrophobic region, between amino acids 114 and 115, of the hepatitis B surface antigen (HBsAg) on viral antigenicity and replication. Hepatitis B virus (HBV) DNA was obtained from patients with HBsAg-positive chronic hepatitis B (CHB) infection and subjected to sequence analysis and comparison to GenBank reference sequences for HBV genotype B (AB073826) and genotype C (AF286594). Insertion mutations detected in the HBsAg region were used to make recombinant expression plasmids via site-directed mutagenesis. After transfecting the recombinant HBsAg into Huh7 cells, the mutants' effects on viral antigenicity and replication were evaluated by chemiluminescence microparticle immunoassay (CMIA) and Southern blot hybridization, respectively. The viral antigenicity of each mutant was predicted by bioinformatic analysis, using the Jameson-Wolf method to predict the antigenic index, the Hopp-Woods method to predict hydrophilicity, the Emini method to predict the probability of a region lying of the protein's surface, and the Karplus-Schulz method to predict the flexibility of the protein backbone. Two CHB patients harbored HBV with insertion mutations in HBsAg: one with two (NT) and one with three (NTT) inserted amino acids between 114 and 115. The NTT recombinant HBsAg mutant showed no impact on viral replication and reacted weakly with anti-HBs in CMIA (P = 0.02). The antigen indices for the insertion of NTT were 1.00, -0.16, and 0.18, and insertion of the three amino acids affected the index values of five proximal amino acid sites (with an average increase of 0.13). The hydrophilic indices for the insertion of NTT were 0.2, -0.4, and -0.4, with no significant effect on the proximal amino acids. The insertion of the three amino acids changed both the surface probability (range: -0.55 to 2.97; affecting eight proximal amino acids) and the flexibility (range: -0.01 to 1.1; affecting five proximal amino

  9. Pre-clinical evaluation of CD38 chimeric antigen receptor engineered T cells for the treatment of multiple myeloma

    DEFF Research Database (Denmark)

    Drent, Esther; Groen, Richard W. J.; Noort, Willy A. Noort

    2016-01-01

    Adoptive transfer of chimeric antigen receptor-transduced T cells is a promising strategy for cancer immunotherapy. The CD38 molecule, with its high expression on multiple myeloma cells, appears a suitable target for antibody therapy. Prompted by this, we used three different CD38 antibody...

  10. Pre-clinical evaluation of CD38 chimeric antigen receptor engineered T cells for the treatment of multiple myeloma

    DEFF Research Database (Denmark)

    Drent, Esther; Groen, Richard W. J.; Noort, Willy A. Noort

    2016-01-01

    Adoptive transfer of chimeric antigen receptor-transduced T cells is a promising strategy for cancer immunotherapy. The CD38 molecule, with its high expression on multiple myeloma cells, appears a suitable target for antibody therapy. Prompted by this, we used three different CD38 antibody sequen...

  11. Low cost tuberculosis vaccine antigens in capsules: expression in chloroplasts, bio-encapsulation, stability and functional evaluation in vitro.

    Directory of Open Access Journals (Sweden)

    Priya Saikumar Lakshmi

    Full Text Available Tuberculosis (TB caused by Mycobacterium tuberculosis is one of the leading fatal infectious diseases. The development of TB vaccines has been recognized as a major public health priority by the World Health Organization. In this study, three candidate antigens, ESAT-6 (6 kDa early secretory antigenic target and Mtb72F (a fusion polyprotein from two TB antigens, Mtb32 and Mtb39 fused with cholera toxin B-subunit (CTB and LipY (a cell wall protein were expressed in tobacco and/or lettuce chloroplasts to facilitate bioencapsulation/oral delivery. Site-specific transgene integration into the chloroplast genome was confirmed by Southern blot analysis. In transplastomic leaves, CTB fusion proteins existed in soluble monomeric or multimeric forms of expected sizes and their expression levels varied depending upon the developmental stage and time of leaf harvest, with the highest-level of accumulation in mature leaves harvested at 6PM. The CTB-ESAT6 and CTB-Mtb72F expression levels reached up to 7.5% and 1.2% of total soluble protein respectively in mature tobacco leaves. Transplastomic CTB-ESAT6 lettuce plants accumulated up to 0.75% of total leaf protein. Western blot analysis of lyophilized lettuce leaves stored at room temperature for up to six months showed that the CTB-ESAT6 fusion protein was stable and preserved proper folding, disulfide bonds and assembly into pentamers for prolonged periods. Also, antigen concentration per gram of leaf tissue was increased 22 fold after lyophilization. Hemolysis assay with purified CTB-ESAT6 protein showed partial hemolysis of red blood cells and confirmed functionality of the ESAT-6 antigen. GM1-binding assay demonstrated that the CTB-ESAT6 fusion protein formed pentamers to bind with the GM1-ganglioside receptor. The expression of functional Mycobacterium tuberculosis antigens in transplastomic plants should facilitate development of a cost-effective and orally deliverable TB booster vaccine with potential

  12. Low Cost Tuberculosis Vaccine Antigens in Capsules: Expression in Chloroplasts, Bio-Encapsulation, Stability and Functional Evaluation In Vitro

    Science.gov (United States)

    Yang, Xiangdong; Lloyd, Bethany; Daniell, Henry

    2013-01-01

    Tuberculosis (TB) caused by Mycobacterium tuberculosis is one of the leading fatal infectious diseases. The development of TB vaccines has been recognized as a major public health priority by the World Health Organization. In this study, three candidate antigens, ESAT-6 (6kDa early secretory antigenic target) and Mtb72F (a fusion polyprotein from two TB antigens, Mtb32 and Mtb39) fused with cholera toxin B-subunit (CTB) and LipY (a cell wall protein) were expressed in tobacco and/or lettuce chloroplasts to facilitate bioencapsulation/oral delivery. Site-specific transgene integration into the chloroplast genome was confirmed by Southern blot analysis. In transplastomic leaves, CTB fusion proteins existed in soluble monomeric or multimeric forms of expected sizes and their expression levels varied depending upon the developmental stage and time of leaf harvest, with the highest-level of accumulation in mature leaves harvested at 6PM. The CTB-ESAT6 and CTB-Mtb72F expression levels reached up to 7.5% and 1.2% of total soluble protein respectively in mature tobacco leaves. Transplastomic CTB-ESAT6 lettuce plants accumulated up to 0.75% of total leaf protein. Western blot analysis of lyophilized lettuce leaves stored at room temperature for up to six months showed that the CTB-ESAT6 fusion protein was stable and preserved proper folding, disulfide bonds and assembly into pentamers for prolonged periods. Also, antigen concentration per gram of leaf tissue was increased 22 fold after lyophilization. Hemolysis assay with purified CTB-ESAT6 protein showed partial hemolysis of red blood cells and confirmed functionality of the ESAT-6 antigen. GM1-binding assay demonstrated that the CTB-ESAT6 fusion protein formed pentamers to bind with the GM1-ganglioside receptor. The expression of functional Mycobacterium tuberculosis antigens in transplastomic plants should facilitate development of a cost-effective and orally deliverable TB booster vaccine with potential for long

  13. Development of oral microencapsulated forms for delivering viral vaccines.

    Science.gov (United States)

    Nechaeva, Elena

    2002-10-01

    Rapid development in biotechnology during the last decade has allowed novel ideas in the development of antiviral vaccines to be considered and provides interesting technological approaches to their realization. Designing of microencapsulated forms for delivering bacterial and viral antigens or antigenic complexes using biodegradable biopolymers is an important novel direction. This approach involves the production of polymeric spherical particles with a diameter of 1 microm to 3 mm, containing isolated viral antigens or whole viral particles. Microencapsulated antigens administered orally are protected from low pH values of the gastric juice, bile acids, their salts and proteolytic enzymes of the gastrointestinal tract. The ability to drastically potentiate the immune response to encapsulated antigens, together with the ability to penetrate into the intestinal and respiratory mucosae upon oral and tracheal administrations, respectively, with induction of local and systemic immune reactions are the special merits of such polymers. However, the majority of data on microencapsulated viral vaccines has so far been obtained in animal models, as well as a limited number of studies on the protective effect they elicit. Certain success in the development of vaccines against a number of human viral infections, such as hepatitis B, cytomegalovirus and rotavirus, gives hope to successful completion of this research. Presumably, such vaccines will be safe and innocuous, simple in administration and capable of inducing both the systemic and local immune responses at the primary portal of viral infection.

  14. Universal real-time PCR assay for quantitation and size evaluation of residual cell DNA in human viral vaccines.

    Science.gov (United States)

    André, Murielle; Reghin, Sylviane; Boussard, Estelle; Lempereur, Laurent; Maisonneuve, Stéphane

    2016-05-01

    Residual host cellular DNA (rcDNA) is one of the principal risk associated with continuous cell lines derived medicines such as viral vaccines. To assess rcDNA degradation, we suggest two quantitative real-time PCR assays designed to separately quantify target sequences shorter and longer than the 200 bp risk limit, the relative abundance of both targets reflecting the extent of rcDNA fragmentation. The conserved multicopy ribosomal 18S RNA gene was targeted to detect host cell templates from most mammalian cell substrates commonly used in the manufacture of human viral vaccines. The detection range of the method was assessed on purified DNA templates from different animal origins. The standard calibrator origin and structural conformation were shown crucial to achieve accurate quantification. Artificial mixtures of PCR products shorter and longer than 200 bp were used as a model to check the ability of the assay to estimate the fragment size distribution. The method was successfully applied to a panel of Vero cell derived vaccines and could be used as a universal method for determination of both content and size distribution of rcDNA in vaccines. Copyright © 2016 International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

  15. Development and Evaluation of Quality Metrics for Bioinformatics Analysis of Viral Insertion Site Data Generated Using High Throughput Sequencing.

    Science.gov (United States)

    Gao, Hongyu; Hawkins, Troy; Jasti, Aparna; Chen, Yu-Hsiang; Mockaitis, Keithanne; Dinauer, Mary; Cornetta, Kenneth

    2014-05-06

    Integration of viral vectors into a host genome is associated with insertional mutagenesis and subjects in clinical gene therapy trials must be monitored for this adverse event. Several PCR based methods such as ligase-mediated (LM) PCR, linear-amplification-mediated (LAM) PCR and non-restrictive (nr) LAM PCR were developed to identify sites of vector integration. Coupling the power of next-generation sequencing technologies with various PCR approaches will provide a comprehensive and genome-wide profiling of insertion sites and increase throughput. In this bioinformatics study, we aimed to develop and apply quality metrics to viral insertion data obtained using next-generation sequencing. We developed five simple metrics for assessing next-generation sequencing data from different PCR products and showed how the metrics can be used to objectively compare runs performed with the same methodology as well as data generated using different PCR techniques. The results will help researchers troubleshoot complex methodologies, understand the quality of sequencing data, and provide a starting point for developing standardization of vector insertion site data analysis.

  16. Entamoeba histolytica: cloning, expression and evaluation of the efficacy of a recombinant amebiasis cysteine proteinase gene (ACP1) antigen in minipig.

    Science.gov (United States)

    He, Guang-Zhi

    2012-02-01

    The amebiasis cysteine proteinase gene (ACP1) encoding an antigen from Entamoeba histolytica, as well as the recombinant ACP1, obtained by cloning and expression of the ACP1 gene in heterologous host Escherichia coli BL-21 (DE3), were used to evaluate their ability to induce immune protective responses in minipig against challenge infection in a minipig -E. histolytica model. There was a 64.52% reduction (P<0.001) in the group of recovery of challenged E. histolytica compared with that in the control group. Specific anti-ACP1 antibodies from immune protected minipig had significantly higher levels of immunoglobulin G (IgG) (P<0.001). Our data indicate recombinant ACP1 may be a potential target as a vaccine antigen. Copyright © 2011 Elsevier Inc. All rights reserved.

  17. Evaluation of circulating cathodic antigen (CCA) urine-tests for diagnosis of Schistosoma mansoni infection in Cameroon.

    Science.gov (United States)

    Tchuem Tchuenté, Louis-Albert; Kueté Fouodo, Césaire Joris; Kamwa Ngassam, Romuald Isaka; Sumo, Laurentine; Dongmo Noumedem, Calvine; Kenfack, Christian Mérimé; Gipwe, Nestor Feussom; Nana, Esther Dankoni; Stothard, J Russell; Rollinson, David

    2012-01-01

    The Kato-Katz is the most common diagnostic method for Schistosoma mansoni infection. However, the day-to-day variability in host egg-excretion and its low detection sensitivity are major limits for its use in low transmission zones and after widespread chemotherapy. We evaluated the accuracy of circulating cathodic antigen (CCA) urine-assay as a diagnostic tool of S. mansoni. In comparison, a low sensitive CCA test (CCA-L) was assessed. THE STUDY WAS CONDUCTED IN THREE SETTINGS: two foci with single S. mansoni infections (settings A and B), and one mixed S. mansoni - S. haematobium focus (setting C). Stool and urine samples were collected from school-children on three consecutive days. Triplicate Kato-Katz readings were performed per stool sample. Each urine sample was tested with one CCA and only the first urine sample was subjected to CCA-L. Urine samples were also examined for S. haematobium eggs using the filtration method and for microhaematuria using urine reagent strips. Overall, 625 children provided three stool and three urine samples. Considering nine Kato-Katz thick smears as 'reference' diagnostic test, the prevalence of S. mansoni was 36.2%, 71.8% and 64.0% in settings A, B and C, respectively. The prevalence of S. haematobium in setting C was 12.0%. The sensitivities of single Kato-Katz, CCA and CCA-L from the first stool or urine samples were 58%, 82% and 46% in setting A, 56.8%, 82.4% and 68.8% in setting B, and 49.0%, 87.7% and 55.5% in setting C. The respective specificities were 100%, 64.7% and 100%; 100%, 62.3% and 91.3%; and 100%, 42.5% and 92.0%. Mixed infection with S. haematobium did not influence the CCA test results for S. mansoni diagnosis. Urine CCA revealed higher sensitivity than CCA-L and triplicate Kato-Katz, and produced similar prevalence as nine Kato-Katz. It seems an attractive method for S. mansoni diagnosis.

  18. Enumeration of antigen-specific CD8+ T lymphocytes by single-platform, HLA tetramer-based flow cytometry: a European multicenter evaluation.

    Science.gov (United States)

    Heijnen, Ingmar A F M; Barnett, David; Arroz, Maria J; Barry, Simon M; Bonneville, Marc; Brando, Bruno; D'hautcourt, Jean-Luc; Kern, Florian; Tötterman, Thomas H; Marijt, Erik W A; Bossy, David; Preijers, Frank W M B; Rothe, Gregor; Gratama, Jan W

    2004-11-01

    HLA class I peptide tetramers represent powerful diagnostic tools for detection and monitoring of antigen-specific CD8(+) T cells. The impetus for the current multicenter study is the critical need to standardize tetramer flow cytometry if it is to be implemented as a routine diagnostic assay. Hence, the European Working Group on Clinical Cell Analysis set out to develop and evaluate a single-platform tetramer-based method that used cytomegalovirus (CMV) as the antigenic model. Absolute numbers of CMV-specific CD8(+) T cells were obtained by combining the percentage of tetramer-binding cells with the absolute CD8(+) T-cell count. Six send-outs of stabilized blood from healthy individuals or CMV-carrying donors with CMV-specific CD8(+) T-cell counts of 3 to 10 cells/microl were distributed to 7 to 16 clinical sites. These sites were requested to enumerate CD8(+) T cells and, in the case of CMV-positive donors, CMV-specific subsets on three separate occasions using the standard method. Between-site coefficients of variation of less than 10% (absolute CD8(+) T-cell counts) and approximately 30% (percentage and absolute numbers of CMV-specific CD8(+) T cells) were achieved. Within-site coefficients of variation were approximately 5% (absolute CD8(+) T-cell counts), approximately 9% (percentage CMV-specific CD8(+) T cells), and approximately 17% (absolute CMV-specific CD8(+) T-cell counts). The degree of variation tended to correlate inversely with the proportion of CMV-specific CD8(+) T-cell subsets. The single-platform MHC tetramer-based method for antigen-specific CD8(+) T-cell counting has been evaluated by a European group of laboratories and can be considered a reproducible assay for routine enumeration of antigen-specific CD8(+) T cells. (c) 2004 Wiley-Liss, Inc.

  19. "Purification and evaluation of somatic, excretory-secretory and Cysteine proteinase antigens of Fasciola Hepatica using IgG-ELISA in diagnosing Fascioliasis "

    Directory of Open Access Journals (Sweden)

    "Rokni MB

    2001-08-01

    Full Text Available Fasciolosis, or liver fluke disease, caused by parasites of the genus Fasciola is emerging as an important disease in man and animals, in the world and Iran, particularly in nortern parts. The economical losses in domestic animals are considerable. In the recent decade there were two major outbreaks of human fasciolosis in the Caspian region, northern part of Iran with 7000-10000 infected cases. Sicne it is impossible to diagnose fasciolosis in acute phase using coprological methods and even in chronic phases its sensitivity is low, evaluating and establishing a reliable and cost-effetive test is indispensable and notewortly.In the present survey, we produced and examined the sensitivity and specificity of liver fluke homogenate (LFH , excretory-secetory (ES and cysteine proteinase (CP antigens of F. hepatica using IgG-ELISA test. A 25-27 kilo Dalton coomassie blue-stained band was observed and using of specific inhibitors indicated that this antigen belongs to the class of cysteine proteinase. The sensitivity of LFH, ES and CP antigen in IgG-ELISa was 100% for each, while their specificity was 97.8%, 98.8% and 98.8% respectively. There was a significant difference in mean OD values between cases of proven fasciolosis and other true negative cases, including healthy control individuals and patients with other parasitic diseases.This present report is the first to demonstrate the purification and evaluation of F. hepatica cysteine proteinase antigen by IgG-ELISA test for the diagnosis of fasciolosis in Iran. In conclusion, the IgG-ELISa using ES and CP show high sensitivity and specificity and would be a valuable tool to diagnose human fasciolosis in Iran, particularly in endemic areas.

  20. T cells specific for lipid antigens.

    Science.gov (United States)

    Mori, Lucia; De Libero, Gennaro

    2012-09-01

    Lipid-specific T cells are important participants in human immune responses. Recognition of lipid antigens contributes to host defense against pathogens that can cause debilitating diseases, including mycobacterial, viral, and parasitic infections. Lipid-specific T cells also play important roles in various autoimmune diseases, atherosclerosis, and in tumor surveillance. A better understanding of the mechanisms that regulate lipid-reactive T-cell functions will enable the development of novel therapies across a wide range of diseases. In recent years, our laboratory has investigated lipid antigen specificities, mechanisms of lipid antigen presentation, molecular interaction of lipid antigens with CD1 antigen-presenting molecules, and the pathogenic and regulatory functions of lipid-specific T cells in a variety of disease settings. In this review, we present recent data that illustrate the critical role played by lipid-specific immune responses in host protection, with a particular focus on human studies.

  1. Evaluation of Vero-cell-derived simian endogenous retrovirus infection in humans by detection of viral genome in clinicopathological samples and commercialized vaccines and by serology of Japanese general population.

    Science.gov (United States)

    Fukumoto, Hitomi; Hishima, Tsunekazu; Hasegawa, Hideki; Saeki, Hidehisa; Kuroda, Makoto; Katano, Harutaka

    2016-05-23

    Vero cells are used in laboratories for the isolation of viruses and the production of vaccines. Recently, the sequence of simian endogenous retrovirus (SERV) was identified in Vero cells (SERVagm-Vero), with homology to exogenously transmitted, pathogenic simian retroviruses (SRVs). Although SERVagm-Vero was shown to be noninfectious to human cells in vitro, SERV infection in humans is controversial. In this study, we evaluated the status of SERV infection in humans by detecting the viral genome in clinicopathological samples and commercialized vaccines, and with a serological survey of the Japanese general population. Real-time polymerase chain reaction (PCR) and reverse transcription-PCR were used to detect the SERVagm-Vero genome. We also examined the seroprevalence of SERV in 1000 individuals in the Japanese population with an enzyme-linked immunoabsorbent assay (ELISA) using mixed SERVagm-Vero gag, pol, and env proteins as antigens. Real-time PCR failed to detect SERVagm-Vero genomic fragments in 783 human clinicopathological samples, and all 13 human cell lines tested were negative for the SERVagm-Vero genome. Thirteen commercialized vaccines, including five Vero-based vaccines, were also negative for the SERVagm-Vero genome on real-time PCR and reverse transcription-PCR. Eight (0.8%) were seropositive on ELISA, and western blotting showed that all eight sera contained anti-pol antibodies. All SERV-seropositive individuals were born before 1965, suggesting that SERV infection in Japan might not be associated with vaccine, because more than 90% of Japanese children born from 1964 to 2012 have received live poliovirus vaccines containing virus produced in Vero cells since the 1980s. We have confirmed that the vaccines we use today are free of SERVagm-Vero. Moreover, SERV infection in humans is very rare and unlikely to be associated with vaccines in the Japanese general population. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. [Presence of Australia antigen in blood donors].

    Science.gov (United States)

    Gota, F

    1980-01-01

    The differential diagnosis of type A and B viral hepatitis is discussed and guidelines for the prevention of post-transfusional hospital hepatitis are proposed. Methods for the immunological demonstration of HBs antigen are illustrated, together with the respective positivity percentages in blood donors.

  3. A Multicenter, Prospective, Randomized Controlled Trial to Evaluate the Additional Benefit of a Multistrain Synbiotic (Prodefen® in the Clinical Management of Acute Viral Diarrhea in Children

    Directory of Open Access Journals (Sweden)

    Emilia García-Menor MD

    2016-11-01

    Full Text Available This randomized, open-label study evaluated the additional benefits of the synbiotic Prodefen® in the clinical management of acute diarrhea of suspected viral origin in children between 6 months and 12 years of age. Study outcomes included the duration of diarrhea, the recovery from diarrhea, and the tolerability and acceptance of the treatment. The proportion of patients without diarrhea over the study period was greater in the synbiotic group than in the control group at all study time points, showing a statistically significant difference on the fifth day (95% vs 79%, p < 0.001. The duration of diarrhea (median and interquartile range was reduced by 1 day in the synbiotic-treated patients (3 [2-5] vs 4 [3-5], p = 0.377. The tolerability of the treatment regimen, as evaluated by the parents, was significantly better in those receiving the synbiotic than in the control group. Overall, 96% of the parents of children receiving the synbiotic reported being satisfied to very satisfied with the treatment regimen. The results of this study indicate that the addition of the synbiotic Prodefen® is a well-tolerated and well-accepted approach that provides an additional benefit to the standard supportive therapy in the management of acute viral diarrhea in children.

  4. Serological responses in calves to vaccines against bovine respiratory syncytial, infectious bovine rhinotracheitis, bovine viral diarrhoea and parainfluenza-3 viruses.

    Science.gov (United States)

    Tollis, M; Di Trani, L; Cordioli, P; Vignolo, E; Di Pasquale, I

    1996-01-01

    The Istituto Superiore di Sanità (ISS), the National Veterinary Services Laboratory in Italy, is in charge of assessing the quality, safety and efficacy of veterinary vaccines before and after licensing. To evaluate the relative potency of several vaccines against bovine respiratory syncytial virus (BRSV), infectious bovine rhinotracheitis virus (IBRV), bovine viral diarrhoea virus (BVDV) and parainfluenza-3 virus (PI3V), the serological responses in vaccinated calves were studied. Vaccination with any of the vaccines under study induced specific antibody titres against the different viral antigens. The differences of the mean antibody titres within and among the test group vaccines were statistically significant. The results confirm and support those obtained by other authors in similar studies, suggesting that serological responses in vaccinated calves can be used as a helpful means of assessing the relative potency of vaccines against viral respiratory diseases of cattle. The criteria allowing such an evaluation are discussed.

  5. Development of a highly specific and sensitive rubella immunoglobulin M antibody capture enzyme immunoassay that uses enzyme-labeled antigen.

    OpenAIRE

    Seppänen, H

    1990-01-01

    An enzyme immunoassay (EIA) for serum immunoglobulin M (IgM) antibodies to rubella virus based on enzyme labeling of viral antigen was developed. The sensitivity of the EIA for the detection of recent rubella virus infection was evaluated by using 115 rubella-IgM-antibody-positive serum specimens, which were confirmed as positive by Rubazyme M (Abbott Diagnostics). In addition, 12 individuals, 2 of whom were exposed to rubella through vaccination and 10 of whom were exposed through natural in...

  6. Meningococcal Antigen Typing System Development and Application to the Evaluation of Effectiveness of Meningococcal B Vaccine and Possible Use for Other Purposes

    Directory of Open Access Journals (Sweden)

    Alexander Domnich

    2015-01-01

    Full Text Available Development of the 4-component meningococcal serogroup B vaccine (4CMenB has required new assays for the reliable evaluation of the expression and cross-reactivity of those specific antigen variants that are predicted to be targeted by bactericidal antibodies elicited by the vaccine in different isolates. Existing laboratory techniques, such as multilocus sequence typing, are poorly suited to this purpose, since they do not provide information on the contribution of single vaccine components and therefore cannot be applied to estimate the potential coverage of the multicomponent vaccine. The hSBA, the only correlate of protection against invasive meningococcal disease accepted thus far, cannot conveniently be used to test large number of strains. To overcome these issues, the meningococcal antigen typing system (MATS has been specifically developed in order to predict 4CMenB coverage of individual meningococcus serogroup B strains. To date, MATS has proved advantageous for several reasons, including its ability to assess both qualitative and quantitative aspects of surface antigens of single strains in a highly reproducible, rapid, and resource-saving manner, while its shortcomings include a possible underestimation of 4CMenB coverage and the use of pooled sera to calculate the positive bactericidal threshold. This paper provides an overview of MATS development and its field application.

  7. Serum IgG responses to food antigens in the italian population evaluated by highly sensitive and specific ELISA test.

    Science.gov (United States)

    Volpi, Nicola; Maccari, Francesca

    2009-01-01

    Using an optimized and validated ELISA method, we performed a serum test for assaying the binding capacity of serum IgG to proteins extracted from approx. 160 different foods to investigate the reactivity of specific IgG antibodies in the Italian population composed of 6,879 subjects (4,551 females and 2,328 males). 44 antigens showed an IgG response greater than 10% and only 14 aliments had an elevated reactivity greater than 20%, in particular, milk, from cow and goat, and several milk derivatives, along with egg albumen and yeasts. The IgG response to the high reactive food antigens depending on the age of the 6880 subjects was also analyzed. We demonstrated a high IgG response in a very large subject group to milk and milk derivatives, and egg albumin antigens, and we conclude that the validated ELISA test may be applied for the serum/plasma IgG antibody level determination as a useful indicator of adverse reactions to food and food hypersensitivity.

  8. Programmatic evaluation of a combined antigen and antibody test for rapid HIV diagnosis in a community and sexual health clinic screening programme.

    Science.gov (United States)

    Taegtmeyer, Miriam; MacPherson, Peter; Jones, Kathy; Hopkins, Mark; Moorcroft, Jay; Lalloo, David G; Chawla, Anu

    2011-01-01

    A substantial proportion of HIV-infected individuals in the UK are unaware of their status and late presentations continue, especially in low prevalence areas. Fourth generation antigen/antibody rapid test kits could facilitate earlier diagnosis of HIV in non-clinical settings but lack data on performance under programmatic conditions. We evaluated the performance of Determine HIV-1/2 Ag/Ab Combo Test (Determine Combo), a rapid test with indicators for both HIV antibodies and p24 antigen, in participants recruited from community outreach and hospital-based sexual health clinics. HIV infection was confirmed using laboratory enzyme-linked immunosorbent assay (EIA), Line Immuno Assay (LIA) and quantitative polymerase chain reaction (PCR). In total, 953 people underwent HIV testing. HIV antibody (Ab) prevalence was 1.8% (17/953). Four false positive rapid tests were identified: two antibody and two p24 antigen (Ag) reactions. Of participants diagnosed as HIV Ab positive, 2/17 (12%) were recent seroconverters based on clinical history and HIV antibody avidity test results. However, none of these were detected by the p24 antigen component of the rapid test kit. There were no other true positive p24 Ag tests. These data lend support to an increasing body of evidence suggesting that 4th generation rapid HIV tests have little additional benefit over 3rd generation HIV kits for routine screening in low prevalence settings and have high rates of false positives. In order to optimally combine community-based case-finding among hard-to-reach groups with reliable and early diagnosis 3rd generation kits should be primarily used with laboratory testing of individuals thought to be at risk of acute HIV infection. A more reliable point of care diagnostic is required for the accurate detection of acute HIV infection under programmatic conditions.

  9. Programmatic evaluation of a combined antigen and antibody test for rapid HIV diagnosis in a community and sexual health clinic screening programme.

    Directory of Open Access Journals (Sweden)

    Miriam Taegtmeyer

    Full Text Available BACKGROUND: A substantial proportion of HIV-infected individuals in the UK are unaware of their status and late presentations continue, especially in low prevalence areas. Fourth generation antigen/antibody rapid test kits could facilitate earlier diagnosis of HIV in non-clinical settings but lack data on performance under programmatic conditions. METHODS AND FINDINGS: We evaluated the performance of Determine HIV-1/2 Ag/Ab Combo Test (Determine Combo, a rapid test with indicators for both HIV antibodies and p24 antigen, in participants recruited from community outreach and hospital-based sexual health clinics. HIV infection was confirmed using laboratory enzyme-linked immunosorbent assay (EIA, Line Immuno Assay (LIA and quantitative polymerase chain reaction (PCR. In total, 953 people underwent HIV testing. HIV antibody (Ab prevalence was 1.8% (17/953. Four false positive rapid tests were identified: two antibody and two p24 antigen (Ag reactions. Of participants diagnosed as HIV Ab positive, 2/17 (12% were recent seroconverters based on clinical history and HIV antibody avidity test results. However, none of these were detected by the p24 antigen component of the rapid test kit. There were no other true positive p24 Ag tests. CONCLUSION: These data lend support to an increasing body of evidence suggesting that 4th generation rapid HIV tests have little additional benefit over 3rd generation HIV kits for routine screening in low prevalence settings and have high rates of false positives. In order to optimally combine community-based case-finding among hard-to-reach groups with reliable and early diagnosis 3rd generation kits should be primarily used with laboratory testing of individuals thought to be at risk of acute HIV infection. A more reliable point of care diagnostic is required for the accurate detection of acute HIV infection under programmatic conditions.

  10. Evaluation of a total hepatitis C virus (HCV) core antigen assay for the detection of antigenaemia in anti-HCV positive individuals.

    Science.gov (United States)

    Valcavi, Pierpaolo; Medici, Maria Cristina; Casula, Francesca; Arcangeletti, Maria Cristina; De Conto, Flora; Pinardi, Federica; Calderaro, Adriana; Chezzi, Carlo; Dettori, Giuseppe

    2004-07-01

    A new, sensitive enzyme immunoassay has been developed for detecting and quantifying total hepatitis C virus (HCV) core antigen in anti-HCV positive or negative sera ("trak-C", Ortho Clinical Diagnostics, Raritan, NJ). The purpose of this study was to evaluate the performance of trak-C as an additional laboratory diagnostic marker of viraemia. The performance was compared to HCV-RNA detection in the "screening" of sera from a large heterogeneous population of hospitalised patients and outpatients. Six hundred and eighteen anti-HCV negative sera, 405 anti-HCV positive/HCV-RNA negative sera, 604 anti-HCV positive/HCV-RNA positive sera and 67 anti-HCV negative sera containing antigens or antibodies potentially interfering with the performance of the assay were analysed. Supplemental HCV antibody testing was performed using a commercial strip immunoblot assay. HCV-RNA was investigated using a qualitative commercial assay. A quantitative commercial RT-PCR was used for the analysis of selected samples. Sensitivity and specificity values were 94.7 and 100%, respectively. The latter was also confirmed when anti-HCV negative samples containing potentially interfering antigens/antibodies were examined. Sensitivity below 100% was probably due to an antigenaemia below the detection limit of trak-C. Besides, because 65.6% of HCV-RNA positive/trak-C negative samples presented specific antibodies against all four RIBA antigens, the hypothesis was raised that, in some cases, the dissociation step efficiency could be sub-optimal. In conclusion, trak-C seems suitable for identifying HCV infection on large based populations. It is a rapid to perform, reliable and specific assay that can be adapted to any laboratory setting. Copyright 2004 Wiley-Liss, Inc.

  11. Cancer antigen 125 and prognosis

    DEFF Research Database (Denmark)

    Høgdall, Estrid Vilma Solyom

    2008-01-01

    PURPOSE OF REVIEW: This review addresses recently reported progress in cancer antigen 125 as a prognostic marker in patients with ovarian cancer. RECENT FINDINGS: Serum cancer antigen 125 levels measured preoperatively in both early and late stage ovarian cancer may be of prognostic value. Before...... cancer antigen 125 determination may be implemented into clinical practice, cut-off levels must be evaluated and internationally defined. Studies examining serum cancer antigen 125 levels after surgery but before, during, or after treatment confirmed that changes in serum levels are of prognostic value....... Furthermore, recent studies have shown that the level of expression of cancer antigen 125 in tissue may be an independent prognostic indicator in late stage ovarian cancer. SUMMARY: Prognostic markers may potentially help to individualize treatment within subgroups of patients. In a recent study the level...

  12. Evaluation of experimental vaccines for bovine viral diarrhea in bovines, ovines and guinea pigs Evaluación de vacunas experimentales para la diarrea viral bovina en bovinos, ovinos y cobayos

    Directory of Open Access Journals (Sweden)

    F. Fernández

    2009-06-01

    Full Text Available The bovine viral diarrhea virus (BVDV infection control should be based on elimination of persistently infected animals and on immunization through vaccination, to prevent fetal infection. However, the efficacy of inactivated BVDV vaccines is variable due to its low immunogenicity. This study evaluated the humoral immune response against homologous and heterologous strains of 7 inactivated BVDV vaccines, in bovines and two experimental models (ovine and guinea pig which might be used to test candidate vaccines. Vaccines formulated with BVDV Singer, Oregon, NADL, and multivalent, induced seroconversion in the three animal models studied, reaching antibody titres higher than 2. Vaccine containing 125C -genotype 2- only induced a low antibody response in ovine, while VS-115 NCP vaccine was not immunogenic. Furthermore, bovine sera at 60 dpv presented homologous as well as heterologous antibody response, indicating a high degree of cross-reactivity among the strains studied. However, when bovine sera were tested against the Argentine field strain 00-693, they showed the lowest levels of cross-reactivity, suggesting the need of continued surveillance to identify and characterize emerging field BVDV strains. Finally, optimal correlations between bovine-ovine and bovine-guinea pig models were observed, indicating that two alternative species could replace bovines when testing the immunogenicity of BVDV candidate vaccines.El control del virus de la diarrea viral bovina (VDVB se basa en la eliminación de animales persistentemente infectados, y la inmunización de hembras para prevenir infecciones fetales. La eficiencia de estas vacunas es variable por su baja inmunogenicidad. Se evaluó la respuesta inmune humoral contra virus homólogos y heterólogos de 7 vacunas experimentales inactivadas del VDVB en bovinos y en dos modelos experimentales (ovinos y cobayos. Las vacunas conteniendo VDVB Singer, Oregon, NADL y polivalentes indujeron seroconversión en

  13. Evaluation of the Seeplex® Meningitis ACE Detection kit for the detection of 12 common bacterial and viral pathogens of acute meningitis.

    Science.gov (United States)

    Shin, So Youn; Kwon, Kye Chul; Park, Jong Woo; Kim, Ji Myung; Shin, So Young; Koo, Sun Hoe

    2012-01-01

    Bacterial meningitis is an infectious disease with high rates of mortality and high frequency of severe sequelae. Early identification of causative bacterial and viral pathogens is important for prompt and proper treatment of meningitis and for prevention of life-threatening clinical outcomes. In the present study, we evaluated the value of the Seeplex Meningitis ACE Detection kit (Seegene Inc., Korea), a newly developed multiplex PCR kit employing dual priming oligonucleotide methods, for diagnosing acute meningitis. Analytical sensitivity of the kit was studied using reference strains for each pathogen targeted by the kit, while it's analytical specificity was studied using the human genome DNA and 58 clinically well-identified reference strains. For clinical validation experiment, we used 27 control cerebrospinal fluid (CSF) samples and 78 clinical CSF samples collected from patients at the time of diagnosis of acute meningitis. The lower detection limits ranged from 10(1) copies/µL to 5×10(1) copies/µL for the 12 viral and bacterial pathogens targeted. No cross-reaction was observed. In the validation study, high detection rate of 56.4% was obtained. None of the control samples tested positive, i.e., false-positive results were absent. The Seeplex Meningitis ACE Detection kit showed high sensitivity, specificity, and detection rate for the identification of pathogens in clinical CSF samples. This kit may be useful for rapid identification of important acute meningitis-causing pathogens.

  14. Prospective evaluation of a whole-blood test using Mycobacterium tuberculosis-specific antigens ESAT-6 and CFP-10 for diagnosis of active tuberculosis

    DEFF Research Database (Denmark)

    Ravn, Pernille; Munk, Martin E; Andersen, Ase B

    2005-01-01

    A new immunodiagnostic test based on the Mycobacterium tuberculosis-specific antigens CFP-10/ESAT-6(QFT-RD1) has been launched as an aid in the diagnosis of latent tuberculosis (TB) infection (LTBI). The aim of this study was to evaluate this test for the diagnosis of active TB. Eighty-two patients....... The accuracy of the QFT-RD1 test will vary with the prevalence of LTBI. We suggest that the QFT-RD1 test could be a very useful supplementary tool for the diagnosis of TB....

  15. Comparative evaluation of viral, nonviral and physical methods of gene delivery to normal and transformed lung epithelial cells.

    Science.gov (United States)

    Gilbert, Jennifer L; Purcell, James; Strappe, Padraig; McCabe, Matthew; O'Brien, Timothy; O'Dea, Shirley

    2008-09-01

    Few studies have directly compared the efficiencies of gene delivery methods that target normal lung cells versus lung tumor cells. We report the first study directly comparing the efficiency and toxicity of viral [adeno-associated virus (AAV2, 5, 6) and lentivirus], nonviral (Effectene, SuperFect and Lipofectamine 2000) and physical [particle-mediated gene transfer (PMGT)] methods of gene delivery in normal mouse lung cells and in mouse adenocarcinoma cells. Lentivirus pseudotyped with the vesicular stomatitis virus glycoprotein was the most efficient gene transfer method for normal mouse airway epithelial cells [25.95 (+/-3.57) %] whereas AAV6 was most efficient for MLE-12 adenocarcinoma cells [68.2 (+/-3.2) %]. PMGT was more efficient in normal mouse airway epithelial cells than AAV5, Lipofectamine 2000 and SuperFect. AAV5 displayed the lowest transfection efficiency at less than 10% in both cell types. PMGT was the only method that resulted in significant toxicity. In summary, for all of the gene delivery methods examined here, lung tumor cells were transfected more easily than normal lung cells. Lipofectamine 2000 is potentially highly selective for lung tumor cells whereas AAV6 and lentivirus vesicular stomatitis virus glycoprotein may be useful for gene delivery strategies that require targeting of both normal and tumor cells.

  16. The impact of a moderate chronic temperature increase on spleen immune-relevant gene transcription depends on whether Atlantic cod (Gadus morhua) are stimulated with bacterial versus viral antigens.

    Science.gov (United States)

    Hori, Tiago S; Gamperl, A Kurt; Nash, Gord; Booman, Marije; Barat, Ashoktaru; Rise, Matthew L

    2013-10-01

    Exposure to elevated temperature is an inherent feature of Atlantic cod (Gadus morhua) sea-cage culture in some regions (e.g., Newfoundland) and may also become an increasingly prevalent challenge for wild fish populations because of accelerated climate change. Therefore, understanding how elevated temperatures impacts the immune response of this commercially important species may help to reduce the potential negative impacts of such challenges. Previously, we investigated the impacts of moderately elevated temperature on the antiviral responses of Atlantic cod (Hori et al. 2012) and reported that elevated temperature modulated the spleen transcriptome response to polyriboinosinic polyribocytidylic acid (pIC, a viral mimic). Herein, we report a complementary microarray study that investigated the impact of the same elevated temperature regime on the Atlantic cod spleen transcriptome response to intraperitoneal (IP) injection of formalin-killed Aeromonas salmonicida (ASAL). Fish were held at two different temperatures (10 °C and 16 °C) prior to immune stimulation and sampled 6 and 24 h post-injection (HPI). In this experiment, we identified 711 and 666 nonredundant ASAL-responsive genes at 6HPI and 24HPI, respectively. These included several known antibacterial genes, including hepcidin, cathelicidin, ferritin heavy subunit, and interleukin 8. However, we only identified 15 differentially expressed genes at 6HPI and 2 at 24HPI (FDR 1%) when comparing ASAL-injected fish held at 10 °C versus 16 °C. In contrast, the same comparisons with pIC-injected fish yielded 290 and 339 differentially expressed genes (FDR 1%) at 6HPI and 24HPI, respectively. These results suggest that moderately elevated temperature has a lesser effect on the Atlantic cod spleen transcriptome response to ASAL (i.e., the antibacterial response) than to pIC (i.e., antiviral response). Thus, the impacts of high temperatures on the cod's immune response may be pathogen dependent.

  17. Evaluation of the immunogenicity and protective efficacy of killed Leishmania donovani antigen along with different adjuvants against experimental visceral leishmaniasis.

    Science.gov (United States)

    Thakur, Ankita; Kaur, Harpreet; Kaur, Sukhbir

    2015-08-01

    Visceral leishmaniasis (VL) caused by Leishmania donovani is a life-threatening disease involving uncontrolled parasitization of vital organs. Drugs to treat leishmaniasis have one or more limitations or insufficiencies in the long run. A safe and efficacious vaccine to control this disease is needed. Killed antigens that could be safer as vaccines have shown limited efficacy in clinical trials. Immunogenic enhancement with appropriate adjuvants may thus be required to elicit protective immunity based on antibodies and effector T-cell functions. Therefore, it is essential to search for adjuvant to enhance the immunogenicity of killed vaccines and to induce protection against leishmaniasis. So, the aim of the present study was to compare the effectiveness of four adjuvants, i.e. alum, saponin, monophosphoryl lipid A, cationic liposome in combination with Killed Leishmania donovani (KLD) antigen against murine VL. Animals were immunized subcutaneously thrice at an interval of 2 weeks with a final volume of 100 μl per dose. Challenge infection was given 2 weeks after last booster. Mice were sacrificed 15 days after last immunization and on 30, 60 and 90 post-infection/challenge days. The protective efficacy of vaccines was revealed by significant reduction in parasite burden and enhanced DTH responses in comparison with the infected controls. Immunized animals also generated significant levels of Th1 cytokines and increased production of IgG2a, thus indicating the generation of a protective Th1 response. All the adjuvants imparted significant protection, but liposomal formulation was most effective followed by KLD + MPL-A, KLD + saponin, KLD + alum and KLD antigen.

  18. Development and evaluation of a Luminex multiplex serology assay to detect antibodies to bovine herpes virus 1, parainfluenza 3 virus, bovine viral diarrhoea virus, and bovine respiratory syncytial virus, with comparison to existing ELISA detection methods.

    Science.gov (United States)

    Anderson, Steve; Wakeley, Phil; Wibberley, Guy; Webster, Kath; Sawyer, Jason

    2011-03-07

    Detection of circulating antibodies to bovine herpes virus 1 (BHV-1), parainfluenza 3 virus (PI3V), bovine viral diarrhoea virus (BVDV) and bovine respiratory syncytial virus (BRSV) using ELISA is widely used for veterinary diagnostics and surveillance. In this paper, the potential of a multiplex serology test based on Luminex technology, where all antibodies are simultaneously detected in a single assay was investigated. The performance of "in-house" separate ELISAs which use relatively crude lysates of cultured virus as capture antigens, was compared to the multiplex assay where the same antigens were covalently bound to the fluorescent beads used in the Luminex platform. A panel of field serum samples was tested by the multiplex assay in parallel with the separate routine ELISAs to provide a comparison between tests. The BHV-1 and PI3V components of the multiplex test showed similar sensitivities and specificities to the separate "in-house" ELISAs. The performance of the BVDV and BRSV components was less successful and was attributed to relatively low signal strength for these antigens, leading to higher assay variability and a reduced ability to distinguish positive and negative samples compared to the "in-house" ELISAs. The results illustrated that antigens commonly used successfully in ELISAs cannot always be transferred for use in alternative assay systems. The use of recombinant BVDV E2 protein was investigated and was shown to lead to an appreciable increase in signal strength compared to the use of crude BVDV antigen in the Luminex system. Crown Copyright © 2011. Published by Elsevier B.V. All rights reserved.

  19. Evaluation of a polyclonal antibody based sandwich ELISA for the detection of faecal antigens in Schistosoma spindale infection in bovines.

    Science.gov (United States)

    Sreenivasa Murthy, G S; D'Souza, Placid E; Shrikrishna Isloor, K

    2013-04-01

    Schistosomosis is a common parasitic infection in animals prevalent in cattle in Asia and Africa, where it is estimated that at least 165 million animals are infected. Out of the 10 species reported to naturally infect cattle only Schistosoma nasale and Schistosoma spindale have received particular attention, because of their recognized veterinary significance. Although animal schistosomes may, under rare conditions favouring intensive transmission, act as important pathogens in endemic areas occur at a subclinical level, causing significant losses due to long term effects on animal growth and productivity. The detection of Schistosoma antigens in serum or stool could be more valuable in diagnosis, hence early treatment before irreparable damage. In this study, fresh adult worms of S. spindale were collected from the mesenteric blood vessels, whole worm antigen was prepared. These were immunized to rabbit and guinea pig to raise antibodies against S. spindale. Polyclonal antibodies of rabbit are further used as primary capture antibodies to coat ELISA plates. The capture of antibodies of guinea pig was conjugation with horse reddish peroxidase was used as secondary antibodies. Sandwich ELISA was performed to detect Schistosoma antigens in faecal samples collected from a total of 86 infected cattle and buffaloes. The working dilutions of capture antibody, detecting antibody and conjugate were found to be 1:32, 1:20 and 1:5,000 respectively by checker board titration method. The dilution of faecal supernatant antigens of S. spindale antibodies was 1:80. Out of 86 faecal samples, 77 samples were positive by Sandwich ELISA indicating 89.54 % infection. Where as in control samples none of the samples was positive. In mixed infection out of 20 samples positive for fasciola, amphistome and hydatid, Out of 20 samples 2 samples were positive indicating 10 % infection rate. The overall sensitivity of this test is 88.65 % and specificity was 90.90 %. It could be concluded

  20. Evaluation of Antigen-Specific IgM and IgG Production during an In Vitro Peripheral Blood Mononuclear Cell Culture Assay.

    Science.gov (United States)

    Matsuda, Yoshiko; Imamura, Ryoichi; Takahara, Shiro

    2017-01-01

    The recent attention given to diseases associated with memory B-cell (mBC)-produced antibodies (Abs) suggests the need for a similar in vitro assay to evaluate the functions of mBCs. Here, we cultured peripheral blood mononuclear cells (PBMCs) with the intent to collect mBC-derived Abs in vitro and maintain their cell-cell contact-dependent interactions with helper T-cells. PBMCs were cultured with interleukin (IL)-21, CpG-oligodeoxynucleotides (ODN), phorbol myristate acetate (PMA), and phytohemagglutinin/leucoagglutinin (PHA-L) in 24-well flat-bottom plates (5 × 10(5) cells/well). A culture supernatant analysis of PBMCs from healthy donors (n = 10) indicated that antigen-specific IgM Ab levels in a PBMC culture supernatant might be better able to demonstrate the antigen sensitization status in a smaller peripheral blood sample, compared to IgG because Epstein-Barr virus-specific IgM mBCs circulate peripherally at a significantly higher frequency once antiviral humoral immunity has stabilized. Thus, our in vitro assay demonstrated the potential significance of antigen-specific IgM Ab production in the culture supernatants. Furthermore, an analysis of cultured PBMCs from allograft kidney recipients (n = 16) sensitized with de novo donor-specific human leukocyte antigen (HLA)-specific Abs (DSAs) showed that IgM-type HLA-specific Abs were detected mainly from the culture supernatants from PBMCs of patients with stable graft function, whereas IgG isotype HLA Abs were detectable only from patients with biopsy-proven antibody-mediated rejection. In other words, these IgG isotype Abs also represented an activated humoral immune response in vivo. Additionally, IgM- and IgG-expressing mBCs from healthy donors (n = 5) were cultured with IL-21, CpG-ODN, and a supernatant produced by stimulating CD19(+) B-cell-depleted PBMCs with PHA-L and PMA in 24-well flat-bottom plates (1 × 10(5) cells/well), and the resulting in vitro analysis provided some

  1. Reduced interleukin-4 receptor α expression on CD8+ T cells correlates with higher quality anti-viral immunity.

    Science.gov (United States)

    Wijesundara, Danushka K; Tscharke, David C; Jackson, Ronald J; Ranasinghe, Charani

    2013-01-01

    With the hope of understanding how interleukin (IL)-4 and IL-13 modulated quality of anti-viral CD8(+) T cells, we evaluated the expression of receptors for these cytokines following a range of viral infections (e.g. pox viruses and influenza virus). Results clearly indicated that unlike other IL-4/IL-13 receptor subunits, IL-4 receptor α (IL-4Rα) was significantly down-regulated on anti-viral CD8(+) T cells in a cognate antigen dependent manner. The infection of gene knockout mice and wild-type (WT) mice with vaccinia virus (VV) or VV expressing IL-4 confirmed that IL-4, IL-13 and signal transducer and activator of transcription 6 (STAT6) were required to increase IL-4Rα expression on CD8(+) T cells, but not interferon (IFN)-γ. STAT6 dependent elevation of IL-4Rα expression on CD8(+) T cells was a feature of poor quality anti-viral CD8(+) T cell immunity as measured by the production of IFN-γ and tumor necrosis factor α (TNF-α) in response to VV antigen stimulation in vitro. We propose that down-regulation of IL-4Rα, but not the other IL-4/IL-13 receptor subunits, is a mechanism by which CD8(+) T cells reduce responsiveness to IL-4 and IL-13. This can improve the quality of anti-viral CD8(+) T cell immunity. Our findings have important implications in understanding anti-viral CD8(+) T cell immunity and designing effective vaccines against chronic viral infections.

  2. Heterosubtypic protection against pathogenic human and avian influenza viruses via in vivo electroporation of synthetic consensus DNA antigens.

    Directory of Open Access Journals (Sweden)

    Dominick J Laddy

    Full Text Available BACKGROUND: The persistent evolution of highly pathogenic avian influenza (HPAI highlights the need for novel vaccination techniques that can quickly and effectively respond to emerging viral threats. We evaluated the use of optimized consensus influenza antigens to provide broad protection against divergent strains of H5N1 influenza in three animal models of mice, ferrets, and non-human primates. We also evaluated the use of in vivo electroporation to deliver these vaccines to overcome the immunogenicity barrier encountered in larger animal models of vaccination. METHODS AND FINDINGS: Mice, ferrets and non-human primates were immunized with consensus plasmids expressing H5 hemagglutinin (pH5HA, N1 neuraminidase (pN1NA, and nucleoprotein antigen (pNP. Dramatic IFN-gamma-based cellular immune responses to both H5 and NP, largely dependent upon CD8+ T cells were seen in mice. Hemaggutination inhibition titers classically associated with protection (>1:40 were seen in all species. Responses in both ferrets and macaques demonstrate the ability of synthetic consensus antigens to induce antibodies capable of inhibiting divergent strains of the H5N1 subtype, and studies in the mouse and ferret demonstrate the ability of synthetic consensus vaccines to induce protection even in the absence of such neutralizing antibodies. After challenge, protection from morbidity and mortality was seen in mice and ferrets, with significant reductions in viral shedding and disease progression seen in vaccinated animals. CONCLUSIONS: By combining several consensus influenza antigens with in vivo electroporation, we demonstrate that these antigens induce both protective cellular and humoral immune responses in mice, ferrets and non-human primates. We also demonstrate the ability of these antigens to protect from both morbidity and mortality in a ferret model of HPAI, in both the presence and absence of neutralizing antibody, which will be critical in responding to the

  3. Evaluation of an in silico predicted specific and immunogenic antigen from the OmcB protein for the serodiagnosis of Chlamydia trachomatis infections

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    Gargouri Jalel

    2008-12-01

    Full Text Available Abstract Background The OmcB protein is one of the most immunogenic proteins in C. trachomatis and C. pneumoniae infections. This protein is highly conserved leading to serum cross reactivity between the various chlamydial species. Since previous studies based on recombinant proteins failed to identify a species specific immune response against the OmcB protein, this study evaluated an in silico predicted specific and immunogenic antigen from the OmcB protein for the serodiagnosis of C. trachomatis infections. Results Using the ClustalW and Antigenic programs, we have selected two predicted specific and immunogenic regions in the OmcB protein: the N-terminal (Nt region containing three epitopes and the C-terminal (Ct region containing two epitopes with high scores. These regions were cloned into the PinPoint Xa-1 and pGEX-6P-1 expression vectors, incorporating a biotin purification tag and a glutathione-S-transferase tag, respectively. These regions were then expressed in E. coli. Only the pGEX-6P-1 has been found suitable for serological studies as its tag showed less cross reactivity with human sera and was retained for the evaluation of the selected antigens. Only the Ct region of the protein has been found to be well expressed in E. coli and was evaluated for its ability to be recognized by human sera. 384 sera were tested for the presence of IgG antibodies to C. trachomatis by our in house microimmunofluorescence (MIF and the developed ELISA test. Using the MIF as the reference method, the developed OmcB Ct ELISA has a high specificity (94.3% but a low sensitivity (23.9. Our results indicate that the use of the sequence alignment tool might be useful for identifying specific regions in an immunodominant antigen. However, the two epitopes, located in the selected Ct region, of the 24 predicted in the full length OmcB protein account for approximately 25% of the serological response detected by MIF, which limits the use of the developed ELISA

  4. Viral Skin Diseases.

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    Ramdass, Priya; Mullick, Sahil; Farber, Harold F

    2015-12-01

    In the vast world of skin diseases, viral skin disorders account for a significant percentage. Most viral skin diseases present with an exanthem (skin rash) and, oftentimes, an accompanying enanthem (lesions involving the mucosal membrane). In this article, the various viral skin diseases are explored, including viral childhood exanthems (measles, rubella, erythema infectiosum, and roseola), herpes viruses (herpes simplex virus, varicella zoster virus, Kaposi sarcoma herpes virus, viral zoonotic infections [orf, monkeypox, ebola, smallpox]), and several other viral skin diseases, such as human papilloma virus, hand, foot, and mouth disease, molluscum contagiosum, and Gianotti-Crosti syndrome. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Novel insights into lipid antigen presentation.

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    De Libero, Gennaro; Mori, Lucia

    2012-03-01

    T cells recognizing lipid antigens are present in large numbers in circulating blood. They exert multiple functions including immunoregulation, tumour surveillance and protection during infection. Here, we review the latest information on the mechanisms of lipid antigen presentation by CD1 molecules. Recent studies have provided insight into CD1 trafficking within the cell, lipid distribution and handling, CD1 maturation, lipid antigen processing and loading. The structural resolution of all human CD1 molecules has revealed unique features that correlate with function. Molecular mechanisms regulating CD1 expression and multiple evasion mechanisms evolved by viral and bacterial pathogens have been disclosed. With rapid progression, these studies have decoded lipid-specific immunity and have revealed the important immunological role of this type of antigen recognition. Copyright © 2012 Elsevier Ltd. All rights reserved.

  6. Expression of class 5 antigens by meningococcal strains obtained from patients in Brazil and evaluation of two new monoclonal antibodies

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    Elizabeth N. De Gaspari

    Full Text Available Determining the profile of antigen expression among meningococci is important for epidemiologic surveillance and vaccine development. To this end, two new mouse monoclonal antibodies (MAbs have been derived against Neisseria meningitidis proteins (class 5. The MAbs were reactive against outer membrane antigens and were bactericidal. Selected anti-class 5 MAbs [(5.1-3E6-2; (5.3-3BH4-C7; (5.4-1BG11-C7; (5.5-3DH-F5G9 also 5F1F4-T3(5.c], and the two new monoclonal antibodies C14F10Br2 (5.8 and 7F11B5Br3 (5.9, were then tested against different meningococcal strains, (63 strains of serogroup A, 60 strains of serogroup C (from 1972 to 1974; and 136 strains of serogroup B (from 1992 meningococci. Our results demonstrated that the expression of class 5 proteins in the N. meningitidis B Brazilian strains studied is highly heterogeneous. The serotypes and subtypes of B:4:P1.15, B:4:P1.9, B:4:P1.7, B:4:P1.3, B:4:P1.14, B:4:P1.16, B:4:NT, and B:NT:NT were detected in N. meningitidis B serogroups.The strains C:2a:P1.2 and A:4.21:P1.9 were dominant in the C and A serogroups, respectively. Serogroup B organisms expressed the class 5 epitopes 5.4 (18%, 5.5 (22%, 5.8 (3.6%, 5.9 (8.0% and 5c (38%. Serogroup C expressed class 5 epitopes 5.1 (81%, 5.4 (35%, 5.5 (33% and 5.9 (5.0%; and serogroup A showed reactivity directed at the class 5 protein 5c (47%; and reactivity was present with the new monoclonal antibody, 5.9 (5.5%. We conclude that the two new MAbs are useful in detecting important group B, class 5 antigens, and that a broad selection of serogroup B, class 5 proteins would be required for an effective vaccine based on the class 5 proteins.

  7. A plate-based histo-blood group antigen binding assay for evaluation of human norovirus receptor binding ability.

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    Moore, Matthew D; Jaykus, Lee-Ann

    2017-09-15

    Human norovirus is a leading cause of gastroenteritis worldwide. Although two in vitro cultivation methods have been reported, they cannot provide mechanistic insights into viral inactivation. Receptor-binding assays supplement these assays and give insight into capsid integrity. We present a streamlined version of a receptor-binding assay with minimal time-to-result while maintaining accuracy and high throughput. We validate assay performance for physical and chemical inactivation treatments of a norovirus GII.4 capsid. The assay produces a high positive/negative ratio (25.3 ± 4.9) in norovirus inactivation. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  8. Evaluation of hepatitis B surface antigen and hepatitis B virus-DNA results in postmortem plasma specimens

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    Nihan Ziyade

    2015-03-01

    Full Text Available Objective: To assess the presence of hepatitis B surface antigen, one of the serologic markers of hepatitis B virus (HBV infection, in postmortem blood samples from autopsy cases using ELISA, and to compare the results with those obtained by PCR, which is the gold standard method in assessing HBV infection. Methods: The HBV test results of the blood samples from 880 autopsy cases determined in our laboratory, were retrospectively studied. Results: When compared with the gold standard method PCR, the sensitivity and specificity of postmortem ELISA were 100% and 84.1%, respectively. Conclusions: The increasingly used molecular diagnostic methods, such as PCR, should be used in cases where serological tests remain insufficient.We think that prospective studies on the comparison of ELISA and PCR assessment of postmortem blood samples with larger material should be carried out.

  9. The use of different concentrations of leishmanial antigen in skin testing to evaluate delayed hypersensitivity in american cutaneous leishmaniasis

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    César A. Cuba Cuba

    1985-12-01

    Full Text Available Three concentrations of Leishmania mexicana amazonensis sonicated whole promastigote antigen (30, 9.6 and 3 ug N in 0.1 ml wereprepared and 0.1 ml of each inoculated intradermally intopatients who live in one endemic leishmaniasis region in Brazil. Patients were divided into groups with active cutaneous leishmaniasis (ACL, healed cutaneous leishmaniasis (HCL, mucosal leishmaniasis (ML, and Controls (C. Skin reactions were recorded by measuring induration 48 hours after inoculation. Skin tests using 9.6 ugN/0.1 mlyielded the best diagnostic resultssince 97% of 30 patients with active lesions (cutaneous or mucosal and 83% with HCL showed reactions of 5 mm orgreater as compared with 4% Controls. Tests using 30ug N/O. 1 ml causedan unacceptable levei of skin reactions with necrosis (10% of ACL patients tested and 17% of HCL, respectively. Tests using 3 ug N/O. 1 ml were less sensitive since only 87% of patients with active lesions and 68% with HCL had reactions of 5mm orgreater. The 3 ug N/O. 1 ml dose was utilized to ask the questions whether skin delayed hypersensitivity decreased with time after the initial lesion and whether mucosal involvement is associated with enhaced hypersensitivity to leishmanial antigen. Decreased delayed hypersensitivity was noted only in those patients who had an initial lesion more than 30 years ago. The mean induration of the reaction in 10 patients with ML was 11.3 mm ± 7.15, in 41 patients with HCL, 9.27 mm ± 6.78 and in 20patients with ACL 10. 7 mm ± 6.10 mm. The percent of patients with 5 mm orgreater induration was ML 80%, HCL 71%, ACL 90%. Thus, we could not confirm an association between enhanced delayed hypersensitivity and mucosal involvement in leishmaniasis.

  10. Development and initial evaluation of a lateral flow dipstick test for antigen detection of Entamoeba histolytica in stool sample.

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    Saidin, Syazwan; Yunus, Muhammad Hafiznur; Othman, Nurulhasanah; Lim, Yvonne Ai-Lian; Mohamed, Zeehaida; Zakaria, Nik Zairi; Noordin, Rahmah

    2017-05-01

    Entamoeba histolytica infection remains a public health concern in developing countries. Early diagnosis of amoebiasis can avoid disease complications, thus this study was aimed at developing a test that can rapidly detect the parasite antigens in stool samples. Rabbits were individually immunized with recombinant pyruvate phosphate dikinase (rPPDK) and E. histolytica excretory-secretory antigens to produce polyclonal antibodies. A rapid dipstick test was produced using anti-rPPDK PAb lined on the dipstick as capture reagent and anti-EhESA PAb conjugated to colloidal gold as the detector reagent. Using E. histolytica-spiked in stool sample of a healthy individual, the detection limit of the dipstick test was found to be 1000 cells ml -1 . Meanwhile when rPPDK was spiked in the stool sample, the minimum concentration detected by the dipstick test was 0.1 μg ml -1 . The performances of the dipstick, commercial Techlab E. histolytica II enzyme-linked immunosorbent assays (ELISA) and real-time PCR were compared using 70 stool samples from patients infected with Entamoeba species (n = 45) and other intestinal pathogens (n = 25). When compared to real-time PCR, the diagnostic sensitivity of the dipstick for detection of E. histolytica was 65.4% (n = 17/26); while the diagnostic specificity when tested with stool samples containing other intestinal pathogens was 92% (23/25). In contrast, Techlab E. histolytica II ELISA detected 19.2% (5/26) of the E. histolytica-positive samples as compared to real-time PCR. The lateral flow dipstick test produced in this study enabled rapid detection of E. histolytica, thus it showed good potential to be further developed into a diagnostic tool for intestinal amoebiasis.

  11. Evaluation of recombinant LigB antigen-based indirect ELISA and latex agglutination test for the serodiagnosis of bovine leptospirosis in India.

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    Deneke, Yosef; Sabarinath, T; Gogia, Neha; Lalsiamthara, Jonathan; Viswas, K N; Chaudhuri, Pallab

    2014-08-01

    Leptospirosis is a zoonotic disease caused by pathogenic spirochetes of the genus Leptospira, causing febrile infection characterized by multi-organ failure in humans and animals. Leptospiral Ig-like protein B (LigB) is a surface-expressed antigen that mediates host cell invasion or attachment. In this study, N-terminal conserved region of LigB protein (46 kDa) was evaluated for its diagnostic potential to detect anti-leptospiral antibodies in the sera of various animal species. Dot blot analysis revealed immunoreactivity of Leptospira-positive sera of cattle, buffalo, dog, sheep and goat to purified LigB protein. We have analyzed 1126 bovine serum samples, collected from Northern and Eastern part of India, by microscopic agglutination test (MAT) and recombinant LigB (rLigB) based ELISA and latex agglutination test (LAT). The sensitivity of rLigB based ELISA for 554 MAT positive sera was 96.9% and the specificity with 572 MAT negative sera was 91.08% whereas LAT showed sensitivity and specificity of 93.68% and 92.31%, respectively. Kappa values of 0.879 and 0.860 for recombinant antigen based ELISA and LAT indicate excellent agreement with the gold standard serological test, MAT, for the detection of anti-leptospiral antibodies in sera. Further, LAT based on rLigB antigen is a simple and rapid test, suitable for serodiagnosis of leptospirosis under field conditions, owing to its portability and longer shelf life. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Evaluation of new monoclonal antibody-based latex agglutination test for detection of cryptococcal polysaccharide antigen in serum and cerebrospinal fluid.

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    Kiska, D L; Orkiszewski, D R; Howell, D; Gilligan, P H

    1994-09-01

    We evaluated the performance of CRYPTO-LEX (Trinity Laboratories, Inc., Raleigh, N. C.), a new mouse immunoglobulin M monoclonal antibody latex agglutination reagent which reacts with the capsular polysaccharide of the four serogroups of Cryptococcus neoformans. This test was compared with CALAS (Meridian Diagnostics, Cincinnati, Ohio) for the ability to detect cryptococcal antigen in serum and cerebrospinal fluid (CSF). A total of 580 clinical specimens (327 serum and 253 CSF samples), primarily from human immunodeficiency virus-infected patients, were tested in this study. Sixty-seven specimens (44 serum and 23 CSF samples) were positive for cryptococcal antigen with both tests, and 511 (282 serum and 229 CSF samples) were negative. The two latex reagents agreed for 326 of 327 serum specimens (44 positives and 282 negatives). One serum specimen with a titer of 1:2 was CALAS positive but CRYPTO-LEX negative. The titer correlation coefficient for the two tests was 0.884 when two highly discordant serum specimens were eliminated from analysis of the data. The two latex tests agreed for 252 of 253 CSF specimens (23 positives and 229 negatives). One specimen with a titer of 1:2 was positive with CALAS and negative by CRYPTO-LEX. The correlation coefficient of the two tests for CSF titers was 0.886. The sensitivity and specificity of CRYPTO-LEX were 97 and 100%, respectively, with a 99.6% correlation with CALAS. These data show that the performance of CRYPTO-LEX is comparable to that of CALAS for detection of cryptococcal antigen in serum and CSF.

  13. COLONOSCOPY AND CARCINOEMBRYONIC ANTIGEN VARIATIONS

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    Rita G SOUSA

    2014-03-01

    Full Text Available Context Colonoscopy is essential for synchronous and metachronous cancer detection. Carcinoembryonic antigen is a colorectal cancer tumor marker, important as a follow-up tool in patients with previous colorectal cancer. False-positive carcinoembryonic antigen elevation results in multiples exams and in patient anxiety. In literature, there is reference to transient carcinoembryonic antigen increase with colonoscopy. Objective To evaluate the influence of bowel preparation and colonoscopy in carcinoembryonic antigen blood levels. Methods We prospectively studied subjects that underwent routine colonoscopy in our institution. Blood samples were collected (1 before bowel cleaning, (2 before colonoscopy and (3 immediately after colonoscopy. Blood carcinoembryonic antigen levels were determined by “Sandwich” immunoassay. The statistical methods used were the paired t-test and ANOVA. Results Thirty-seven patients (22M/15F were included; age range 28-84 (mean 56 years. Mean carcinoembryonic antigen values were 1.9, 2 and 1.8 for (1, (2 and (3, respectively. An increase in value (2 compared with (1 was observed in 20/37 patients (P = 0.018, mainly in younger patients and in patients requiring more endoluminal interventions. In 29/37 patients, the CEA value decreased from (2 to (3 (P = 1.3x10-7. Conclusions A trend for carcinoembryonic antigen increase after bowel cleaning was observed, especially in younger patients and in patients with more endoluminal interventions, but without clinical meaning.

  14. Evaluation of Ki-67 antigen expression in the vaginal epithelium of castrated female rats treated with raloxifene and tamoxifen: a comparative study.

    Science.gov (United States)

    de Moura-Leal, Adinaide C; de Sousa, João Macedo; de Moura-Leal, Heyder; de Sousa, Gabriela V; Alencar, Airlane P; Borges, Umbelina Soares; Pinto, Giovanny R; da Silva, Benedito B

    2016-06-01

    The aim of this study was to evaluate the effects of raloxifene and tamoxifen on Ki-67 antigen expression in the vaginal epithelium of castrated rats. Thirty-nine virgin, adult, castrated female Wistar-Hannover rats were randomly divided into three groups: Group I (control, n = 13), Group II (raloxifene, n  = 13) and Group III (tamoxifen, n = 13). After confirmation of their hypoestrogenic state, the rats were given 0.5 ml of propylene glycol (vehicle), 750 μg of raloxifene or 250 μg of tamoxifen, respectively, by gavage, for 30 days. On the 31st day, the rats were euthanized and their vaginas removed and fixed in 10% buffered formalin for of Ki-67 immunohistochemical evaluation. Data were analyzed using Levene's test and Tukey's method (p raloxifene and control groups.

  15. Detection of genome, antigen, and antibodies in oral fluids from pigs infected with foot-and-mouth disease virus.

    Science.gov (United States)

    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.

  16. Evaluation of Two Techniques for Viral Load Monitoring Using Dried Blood Spot in Routine Practice in Vietnam (French National Agency for AIDS and Hepatitis Research 12338).

    Science.gov (United States)

    Taieb, Fabien; Tram, Tran Hong; Ho, Hien Thi; Pham, Van Anh; Nguyen, Lan; Pham, Ban Hien; Tong, Linh An; Tuaillon, Edouard; Delaporte, Eric; Nguyen, Anh Tuan; Bui, Duc Duong; Do, NhanThi; Madec, Yoann

    2016-09-01

    Background.  Although it is the best method to detect early therapeutic failure, viral load (VL) monitoring is still not widely available in many resource-limited settings because of difficulties in specimen transfer, personnel shortage, and insufficient laboratory infrastructures. Dried blood spot (DBS) use, which was introduced in the latest World Health Organization recommendations, can overcome these difficulties. This evaluation aimed at validating VL measurement in DBS, in a laboratory without previous DBS experience and in routine testing conditions. Methods.  Human immunodeficiency virus (HIV)-infected adults were observed in a HIV care site in Hanoi, and each patient provided 2 DBS cards with whole blood spots and 2 plasma samples. Viral load was measured in DBS and in plasma using the COBAS Ampliprep/TaqMan and the Abbott RealTime assays. To correctly identify those with VL ≥ 1000 copies/mL, sensitivity and specificity were estimated. Results.  A total of 198 patients were enrolled. With the Roche technique, 51 plasma VL were ≥1000 copies/mL; among these, 28 presented a VL in DBS that was also ≥1000 copies/mL (sensitivity, 54.9; 95% confidence interval [CI], 40.3-68.9). On the other hand, all plasma VL < 1000 copies/mL were also <1000 copies/mL in DBS (specificity, 100; 95% CI, 97.5-100). With the Abbott technique, 45 plasma VL were ≥1000 copies/mL; among these, 42 VL in DBS were also ≥1000 copies/mL (sensitivity, 93.3%; 95% CI, 81.7-98.6); specificity was 94.8 (95% CI, 90.0-97.7). Conclusions.  The Abbott RealTime polymerase chain reaction assay provided adequate VL results in DBS, thus allowing DBS use for VL monitoring.

  17. Evaluation of the Tetracore Orthopox BioThreat® antigen detection assay using laboratory grown orthopoxviruses and rash illness clinical specimens.

    Science.gov (United States)

    Townsend, Michael B; MacNeil, Adam; Reynolds, Mary G; Hughes, Christine M; Olson, Victoria A; Damon, Inger K; Karem, Kevin L

    2013-01-01

    The commercially available Orthopox BioThreat® Alert assay for orthopoxvirus (OPV) detection is piloted. This antibody-based lateral-flow assay labels and captures OPV viral agents to detect their presence. Serial dilutions of cultured Vaccinia virus (VACV) and Monkeypox virus (MPXV) were used to evaluate the sensitivity of the Tetracore assay by visual and quantitative determinations; specificity was assessed using a small but diverse set of diagnostically relevant blinded samples from viral lesions submitted for routine OPV diagnostic testing. The BioThreat® Alert assay reproducibly detected samples at concentrations of 10(7)pfu/ml for VACV and MPXV and positively identified samples containing 10(6)pfu/ml in 4 of 7 independent experiments. The assay correctly identified 9 of 11 OPV clinical samples and had only one false positive when testing 11 non-OPV samples. Results suggest applicability for use of the BioThreat® Alert assay as a rapid screening assay and point of care diagnosis for suspect human monkeypox cases. Copyright © 2012 Elsevier B.V. All rights reserved.

  18. Evaluation of an ELISA using recombinant Ssλ20ΔB3 antigen for the serological diagnosis of Sarcoptes scabiei infestation in domestic and wild rabbits.

    Science.gov (United States)

    Casais, Rosa; Millán, Javier; Rosell, Joan Maria; Dalton, Kevin P; Prieto, José Miguel

    2015-12-15

    An ELISA, based on the Sarcoptes scabiei Ssλ20ΔB3 inmunodominant antigen, was evaluated for the detection of antibodies to S. scabiei in experimentally infested (n=10), farm (n=109), and wild (n=78) rabbit sera. The S. scabiei antigen Ssλ20ΔB3, a major structural protein present over the entire mite's body, was produced as a recombinant protein in Escherichia coli and purified for its use in the ELISA. The resulting ELISA showed, in experimentally infested domestic rabbits, detectable specific antibody responses (IgG) above the cut off level from week three post-infestation indicating that the assay is able to detect positive rabbits very early during the course of the infestation. The ELISA was validated on a panel of 109 domestic breeding rabbit sera collected from 26 Spanish farms, of which 41 were obtained from rabbits with skin lesions compatible with sarcoptic mange, 26 with skin lesions compatible with psoroptic mange, and 42 from unexposed individuals from mange-free farms. The ELISA in this group was characterized by 95% sensitivity, 97% specificity, and a high degree of repeatability. In the psoroptic mange compatible lesions group, included in the study as control group for cross-reactivity with the closely related mite Psoroptes cuniculi, cross-reacting antibodies to Ssλ20ΔB3 S. scabiei antigen were detected in 42.30% of the rabbit sera. However, mean% OD values of the sarcoptic-mange group (55.61 ± 39.20%) were significantly higher (prabbits from Mallorca Island. The sensitivity of the assay for this group was 100% (4 out of the 4 rabbits with sarcoptic mange compatible lesions and presence of S. scabiei mites were seropositive) and the specificity was 90% (67 out of 74 wild rabbits without detectable mange lesions were seronegative). Although, the total number of tested samples from experimentally infested, farm and wild rabbits was limited, our study showed that the ELISA is able to differentiate between infested and non-infested animals in

  19. Viral lysis of

    NARCIS (Netherlands)

    Lønborg, C.; Middelboe, M.; Brussaard, C.P.D.

    2013-01-01

    The viral mediated transformation of phytoplankton organic carbon to dissolved forms (“viral shunt”) has been suggested as a major source of dissolved organic carbon (DOC) in marine systems. Despite the potential implications of viral activity on the global carbon fluxes, studies investigating

  20. Obtenção e avaliação de antígenos de Aspergillus fumigatus Obtention and evaluation of Aspergillus fumigatus antigens extraction

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    Vanda de Sá Lirio

    1992-08-01

    Full Text Available Antígenos de três amostras de A. fumigatus (354, 356, JIG e antissoro contra a mistura destes antígenos foram produzidos e avaliados imunoquimicamente. Os antígenos de filtrado de cultura foram obtidos após concentração com acetona conforme adaptação da técnica descrita por Coleman & Kaufman. Em prova de ID obteve-se 100% de positividade com os soros de pacientes com aspergilose estudados. Com relação aos soros heterólogos encontramos reatividade com soro de um paciente com candidíase e com soro de um paciente com histoplasmose; foi encontrado padrão idêntico de resposta quando se utilizou o antígeno de referência. O antissoro foi titulado por ID, CIE e RFC MI contra o antígeno específico apresentando títulos respectivos de 1:32, 1:32 e 1:128, e utilizado para reagir contra o mesmo antígeno por IEF, demonstrando 8 linhas de precipitação, sendo 5 na região anódica e 3 na região catódica. O perfil de bandeamento do antígeno em eletroforese utilizando gel de poliacrilamida (SDS-PAGE a 12,5% apresentou-se complexo com 26 sub-unidades protéicas, cujos pesos moleculares variaram de 18 a > 100kDa. Quando estes componentes foram eletrotransferidos e reagidos com o antissoro específico ("immunoblotting", verificou-se imunogenicidade em todas as frações bandeadas.Antigens from three strains of Aspergillus fumigatus (354, 356, and JIG and an antiserum against the mixing of these antigens have been produced, and evaluated immunochemically. The antigens were obtained through a modified Coleman & Kaufman technique (culture filtrate concentrated by acetone. Analysis by the immunodiffusion test (ID against homologous serum has yielded 100% sensitivity (with the studied sera. Concerning heterologous sera we found reactivity with a serum of a patient of candidiasis and another with histoplasmosis. The same result was obtained with a reference antigen in immunodiffusion, showing similar standards of response. Titration of the

  1. Avaliação de ensaio molecular para determinação de carga viral em indivíduos sorologicamente negativos para o HIV-1 Evaluation of a molecular assay for determining viral load on HIV-1 antibody negative patients

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    José Moreira Pereira

    2002-01-01

    Full Text Available O teste de carga viral foi concebido para acompanhar a evolução e o tratamento do paciente com diagnóstico confirmado de HIV-1. Contudo, sua especificidade diagnóstica não foi ainda avaliada em pessoas que apresentam um teste sorológico negativo. Mesmo assim, ele tem sido erroneamente utilizado para o diagnóstico da infecção primária pelo HIV-1. Este trabalho relata quatro pacientes em que a carga viral plasmática NucliSens (Organon Teknika foi repetidamente positiva na ausência de anticorpos para HIV e chama atenção para o fato de que a carga viral abaixo de 10 mil cópias/ml é de difícil interpretação, como tem sido assinalado em numerosos artigos, em que foram utilizadas outras metodologias.The plasma viral load test for HIV-1,a exquisitely high sensitive assay, were neither developed nor evaluated for the diagnosis of primary HIV infection; therefore, their diagnostic specificity is not well delineated when applied to persons who are negative for HIV antibody. This article reported four cases of false positive results obtained by using NucliSens viral load assay (Organon Teknika and emphasize the importance that low positive plasma viral load (< 10 000 copies/ml may be difficult to interpret how has been assinalated in numerous articles in the medical literature, using other methodologies.

  2. [Retrospective evaluation of PSA density for selection of biopsy candidates with prostate specific antigen in the gray zone].

    Science.gov (United States)

    Tochigi, T; Kawamura, S; Numahata, K; Tokuyama, S; Kuwahara, M; Horaguchi, T; Satou, S

    2001-09-01

    We examined the usefulness of prostate specific antigen density (PSAD) for selection of biopsy candidate with prostate specific antigen levels between 4.1 and 10.0 ng./ml. in prostate cancer screening retrospectively. The screening was conducted on male candidates in Natori city, aged 55 years or older, for 6 years from 1994 through 1999. We could analyze serum PSA levels and PSA density in 118 men with PSA levels between 4.1 and 10.0 ng./ml. All of 118 men underwent ultrasound guided systematic prostate biopsy regardless of findings of digital rectal examination and transrectal ultrasound. Prostate volume was estimated by transrectal ultrasound measurements using the prolate ellipse formula (pi/6 x length x width x height). PSAD was calculated by dividing serum PSA level by prostate volume. Serum PSA levels were determined by Tandem-R assay. In 118 men, twenty-five men had prostate cancer. There was no significant difference in mean PSA between those with prostate cancer and those without prostate cancer, but the difference was significant in the mean PSA density (mean 0.26 and 0.16, respectively, p PSA and PSAD demonstrated superior benefit for PSAD in 118 men. A sensitivity, a specificity, a positive predictive value and a negative predictive value of PSAD cut-off of 0.15 were 88%, 52.7%, 33.3% and 94.2%. PSAD cut-off of 0.18 showed the highest sum of sensitivity and specificity, which gave a sensitivity of 80%, a specificity of 72%, a positive predictive value of 43.5% and a negative predictive value of 93.1%. PSAD cut-off of 0.15 would seem to be preferable to cut-off of 0.18 because of less cancer missing. Although further studies are needed to determine optimal cut-off value to be used in clinical practice, PASD seems to be useful for the selection of biopsy candidates with PSA levels of 4.1 to 10.0 ng./ml. in the prostate cancer screening.

  3. Evaluation of the Cell Proliferation Process of Ovarian Follicles in Hypothyroid Rats by Proliferation Cell Nuclear Antigen Immunohistochemical Technique

    Directory of Open Access Journals (Sweden)

    M. Moghaddam Dorafshani

    2012-10-01

    Full Text Available Introduction & Objective: The normal females reproductive function , needs hypothalamus-hypophysis-ovarian extensive hormonal messages. Primary hypothyroidism is characterized by reduced production and secretion of thyroid hormones. During follicular growth PCNA (Proliferating Cell Nuclear Antigen and cycklin D complex play an important role in regulating cell proliferation .This study aimed to determine the cell proliferation index and how this process changes induced by thyroid hormone decreased in rat ovarian follicles.Materials & Methods: In this experimental study, 20 Wistar female rats were divided into experimental and control groups. Experimental group was chemically thyroidectomized by administering propylthiouracil (PTU (500 mg per liter of drinking water. The control group received normal drinking water. After three weeks rats were killed and their ovaries dissected and fixed for the histological preparation. Cell proliferation was determined by PCNA and stereological methods were used for counting cells.Results: Cell proliferation index showed a significant decrease in the frequency of follicular growth from prenatal to graafian follicles in hypothyroidism groups(P0.05 . PCNA expression determined that Primary follicle growth begins earlier. Positive PCNA cells were not observed in primordial follicles of the groups.Conclusion: According to the results of our study, this hypothesis is raised that granulosa cells in growing follicles may be increased by follicle adjacent cells in ovarian stroma . Hormonal changes following the reduction of thyroid hormones may greatly affect the cell proliferation index and lead to faster follicle degeneration.(Sci J Hamadan Univ Med Sci 2012; 19 (3:5-15

  4. Prospective evaluation of RT-PCR on sputum versus culture, urinary antigens and serology for Legionnaire's disease diagnosis.

    Science.gov (United States)

    Botelho-Nevers, Elisabeth; Grattard, Florence; Viallon, Alain; Allegra, Séverine; Jarraud, Sophie; Verhoeven, Paul; Marcuccilli, Adrien; Lucht, Fréderic; Pozzetto, Bruno; Berthelot, Philippe

    2016-08-01

    Legionnaires' disease (LD) is a severe disease associated with community and hospital-acquired pneumonia, frequently under diagnosed. The main aim of our study was to determine the value of PCR for the diagnosis of LD in routine clinical practice. In a prospective study, from March 2007 to April 2010, the value of PCR on non-invasive respiratory specimens (NIRS) was compared to those of the other available tools for LD diagnosis in patients hospitalized for pneumonia. Among 254 consecutive cases of pneumonia included, 24 cases were LD (19 confirmed and 5 probable) representing the first documented microbiological etiology. Molecular diagnosis of LD was performed on NIRS by using 16S rRNA PCR, and secondarily mip PCR, with no discrepant results between the 2 methods: it was found positive in 14 cases and led to identify 2 supplementary probable cases of LD. Based on clinical and at least 2 positive LD tests, PCR yielded a better diagnostic value than antigen urinary test (12 vs 10 cases). These results revealed that molecular diagnosis of LD on NIRS is reliable and may contribute to better identify cases of LD. Copyright © 2016 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

  5. Evaluation of viral load thresholds for predicting new WHO Stage 3 and 4 events in HIV-infected children receiving highly active antiretroviral therapy

    Science.gov (United States)

    Siberry, George K; Harris, D. Robert; Oliveira, Ricardo Hugo; Krauss, Margot R.; Hofer, Cristina B.; Tiraboschi, Adriana Aparecida; Marques, Heloisa; Succi, Regina C.; Abreu, Thalita; Negra, Marinella Della; Mofenson, Lynne M.; Hazra, Rohan

    2012-01-01

    Background This study evaluated a wide range of viral load (VL) thresholds to identify a cut-point that best predicts new clinical events in children on stable highly-active antiretroviral therapy (HAART). Methods Cox proportional hazards modeling was used to assess the adjusted risk of World Health Organization stage 3 or 4 clinical events (WHO events) as a function of time-varying CD4, VL, and hemoglobin values in a cohort study of Latin American children on HAART ≥ 6 months. Models were fit using different VL cut-points between 400 and 50,000 copies/mL, with model fit evaluated on the basis of the minimum Akaike Information Criterion (AIC) value, a standard model fit statistic. Results Models were based on 67 subjects with WHO events out of 550 subjects on study. The VL cutpoints of > 2600 copies/mL and > 32,000 copies/mL corresponded to the lowest AIC values and were associated with the highest hazard ratios [2.0 (p = 0.015) and 2.1 (p = 0.0058), respectively] for WHO events. Conclusions In HIV-infected Latin American children on stable HAART, two distinct VL thresholds (> 2,600 copies/mL and > 32,000 copies/mL) were identified for predicting children at significantly increased risk of HIV-related clinical illness, after accounting for CD4 level, hemoglobin level, and other significant factors. PMID:22343177

  6. Genomic Loads and Genotypes of Respiratory Syncytial Virus: Viral Factors during Lower Respiratory Tract Infection in Chilean Hospitalized Infants.

    Science.gov (United States)

    Espinosa, Yazmín; San Martín, Camila; Torres, Alejandro A; Farfán, Mauricio J; Torres, Juan P; Avadhanula, Vasanthi; Piedra, Pedro A; Tapia, Lorena I

    2017-03-21

    The clinical impact of viral factors (types and viral loads) during respiratory syncytial virus (RSV) infection is still controversial, especially regarding newly described genotypes. In this study, infants with RSV bronchiolitis were recruited to describe the association of these viral factors with severity of infection. RSV antigenic types, genotypes, and viral loads were determined from hospitalized patients at Hospital Roberto del Río, Santiago, Chile. Cases were characterized by demographic and clinical information, including days of lower respiratory symptoms and severity. A total of 86 patients were included: 49 moderate and 37 severe cases. During 2013, RSV-A was dominant (86%). RSV-B predominated in 2014 (92%). Phylogenetic analyses revealed circulation of GA2, Buenos Aires (BA), and Ontario (ON) genotypes. No association was observed between severity of infection and RSV group (p = 0.69) or genotype (p = 0.87). After a clinical categorization of duration of illness, higher RSV genomic loads were detected in infants evaluated earlier in their disease (p < 0.001) and also in infants evaluated later, but coursing a more severe infection (p = 0.04). Although types and genotypes did not associate with severity in our children, higher RSV genomic loads and delayed viral clearance in severe patients define a group that might benefit from new antiviral therapies.

  7. Recombinant hepatitis B surface antigen production in Aspergillus niger: evaluating the strategy of gene fusion to native glucoamylase.

    Science.gov (United States)

    James, Er; van Zyl, Wh; van Zyl, Pj; Görgens, Jf

    2012-10-01

    This study demonstrates the potential of Aspergillus niger as a candidate expression system for virus-like particle production using gene fusion. Hepatitis B surface antigen (HBsAg) production, targeted through the secretory pathway in A. niger, resulted in completely assembled and properly folded HBsAg. This was achieved by implementing a gene fusion strategy using the highly expressed catalytic domain of the native glucoamylase gene (GlaA ( G2 )) fused to the HBsAg S gene. The inducible glucoamylase promoter (GlaA ( p )) was used to control transcription in the A. niger D15 host. The gene fusion strategy was designed for cleavage of the fused product by the KEX2-like protease, resulting in intracellular accumulation of HBsAg and extracellular secretion of glucoamylase. Immunodetection using a monoclonal HBsAg antibody could not detect the fused GlaA ( G2 ) ::S product in intracellular and extracellular fractions, indicating that full assembly and maturation of HBsAg occurred after cleavage of the fused product in the Golgi complex. Several breakdown products showing an immunoreactive response to the glucoamylase polyclonal antibody indicated a level of intracellular degradation. The choice of carbon source used in cultivation significantly affected HBsAg production levels through induction of the glucoamylase promoter. The highest specific HBsAg production was observed during growth on inducing substrates of starch and its degradation products (maltodextrin and maltose), although residual glucose accumulation in the mid-exponential phase reduced HBsAg production. HBsAg production in starch-based cultures may be improved further by optimization of the rates of starch hydrolysis by glucoamylase and subsequent glucose consumption by the host.

  8. Evaluation of a rapid method for the detection of streptococcal group A antigen directly from throat swabs.

    Science.gov (United States)

    Venezia, R A; Ryan, A; Alward, S; Kostun, W A

    1985-01-01

    Throat swabs from 196 pediatric patients were processed by a direct extraction-latex agglutination method (Group A Strep Direct Antigen Identification Test [DAI]) that detects group A streptococci in the specimen. The method requires a 45-min enzymatic extraction period at 37 degrees C and a 4-min reaction period with antibody-linked latex particles. The results were compared with those of the culture and fluorescent antibody methods and the clinical presentation of the patient for pharyngitis. Ninety-three percent of the specimens resulted in agreement by all tests, and 28% were culture positive for group A streptococci. Compared with the culture method, the DAI had a sensitivity and a specificity of 83% and 99%, respectively. The positive predictive values were 98% versus the culture method and 93% versus the fluorescent antibody method, whereas the negative predictive values were 94% versus both other methods. Of the 14 discrepant results when both clinical presentation of an acute pharyngitis and the test results were compared, the culture method provided the best correlation. An additional 64 specimens were processed by the DAI and another direct extraction-latex agglutination method (Culturette Ten-Minute Group A Strep ID Test), and the results were compared with those of the culture method. This group had a 40.6% culture isolation rate for group A streptococci. The sensitivity and specificity of the DAI and Strep ID methods versus the culture method were 81 and 100%, and 77 and 97%, respectively. These results indicate that the DAI is accurate for diagnosing group A streptococcal pharyngitis directly from throat swabs. However, negative results in the presence of a symptomatic patient must be confirmed by standard culture techniques. PMID:3884656

  9. Evaluation of the effects of long-term storage of bovine ear notch samples on the ability of 2 diagnostic assays to identify calves persistently infected with bovine viral diarrhoea virus

    Directory of Open Access Journals (Sweden)

    F. Khan

    2011-04-01

    Full Text Available Research aimed at optimising diagnostic laboratory procedures is central to the development of effective bovine viral diarrhoea virus (BVDV control programmes. BVDV is a singlestranded RNA virus that crosses the placenta to infect foetuses, resulting in reproductive losses due to foetal death or persistently infected calves that die early in life. Persistently infected animals are widely accepted to be the primary reservoir of BVDV and the largest source of infection. This poses important challenges to overall animal/herd health and can cause major losses to the cattle industry. Long-term storage of bovine ear notch samples from calves persistently infected with BVDV may adversely affect the ability of diagnostic assays to detect the virus efficiently. In order to test this hypothesis, ear notch samples from 7 animals were divided into 2 groups. One set was subjected to prompt formalin fixation and the other set stored either as fresh samples without preservatives at –2 °C, or soaked overnight in phosphate buffered saline followed by freezing of the supernatant fluid at –2 °C. Frozen ear notches and ear notch supernatant yielded positive results with an antigen- capture, enzyme linked immunosorbent assay (AC-ELISA for the duration of the study (6 months and optical density (OD values remained significantly within range. There was no significant difference between storing fresh ear notch samples or PBS at –2 °C. However, positive immunohistochemistry (IHC staining on formalin fixed ear notches started to fade between Day 17 and Day 29 when stored at room temperature. It was concluded that fresh ear notches could safely be stored at –2 °C for a period of 6 months prior to testing for BVD viral antigens.

  10. Evaluation of the effects of long-term storage of bovine ear notch samples on the ability of 2 diagnostic assays to identify calves persistently infected with bovine viral diarrhoea virus.

    Science.gov (United States)

    Khan, F; Vorster, J H; van Vuuren, M; Mapham, P

    2011-03-01

    Research aimed at optimising diagnostic laboratory procedures is central to the development of effective bovine viral diarrhoea virus (BVDV) control programmes. BVDV is a single-stranded RNA virus that crosses the placenta to infect foetuses, resulting in reproductive losses due to foetal death or persistently infected calves that die early in life. Persistently infected animals are widely accepted to be the primary reservoir of BVDV and the largest source of infection. This poses important challenges to overall animal/herd health and can cause major losses to the cattle industry. Long-term storage of bovine ear notch samples from calves persistently infected with BVDV may adversely affect the ability of diagnostic assays to detect the virus efficiently. In order to test this hypothesis, ear notch samples from 7 animals were divided into 2 groups. One set was subjected to prompt formalin fixation and the other set stored either as fresh samples without preservatives at -2 degrees C, or soaked overnight in phosphate buffered saline followed by freezing of the supernatant fluid at -2 degrees C. Frozen ear notches and ear notch supernatant yielded positive results with an antigen-capture, enzyme linked immunosorbent assay (AC-ELISA) for the duration of the study (6 months) and optical density (OD) values remained significantly within range. There was no significant difference between storing fresh ear notch samples or PBS at -2 degrees C. However, positive immunohistochemistry (IHC) staining on formalin fixed ear notches started to fade between Day 17 and Day 29 when stored at room temperature. It was concluded that fresh ear notches could safely be stored at -2 degrees C for a period of 6 months prior to testing for BVD viral antigens.

  11. Lower respiratory tract viral infections: Diagnostic role of exfoliative cytology.

    Science.gov (United States)

    Martínez-Girón, Rafael; Pantanowitz, Liron

    2017-07-01

    Viral lower respiratory tract infections (VLRTI) remain one of the most common causes of morbidity and mortality worldwide. For many years, the diagnosis of VLRTI was based on laboratory techniques such as viral isolation in cell culture, antigen detection by direct fluorescent antibody staining, and rapid enzyme immunoassay. Radiological imaging and morphology also play an important role in diagnosing these infections. Exfoliative cytology provides a simple, rapid, inexpensive, and valuable means to diagnose and manage VLRTI. Here we review viral-associated cytomorphological changes seen in exfoliated cells of the lower respiratory tract. Diagn. Cytopathol. 2017;45:614-620. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  12. Evaluation of a quadrivalent inactivated vaccine for the protection of cattle against diseases due to common viral infections : research report

    Directory of Open Access Journals (Sweden)

    J.R. Patel

    2004-06-01

    Full Text Available Efficacy of an inactivated quadrivalent vaccine containing infectious bovine rhinotracheitis (IBR virus, parainfluenza type 3 (PI3 virus, bovine virus diarrhoea virus (BVDV and bovine respiratory syncytial virus (BRSV was assessed in naive bovine calves to evaluate short-term (4-18 weeks and long-term (24-38 weeks protection following the basic intramuscular vaccination regime of 2 inoculations a month apart. Vaccination was staggered between the long-term and the short-term groups by about 5 months so that both groups, along with a matched group of 6 unvaccinated (control calves, could be challenged at the same time. Sequential challenges at intervals of 3-8 weeks were done in the order: IBR virus (intranasally, IN, PI3 virus (IN and intratracheally, IT, pestiviruses (IN and BRSV (IN and IT. The IBR virus challenge produced febrile rhinotracheitis (FRT in control calves but both the severity and the duration of FRT was significantly reduced in both vaccinated groups. The amount and the duration of IBR virus shed by the vaccinated groups was significantly reduced compared to the control group. Although PI3 virus, pooled pestivirus and BRSV challenges did not result in a noteworthy disease, challenge virus shedding (amount and duration from the upper (all 3 viruses and the lower (BRSV respiratory tracts was significantly reduced in vaccinated groups. After pestivirus challenge, sera and leukocytes from all control calves were infectious for 6-9 days whereas virus was recovered only from leukocytes in vaccinated calves and only for 1.6-2.7 days. Thus a standard course of the quadrivalent vaccine afforded a significant protection against IBR virus, PI3 virus, BVDV and BRSV for at least 6 months.

  13. Multicenter Evaluation of a New Automated Fourth-Generation Human Immunodeficiency Virus Screening Assay with a Sensitive Antigen Detection Module and High Specificity

    OpenAIRE

    Weber, Bernard; Gürtler, Lutz; Thorstensson, Rigmor; Michl, Ulrike; Mühlbacher, Annelies; Bürgisser, Philippe; Villaescusa, Roberto; Eiras, Adolfo; Gabriel, Christian; Stekel, Herbert; Tanprasert, Srivilai; Oota, Sinenaart; Silvestre, Maria-Jose; Marques, Cristina; Ladeira, Maria

    2002-01-01

    Fourth-generation assays for the simultaneous detection of human immunodeficiency virus (HIV) antigen and antibody that were available on the international market until now have antigen detection modules with relatively poor sensitivity and produce a higher rate of false-positive results than third-generation enzyme immunoassays (EIAs). The new Cobas Core HIV Combi EIA with an improved sensitivity for HIV p24 antigen was compared to alternative fourth- and third-generation assays, the p24 ant...

  14. Evaluation of pH-sensitive fusogenic polymer-modified liposomes co-loaded with antigen and α-galactosylceramide as an anti-tumor vaccine.

    Science.gov (United States)

    Okazaki, Seiji; Iwasaki, Tadashi; Yuba, Eiji; Watarai, Shinobu

    2017-12-28

    pH-Sensitive fusogenic polymer-modified (pH-sensitive) liposomes co-loaded with tumor model antigen, ovalbumin (OVA), and adjuvant, α-galactosylceramide (α-GalCer) were fabricated and administered subcutaneously into mice. The ability of pH-sensitive liposomes containing OVA and α-GalCer to stimulate cellular and humoral immune responses in vivo was compared with OVA-encapsulating pH-sensitive liposomes as well as with OVA alone. After immunization, significant OVA-specific antibodies were detected in the serum. When sera were analyzed for isotype distribution, antigen-specific IgG1 antibody responses were noted in mice immunized with OVA alone, whereas immunization with OVA-containing pH-sensitive liposomes and with pH-sensitive liposomes containing OVA and α-GalCer resulted in the induction of OVA-specific IgG1 and IgG2b antibody responses. Moreover, more substantial production of IFN-γ and IL-4 was demonstrated in spleen cells from mice immunized with pH-sensitive liposomes having OVA and α-GalCer than OVA-containing pH-sensitive liposomes in vitro. Spleen cells from the immunized mice showed strong cytotoxic activity against E.G7-OVA tumor cells. In addition, prophylactic vaccination efficacy against tumor formation was evaluated. In all mice immunized with pH-sensitive liposomes having OVA and α-GalCer, immunization provided substantial protection from tumor formation. The therapeutic efficacy of pH-sensitive liposomes containing OVA and α-GalCer against already established E.G7-OVA tumors was also investigated. Tumor growth was reduced significantly in all mice treated with pH-sensitive liposomes having OVA and α-GalCer. The provided evidence on the advantage of antigen and α-GalCer co-encapsulation into pH-sensitive liposomes should be considered in the design of future cancer vaccines for prophylactic and therapeutic purposes.

  15. Evaluation of the antibody response to Anopheles salivary antigens as a potential marker of risk of malaria.

    Science.gov (United States)

    Remoue, Franck; Cisse, Badara; Ba, Fatou; Sokhna, Cheikh; Herve, Jean-Pierre; Boulanger, Denis; Simondon, François

    2006-04-01

    The evaluation of human immune responses to arthropod bites may be a useful marker of exposure to vector-borne diseases, with applications to malaria, the most serious parasitic infection in humans. The specific antibody (Ab) IgG response to saliva obtained from Anopheles gambiae mosquitoes was evaluated in young children from an area of seasonal malaria transmission in Senegal. Specific IgG was higher in children who developed clinical Plasmodium falciparum malaria within the 3 months that followed than in those who did not (P<0.05), and it increased significantly (P<0.0001) with the level of Anopheles exposure, as evaluated by conventional entomological methods. These results suggest that evaluation of antisalivary Ab responses could be a useful approach for identifying a marker for the risk of malaria transmission.

  16. Evaluation of an IgE ELISA with Culicoides spp. extracts and recombinant salivary antigens for diagnosis of insect bite hypersensitivity in Warmblood horses.

    Science.gov (United States)

    Peeters, L M; Janssens, S; Goddeeris, B M; De Keyser, K; Wilson, A D; Kaufmann, C; Schaffartzik, A; Marti, E; Buys, N

    2013-10-01

    Insect bite hypersensitivity (IBH) in horses represents an immunoglobulin E (IgE)-mediated hypersensitivity to salivary antigens from biting midges (Culicoides spp.). The aim of this study was to evaluate and compare the performances of IgE ELISAs using recombinant Culicoides spp. Obsoletus group salivary gland antigens or crude whole body extracts ('ObsWBE'), C. nubeculosus recombinant proteins (Culn1, 3, 4, 5, 7, 8 and 10) and Obsoletus group recombinant proteins (Culo1 and 2). IgE levels were measured in plasma of 343 Warmblood horses classified as IBH-affected (n=167) and IBH-unaffected (n=176) according to the owners' descriptions. IBH-affected horses were subdivided based on the severity of their clinical signs at sampling and whether or not their IBH history was considered to be classical. The accuracies of the tests increased when clinical signs at sampling were more pronounced or when the IBH history could be considered as classical. A combination of IgE levels against the three best performing Culicoides spp. recombinant proteins (Culn4, Culo1 and Culo2) and ObsWBE resulted in the best performing test. When IBH-affected horses showing a classical history of the disease and severe clinical signs were compared with IBH-unaffected horses, the Youden's index at the optimal cut-off for the three tests in combination was 0.67. This optimal cut-off had a sensitivity of 70%, a specificity of 97% and a total accuracy of 92%. The performance of the IgE ELISA was affected by the severity of IBH clinical signs at sampling and was improved when IgE levels against several recombinant proteins were combined. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. Sharing the burden: antigen transport and firebreaks in immune responses.

    Science.gov (United States)

    Handel, Andreas; Yates, Andrew; Pilyugin, Sergei S; Antia, Rustom

    2009-05-06

    Communication between cells is crucial for immune responses. An important means of communication during viral infections is the presentation of viral antigen on the surface of an infected cell. Recently, it has been shown that antigen can be shared between infected and uninfected cells through gap junctions, connexin-based channels, that allow the transport of small molecules. The uninfected cell receiving antigen can present it on its surface. Cells presenting viral antigen are detected and killed by cytotoxic T lymphocytes. The killing of uninfected cells can lead to increased immunopathology. However, the immune response might also profit from killing those uninfected bystander cells. One benefit might be the removal of future 'virus factories'. Another benefit might be through the creation of 'firebreaks', areas void of target cells, which increase the diffusion time of free virions, making their clearance more likely. Here, we use theoretical models and simulations to explore how the mechanism of gap junction-mediated antigen transport (GMAT) affects the dynamics of the virus and immune response. We show that under the assumption of a well-mixed system, GMAT leads to increased immunopathology, which always outweighs the benefit of reduced virus production due to the removal of future virus factories. By contrast, a spatially explicit model leads to quite different results. Here we find that the firebreak mechanism reduces both viral load and immunopathology. Our study thus shows the potential benefits of GMAT and illustrates how spatial effects may be crucial for the quantitative understanding of infection dynamics and immune responses.

  18. Design, synthesis and biological evaluation of sugar-derived esters, [alpha]-ketoesters and [alpha]-ketoamides as inhibitors for Mycobacterium tuberculosis antigen 85C

    Energy Technology Data Exchange (ETDEWEB)

    Sanki, Aditya K.; Boucau, Julie; Umesiri, Francis E.; Ronning, Donald R.; Sucheck, Steven J.

    2010-08-16

    Peptide-based 1,2-dicarbonyl compounds have emerged as potent inhibitors for serine proteases. Herein, we have designed and synthesized D-arabinose and D-trehalose-based esters, {alpha}-ketoesters and {alpha}-ketoamides, and evaluated their inhibitory activity against Mycobacterium tuberculosis (Mtb) antigen 85C (ag85C), an acyltransferase in the serine hydrolase superfamily. In addition the compounds were evaluated for the ability to inhibit the growth of Mycobacterium smegmatis ATCC 14468, a non-pathogenic surrogate for Mtb. Among the synthetic analogs evaluated only the methyl ester1 derived from D-arabinose was found to inhibit the acyltransferase activity of ag85C (IC{sub 50} = 25 mM). Based on this weak inhibitory activity it was not surprising that none of the compounds inhibits the growth of M. smegmatis. In spite of the weak inhibitory activity of 1, X-ray crystallography on crystals of ag85C soaked with 1 suggested the formation of a covalent ester adduct between 1 and the Ser124 side chain hydroxyl moiety found within the catalytic site of ag85C; however, some of the active site electron density appears to result from bound glycerol. The lack of activity associated with the {alpha}-ketoester and {alpha}-ketoamide derivatives of D-trehalose may be the result of intramolecular cyclization of the {alpha}-keto moiety with the nearby C-4/4' hydroxyls leading to the formation of stable bicyclo-ester and amide derivatives.

  19. Viral Entry into Cells

    Science.gov (United States)

    D'Orsogna, Maria R.

    2010-09-01

    Successful viral infection of a healthy cell requires complex host-pathogen interactions. In this talk we focus on the dynamics specific to the HIV virus entering a eucaryotic cell. We model viral entry as a stochastic engagement of receptors and coreceptors on the cell surface. We also consider the transport of virus material to the cell nucleus by coupling microtubular motion to the concurrent biochemical transformations that render the viral material competent for nuclear entry. We discuss both mathematical and biological consequences of our model, such as the formulation of an effective integrodifferential boundary condition embodying a memory kernel and optimal timing in maximizing viral probabilities.

  20. Evaluation of a New Device for Simplifying and Standardizing Stool Sample Preparation for Viral Molecular Testing with Limited Hands-On Time.

    Science.gov (United States)

    Feghoul, Linda; Salmona, Maud; Cherot, Janine; Fahd, Mony; Dalle, Jean-Hugues; Vachon, Carole; Perrod, Aurélie; Bourgeois, Philippe; Scieux, Catherine; Baruchel, André; Simon, François; LeGoff, Jérôme

    2016-04-01

    Sensitive molecular assays have greatly improved the diagnosis of viral gastroenteritis. However, the proper preparation of stool samples for clinical testing remains an issue. bioMérieux has developed a stool preprocessing device (SPD) that includes a spoon for calibrated sampling and a vial containing buffer, glass beads, and two filters. The resulting stool filtrate is used for nucleic acid extraction. The purpose of this study was to evaluate the performance of the SPD for the quantification of human adenovirus (HAdV) DNA in stool samples collected from hematopoietic stem cell transplant (HSCT) recipients. HAdV DNA was quantified with the Adenovirus R-gene kit. The suitability of the device to reproducibly quantify HAdV DNA in stools using different extraction platforms (easyMAG and QIAsymphony) was determined using archived HAdV-positive stool samples. Coefficients of variation of HAdV DNA quantifications ranged from 1.79% to 1.83%, and no difference in quantification was observed between the two extraction systems. The HAdV DNA limit of quantification using the SPD was 3.75 log10copies/g of stool. HAdV DNA quantification using the SPD was then compared to that of the routine preprocessing technique on 75 fresh stool samples collected prospectively from pediatric HSCT recipients at risk for HAdV infections. Thirty-eight samples were HAdV DNA positive with both the SPD and routine preprocessing methods. HAdV DNA loads were on average 1.14-log10copies/g of stool higher with the SPD (Pstool samples in DNA. The use of the SPD for the detection of other gastrointestinal infections warrants further evaluation. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  1. Hepatitis B Viral Markers in Surface Antigen Negative Blood Donors ...

    African Journals Online (AJOL)

    Of the 20 who were anti-HBc positive, seven had tattoo/traditional marks on their body and one had previous history of blood transfusion. Conclusion: This study has shown that some potential blood units containing HBV are being transfused to patients unknowingly by screening for HBsAg only. Screening for other markers ...

  2. Hepatitis B Viral Markers in Surface Antigen Negative Blood Donors

    African Journals Online (AJOL)

    user1

    The majority of the donors (55.8%) were in the age bracket 21–30 years. Most of the donors (55.9%) were unmarried. The majority had not received blood transfusion ((98.6%) nor received hepatitis B virus vaccine (95.2%) in the past. Of the 495 screened blood donors,. 457 were negative to HBsAg, HIV, HCV and syphilis).

  3. Hepatitis B Viral Markers in Surface Antigen Negative Blood Donors

    African Journals Online (AJOL)

    user1

    seven had tattoo/traditional marks on their body and one had previous history of blood transfusion. CONCLUSION: This study has shown that some potential blood units containing HBV are being transfused to patients unknowingly by screening for HBsAg only. Screening for other markers of hepatitis B virus may increase ...

  4. Antigenic behaviour of several Aujeszky's disease viral strains

    OpenAIRE

    Echeverría, María Gabriela; Nosetto, Edgardo Omar

    1999-01-01

    Desde que la Enfermedad de Aujeszky fue diagnosticada por primera vez en Argentina en 1978, fueron sucediéndose numerosos brotes en diferentes lugares del país. El objetivo de este trabajo fue comparar diferentes cepas del virus de la Pseudorrabia porcina. Para ello utilizamos 5 cepas argentinas (3 aisladas en nuestro laboratorio) y 4 de referencia internacional. Las mismas fueron caracterizadas mediante pruebas físi- co-químicas; sus proteínas comparadas por electroforesis en SDS-PAGE; su an...

  5. Kinetics of Antigen Expression and Epitope Presentation during Virus Infection

    Science.gov (United States)

    Croft, Nathan P.; Smith, Stewart A.; Wong, Yik Chun; Tan, Chor Teck; Dudek, Nadine L.; Flesch, Inge E. A.; Lin, Leon C. W.; Tscharke, David C.; Purcell, Anthony W.

    2013-01-01

    Current knowledge about the dynamics of antigen presentation to T cells during viral infection is very poor despite being of fundamental importance to our understanding of anti-viral immunity. Here we use an advanced mass spectrometry method to simultaneously quantify the presentation of eight vaccinia virus peptide-MHC complexes (epitopes) on infected cells and the amounts of their source antigens at multiple times after infection. The results show a startling 1000-fold range in abundance as well as strikingly different kinetics across the epitopes monitored. The tight correlation between onset of protein expression and epitope display for most antigens provides the strongest support to date that antigen presentation is largely linked to translation and not later degradation of antigens. Finally, we show a complete disconnect between the epitope abundance and immunodominance hierarchy of these eight epitopes. This study highlights the complexity of viral antigen presentation by the host and demonstrates the weakness of simple models that assume total protein levels are directly linked to epitope presentation and immunogenicity. PMID:23382674

  6. VaxiJen: a server for prediction of protective antigens, tumour antigens and subunit vaccines

    Directory of Open Access Journals (Sweden)

    Flower Darren R

    2007-01-01

    Full Text Available Abstract Background Vaccine development in the post-genomic era often begins with the in silico screening of genome information, with the most probable protective antigens being predicted rather than requiring causative microorganisms to be grown. Despite the obvious advantages of this approach – such as speed and cost efficiency – its success remains dependent on the accuracy of antigen prediction. Most approaches use sequence alignment to identify antigens. This is problematic for several reasons. Some proteins lack obvious sequence similarity, although they may share similar structures and biological properties. The antigenicity of a sequence may be encoded in a subtle and recondite manner not amendable to direct identification by sequence alignment. The discovery of truly novel antigens will be frustrated by their lack of similarity to antigens of known provenance. To overcome the limitations of alignment-dependent methods, we propose a new alignment-free approach for antigen prediction, which is based on auto cross covariance (ACC transformation of protein sequences into uniform vectors of principal amino acid properties. Results Bacterial, viral and tumour protein datasets were used to derive models for prediction of whole protein antigenicity. Every set consisted of 100 known antigens and 100 non-antigens. The derived models were tested by internal leave-one-out cross-validation and external validation using test sets. An additional five training sets for each class of antigens were used to test the stability of the discrimination between antigens and non-antigens. The models performed well in both validations showing prediction accuracy of 70% to 89%. The models were implemented in a server, which we call VaxiJen. Conclusion VaxiJen is the first server for alignment-independent prediction of protective antigens. It was developed to allow antigen classification solely based on the physicochemical properties of proteins without

  7. Evaluation of urine-circulating cathodic antigen (Urine-CCA) cassette test for the detection of Schistosoma mansoni infection in areas of moderate prevalence in Ethiopia.

    Science.gov (United States)

    Erko, Berhanu; Medhin, Girmay; Teklehaymanot, Tilahun; Degarege, Abraham; Legesse, Mengistu

    2013-08-01

    To evaluate the diagnostic performance of antigen detecting urine-CCA cassette test for the detection of Schistosoma mansoni infection in areas of moderate prevalence in Ethiopia. Stool specimens were collected from 620 schoolchildren on three consecutive days. The samples were microscopically examined using double Kato slides; midstream urine specimens were also collected for three consecutive days and tested for S. mansoni. The sensitivity of the urine-CCA cassette test was determined using combined results of six Kato-Katz thick smears and three urine-CCA cassette tests as gold standard. The specificity of the urine-CCA cassette test was evaluated in an area where schistosomiasis is not endemic. Prevalence of S. mansoni infection as determined by single urine-CCA cassette test was 65.9%, by single Kato-Katz smear 37.3% and by six Kato-Katz thick smears 53.1% (P CCA cassette test was significantly (P CCA cassette test showed 100% specificity in endemic settings. In moderate and high prevalence areas, urine-CCA cassette test is more sensitive than the Kato-Katz method and can be used for screening and mapping of S. mansoni infection. © 2013 Blackwell Publishing Ltd.

  8. Viral Haemorrhagic Septicaemia Virus

    DEFF Research Database (Denmark)

    Olesen, Niels Jørgen; Skall, Helle Frank

    2013-01-01

    This chapter covers the genetics (genotypes and serotypes), clinical signs, host species, transmission, prevalence, diagnosis, control and prevention of viral haemorrhagic septicaemia virus.......This chapter covers the genetics (genotypes and serotypes), clinical signs, host species, transmission, prevalence, diagnosis, control and prevention of viral haemorrhagic septicaemia virus....

  9. Novel viral translation strategies.

    Science.gov (United States)

    Au, Hilda H T; Jan, Eric

    2014-01-01

    Viral genomes are compact and encode a limited number of proteins. Because they do not encode components of the translational machinery, viruses exhibit an absolute dependence on the host ribosome and factors for viral messenger RNA (mRNA) translation. In order to recruit the host ribosome, viruses have evolved unique strategies to either outcompete cellular transcripts that are efficiently translated by the canonical translation pathway or to reroute translation factors and ribosomes to the viral genome. Furthermore, viruses must evade host antiviral responses and escape immune surveillance. This review focuses on some recent major findings that have revealed unconventional strategies that viruses utilize, which include usurping the host translational machinery, modulating canonical translation initiation factors to specifically enhance or repress overall translation for the purpose of viral production, and increasing viral coding capacity. The discovery of these diverse viral strategies has provided insights into additional translational control mechanisms and into the viral host interactions that ensure viral protein synthesis and replication. © 2014 John Wiley & Sons, Ltd.

  10. Discovering hidden viral piracy.

    Science.gov (United States)

    Kim, Eddo; Kliger, Yossef

    2005-12-01

    Viruses and developers of anti-inflammatory therapies share a common interest in proteins that manipulate the immune response. Large double-stranded DNA viruses acquire host proteins to evade host defense mechanisms. Hence, viral pirated proteins may have a therapeutic potential. Although dozens of viral piracy events have already been identified, we hypothesized that sequence divergence impedes the discovery of many others. We developed a method to assess the number of viral/human homologs and discovered that at least 917 highly diverged homologs are hidden in low-similarity alignment hits that are usually ignored. However, these low-similarity homologs are masked by many false alignment hits. We therefore applied a filtering method to increase the proportion of viral/human homologous proteins. The homologous proteins we found may facilitate functional annotation of viral and human proteins. Furthermore, some of these proteins play a key role in immune modulation and are therefore therapeutic protein candidates.

  11. Evaluation of the efficacy of a recombinant Entamoeba histolytica cysteine proteinase (EhCP112) antigen in minipig.

    Science.gov (United States)

    He, Guang-Zhi; Deng, Shu-Xuan; An, Chuan-Wei

    2012-06-01

    Cysteine proteinases 112 (EhCP112) of Entamoeba histolytica are considered important for ameba pathogenicity. The recombinant gene was obtained by cloning and expression of the EhCP112 gene in heterologous host Escherichia coli BL-21 (DE3), were used to evaluate their ability to induce immune protective responses in minipig against challenge infection in a minipig-E. histolytica model. There was a 46.29% reduction (Pcoli, to immunize a minipig model of E. histolytica, and there is significant protection. This study may help to understand the EhCP112 for human in the future. Copyright © 2012 Elsevier Inc. All rights reserved.

  12. Development and comparative evaluation of a multiple-antigen RAST as a screening test for inhalant allergy.

    Science.gov (United States)

    Ownby, D R; Anderson, J A; Jacobs, G L; Homburger, H A

    1984-04-01

    We have developed a modified in vitro test for IgE antibodies, the multi-RAST, to detect antibodies of different allergen specificities simultaneously in a single tube. The multi-RAST is as sensitive for detecting low concentrations of individual IgE antibodies as the discrete RAST. We also evaluated the multi-RAST as a screening test for respiratory allergy to inhalant allergens in children by comparing the results of the multi-RAST performed by use of a mixture of SRP, TGP, and DF allergen-immunosorbents with the results of skin tests, discrete RAST tests for the same allergens, serum total IgE concentrations, and nasal smears for eosinophils in 100 children referred for allergic-disease evaluation. The results of the multi-RAST were more sensitive, specific, and efficient than the results of tests for serum IgE concentration and nasal eosinophils in establishing the diagnosis of inhalant allergy; the multi-RAST was the only diagnostic test that yielded results that were significantly associated with the clinician's impression of allergy. We conclude that the multi-RAST is a useful and cost-effective screening test for inhalant allergy in children.

  13. Intracerebral Borna disease virus infection of bank voles leading to peripheral spread and reverse transcription of viral RNA.

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    Paula Maria Kinnunen

    Full Text Available Bornaviruses, which chronically infect many species, can cause severe neurological diseases in some animal species; their association with human neuropsychiatric disorders is, however, debatable. The epidemiology of Borna disease virus (BDV, as for other members of the family Bornaviridae, is largely unknown, although evidence exists for a reservoir in small mammals, for example bank voles (Myodes glareolus. In addition to the current exogenous infections and despite the fact that bornaviruses have an RNA genome, bornavirus sequences integrated into the genomes of several vertebrates millions of years ago. Our hypothesis is that the bank vole, a common wild rodent species in traditional BDV-endemic areas, can serve as a viral host; we therefore explored whether this species can be infected with BDV, and if so, how the virus spreads and whether viral RNA is transcribed into DNA in vivo.We infected neonate bank voles intracerebrally with BDV and euthanized them 2 to 8 weeks post-infection. Specific Ig antibodies were detectable in 41%. Histological evaluation revealed no significant pathological alterations, but BDV RNA and antigen were detectable in all infected brains. Immunohistology demonstrated centrifugal spread throughout the nervous tissue, because viral antigen was widespread in peripheral nerves and ganglia, including the mediastinum, esophagus, and urinary bladder. This was associated with viral shedding in feces, of which 54% were BDV RNA-positive, and urine at 17%. BDV nucleocapsid gene DNA occurred in 66% of the infected voles, and, surprisingly, occasionally also phosphoprotein DNA. Thus, intracerebral BDV infection of bank vole led to systemic infection of the nervous tissue and viral excretion, as well as frequent reverse transcription of the BDV genome, enabling genomic integration. This first experimental bornavirus infection in wild mammals confirms the recent findings regarding bornavirus DNA, and suggests that bank voles are

  14. Intracerebral Borna disease virus infection of bank voles leading to peripheral spread and reverse transcription of viral RNA.

    Science.gov (United States)

    Kinnunen, Paula Maria; Inkeroinen, Hanna; Ilander, Mette; Kallio, Eva Riikka; Heikkilä, Henna Pauliina; Koskela, Esa; Mappes, Tapio; Palva, Airi; Vaheri, Antti; Kipar, Anja; Vapalahti, Olli

    2011-01-01

    Bornaviruses, which chronically infect many species, can cause severe neurological diseases in some animal species; their association with human neuropsychiatric disorders is, however, debatable. The epidemiology of Borna disease virus (BDV), as for other members of the family Bornaviridae, is largely unknown, although evidence exists for a reservoir in small mammals, for example bank voles (Myodes glareolus). In addition to the current exogenous infections and despite the fact that bornaviruses have an RNA genome, bornavirus sequences integrated into the genomes of several vertebrates millions of years ago. Our hypothesis is that the bank vole, a common wild rodent species in traditional BDV-endemic areas, can serve as a viral host; we therefore explored whether this species can be infected with BDV, and if so, how the virus spreads and whether viral RNA is transcribed into DNA in vivo.We infected neonate bank voles intracerebrally with BDV and euthanized them 2 to 8 weeks post-infection. Specific Ig antibodies were detectable in 41%. Histological evaluation revealed no significant pathological alterations, but BDV RNA and antigen were detectable in all infected brains. Immunohistology demonstrated centrifugal spread throughout the nervous tissue, because viral antigen was widespread in peripheral nerves and ganglia, including the mediastinum, esophagus, and urinary bladder. This was associated with viral shedding in feces, of which 54% were BDV RNA-positive, and urine at 17%. BDV nucleocapsid gene DNA occurred in 66% of the infected voles, and, surprisingly, occasionally also phosphoprotein DNA. Thus, intracerebral BDV infection of bank vole led to systemic infection of the nervous tissue and viral excretion, as well as frequent reverse transcription of the BDV genome, enabling genomic integration. This first experimental bornavirus infection in wild mammals confirms the recent findings regarding bornavirus DNA, and suggests that bank voles are capable of

  15. Evaluation of cell population data on the UniCel DxH 800 Coulter Cellular Analysis system as a screening for viral infection in children.

    Science.gov (United States)

    Jung, Y-J; Kim, J-H; Park, Y-J; Kahng, J; Lee, H; Lee, K-Y; Kim, M Y; Han, K; Lee, W

    2012-06-01

    The utility of WBC cell population data (CPD) for the differential diagnosis of viral infection from normal control, bacterial infection, and tuberculosis in children was investigated. A data set of 602 total whole-blood samples were analyzed on the DxH 800 System for complete blood cell count (CBC) with leukocyte differential from children with the following sample breakdown: 77 confirmed diagnoses of viral infections (Epstein-Barr virus; 30, influenza A; 19, rota virus; 11, other viruses;17), 54 normal control, 71 bacterial infection, 17 TB patients, and 383 with various diseases. The mean (MN) and standard deviation (SD) of the volume (V), conductivity (C), five light-scatter measurements, and 14 calculated parameters were obtained for the leukocytes. Using a combination of the CBC and CPD parameter values, a decision rule, composed of 21 parameters, for the screening of viral infection in children was developed. Using this decision rule, 74 of 77 (96.1%) viral infections, two of 54 (3.7%) normal samples, one of 17 (5.9%) TB, and six of 71 (8.5%) bacterial infection samples were identified. The sensitivity was 96.1%, and specificity for normal control was 96.3% with an overall specificity of 93.7%. Fifty-nine samples of 383 samples (15.4%) collected from in-patient children with various diseases without confirmation of viral infection were included in this decision rule. In conclusion, the implementation of leukocytes CPD parameters can be useful in the detection of viral infection in children. © 2012 Blackwell Publishing Ltd.

  16. Evaluation of circulating cathodic antigen (CCA) strip for diagnosis of urinary schistosomiasis in Hassoba school children, Afar, Ethiopia.

    Science.gov (United States)

    Ayele, B; Erko, B; Legesse, M; Hailu, A; Medhin, G

    2008-03-01

    A total of 206 urine samples collected from Hassoba Elementary schoolchildren, Afar, Ethiopia, a low Schistosoma haematobium endemic setting, was diagnosed to evaluate the performance of CCA strip using double references, urine filtration technique and urinalysis dipstick (Combur 1.0 Test) that detect schistosome eggs and blood in urine, respectively. The former was used as a gold standard reference method. Sensitivity, specificity, positive and negative predictive values for the CCA were 52%, 63.8%, 56.7% and 59% respectively, with reference to urine filtration technique whereas these parameters were 50.4%, 62.4%, 55.6% and 57.5% respectively, with reference to Combur 10 Test. 47 S. haematobium egg-positive children were found negative by CCA strip while 38 egg-negative children were found positive by CCA strip. Moreover, among the pre-tests done in duplicate, inconsistent results were also recorded. Assays were also compared with regard to the cost of equipment and reagents, speed and simplicity of use. Though CCA strip was found to be rapid and could be performed with minimal training, it was found to be expensive (US $ 4.95 per test) to use it for large-scale field use even if its diagnostic value would have been satisfactory. Further development and standardization of the CCA strip are required for its applicability for field use. It is also recommended that its cost per strip should be substantially cut down if it is to be used in poor schistosomiasis endemic countries.

  17. Monitoring T-Cell Responses in Translational Studies: Optimization of Dye-Based Proliferation Assay for Evaluation of Antigen-Specific Responses

    Directory of Open Access Journals (Sweden)

    Anja Ten Brinke

    2017-12-01

    Full Text Available Adoptive therapy with regulatory T cells or tolerance-inducing antigen (Ag-presenting cells is innovative and promising therapeutic approach to control undesired and harmful activation of the immune system, as observed in autoimmune diseases, solid organ and bone marrow transplantation. One of the critical issues to elucidate the mechanisms responsible for success or failure of these therapies and define the specificity of the therapy is the evaluation of the Ag-specific T-cell responses. Several efforts have been made to develop suitable and reproducible assays. Here, we focus on dye-based proliferation assays. We highlight with practical examples the fundamental issues to take into consideration for implementation of an effective and sensitive dye-based proliferation assay to monitor Ag-specific responses in patients. The most critical points were used to design a road map to set up and analyze the optimal assay to assess Ag-specific T-cell responses in patients undergoing different treatments. This is the first step to optimize monitoring of tolerance induction, allowing comparison of outcomes of different clinical studies. The road map can also be applied to other therapeutic interventions, not limited to tolerance induction therapies, in which Ag-specific T-cell responses are relevant such as vaccination approaches and cancer immunotherapy.

  18. Evaluation of the use of decision-support software in carcino-embryonic antigen (CEA-based follow-up of patients with colorectal cancer

    Directory of Open Access Journals (Sweden)

    Verberne Charlotte J

    2012-03-01

    Full Text Available Abstract Background The present paper is a first evaluation of the use of "CEAwatch", a clinical support software system for surgeons for the follow-up of colorectal cancer (CRC patients. This system gathers Carcino-Embryonic Antigen (CEA values and automatically returns a recommendation based on the latest values. Methods Consecutive patients receiving follow-up care for CRC fulfilling our in- and exclusion criteria were identified to participate in this study. From August 2008, when the software was introduced, patients were asked to undergo the software-supported follow-up. Safety of the follow-up, experiences of working with the software, and technical issues were analyzed. Results 245 patients were identified. The software-supported group contained 184 patients; the control group contained 61 patients. The software was safe in finding the same amount of recurrent disease with fewer outpatient visits, and revealed few technical problems. Clinicians experienced a decrease in follow-up workload of up to 50% with high adherence to the follow-up scheme. Conclusion CEAwatch is an efficient software tool helping clinicians working with large numbers of follow-up patients. The number of outpatient visits can safely be reduced, thus significantly decreasing workload for clinicians.

  19. Plant-derived antigens as mucosal vaccines.

    Science.gov (United States)

    Mason, H S; Herbst-Kralovetz, M M

    2012-01-01

    During the last two decades, researchers have developed robust systems for recombinant subunit vaccine production in plants. Stably and transiently transformed plants have particular advantages that enable immunization of humans and animals via mucosal delivery. The initial goal to immunize orally by ingestion of plant-derived antigens has proven difficult to attain, although many studies have demonstrated antibody production in both humans and animals, and in a few cases, protection against pathogen challenge. Substantial hurdles for this strategy are low-antigen content in crudely processed plant material and limited antigen stability in the gut. An alternative is intranasal delivery of purified plant-derived antigens expressed with robust viral vectors, especially virus-like particles. The use of pattern recognition receptor agonists as adjuvants for mucosal delivery of plant-derived antigens can substantially enhance serum and mucosal antibody responses. In this chapter, we briefly review the methods for recombinant protein expression in plants, and describe progress with human and animal vaccines that use mucosal delivery routes. We do not attempt to compile a comprehensive list, but focus on studies that progressed to clinical trials or those that showed strong indications of efficacy in animals. Finally, we discuss some regulatory concerns regarding plant-based vaccines.

  20. Human Tumor Antigens Yesterday, Today, and Tomorrow.

    Science.gov (United States)

    Finn, Olivera J

    2017-05-01

    The question of whether human tumors express antigens that can be recognized by the immune system has been answered with a resounding YES. Most were identified through spontaneous antitumor humoral and cellular immune responses found in cancer patients and include peptides, glycopeptides, phosphopeptides, viral peptides, and peptides resulting from common mutations in oncogenes and tumor-suppressor genes, or common gene fusion events. Many have been extensively tested as candidates for anticancer vaccines. More recently, attention has been focused on the potentially large number of unique tumor antigens, mutated neoantigens, that are the predicted products of the numerous mutations revealed by exome sequencing of primary tumors. Only a few have been confirmed as targets of spontaneous immunity and immunosurveillance, and even fewer have been tested in preclinical and clinical settings. The field has been divided for a long time on the relative importance of shared versus mutated antigens in tumor surveillance and as candidates for vaccines. This question will eventually need to be answered in a head to head comparison in well-designed clinical trials. One advantage that shared antigens have over mutated antigens is their potential to be used in vaccines for primary cancer prevention. Cancer Immunol Res; 5(5); 347-54. ©2017 AACR. ©2017 American Association for Cancer Research.

  1. Evaluation of a new cryptococcal antigen lateral flow immunoassay in serum, cerebrospinal fluid and urine for the diagnosis of cryptococcosis: a meta-analysis and systematic review.

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    Hua-Rong Huang

    Full Text Available A new lateral flow immunoassay (LFA for the detection of cryptococcal antigen was developed.We aimed to systematically review all relevant studies to evaluate the diagnostic accuracy of the cryptococcal antigen LFA on serum, CSF and urine specimens.We searched public databases including PubMed, Web of Science, Elsevier Science Direct and Cochrane Library for the English-language literature published up to September 2014. We conducted meta-analyses of sensitivity, specificity, positive likelihood ratio (PLR, negative likelihood ratio (NLR and diagnostic odds ratios (DOR and SROC of LFA in serum and CSF, respectively. The sensitivity of LFA in urine was also analyzed. Subgroup analyses were carried out to analyze the potential heterogeneity.12 studies were included in this study. The pooled sensitivity and specificity values of LFA in serum were 97.6% (95% CI, 95.6% to 98.9% and 98.1% (95% CI, 97.4% to 98.6%, respectively. The average PLR of LFA in serum was 43.787 (95% CI, 22.60-84.81 and the NLR was 0.03 (95% CI, 0.01-0.09. The pooled DOR was 2180.30 (95% CI, 868.92-5471.00 and the AUC was 0.9968. The pooled sensitivity and specificity values of LFA in CSF were 98.9% (95% CI, 97.9% to 99.5% and 98.9% (95% CI, 98.0% to 99.5%, respectively. The average PLR of LFA in serum was 48.83 (95% CI, 21.59-110.40 and the NLR was 0.02 (95% CI, 0.01-0.04. The pooled DOR was 2931.10 (95% CI, 1149.20-7475.90 and the AUC was 0.9974. The pooled sensitivity value of LFA in urine was 85.0% (95% CI, 78.7% to 90.1%.The study demonstrates a very high accuracy of LFA in serum and CSF for the diagnosis of cryptococcosis in patients at risk. LFA in urine can be a promising sample screening tool for early diagnosis of cryptococcosis.

  2. Mycoplasma pneumoniae: IgG and IgM antibody response in presence of different antigens. Evaluation of commercial tests

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    Mariella Tonella

    2011-12-01

    Full Text Available Background. P1 adhesin protein (170-kDa is responsible for the interaction between M. pneumoniae and the host. Such protein is the target of specific antibodies produced by the host in response to M. pneumoniae infection. Objectives. The aim of this study was to evaluate the performance of serological tests for Mycoplasma pneumoniae IgG and IgM on chemiluminescence analyzer LIAISON® and to demonstrate how the determination of specific antibodies is the main tool for accurate diagnosis of Mycoplasma infection. Study Design. Eighty-one unselected samples were assayed by ELISA kits Vircell IgG and IgM (bacterial lysate and LIAISON® Mycoplasma IgG and IgM (recombinant protein P1. The discordant samples were tested with ELISA Savyon Serorecombinant IgG and IgM (recombinant protein P1 and reclassified according to the consensus rule. Results. Fifty-one samples were concordant between ELISA Vircell IgG and LIAISON® Mycoplasma IgG. After resolution of the 30 discordant: twenty-eight samples were classified concordant negative and one positive. Seventy-seven samples were concordant between ELISA Vircell IgM and LIAISON® Mycoplasma IgM. After resolution of 4 discordant samples: three samples were classified as negative. Conclusions. The higher correlation between LIAISON® and Savyon Serorecombinant is determined by the use of recombinant protein P1 in the solid phase and the type of cut-off calculation allows high specificity and sensitivity. The Mycoplasma pneumoniae IgG and IgM kits, performed in full automation on LIAISON® analyzer, show characteristics comparable to other kits and an easy handling of the sample.

  3. The evaluation of yeast derivatives as adjuvants for the immune response to the Bm86 antigen in cattle

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    Machado Héctor

    2001-05-01

    Full Text Available Abstract Background The Gavac™ vaccine against the cattle tick Boophilus microplus has proven its efficacy in a large number of controlled and field experiments. However, this vaccine could be further improved by searching for new alternative adjuvants that would induce a stronger long-lasting immune response. We conducted several experiments to assay the adjuvant effect of fractions of the recombinant yeast Pichia pastoris in mouse and cattle models. In previous experiments, the combination of the yeast membrane with saponin was the most effective formulation in stimulating the humoral immune response in mice, eliciting a response higher than Montanide 888. The response was predominantly of the IgG1 isotype. Here, we evaluated the response in cattle and compared the results with that obtained in mice. Results Bm86 on the membrane of P. pastoris plus saponin produced high antibody titers in cattle, though the protection level against tick infestations was lower when compared to Gavac™, probably due to a decrease in the IgG1/IgG2 ratio. The predictive value of the mouse model was studied through correlation analysis between the isotype levels in mice and the efficacy of formulations in cattle. Good correlation was established between the level of antibodies in mice and cattle, and between the amount of anti -Bm86 IgG1 in mice and the degree of protection in cattle. Conclusion Mouse model have the potential to predict immunogenicity and efficacy of formulations in cattle. These results also support the use of the yeast expression system for recombinant vaccine formulations, enabling the prediction of more cost - effective formulations.

  4. Sequeale of acute viral hepatitis type B.

    Science.gov (United States)

    Lesnicar, J; Ferluga, D; Lesnicar, G

    1977-08-01

    In 1976 we undertook to evaluate the incidence of chronic liver lesions in 161 patients treated in hospital during the years 1970-1975 for their serologically established acute viral hepatitis type B (AVH-B). At systematic control examination mode in 1976, after a period from 1-5 years since the acute onset of disease, it was established that in 133 individuals (82.6%) the antigen HBs had disappeared from blood and the BLT had become normal. Persistent HBs antigenemia was established in 20 (12.4%) individuals. In 15 (9.2%) patients persistent HBs antigenemia was accompanied by pathologic BLT, in 5 (3.1%) cases liver function became returned to normal yet with the persistent HBs antigenemia after their recovery from A VH-B. In 8 (4.9%) patients pathologic BLT persisted although HBsAg had disappeared from blood. Among 28 persons with persistent pathological BLT or with persistent HBs antigenemia out of a total of 161 patients who had had A VH-B, there were 11 (6.8%) cases with the bioptically proved CPHf, 8 (5.0%) cases with CPH, 5 (3.1%) cases with CAH, while 4 (2.5%) patients showed fatty liver metamorphosis or had by light microskopy completely normal liver. CAH was established only in cases with persistent BHs antigenemia and pathological BLT. The incidence of the chronic liver lesion and of the persistent antigenemia was among our patients who had had A VH-B in inverse ratio to the intensity of their initial infection. Our study suggests that no prodisposition for persistent HBs antigenemia is created by the prednisolone therapy.

  5. Evaluacion de la tecnica de contraimmunoelectroforesis para determinar la potencia antigena de las vacunas antirrabicas Evaluation ot the counterimmunoelectrophoresis technique to determine the antigenic potency of antirrabic vaccines

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    Graciela Miceli

    1993-12-01

    paper presents the results obtained when using a counterimmunoelectrophoresis (CIE techniques to determine the antigens content in samples of 84 and 40 samples of lots of suckling mouse rabies and tissue culture rabies vaccines, respectively. The evaluation of rabies vaccines under, at, near and up to 0.3 UI by the NIH and CIE tests showed that for practical use, a CIE titer of 1:4 would match the minimum of NIH potency test requirement. Low degree of CIE test variability within our laboratory was observed, when two working reference and 7 lots of rabies vaccines from different origin and activity were tested in five independent assays. All titres fell within one double dilution what is an indication of its reproducibility. Good sensitivity to detect antigen deterioration in thermal degradation test was observed when samples of 3 lots of liquid suckling mouse rabies vaccine were held stored at 4ºC or at 37ºC for 4 weeks and assays by CIE and NIH tests. Finally, CIE test could apply by manufactures to follow the concentration procedures to determine the final end point which could match a desired antigenic value in the NIH potency test.

  6. Development and evaluation of a replicon particle vaccine expressing the E2 glycoprotein of bovine viral diarrhea virus (BVDV in cattle

    Directory of Open Access Journals (Sweden)

    Loy John Dustin

    2013-01-01

    Full Text Available Abstract Background Bovine viral diarrhea virus is one of the most significant and costly viral pathogens of cattle worldwide. Alphavirus-derived replicon particles have been shown to be safe and highly effective vaccine vectors against a variety of human and veterinary pathogens. Replicon particles are non-propagating, DIVA compatible, and can induce both humoral and cell mediated immune responses. This is the first experiment to demonstrate that Alphavirus-based replicon particles can be utilized in a standard prime/boost vaccination strategy in calves against a commercially significant bovine pathogen. Findings Replicon particles that express bovine viral diarrhea virus sub-genotype 1b E2 glycoprotein were generated and expression was confirmed in vitro using polyclonal and monoclonal antibodies specific to E2. Vaccine made from particles was generated in Vero cells and administered to BVDV free calves in a prime/boost regimen at two dosage levels. Vaccination resulted in neutralizing antibody titers that cross-neutralized both type 1 and type 2 BVD genotypes following booster vaccination. Additionally, high dose vaccine administration demonstrated some protection from clinical disease and significantly reduced the degree of leukopenia caused by viral infection. Conclusions Replicon particle vaccines administered in a prime/boost regimen expressing BVDV E2 glycoprotein can induce cross-neutralizing titers, reduce leukopenia post challenge, and mitigate clinical disease in calves. This strategy holds promise for a safe and effective vaccine to BVDV.

  7. EVALUATION OF MURINE NOROVIRUS, FELINE CALICIVIRUS, POLIOVIRUS, AND MS2 AS SURROGATES FOR HUMAN NOROVIRUS IN a Model of Viral Persistence in SURFACE Water AND GROUNDWATER

    Science.gov (United States)

    Human noroviruses (NoV) are a significant cause of non bacterial gastroenteritis worldwide with contaminated drinking water a potential transmission route. The absence of a cell culture infectivity model for NoV necessitates the use of molecular methods and/or viral surrogate mod...

  8. Insulated Foamy Viral Vectors

    Science.gov (United States)

    Browning, Diana L.; Collins, Casey P.; Hocum, Jonah D.; Leap, David J.; Rae, Dustin T.; Trobridge, Grant D.

    2016-01-01

    Retroviral vector-mediated gene therapy is promising, but genotoxicity has limited its use in the clinic. Genotoxicity is highly dependent on the retroviral vector used, and foamy viral (FV) vectors appear relatively safe. However, internal promoters may still potentially activate nearby genes. We developed insulated FV vectors, using four previously described insulators: a version of the well-studied chicken hypersensitivity site 4 insulator (650cHS4), two synthetic CCCTC-binding factor (CTCF)-based insulators, and an insulator based on the CCAAT box-binding transcription factor/nuclear factor I (7xCTF/NF1). We directly compared these insulators for enhancer-blocking activity, effect on FV vector titer, and fidelity of transfer to both proviral long terminal repeats. The synthetic CTCF-based insulators had the strongest insulating activity, but reduced titers significantly. The 7xCTF/NF1 insulator did not reduce titers but had weak insulating activity. The 650cHS4-insulated FV vector was identified as the overall most promising vector. Uninsulated and 650cHS4-insulated FV vectors were both significantly less genotoxic than gammaretroviral vectors. Integration sites were evaluated in cord blood CD34+ cells and the 650cHS4-insulated FV vector had fewer hotspots compared with an uninsulated FV vector. These data suggest that insulated FV vectors are promising for hematopoietic stem cell gene therapy. PMID:26715244

  9. Eficácia da vacina anti-rábica ERA em camundongos, frente a quatro diferentes variantes antigênicas do vírus da raiva Evaluation of ERA anti-rabies vaccine against four different antigenic strains of rabies virus in mice

    Directory of Open Access Journals (Sweden)

    Elcio Benedito Erbolato

    1989-12-01

    Full Text Available Estudou-se a eficácia da vacina anti-rábica preparada em cultura primária de tecido renal de suínos, a partir da amostra ERA, na prevenção da raiva em camundongos, frente a quatro cepas antigenicamente distintas do vírus rábico, duas originadas de cão. C/SP e C/NG, uma originada de morcego, DR-19, e uma cepa fixa, CVS (Challenge Vírus Standard. O perfil antigênico desta cepa foi determinado pela técnica dos anticorpos antirrábicos monoclonais antinucleocapside. Os animais foram vacinados, aos 21 dias de idade, por via intramuscular na face interna da coxa, com uma única dose de 0,05 ml de vacina e desafiados aos 42 dias de idade, em conjunto com os animais do grupo testemunho, por via intramuscular na face interna da coxa, com 0,05 ml da suspensão da cepa viral correspondente. Os resultados obtidos permitiram constatar que a vacina ERA protegeu 100% dos animais desafiados com as cepas C/SP, C/NG e DR-19 e 83% dos animais desafiados com à cepa CVS, enquanto que a mortalidade no grupo testemunho variou entre 70 e 90%.ERA anti-rabies vaccine prepared in kidney tissue culture was evaluated against four different antigenic strains of rabies virus in mice: two of them dog strains, C/SP and C/NG, another a bat vampire strain, DR-19, and the CVS strain. The CVS antigenical characteristics were determined by means the antinucleocapsid monoclonal antibodies technique. Twenty one days old mice were vaccinated, intramuscularly, in the inner side of the thigh, with 0.05 ml of vaccine and challenged at 42 days old, together with those of the control group, intramuscularly, in the inner side of the thigh, with 0.05 ml of the corresponding viral strain dilution. The ERA anti-rabies vaccine protected 100% of all the mice challenged with C/SP, C/NG and DR-19 strains and 83% of those challenged with CVS. The control groups mortality rate varied between 70 and 90%.

  10. Sperm antigens in fertilization.

    Science.gov (United States)

    Saling, P M

    1990-02-01

    A review of sperm antigens involved in fertilization includes a description of sperm differentiation, seminal fluid components that coat sperm, sperm antigens involved in binding to the zona pellucida (ZP), antigens involved in the acrosome reaction, in zona pellucida penetration, and those active in fusion with the ova membrane. Sperm antigens are located in certain domains of the cell, and they are altered during capacitation and passage through the female tract. Caltrin and acrosome-stabilizing factor are applied by seminal fluid. At least 2 antigens have been studied that occur in sterile women, although one cross reacts with milk proteins. Some antigens active in ZP binding are trypsin, proacrosin, acrosin, PH-20 from guinea pigs, and rabbit sperm autoantigen I. Antigens involved in the acrosome reaction, such as M42, are likely to cross react with other body proteins that also entail exocytosis. A mouse antigen involved in ZP penetration, MS 207 is well characterized. PH-30 from guinea pigs and M29 from mouse participate in sperm-egg membrane fusion, as does fertilization antigen I from human and mouse sperm which is know to cause infertility. Oddly, patients' sera react with polymers but not monomers of this antigen. Studies with antisperm antibodies suggest that it will not be necessary to agglutinate all sperm to block fertility, only to inhibit a single sperm epitope and function. It will probably be feasible to inhibit multiple successive events, and possibly to induce temporary immunity.

  11. Structural analysis of hepatitis B surface antigen by monoclonal antibodies.

    Science.gov (United States)

    Ben-Porath, E; Wands, J R; Marciniak, R A; Wong, M A; Hornstein, L; Ryder, R; Canlas, M; Lingao, A; Isselbacher, K J

    1985-10-01

    A method has been developed for the analysis of hepatitis B surface antigen (HBsAg) antigenic structure at the molecular level that creates "fingerprints" or "signatures" of various hepatitis B viral (HBV) strains. This technique employs high affinity IgM and IgG monoclonal antibodies (anti-HBs) directed against distinct and separate determinants on HBsAg. In performing this antigenic structural analysis, separate binding curves for different monoclonal anti-HBs are generated by measuring immunoreactivity in serial dilutions of HBsAg-positive serum by radioimmunoassay. Since the HBsAg concentration in serum is unknown, the binding profiles of groups of samples are aligned by an iterative least-squares procedure to generate the numerical signature characteristic of the viral strain. The numerical signatures are then displayed on a computer-graphic plot. The signature profiles of HBsAg subtypes are a true reflection of their antigenic structure, and in vertical and horizontal transmission studies the molecular characteristics of the viral epitopes are conserved. By signature analysis we found substantial antigenic heterogeneity among the ayw3 strain both in the U.S. and France, as well as in populations of the Far East and Africa. Populations in Ethiopia, Gambia, and the Philippines were infected with two antigenically distinct HBV strains. In some newly identified HBV strains, it was found that epitopes identified by some monoclonal antibodies were absent or substantially reduced, which suggested that a genetic mutation may have occurred. Thus this study suggests that there is far more antigenic heterogeneity in HBV than previously recognized. These variants are antigenically distinct from each other at the epitope level, and were heretofore unrecognized by polyvalent anti-HBsAg antibodies.

  12. Preclinical evaluation of NF-kappa B-triggered dendritic cells expressing the viral oncogenic driver of Merkel cell carcinoma for therapeutic vaccination

    DEFF Research Database (Denmark)

    Gerer, Kerstin F.; Erdmann, Michael; Hadrup, Sine Reker

    2017-01-01

    Background: Merkel cell carcinoma (MCC) is a rare but very aggressive skin tumor that develops after integration of a truncated form of the large T-antigen (truncLT) of the Merkel cell polyomavirus (MCV) into the host's genome. Therapeutic vaccination with dendritic cells (DCs) loaded with tumor ...

  13. Electrochemiluminescence evaluation for carbohydrate antigen 15-3 based on the dual-amplification of ferrocene derivative and Pt/BSA core/shell nanospheres.

    Science.gov (United States)

    Ke, Hong; Zhang, Xin; Huang, Chusen; Jia, Nengqin

    2018-04-30

    Herein, a novel methodology for ultrasensitive and facile breast cancer biomarker carbohydrate antigen 15-3 (CA15-3) evaluation was proposed by fabricating a sandwiched ECL immunosensor. In the protocol, MOCs-Fc and Pt@BSA-luminol nanohybrids were successfully synthesized and further employed to achieve dual-amplification strategy for luminol-H2O2 system. Notably, inherent porous microstructure and large specific surface area from MOCs enabled a high loading of Fc, which succeeded in catalyzing luminol-H2O2 ECL emission and therefore enhancing ECL response. In addition, higher sensitivity could be realized due to the excellent electronic conductivity of MOCs. Furthermore, the as-obtained Pt@BSA- luminol was not only employed as capture probe to recognize CA15-3 after hybridization with Ab2, but also played a crucial role in acting as ECL signal probe due to the presence of massive luminol. It is of vital importance that Pt@BSA core/shell nanospheres showed admirable catalytic effect towards H2O2, which resulted in more excited state luminol and stronger ECL intensity. Therefore, the synergistic amplification strategy of MOCs-Fc and Pt@BSA nanohybrids offered an extremely enhanced ECL signal. The well-established applicable ECL immune- sensing platform displayed favorable analytical performance for CA15-3. In summary, the proposed ECL immunosensor opened a new era for sensitive CA15-3 evaluation and offered a promising platform for clinical breast cancer diagnostics. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Evaluation of the Prostate Imaging Reporting and Data System for Magnetic Resonance Imaging Diagnosis of Prostate Cancer in Patients with Prostate-specific Antigen <20 ng/ml.

    Science.gov (United States)

    Wang, Xuan; Wang, Jian-Ye; Li, Chun-Mei; Zhang, Ya-Qun; Wang, Jian-Long; Wan, Ben; Zhang, Wei; Chen, Min; Li, Sa-Ying; Wan, Gang; Liu, Ming

    2016-06-20

    The European Society of Urogenital Radiology has built the Prostate Imaging Reporting and Data System (PI-RADS) for standardizing the diagnosis of prostate cancer (PCa). This study evaluated the PI-RADS diagnosis method in patients with prostate-specific antigen (PSA) PSA PI-RADS. The diagnostic accuracy of the PI-RADS score was evaluated using histopathology of prostate biopsies as the reference standard. PCa was histologically diagnosed in 169 (21.2%) regions. Increased PI-RADS score correlated positively with increased cancer detection rate. The cancer detection rate for scores 1 to 5 was 2.8%, 15.0%, 34.6%, 52.6%, and 88.9%, respectively, using T2WI and 12.0%, 20.2%, 48.0%, 85.7%, and 93.3%, respectively, using DWI. For T2WI + DWI, the cancer detection rate was 1.5% (score 2), 13.5% (scores 3-4), 41.3% (scores 5-6), 75.9% (scores 7-8), and 92.3% (scores 9-10). The area under the curve for cancer detection was 0.700 (T2WI), 0.735 (DWI) and 0.749 (T2WI + DWI). The sensitivity and specificity were 53.8% and 89.2%, respectively, when using scores 5-6 as the cutoff value for T2WI + DWI. The PI-RADS score correlates with the PCa detection rate in patients with PSA <20 ng/ml. The summed score of T2WI + DWI has the highest accuracy in detection of PCa. However, the sensitivity should be further improved.

  15. Viral pathogenesis in diagrams

    National Research Council Canada - National Science Library

    Tremblay, Michel; Berthiaume, Laurent; Ackermann, Hans-Wolfgang

    2001-01-01

    .... The 268 diagrams in Viral Pathogenesis in Diagrams were selected from over 800 diagrams of English and French virological literature, including one derived from a famous drawing by Leonardo da Vinci...

  16. Viral Gastroenteritis (Stomach Flu)

    Science.gov (United States)

    ... Viral gastroenteritis (stomach flu) Symptoms & causes Diagnosis & treatment Advertisement Mayo Clinic does not endorse companies or products. ... a Job Site Map About This Site Twitter Facebook Google YouTube Pinterest Mayo Clinic is a not- ...

  17. Viral hemorrhagic septicemia

    Science.gov (United States)

    Batts, William N.; Winton, James R.

    2012-01-01

    Viral hemorrhagic septicemia (VHS) is one of the most important viral diseases of finfish worldwide. In the past, VHS was thought to affect mainly rainbow trout Oncorhynchus mykiss reared at freshwater facilities in Western Europe where it was known by various names including Egtved disease and infectious kidney swelling and liver degeneration (Wolf 1988). Today, VHS is known as an important source of mortality for cultured and wild fish in freshwater and marine environments in several regions of the northern hemisphere (Dixon 1999; Gagné et al. 2007; Kim and Faisal 2011; Lumsden et al. 2007; Marty et al. 1998, 2003; Meyers and Winton 1995; Skall et al. 2005b; Smail 1999; Takano et al. 2001). Viral hemorrhagic septicemia is caused by the fish rhabdovirus, viral hemorrhagic septicemia virus (VHSV), a member of the genus Novirhabdovirus of the family Rhabdoviridae

  18. HIV Viral Load

    Science.gov (United States)

    ... Chains Sex Hormone Binding Globulin (SHBG) Shiga toxin-producing Escherichia coli Sickle Cell Tests Sirolimus Smooth Muscle ... used each time. Will exercise, nutrition, and other lifestyle modifications help decrease my HIV viral load? There ...

  19. Carbohydrate antigen microarrays.

    Science.gov (United States)

    Wang, Denong

    2012-01-01

    This chapter describes one of my laboratory's working protocols for carbohydrate-based microarrays. Using a standard microarray spotter, we print carbohydrate antigens directly on the nitrocellulose-coated bioarray substrates. Because these substrates support noncovalent immobilization of many spotted antigens, in general no chemical modification of the antigen is needed for microarray production. Thus, this bioarray platform is technically simple and applicable for high-throughput construction of carbohydrate antigen microarrays. A number of nitrocellulose-coated glass slides with different technical characteristics are commercially available. Given the structural diversity of carbohydrate antigens, examining each antigen preparation to determine the efficacy of its immobilization in a given type of substrate and the surface display of the desired glycoepitopes in a microarray assay is essential.

  20. Myeloid C-Type Lectin Receptors in Viral Recognition and Antiviral Immunity.

    Science.gov (United States)

    Monteiro, João T; Lepenies, Bernd

    2017-03-22

    Recognition of viral glycans by pattern recognition receptors (PRRs) in innate immunity contributes to antiviral immune responses. C-type lectin receptors (CLRs) are PRRs capable of sensing glycans present in viral pathogens to activate antiviral immune responses such as phagocytosis, antigen processing and presentation, and subsequent T cell activation. The ability of CLRs to elicit and shape adaptive immunity plays a critical role in the inhibition of viral spread within the host. However, certain viruses exploit CLRs for viral entry into host cells to avoid immune recognition. To block CLR interactions with viral glycoproteins, antiviral strategies may involve the use of multivalent glycan carrier systems. In this review, we describe the role of CLRs in antiviral immunity and we highlight their dual function in viral clearance and exploitation by viral pathogens.

  1. A subset of human immunodeficiency virus type 1 long-term non-progressors is characterized by the unique presence of ancestral sequences in the viral population.

    Science.gov (United States)

    Bello, Gonzalo; Casado, Concepción; Sandonis, Virginia; Alonso-Nieto, Manuela; Vicario, José Luis; García, Soledad; Hernando, Victoria; Rodríguez, Carmen; del Romero, Jorge; López-Galíndez, Cecilio

    2005-02-01

    Within human immunodeficiency virus type 1 (HIV-1)-infected patients, there are those who have been infected for more than 10 years with a CD4+ cell count of >500 cells microl(-1) and who remain asymptomatic without antiretroviral therapy; these patients are designated long-term non-progressors (LTNPs). In a set of 16 LTNPs, viral dating, DNA viral load, quasispecies heterogeneity and antibody (Ab) titres against gp160 and beta2 microglobulin (beta2m) were determined. Plasma viral RNA and CD4+ and CD8+ T-cell numbers were estimated in more than three samples per patient. Host genetic characteristics, such as Delta32-CCR5 genotype and human leukocyte antigen (HLA) genotype and supertypes, and clinical-epidemiological factors were evaluated. Dating of global populations and of DNA and RNA viral quasispecies identified two subsets of patients: one displaying only ancestral sequences and the other displaying predominantly modern sequences. The ancestral patients displayed a significant reduction in RNA and DNA viral loads, quasispecies heterogeneity, CD8+ cell number, anti-gp160 Ab titres and beta2m level, and they were also associated with better use of safe-sex practices and higher presence of the HLA sB58 supertype than the modern subset. Viral dating has therefore permitted the segregation of LTNPs into two subsets that show very different virological, immunological, host and clinical-epidemiological characteristics. Moreover, whereas the modern subset displayed low levels of virus replication, the ancestral group displayed not only a very limited virus replication, often to undetectable levels, but also very slow or arrested viral evolution, maintaining the close relationship of the viral population to the transmitted virus.

  2. Antigens of Streptococcus sanguis

    Science.gov (United States)

    Rosan, Burton

    1973-01-01

    An antigenic analysis of the alpha-hemolytic streptococci isolated from dental plaque was performed by use of antisera against a strain of Streptococcus sanguis (M-5) which was isolated from dental plaque. Immunoelectrophoretic and Ouchterlony tests of Rantz and Randall extracts of 45 strains gave positive reactions with the M-5 antisera. These strains represented 60% of the strains tested. The number of antigens which could be identified in these extracts varied from one to five and were designated a to e. The a antigen was found in 36 of the strains tested, including reference strains of S. sanguis and the group H streptococci. The strains reacting with the M-5 antisera were divided into two majors types: type I consisted of 23 strains in which the a antigen was found alone or with one or more of the c, d, and e antigens; type II consisted of 13 strains in which both the a and b antigens were found with or without one or more of the c, d, and e antigens. The remaining strains contained, either singly or in combination, the b, c, d, and e antigens but not the a antigen. Biochemical tests of representatives of each serotype and reference strains indicated that strains reacting with M-5 antisera were S. sanguis. These findings suggest that S. sanguis strains share common physiological and serological properties. Images PMID:4633291

  3. AntigenMap 3D: an online antigenic cartography resource.

    Science.gov (United States)

    Barnett, J Lamar; Yang, Jialiang; Cai, Zhipeng; Zhang, Tong; Wan, Xiu-Feng

    2012-05-01

    Antigenic cartography is a useful technique to visualize and minimize errors in immunological data by projecting antigens to 2D or 3D cartography. However, a 2D cartography may not be sufficient to capture the antigenic relationship from high-dimensional immunological data. AntigenMap 3D presents an online, interactive, and robust 3D antigenic cartography construction and visualization resource. AntigenMap 3D can be applied to identify antigenic variants and vaccine strain candidates for pathogens with rapid antigenic variations, such as influenza A virus. http://sysbio.cvm.msstate.edu/AntigenMap3D

  4. Bile acids for viral hepatitis

    DEFF Research Database (Denmark)

    Chen, Weikeng; Liu, J; Gluud, C

    2003-01-01

    The viral hepatitides are common causes of liver diseases globally. Trials have assessed bile acids for patients with viral hepatitis, but no consensus was reached regarding their usefulness.......The viral hepatitides are common causes of liver diseases globally. Trials have assessed bile acids for patients with viral hepatitis, but no consensus was reached regarding their usefulness....

  5. Obtaining of three recombinant antigens of Entamoeba histolytica and evaluation of their immunogenic ability without adjuvant in a hamster model of immunoprotection.

    Science.gov (United States)

    González-Vázquez, María Cristina; Carabarin-Lima, Alejandro; Baylón-Pacheco, Lidia; Talamás-Rohana, Patricia; Rosales-Encina, José Luis

    2012-05-01

    A 30-kDa surface collagen binding protein peroxiredoxin of Entamoeba histolytica (EhCBP30) was evaluated either alone or fused to the chaperone (CHP) or ATPase (ATP) domains of heat shock protein 70 of Trypanosoma cruzi (TcHSP70) as a vaccine candidate in a hamster model of experimental amoebic liver abscess (ALA) development. Three constructs were produced containing the EhCBP30 DNA sequence, one expressing EhCBP30 and two expressing EhCBP30 fused to either CHP or ATP domains of TcHSP70. High purity recombinant proteins rEhCBP30, rEhCBP30-CHP and rEhCBP30-ATP with N-terminal His tag were obtained by single step affinity purification. Hamsters were immunized without adjuvant with the antigenic recombinant proteins and then challenged intrahepatically with E. histolytica trophozoites. A 70% decrease in ALA development was detected in hamsters immunized with rEhCBP30 and rEhCBP30-CHP, while animals immunized with rEhCBP30-ATP did not show a statistically significant decrease in ALA formation compared with non-immunized animals. Histological analysis of liver tissue showed that the inflammatory infiltrate was discrete or moderate in hamsters immunized with rEhCBP30 or rEhCBP30-CHP compared with that observed in control hamsters or hamsters immunized with rEhCBP30-ATP. These results suggest that rEhCBP30 and rEhCBP30-CHP are able to induce an effective immune response that may protect hamsters against ALA development. Copyright © 2011 Elsevier B.V. All rights reserved.

  6. Evaluation of local immune response to Fasciola hepatica experimental infection in the liver and hepatic lymph nodes of goats immunized with Sm14 vaccine antigen

    Directory of Open Access Journals (Sweden)

    Ricardo E Mendes

    2010-08-01

    Full Text Available Protection against Fasciola hepatica in goats immunized with a synthetic recombinant antigen from Schistosoma mansoni fatty acid-binding protein 14 (rSm14 was investigated by assessing worm burdens, serum levels of hepatic enzymes, faecal egg count and hepatic damage, which was evaluated using gross and microscopic morphometric observation. The nature of the local immune response was assessed by examining the distribution of CD2+, CD4+, CD8+ and γ´+ T lymphocytes along with IgG+, IL-4+ and IFN-γ+ cells in the liver and hepatic lymph nodes (HLN. The goats used consisted of group 1 (unimmunized and uninfected, group 2 [infected control - immunized with Quillaia A (Quil A] and group 3 (immunized with rSm14 in Quil A and infected, each containing seven animals. Immunization with rSm14 in Quil A adjuvant induced a reduction in gross hepatic lesions of 56.6% (p < 0.001 and reduced hepatic and HLN infiltration of CD2+, CD4+, CD8+ and γ´+ T lymphocytes as well as IL-4+ and IFN-γ+ cells (p < 0.05. This is the first report of caprine immunization against F. hepatica using a complete rSm14 molecule derived from S. mansoni. Immunization reduced hepatic damage and local inflammatory infiltration into the liver and HLN. However, considering that Quil A is not the preferential/first choice adjuvant for Sm14 immunization, further studies will be undertaken using the monophosphoryl lipid A-based family of adjuvants during clinical trials to facilitate anti-Fasciolavaccine development.

  7. Evaluation of a simultaneous detection kit for the glutamate dehydrogenase antigen and toxin A/B in feces for diagnosis of Clostridium difficile infection.

    Science.gov (United States)

    Kawada, Miki; Annaka, Megumi; Kato, Haru; Shibasaki, Sumie; Hikosaka, Keiko; Mizuno, Hiroshi; Masuda, Yoshishige; Inamatsu, Takashi

    2011-12-01

    Rapid detection kits for toxin A/B in feces are widely used as a diagnostic tool for Clostridium difficile infection (CDI). Their low sensitivity, however, has been considered a problem. In this study, we evaluated a new rapid diagnostic kit for simultaneous detection of the glutamate dehydrogenase (GDH) antigen and toxin A/B, C. DIFF QUIK CHEK COMPLETE. A total of 60 stool specimens from 60 patients with antibiotic-associated diarrhea were examined. Using C. difficile culture as the reference method, the GDH portion of this kit indicated a sensitivity, specificity, and negative predictive value of 100, 93.3, and 100%, respectively. The toxin A/B portion showed a sensitivity and specificity of 78.6 and 96.9%, respectively, compared to the culture results of toxin B-positive C. difficile (toxigenic culture). Of the 23 specimens that showed "dual positives" for GDH and toxin A/B, 22 were toxigenic culture positive, whereas C. difficile culture was negative in all the 28 specimens that showed "dual negatives" for GDH and toxin A/B. Of the nine "GDH-positive and toxin A/B-negative" specimens, six exhibited positive results by toxigenic culture. Results showing "dual positives" and "dual negatives" for GDH and toxin A/B can be reported as "true positive" and "true negative," respectively, whereas additional testing for confirmation, such as toxigenic culture, is required for specimens with discrepant results. Diagnostic algorithms, utilizing the simultaneous detection kit for GDH and toxin A/B as an initial screening test, may be useful for accurate and efficient diagnosis of CDI as well as the control of healthcare-associated infections.

  8. Evaluation of circulating cathodic antigen (CCA) urine-cassette assay as a survey tool for Schistosoma mansoni in different transmission settings within Bugiri District, Uganda.

    Science.gov (United States)

    Adriko, M; Standley, C J; Tinkitina, B; Tukahebwa, E M; Fenwick, A; Fleming, F M; Sousa-Figueiredo, J C; Stothard, J R; Kabatereine, N B

    2014-08-01

    Diagnosis of schistosomiasis at the point-of-care (POC) is a growing topic in neglected tropical disease research. There is a need for diagnostic tests which are affordable, sensitive, specific, user-friendly, rapid, equipment-free and delivered to those who need it, and POC is an important tool for disease mapping and guiding mass deworming. The aim of present study was to evaluate the relative diagnostic performance of two urine-circulating cathodic antigen (CCA) cassette assays, one commercially available and the other in experimental production, against results obtained using the standard Kato-Katz faecal smear method (six thick smears from three consecutive days), as a 'gold-standard', for Schistosoma mansoni infection in different transmission settings in Uganda. Our study was conducted among 500 school children randomly selected across 5 schools within Bugiri district, adjacent to Lake Victoria in Uganda. Considering results from the 469 pupils who provided three stool samples for the six Kato-Katz smears, 293 (76%) children had no infection, 109 (23%) were in the light intensity category, while 42 (9%) and 25 (5%) were in the moderate and heavy intensity categories respectively. Following performance analysis of CCA tests in terms of sensitivity, specificity, negative and positive predictive values, overall performance of the commercially available CCA test was more informative than single Kato-Katz faecal smear microscopy, the current operational field standard for disease mapping. The current CCA assay is therefore a satisfactory method for surveillance of S. mansoni in an area where disease endemicity is declining due to control interventions. With the recent resolution on schistosomiasis elimination by the 65th World Health Assembly, the urine POC CCA test is an attractive tool to augment and perhaps replace the Kato-Katz sampling within ongoing control programmes. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. HBeAg and not genotypes predicts viral load in patients with hepatitis B in Denmark: a nationwide cohort study

    DEFF Research Database (Denmark)

    Krarup, Henrik Bygum; Andersen, Stig; Madsen, Poul Henning

    2011-01-01

    To explore the influence of HBV genotype on viral load in patients with HBV infection, and to investigate the relation to gender, age and country of origin or antibodies against hepatitis Be antigen (anti-HBe).......To explore the influence of HBV genotype on viral load in patients with HBV infection, and to investigate the relation to gender, age and country of origin or antibodies against hepatitis Be antigen (anti-HBe)....

  10. HBeAg and not genotypes predicts viral load in patients with hepatitis B in Denmark: A nationwide cohort study

    DEFF Research Database (Denmark)

    Krarup, Henrik; Andersen, Stig; Madsen, Poul Henning

    2011-01-01

    To explore the influence of HBV genotype on viral load in patients with HBV infection, and to investigate the relation to gender, age and country of origin or antibodies against hepatitis Be antigen (anti-HBe).......To explore the influence of HBV genotype on viral load in patients with HBV infection, and to investigate the relation to gender, age and country of origin or antibodies against hepatitis Be antigen (anti-HBe)....

  11. Use of a large national database for comparative evaluation of the effect of a bronchiolitis/viral pneumonia clinical care guideline on patient outcome and resource utilization.

    Science.gov (United States)

    Todd, James; Bertoch, David; Dolan, Susan

    2002-11-01

    To use a large national comparative database to measure the internal effect of a set of evidence-based bronchiolitis/viral pneumonia clinical care guidelines on clinical practice at a children's hospital, and to compare these changes with those at other children's hospitals. Prospective cohort study with retrospective and concurrent (other hospital) controls. The Children's Hospital, Denver, Colo. Hospitalized children with bronchiolitis and/or viral pneumonia. Our clinical guidelines focused on clear admission and discharge criteria, individualized transition-anticipating orders, and "prove it or don't use it" criteria for the use of respiratory syncytial virus testing, bronchodilators, chest physiotherapy, and ribavirin. The effect of guideline implementation was determined by comparative measurement of internal changes in utilization and outcome (nosocomial infection rate) across time and by external comparison with other children's hospitals using standardized data from the Pediatric Health Information System database of the Child Health Corporation of America (Shawnee Mission, Kan). Overall, 10 636 bronchiolitis/viral pneumonia cases were studied: 1302 at the index hospital and 9334 at the 7 comparison hospitals. Internally, the index hospital's residents and attending physicians responded favorably to the bronchiolitis/viral pneumonia care guidelines, resulting in decreases in targeted resource utilization. There were no fatalities, and the number of days in the intensive care unit decreased even though the mean severity of admitted cases increased significantly. Targeted utilization was favorably affected, whereas untargeted utilization was not. Nosocomial infections did not increase with a decreased use of respiratory syncytial virus testing. The index hospital differed favorably from other children's hospitals in several categories. Evidence-based care guidelines can successfully influence utilization and clinical outcome.

  12. Serological evaluation of relationship between viral pathogens (BHV-1, BVDV, BRSV, PI-3V, and Adeno3) and dairy calf pneumonia by indirect ELISA.

    Science.gov (United States)

    Roshtkhari, Fatemeh; Mohammadi, Gholamreza; Mayameei, Ashraf

    2012-06-01

    In this study, viral pathogens associated with nine outbreaks of naturally occurring dairy calf pneumonia in Mashhad area of Khorasan Razavi province from September 2008 to May 2009 were assessed. Five diseased calves from each farm were chosen for examination. Acute and convalescent serum samples were taken from calves with signs of respiratory disease. Sera were analyzed for antibodies to bovine viral diarrhea virus (BVDV), bovine herpesvirus type 1 (BHV-1), bovine respiratory syncytial virus (BRSV), parainfluenza virus type 3 (PI-3V), and bovine adenovirus-3 (BAV-3) by indirect ELISA kits. Among 42 serum samples collected at sample 1, seroprevalence values for viruses BHV-1, BVDV, BRSV, PI-3V, and BAV-3 were 61.9% (26), 57.1% (24), 64.2% (27), 90% (38), and 61.9% (26), respectively. Seroconversion to BVDV, BRSV, PI-3V, and BAV-3 occurred in 11.9% (5), 16.6% (7), 26.1% (11), and 21.4% (9) of animals, and 52.3% (22) had generated antibodies against one or more viral infections at sample 2. In addition, no significant relationship between seroprevalence of BHV-1, BVDV, BRSV, PI-3V, and BAV-3 and dairy herd size was observed (P > 0.05). According to serological findings, BHV-1, BVDV, BRSV, PI-3V, and BAV-3 are common pathogens of the dairy calf pneumonia in dairy herds in Mashhad area of Khorasan Razavi province, Iran.

  13. Mucosal Vaccine for Prevention of Viral Disease in Animal

    Directory of Open Access Journals (Sweden)

    Sudarisman

    2006-12-01

    Full Text Available The major obstacle in combating infectious viral diseases in animals is the lack of effective vaccines . A large number of viral pathogens are mucosaly transmitted and must cross mucosal barriers to infect the host . The mucosal surfaces of the gastrointestinal and respiratory tracts represent the principal portals of entry for most animal viral pathogens . Current inactivated viral vaccines administered by intramuscular injection elicit primarily circulating antibodies . The best defense against these predominantly mucosal viral pathogens would be vaccines capable inducing both systemic and mucosal immunity which is a cost effective disease prevention tool . For most viral pathogens, induction of mucosal immunity appears most appropriate based on the routes of infection . The effectiveness of vaccine delivery to mucosal surfaces including respiratory tract may be most useful for prevention of the upper ways where secretory antibody is most important for protection against viral infection . Most external mucosal surfaces are replete with organized follicles and scattered antigen-reactive or sensitized lymphoid elements, including B cells, T lymphocytes, T cell subsets . plasma cells and a variety of other cellular elements involved in the induction and maintenance of immune response . Thus, a better understanding of the mucosal immune system is needed before effiective mucosal vaccines can be developed.

  14. Viral Evasion and Subversion Mechanisms of the Host Immune System

    Directory of Open Access Journals (Sweden)

    Mehran Ghaemi-Bafghi

    2013-10-01

    Full Text Available Viruses are the most abundant and versatile pathogens which challenge the immune system and cause major threats to human health. Viruses employ differ¬ent mechanisms to evade host immune responses that we describe them under the following headings: Inhibition of humoral responses, Interference with interferons, Inhibition and modulation of cytokines and chemokines, Inhibitors of apoptosis, Evading CTLs and NKs, and modulating MHC function.Viruses inhibit humoral immunity in different ways which contains change of viral antigens, production of regulatory proteins of complement system and receptors of the Fc part of antibodies. Viruses block interferon production and function via interruption of cell signaling JAK/STAT pathway, Inhibition of eIF-2α phosphorylation and translational arrest and 2'5'OS/RNAse L system. Also, Poxviruses produce soluble versions of receptors for interferons. One of the most important ways of viral evasion is inhibition and manipulation of cytokines; for example, Herpsviruses and Poxviruses produce viral cytokines (virokines and cytokine receptors (viroceptors. In addition, viruses change maturation and expression of MHC I and MHC II molecules to interrupt viral antigens presentation and hide them from immune system recognition. Also, they inhibit NK cell functions.In this review, we provide an overview of the viral evasion mechanisms of immune system. Since most viruses have developed strategies for evasion of immune system, if we know these mechanisms in detail we can fight them more successfully.

  15. VIRAL DISEASES IN SEA FISH

    OpenAIRE

    Ivančica Strunjak-Perović; Mato Hacmanjek; Rozelinda Čož-Rakovac; Emin Teskeredžić

    1996-01-01

    Adequate knowledge on fish diseases caused by viruses is still lacking. Up until now, in fish which live their entire life cycle or part of it in the sea, some viral diseases have been determined (lymphoeytis, viral necrosis of crythrocytes, ciravosti cod syndrome, encephalitis, viral hemoragic septichemistry, viral hematopoetic necrosis, viral gusteraca necrosis, chum renviral infection, branchionephritis, rabdociral eel infection). Some of these diseases primarily occur in the freshwater ph...

  16. Diagnostic Values of Carcinoembryonic Antigen, Cancer Antigen 15-3 and Cancer Antigen 125 Levels in Nipple Discharge.

    Science.gov (United States)

    Zhao, Song; Gai, Xiaodong; Wang, Yongmei; Liang, Weili; Gao, Haidong; Zhang, Kai; Wang, Huimin; Liu, Yanhong; Wang, Jianli; Ma, Rong

    2015-12-31

    An expedient and cost-effective diagnostic tool is needed to complement galactography and exfoliative cytology for detection of benign or malignant breast diseases with nipple discharge. The aim of this prospective study is to explore the utility of carcinoembryonic antigen, cancer antigen 15-3 and cancer antigen 125 levels in nipple discharge for the diagnosis of various breast diseases. We evaluated the pre-operative tumor marker levels in 153 nipple discharge samples collected from one or both breasts of 142 women undergoing surgery. Patients with nipple discharge underwent auxiliary examination (ultrasonography, exfoliative cytology, ductoscopy and galactography). Statistically higher levels of carcinoembryonic antigen and cancer antigen 15-3 were found in patients in the malignant group as compared to those in the benign group. No statistically significant difference in the level of cancer antigen 125 (P = 0.895). Sensitivities of carcinoembryonic antigen and cancer antigen 15-3 for diagnosing breast cancer were 74.42% and 58.14%, and specificities were 87.27% and 80.00% where as the cutoff values with max-sum of sensitivity and specificity were 224.3 ng/ml and 1368.2 U/ml, respectively. The following sensitivities for telling malignant from benign could be determined: exfoliative cytology 46.67%, ultrasonography 76.74%, galactography 75.00%, and ductoscopy 0%. Exfoliative cytology was found to be a valuable alternative method for differentiating benign from malignancy. Thus, tumor marker analysis of nipple discharge fluid for carcinoembryonic antigen and cancer antigen 15-3 would enhance the accurate assessment and treatment planning for patients with nipple discharge.

  17. Antigen presentation pathways to class I and class II MHC-restricted T lymphocytes.

    Science.gov (United States)

    Braciale, T J; Morrison, L A; Sweetser, M T; Sambrook, J; Gething, M J; Braciale, V L

    1987-08-01

    intracellularly with newly synthesized or recycling class I MHC molecules. An understanding of the molecular basis of this phenomenon will require detailed information on the expression, intracellular trafficking, and transport of class I and class II MHC molecules in the antigen-presenting cell. An unresolved issue, at least in the case of viral antigens, is the nature and form of the antigenic moieties presented by the exogenous and endogenous pathways. In the case of viral antigen presentation to class II MHC-restricted T lymphocytes, there is strong, albeit indirect, evidence for processing of antigen and recognition of fragments of viral polypeptides (Lamb et al. 1982, Hackett et al. 1983).(ABSTRACT TRUNCATED AT 400 WORDS)

  18. Comparative evaluation of recombinant LigB protein and heat-killed antigen-based latex agglutination test with microscopic agglutination test for diagnosis of bovine leptospirosis.

    Science.gov (United States)

    Nagalingam, Mohandoss; Thirumalesh, Sushma Rahim Assadi; Kalleshamurthy, Triveni; Niharika, Nakkala; Balamurugan, Vinayagamurthy; Shome, Rajeswari; Sengupta, Pinaki Prasad; Shome, Bibek Ranjan; Prabhudas, Krishnamsetty; Rahman, Habibur

    2015-10-01

    This study aimed to develop latex agglutination test (LAT) using recombinant leptospiral immunoglobulin-like protein (LigB) (rLigB) antigen and compare its diagnostic efficacy with LAT using conventional heat-killed leptospiral antigen and microscopic agglutination test (MAT) in diagnosing bovine leptospirosis. The PCR-amplified 1053-bp ligB gene sequences from Leptospira borgpetersenii Hardjo serovar were cloned in pET 32 (a) vector at EcoRI and NotI sites and expressed in BL21 E. coli cells as fusion protein with thioredoxin (-57 kDa) and characterized by SDS-PAGE and immunoblot. Out of 390 serum samples [cattle (n = 214), buffaloes (n = 176)] subjected to MAT, 115 samples showed reciprocal titre≥100 up to 1600 against one or more serovars. For recombinant LigB protein/antigen-based LAT, agglutination was observed in the positive sample, while no agglutination was observed in the negative sample. Similarly, heat-killed leptospiral antigen was prepared from and used in LAT for comparison with MAT. A two-sided contingency table was used for analysis of LAT using both the antigens separately against MAT for 390 serum samples. The sensitivity, specificity and positive and negative predictive values of recombinant LigB LAT were found to be 75.65, 91.27, 78.38 and 89.96 %, respectively, and that of heat-killed antigen-based LAT were 72.17, 89.82, 74.77 and 88.53 %, respectively, in comparison with MAT. This developed test will be an alternative/complementary to the existing battery of diagnostic assays/tests for specific detection of pathogenic Leptospira infection in bovine population.

  19. Evaluation of antigen detection and polymerase chain reaction for diagnosis of amoebic liver abscess in patients on anti-amoebic treatment

    Directory of Open Access Journals (Sweden)

    Jaiswal Virendra

    2012-08-01

    Full Text Available Abstract Background Diagnosis of amoebic liver abscess (ALA in patients on anti-amoebic drugs is difficult. There is scanty data on this issue using Entamoeba histolytica (E. histolytica lectin antigen and polymerase chain reaction (PCR. We studied utility of lectin antigen, PCR, and IgG antibody in diagnosis of liver abscess in patients on anti-amoebic treatment. Liver aspirate of 200 patients, of which 170 had anti-amoebic drug prior to drainage, was tested for E. histolytica lectin antigen by (ELISA, PCR, bacterial culture, and serum IgG antibody by (ELISA. Classification of abscesses was based on result of anti-amoebic IgG antibody and bacterial culture, E. histolytica PCR and bacterial culture, and E. histolytica lectin antigen and bacterial culture. Findings Using anti-amoebic IgG antibody and bacterial culture, 136/200 (68.0% were classified as ALA, 12/200 (6.0% as pyogenic liver abscess (PLA, 29/200 (14.5% as mixed infection, and 23/200 (11.5% remained unclassified. Using amoebic PCR and bacterial culture 151/200 (75.5% were classified as ALA, 25/200 (12.5% as PLA, 16/200 (8.0% as mixed infection, and 8/200 (4.0% remained unclassified. With E. histolytica lectin antigen and bacterial culture, 22/200 (11.0% patients were classified as ALA, 39/200 (19.5% as PLA, 2/200 (1.0% as mixed infection, and 137/200 (68.5% remained unclassified. Conclusions E. histolytica lectin antigen was not suitable for classification of ALA patients who had prior anti-amoebic treatment. However, PCR may be used as alternative test to anti-amoebic antibody in diagnosis of ALA.

  20. An investigation of an outbreak of viral hepatitis B in Modasa town, Gujarat, India

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    Disha A Patel

    2012-01-01

    Full Text Available Background: Most outbreaks of viral hepatitis in India are caused by hepatitis E. Recently in the year 2009, Modasa town of Sabarkantha district in Gujarat witnessed the outbreak of hepatitis B. Purpose: An attempt was made to study the outbreak clinically and serologically, to estimate the seropositivity of hepatitis B Virus among the cases and their contacts and to know the seroprevalence of hepatitis B envelope antigen (HBeAg and IgM antibody against hepatitis B core antigen (IgM HBcAb out of all the Hepatitis B surface Antigen (HBsAg positive ones. Materials and Methods: Eight hundred and fifty-six (856 cases and 1145 contacts were evaluated for hepatitis B markers namely HBsAg, HBeAg and IgM HBcAb by enzyme-linked immuno Sorbent Assay (ELISA test. Results: This outbreak of viral hepatitis B in Modasa, Gujarat was most likely due to unsafe injection practices. Evidence in support of this was collected by Government authorities. Most of the patients and approximately 40% of the surveyed population gave history of injections in last 1.5-6 months. Total 664/856 (77.57% cases and 20/1145 (1.75% contacts were found to be positive for HBsAg. 53.41% of the positive cases and 52.93% of the positive contacts were HBeAg-positive and thus in a highly infectious stage. Conclusions: Inadequately sterilized needles and syringes are an important cause of transmission of hepatitis B in India. Our data reflects the high positivity rate of a hepatitis B outbreak due to such unethical practices. There is a need to strengthen the routine surveillance system, and to organise a health education campaign targeting all health care workers including private practitioners, especially those working in rural areas, as well as the public at large, to take all possible measures to prevent this often fatal infection.

  1. Production and Evaluation of Toxoplasma gondii Recombinant Sur­face Antigen 1 (SAG1 for Serodiagnosis of Acute and ChronicToxop­lasma Infection in Human Sera

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    M(Monavar Selseleh

    2012-09-01

    Full Text Available Background: The assays currently available for the detection of specific anti-Toxoplasma antibodies may vary in their abilities to detect serum immunoglobulins, due to the Lack of a purified standardized antigen. The aim of this study was evaluation the recombinant Toxoplasma gondii SAG1 antigen for the serodiagnosis of acute and chronic toxoplasmosis. Methods: This study describes an ELISA using recombinant SAG1 for detection of IgM and IgG antibodies against Toxoplasma gondii in human sera. Genomic DNA of T. gondii (RH Strain was isolated and PCR reaction was performed. Recovered DNA was cloned into PTZ57R cloning vector. The recombinant plasmid was detected by restriction analysis. The SAG1 gene was subcloned in the pET- 28a expression vector. Protein production was then induced with 1 mM isopropyl-D - thiogalactopyrano­side (IPTG. A total of 204 sera were tested using a commercial IgG and IgM ELISA kit (Trinity, USA as gold standard prior to testing them with the recombinant antigen. Results: Tested sera were divided into the following groups:(a The 74 T. gondii IgG positive (b 70 T.gondii IgM positive (c 60 sera who had no serological evidence of toxoplasmosis as negative sera.To determine the specificity of the test, we used other parasitic diseases including echinococusis (N=5, malaria (N=14, leishmania­sis (N=7,fasciolasis (N=4 , sterengyloidiasis (N=1 . Sensitivity and specificity of the generated recombinant IgG ELISA in comparison with commercial ELISA (Com ELISA were 93% and 95%, and the sensitivity and specificity of the generated recombinant IgM ELISA were 87% and 95% respectively. Conclusion: The results acquired here show that this antigen is useful for diagnostic purposes and could be replaced by lysed, whole cell antigens for diagnosis of chronic toxoplasmosis.

  2. Modeling Viral Capsid Assembly

    Science.gov (United States)

    2014-01-01

    I present a review of the theoretical and computational methodologies that have been used to model the assembly of viral capsids. I discuss the capabilities and limitations of approaches ranging from equilibrium continuum theories to molecular dynamics simulations, and I give an overview of some of the important conclusions about virus assembly that have resulted from these modeling efforts. Topics include the assembly of empty viral shells, assembly around single-stranded nucleic acids to form viral particles, and assembly around synthetic polymers or charged nanoparticles for nanotechnology or biomedical applications. I present some examples in which modeling efforts have promoted experimental breakthroughs, as well as directions in which the connection between modeling and experiment can be strengthened. PMID:25663722

  3. Interval Between Infections and Viral Hierarchy Are Determinants of Viral Interference Following Influenza Virus Infection in a Ferret Model

    Science.gov (United States)

    Laurie, Karen L.; Guarnaccia, Teagan A.; Carolan, Louise A.; Yan, Ada W. C.; Aban, Malet; Petrie, Stephen; Cao, Pengxing; Heffernan, Jane M.; McVernon, Jodie; Mosse, Jennifer; Kelso, Anne; McCaw, James M.; Barr, Ian G.

    2015-01-01

    Background. Epidemiological studies suggest that, following infection with influenza virus, there is a short period during which a host experiences a lower susceptibility to infection with other influenza viruses. This viral interference appears to be independent of any antigenic similarities between the viruses. We used the ferret model of human influenza to systematically investigate viral interference. Methods. Ferrets were first infected then challenged 1–14 days later with pairs of influenza A(H1N1)pdm09, influenza A(H3N2), and influenza B viruses circulating in 2009 and 2010. Results. Viral interference was observed when the interval between initiation of primary infection and subsequent challenge was virus specific and occurred between antigenically related and unrelated viruses. Coinfections occurred when 1 or 3 days separated infections. Ongoing shedding from the primary virus infection was associated with viral interference after the secondary challenge. Conclusions. The interval between infections and the sequential combination of viruses were important determinants of viral interference. The influenza viruses in this study appear to have an ordered hierarchy according to their ability to block or delay infection, which may contribute to the dominance of different viruses often seen in an influenza season. PMID:25943206

  4. [Viral hepatitis in travellers].

    Science.gov (United States)

    Abreu, Cândida

    2007-01-01

    Considering the geographical asymmetric distribution of viral hepatitis A, B and E, having a much higher prevalence in the less developed world, travellers from developed countries are exposed to a considerable and often underestimated risk of hepatitis infection. In fact a significant percentage of viral hepatitis occurring in developed countries is travel related. This results from globalization and increased mobility from tourism, international work, humanitarian and religious missions or other travel related activities. Several studies published in Europe and North America shown that more than 50% of reported cases of hepatitis A are travel related. On the other hand frequent outbreaks of hepatitis A and E in specific geographic areas raise the risk of infection in these restricted zones and that should be clearly identified. Selected aspects related with the distribution of hepatitis A, B and E are reviewed, particularly the situation in Portugal according to the published studies, as well as relevant clinical manifestations and differential diagnosis of viral hepatitis. Basic prevention rules considering enteric transmitted hepatitis (hepatitis A and hepatitis E) and parenteral transmitted (hepatitis B) are reviewed as well as hepatitis A and B immunoprophylaxis. Common clinical situations and daily practice "pre travel" advice issues are discussed according to WHO/CDC recommendations and the Portuguese National Vaccination Program. Implications from near future availability of a hepatitis E vaccine, a currently in phase 2 trial, are highlighted. Potential indications for travellers to endemic countries like India, Nepal and some regions of China, where up to 30% of sporadic cases of acute viral hepatitis are caused by hepatitis E virus, are considered. Continued epidemiological surveillance for viral hepatitis is essential to recognize and control possible outbreaks, but also to identify new viral hepatitis agents that may emerge as important global health

  5. Immigration and viral hepatitis.

    Science.gov (United States)

    Sharma, Suraj; Carballo, Manuel; Feld, Jordan J; Janssen, Harry L A

    2015-08-01

    WHO estimates reveal that the global prevalence of viral hepatitis may be as high as 500 million, with an annual mortality rate of up to 1.3 million individuals. The majority of this global burden of disease is borne by nations of the developing world with high rates of vertical and iatrogenic transmission of HBV and HCV, as well as poor access to healthcare. In 2013, 3.2% of the global population (231 million individuals) migrated into a new host nation. Migrants predominantly originate from the developing countries of the south, into the developed economies of North America and Western Europe. This mass migration of individuals from areas of high-prevalence of viral hepatitis poses a unique challenge to the healthcare systems of the host nations. Due to a lack of universal standards for screening, vaccination and treatment of viral hepatitis, the burden of chronic liver disease and hepatocellular carcinoma continues to increase among migrant populations globally. Efforts to increase case identification and treatment among migrants have largely been limited to small outreach programs in urban centers, such that the majority of migrants with viral hepatitis continue to remain unaware of their infection. This review summarizes the data on prevalence of viral hepatitis and burden of chronic liver disease among migrants, current standards for screening and treatment of immigrants and refugees, and efforts to improve the identification and treatment of viral hepatitis among migrants. Copyright © 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

  6. Viral meningitis and encephalitis.

    Science.gov (United States)

    Tuppeny, Misti

    2013-09-01

    Meningitis is an inflammation of the meninges, whereas encephalitis is inflammation of the parenchymal brain tissue. The single distinguishing element between the 2 diagnoses is the altered state of consciousness, focal deficits, and seizures found in encephalitis. Consequently meningoencephalitis is a term used when both findings are present in the patient. Viral meningitis is not necessarily reported as it is often underdiagnosed, whereas encephalitis cases are on the increase in various areas of North America. Improved imaging and viral diagnostics, as well as enhanced neurocritical care management, have improved patient outcomes to date. Copyright © 2013 Elsevier Inc. All rights reserved.

  7. Neutrophils and viral-induced neurologic disease.

    Science.gov (United States)

    Grist, Jonathan J; Marro, Brett; Lane, Thomas E

    2016-06-08

    Infection of the central nervous system (CNS) by neurotropic viruses represents an increasing worldwide problem in terms of morbidity and mortality for people of all ages. Although unique structural features of the blood-brain-barrier (BBB) provide a physical and physiological barrier, a number of neurotropic viruses are able to enter the CNS resulting in a variety of pathological outcomes. Nonetheless, antigen-specific lymphocytes are ultimately able to accumulate within the CNS and contribute to defense by reducing or eliminating the invading viral pathogen. Alternatively, infiltration of activated cells of the immune system may be detrimental, as these cells can contribute to neuropathology that may result in long-term cellular damage or death. More recently, myeloid cells e.g. neutrophils have been implicated in contributing to both host defense and disease in response to viral infection of the CNS. This review highlights recent studies using coronavirus-induced neurologic disease as a model to determine how neutrophils affect effective control of viral replication as well as demyelination. Copyright © 2016 Elsevier Inc. All rights reserved.

  8. [Liver hemosiderosis study in chronic viral hepatitis].

    Science.gov (United States)

    Cojocariu, Camelia; Trifan, Anca; Mihailovici, Maria Sultana; Danciu, M; Stanciu, C

    2008-01-01

    In chronic viral hepatitis the histopathological exam can reveal the presence of liver iron deposits in 10 to 73% of patients. Iron deposits are usually found in Kupffer cells, in endothelial cells and portal macrophages, and extremely rarely in hepatocytes. To evaluate the incidence of hepatic hemosiderosis in chronic viral hepatitis. 549 morphopathological features of liver biopsy specimens performed in the Gastroenterology and Hepatology Institute IaSi, between January 1 2003 and December 31 2007 have been analyzed. Semiquantitative assessment of the degree of hepatic iron overload was performed and the localization of haemosiderin deposits: at the level of hepatocytes, the reticuloendothelial system or mixedly. The same anatomopathologist examined the blades and interpreted the results. The medium age of patients who underwent liver biopsy was 45.08 years +/- 10.045. Positive iron staining was found in 22.8% of cases, more frequently in males (31%), and in 91.82% of cases iron deposits were grade 1-2. The association of alcoholic etiology did not influence the incidence of hemosiderosis: 23% in patients with hepatitis and no ethanol exposure vs 25% in cases of strictly viral etiology. Deposits of haemosiderin were more frequent in viral hepatitis B (38.6%) than in viral hepatitis C (26.9%). In 34% of cases stainable iron was found only in reticuloendothelial system and in 46% of cases both in Kupffer cells and hepatocytes. Almost a quarter of chronic viral hepatitis cases are associated with liver deposits of haemosiderin, with features of secondary iron overload (deposits localized in the mesenchymal areas or mixedly). There is a higher risk of hemosiderosis in men, especially for those between 30 and 50. Liver iron overload levels in chronic viral hepatitis are, in most cases, low or medium, and the association with an alcoholic etiology does not influence the incidence of hemosiderosis in chronic viral hepatitis.

  9. Use of antigenic cartography in vaccine seed strain selection.

    Science.gov (United States)

    Fouchier, Ron A M; Smith, Derek J

    2010-03-01

    Human influenza A viruses are classic examples of antigenically variable pathogens that have a seemingly endless capacity to evade the host's immune response. The viral hemagglutinin (HA) and neuraminidase (NA) proteins are the main targets of our antibody response to combat infections. HA and NA continuously change to escape from humoral immunity, a process known as antigenic drift. As a result of antigenic drift, the human influenza vaccine is updated frequently. The World Health Organization (WHO) coordinates a global influenza surveillance network that, by the hemagglutination inhibition (HI) assay, routinely characterizes the antigenic properties of circulating strains in order to select new seed viruses for such vaccine updates. To facilitate a quantitative interpretation and easy visualization of HI data, a new computational technique called "antigenic cartography" was developed. Since its development, antigenic cartography has been applied routinely to assist the WHO with influenza surveillance activities. Until recently, antigenic variation was not considered a serious issue with influenza vaccines for poultry. However, because of the diversification of the Asian H5N1 lineage since 1996 into multiple genetic clades and subclades, and because of the long-term use of poultry vaccines against H5 in some parts of the world, this issue needs to be re-addressed. The antigenic properties of panels of avian H5N1 viruses were characterized by HI assay, using mammalian or avian antisera, and analyzed using antigenic cartography methods. These analyses revealed antigenic differences between circulating H5N1 viruses and the H5 viruses used in poultry vaccines. Considerable antigenic variation was also observed within and between H5N1 clades. These observations have important implications for the efficacy and long-term use of poultry vaccines.

  10. Enumeration of antigen-specific CD8+ T lymphocytes by single-platform, HLA tetramer-based flow cytometry: a European multicenter evaluation.

    NARCIS (Netherlands)

    Heijnen, I.; Barnett, D.; Arroz, M.J.; Barry, S.M.; Bonneville, M.; Brando, B.; D'Hautcourt, J.L.; Kern, F.; Totterman, T.H.; Marijt, E.W.; Bossy, D.; Preijers, F.W.M.B.; Rothe, G.; Gratama, J.W.

    2004-01-01

    BACKGROUND: HLA class I peptide tetramers represent powerful diagnostic tools for detection and monitoring of antigen-specific CD8(+) T cells. The impetus for the current multicenter study is the critical need to standardize tetramer flow cytometry if it is to be implemented as a routine diagnostic

  11. The evaluation of a physical method for the quantification of inactivated poliovirus particles and its relationship to D-antigenicity and potency testing in rats.

    NARCIS (Netherlands)

    T.R. Doel; A.D.M.E. Osterhaus (Albert); A.L. van Wezel; G. van Steenis (Bert)

    1984-01-01

    textabstractThe use of a density gradient procedure for the quantification of intact, inactivated poliovirus particles in vaccine preparations is described. The procedure is both sensitive and highly reproducible and the results correlate with those of potency tests in rats and with D-antigen

  12. Evaluation of enzyme-linked immunosorbent assays based on monoclonal antibodies for the serology and antigen detection in canine parvovirus infections.

    NARCIS (Netherlands)

    G.F. Rimmelzwaan (Guus); N. Juntti; B. Klingeborn; J. Groen (Jan); F.G.C.M. Uytdehaag (Fons); A.D.M.E. Osterhaus (Albert)

    1990-01-01

    textabstractAn enzyme-linked immunosorbent assay (ELISA) system was developed for the detection of canine parvovirus (CPV) or CPV antigen in dog faeces and two other ELISA systems were developed for the detection of CPV-specific antibodies in dog sera. The ELISA's were based on the use of

  13. Real-time PCR versus viral culture on urine as a gold standard in the diagnosis of congenital cytomegalovirus infection

    NARCIS (Netherlands)

    de Vries, Jutte J. C.; van der Eijk, Annemiek A.; Wolthers, Katja C.; Rusman, Lisette G.; Pas, Suzan D.; Molenkamp, Richard; Claas, Eric C.; Kroes, Aloys C. M.; Vossen, Ann C. T. M.

    2012-01-01

    Background: Cytomegalovirus (CMV) infection is the most common cause of congenital infection. Whereas CMV PCR has replaced viral culture and antigen detection in immunocompromised patients because of higher sensitivity, viral culture of neonatal urine is still referred to as the gold standard in the

  14. Characterization of antigen-specific B cells using nominal antigen-coated flow-beads.

    Directory of Open Access Journals (Sweden)

    Nicolas Degauque

    Full Text Available In order to characterize the reactivity of B cells against nominal antigens, a method based on the coupling of antigens onto the surface of fluorescent core polystyrene beads was developed. We first demonstrate that murine B cells with a human MOG-specific BCR are able to interact with MOG-coated beads and do not recognize beads coated with human albumin or pp65. B cells purified from human healthy volunteer blood or immunized individuals were tested for their ability to interact with various nominal antigens, including viral, vaccine, self and alloantigens, chosen for their usefulness in studying a variety of pathological processes. A substantial amount of B cells binding self-antigen MOG-coated beads can be detected in normal blood. Furthermore, greater frequencies of B cell against anti-Tetanic Toxin or anti-EBNA1 were observed in primed individuals. This method can reveal increased frequencies of anti-HLA committed B cells in patients with circulating anti-HLA antibodies compared to unsensitized patients and normal individuals. Of interest, those specific CD19 cells were preferentially identified within CD27(-IgD(+ (i-e naïve subset. These observations suggest that a broad range of medical situations could benefit from a tool that allows the detection, the quantification and the characterization of antigen-specific blood B cells.

  15. Role of T cell competition in the induction of cytotoxic T lymphocyte activity during viral vector-based immunization regimens.

    NARCIS (Netherlands)

    Lambeck, A.J.A.; Nijman, H.W.; Hoogeboom, B.N.; Regts, J.; Mare, A. de; Wilschut, J.; Daemen, T.

    2010-01-01

    T cell competition between antigen- and vector-specific T cells may determine the outcome of viral vector-based immunization regimens, as we previously proposed. Here, we unravelled the interplay between antigen- and vector-specific immunity, using recombinant Semliki Forest virus (rSFV). Priming of

  16. HIV and Viral Hepatitis

    Science.gov (United States)