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  1. Prostaglandin E2 Reduces the Release and Infectivity of New Cell-Free Virions and Cell-To-Cell HIV-1 Transfer

    PubMed Central

    Serramía, María Jesús; Martínez-Bonet, Marta; Muñoz-Fernández, María Ángeles

    2014-01-01

    Background The course of human immunodeficiency virus type-1 (HIV-1) infection is influenced by a complex interplay between viral and host factors. HIV infection stimulates several proinflammatory genes, such as cyclooxigense-2 (COX-2), which leads to an increase in prostaglandin (PG) levels in the plasma of HIV-1-infected patients. These genes play an indeterminate role in HIV replication and pathogenesis. The effect of prostaglandin E2 (PGE2) on HIV infection is quite controversial and even contradictory, so we sought to determine the role of PGE2 and the signal transduction pathways involved in HIV infection to elucidate possible new targets for antiretrovirals. Results Our results suggest that PGE2 post-infection treatment acts in the late stages of the viral cycle to reduce HIV replication. Interestingly, viral protein synthesis was not affected, but a loss of progeny virus production was observed. No modulation of CD4 CXCR4 and CCR5 receptor expression, cell proliferation, or activation after PGE2 treatment was detected. Moreover, PGE2 induced an increase in intracellular cAMP (cyclic AMP) levels through the EP2/EP4 receptors. PGE2 effects were mimicked by dbcAMP and by a specific Epac (exchange protein directly activated by cyclic AMP) agonist, 8-Cpt-cAMP. Treatment with PGE2 increased Rap1 activity, decreased RhoA activity and subsequently reduced the polymerization of actin by approximately 30% compared with untreated cells. In connection with this finding, polarized viral assembly platforms enriched in Gag were disrupted, altering HIV cell-to-cell transfer and the infectivity of new virions. Conclusions Our results demonstrate that PGE2, through Epac and Rap activation, alters the transport of newly synthesized HIV-1 components to the assembly site, reducing the release and infectivity of new cell-free virions and cell-to-cell HIV-1 transfer. PMID:24586238

  2. Apolipoprotein E on Hepatitis C Virion Facilitates Infection through Interaction with Low Density Lipoprotein Receptor

    PubMed Central

    Owen, David M.; Huang, Hua; Ye, Jin; Gale, Michael

    2009-01-01

    Hepatitis C virus (HCV) infection is a major cause of liver disease. HCV associates with host apolipoproteins and enters hepatocytes through complex processes involving some combination of CD81, claudin-I, occludin, and scavenger receptor BI. Here we show that infectious HCV resembles very low density lipoprotein (VLDL) and that entry involves co-receptor function of the low density lipoprotein receptor (LDL-R). Blocking experiments demonstrate that β-VLDL itself or anti-apolipoprotein E (apoE) antibody can block HCV entry. Knockdown of the LDL-R by treatment with 25-hydroxycholesterol or siRNA ablated ligand uptake and reduced HCV infection of cells, whereas infection was rescued upon cell ectopic LDL-R expression. Analyses of gradient-fractionated HCV demonstrate that apoE is associated with HCV virions exhibiting peak infectivity and dependence upon the LDL-R for cell entry. Our results define the LDL-R as a cooperative HCV co-receptor that supports viral entry and infectivity through interaction with apoE ligand present in an infectious HCV/lipoprotein complex comprising the virion. Disruption of HCV/LDL-R interactions by altering lipoprotein metabolism may therefore represent a focus for future therapy. PMID:19751943

  3. A rev1-vpu polymorphism unique to HIV-1 subtype A and C strains impairs envelope glycoprotein expression from rev-vpu-env cassettes and reduces virion infectivity in pseudotyping assays

    SciTech Connect

    Kraus, Matthias H.; Parrish, Nicholas F.; Shaw, Katharina S.; Decker, Julie M.; Keele, Brandon F.; Salazar-Gonzalez, Jesus F.; Grayson, Truman; McPherson, David T.; Ping, Li-Hua; Anderson, Jeffrey A.; Swanstrom, Ronald; Williamson, Carolyn; Shaw, George M.; Hahn, Beatrice H.

    2010-02-20

    Functional studies of HIV-1 envelope glycoproteins (Envs) commonly include the generation of pseudoviruses, which are produced by co-transfection of rev-vpu-env cassettes with an env-deficient provirus. Here, we describe six Env constructs from transmitted/founder HIV-1 that were defective in the pseudotyping assay, although two produced infectious virions when expressed from their cognate proviruses. All of these constructs exhibited an unusual gene arrangement in which the first exon of rev (rev1) and vpu were in the same reading frame without an intervening stop codon. Disruption of the rev1-vpu fusion gene by frameshift mutation, stop codon, or abrogation of the rev initiation codon restored pseudovirion infectivity. Introduction of the fusion gene into wildtype Env cassettes severely compromised their function. The defect was not due to altered env and rev transcription or a dominant negative effect of the expressed fusion protein, but seemed to be caused by inefficient translation at the env initiation codon. Although the rev1-vpu polymorphism affects Env expression only in vitro, it can cause problems in studies requiring Env complementation, such as analyses of co-receptor usage and neutralization properties, since 3% of subtype A, 20% of subtype C and 5% of CRF01{sub A}/E viruses encode the fusion gene. A solution is to eliminate the rev initiation codon when amplifying rev-vpu-env cassettes since this increases Env expression irrespective of the presence of the polymorphism.

  4. Quantitative Correlation between Infectivity and Gp120 Density on HIV-1 Virions Revealed by Optical Trapping Virometry.

    PubMed

    DeSantis, Michael C; Kim, Jin H; Song, Hanna; Klasse, Per Johan; Cheng, Wei

    2016-06-17

    The envelope glycoprotein (Env) gp120/gp41 is required for HIV-1 infection of host cells. Although in general it has been perceived that more Env gives rise to higher infectivity, the precise quantitative dependence of HIV-1 virion infectivity on Env density has remained unknown. Here we have developed a method to examine this dependence. This method involves 1) production of a set of single-cycle HIV-1 virions with varied density of Env on their surface, 2) site-specific labeling of Env-specific antibody Fab with a fluorophore at high efficiency, and 3) optical trapping virometry to measure the number of gp120 molecules on individual HIV-1 virions. The resulting gp120 density per virion is then correlated with the infectivity of the virions measured in cell culture. In the presence of DEAE-dextran, the polycation known to enhance HIV-1 infectivity in cell culture, virion infectivity follows gp120 density as a sigmoidal dependence and reaches an apparent plateau. This quantitative dependence can be described by a Hill equation, with a Hill coefficient of 2.4 ± 0.6. In contrast, in the absence of DEAE-dextran, virion infectivity increases monotonically with gp120 density and no saturation is observed under the experimental conditions. These results provide the first quantitative evidence that Env trimers cooperate on the virion surface to mediate productive infection by HIV-1. Moreover, as a result of the low number of Env trimers on individual virions, the number of additional Env trimers per virion that is required for the optimal infectivity will depend on the inclusion of facilitating agents during infection. PMID:27129237

  5. Dynamic Antibody Specificities and Virion Concentrations in Circulating Immune Complexes in Acute to Chronic HIV-1 Infection ▿ †

    PubMed Central

    Liu, Pinghuang; Overman, R. Glenn; Yates, Nicole L.; Alam, S. Munir; Vandergrift, Nathan; Chen, Yue; Graw, Frederik; Freel, Stephanie A.; Kappes, John C.; Ochsenbauer, Christina; Montefiori, David C.; Gao, Feng; Perelson, Alan S.; Cohen, Myron S.; Haynes, Barton F.; Tomaras, Georgia D.

    2011-01-01

    Understanding the interactions between human immunodeficiency virus type 1 (HIV-1) virions and antibodies (Ab) produced during acute HIV-1 infection (AHI) is critical for defining antibody antiviral capabilities. Antibodies that bind virions may prevent transmission by neutralization of virus or mechanically prevent HIV-1 migration through mucosal layers. In this study, we quantified circulating HIV-1 virion-immune complexes (ICs), present in approximately 90% of AHI subjects, and compared the levels and antibody specificity to those in chronic infection. Circulating HIV-1 virions coated with IgG (immune complexes) were in significantly lower levels relative to the viral load in acute infection than in chronic HIV-1 infection. The specificities of the antibodies in the immune complexes differed between acute and chronic infection (anti-gp41 Ab in acute infection and anti-gp120 in chronic infection), potentially suggesting different roles in immunopathogenesis for complexes arising at different stages of infection. We also determined the ability of circulating IgG from AHI to bind infectious versus noninfectious virions. Similar to a nonneutralizing anti-gp41 monoclonal antibody (MAb), purified plasma IgG from acute HIV-1 subjects bound both infectious and noninfectious virions. This was in contrast to the neutralizing antibody 2G12 MAb that bound predominantly infectious virions. Moreover, the initial antibody response captured acute HIV-1 virions without selection for different HIV-1 envelope sequences. In total, this study demonstrates that the composition of immune complexes are dynamic over the course of HIV-1 infection and are comprised initially of antibodies that nonselectively opsonize both infectious and noninfectious virions, likely contributing to the lack of efficacy of the antibody response during acute infection. PMID:21865397

  6. Virion encapsidated HIV-1 Vpr induces NFAT to prime non-activated T cells for productive infection

    PubMed Central

    Höhne, Kristin; Businger, Ramona; van Nuffel, Anouk; Bolduan, Sebastian; Koppensteiner, Herwig; Baeyens, Ann; Vermeire, Jolien; Malatinkova, Eva; Verhasselt, Bruno; Schindler, Michael

    2016-01-01

    The majority of T cells encountered by HIV-1 are non-activated and do not readily allow productive infection. HIV-1 Vpr is highly abundant in progeny virions, and induces signalling and HIV-1 LTR transcription. We hence hypothesized that Vpr might be a determinant of non-activated T-cell infection. Virion-delivered Vpr activated nuclear factor of activated T cells (NFAT) through Ca2+ influx and interference with the NFAT export kinase GSK3β. This leads to NFAT translocation and accumulation within the nucleus and was required for productive infection of unstimulated primary CD4+ T cells. A mutagenesis approach revealed correlation of Vpr-mediated NFAT activation with its ability to enhance LTR transcription and mediate cell cycle arrest. Upon NFAT inhibition, Vpr did not augment resting T-cell infection, and showed reduced G2/M arrest and LTR transactivation. Altogether, Vpr renders unstimulated T cells more permissive for productive HIV-1 infection and stimulates activation of productively infected as well as virus-exposed T cells. Therefore, it could be involved in the establishment and reactivation of HIV-1 from viral reservoirs and might have an impact on the levels of immune activation, which are determinants of HIV-1 pathogenesis. PMID:27383627

  7. The phosphorylation of HIV-1 Gag by atypical protein kinase C facilitates viral infectivity by promoting Vpr incorporation into virions

    PubMed Central

    2014-01-01

    Background Human immunodeficiency virus type 1 (HIV-1) Gag is the main structural protein that mediates the assembly and release of virus-like particles (VLPs) from an infected cell membrane. The Gag C-terminal p6 domain contains short sequence motifs that facilitate virus release from the plasma membrane and mediate incorporation of the viral Vpr protein. Gag p6 has also been found to be phosphorylated during HIV-1 infection and this event may affect virus replication. However, the kinase that directs the phosphorylation of Gag p6 toward virus replication remains to be identified. In our present study, we identified this kinase using a proteomic approach and further delineate its role in HIV-1 replication. Results A proteomic approach was designed to systematically identify human protein kinases that potently interact with HIV-1 Gag and successfully identified 22 candidates. Among this panel, atypical protein kinase C (aPKC) was found to phosphorylate HIV-1 Gag p6. Subsequent LC-MS/MS and immunoblotting analysis with a phospho-specific antibody confirmed both in vitro and in vivo that aPKC phosphorylates HIV-1 Gag at Ser487. Computer-assisted structural modeling and a subsequent cell-based assay revealed that this phosphorylation event is necessary for the interaction between Gag and Vpr and results in the incorporation of Vpr into virions. Moreover, the inhibition of aPKC activity reduced the Vpr levels in virions and impaired HIV-1 infectivity of human primary macrophages. Conclusion Our current results indicate for the first time that HIV-1 Gag phosphorylation on Ser487 is mediated by aPKC and that this kinase may regulate the incorporation of Vpr into HIV-1 virions and thereby supports virus infectivity. Furthermore, aPKC inhibition efficiently suppresses HIV-1 infectivity in macrophages. aPKC may therefore be an intriguing therapeutic target for HIV-1 infection. PMID:24447338

  8. DHX9/RHA Binding to the PBS-Segment of the Genomic RNA during HIV-1 Assembly Bolsters Virion Infectivity.

    PubMed

    Boeras, Ioana; Song, Zhenwei; Moran, Andrew; Franklin, Jarryd; Brown, William Clay; Johnson, Marc; Boris-Lawrie, Kathleen; Heng, Xiao

    2016-06-01

    Cellular RNA-binding proteins incorporated into virions during human immunodeficiency virus type 1 (HIV-1) assembly promote the replication efficiency of progeny virions. Despite its critical role in bolstering virion infectivity, the molecular basis for the incorporation of DHX9/RNA helicase A (RHA) to virions remains unclear. Here, cell-based experiments demonstrate that the truncation of segments of the HIV-1 5'-untranslated region (5'-UTR) distinct from the core encapsidation sequence eliminated virion incorporation of RHA, indicating that RHA recruitment is mediated by specific interactions with the HIV-1 5'-UTR. In agreement with biological data, isothermal titration calorimetry determined that the dimer conformation of the 5'-UTR binds one RHA molecule per RNA strand, and the interaction is independent of nucleocapsid protein binding. NMR spectra employing a deuterium-labeling approach enabled resolution of the dimeric 5'-UTR in complex with the RHA N-terminal domain. The structure of the large molecular mass complex was dependent on RHA binding to a double-stranded region of the primer binding site (PBS)-segment of the 5'-UTR. A single A-to-C substitution was sufficient to disrupt biophysical conformation and attenuate virion infectivity in cell-based assays. Taken together, our studies demonstrate the structural basis for HIV-1 genomic RNA to recruit beneficial cellular cofactor to virions. The support of progeny virion infectivity by RHA is attributable to structure-dependent binding at the PBS-segment of the HIV-1 5'-UTR during virus assembly. PMID:27107641

  9. HIV-1 Nef promotes infection by excluding SERINC5 from virion incorporation

    PubMed Central

    Rosa, Annachiara; Chande, Ajit; Ziglio, Serena; Sanctis, Veronica De; Bertorelli, Roberto; Goh, Shih Lin; McCauley, Sean M.; Nowosielska, Anetta; Antonarakis, Stylianos E.; Luban, Jeremy; Santoni, Federico Andrea; Pizzato, Massimo

    2016-01-01

    HIV-1 Nef, a protein important for the development of AIDS, has well-characterized effects on host membrane trafficking and receptor downregulation. By an unidentified mechanism, Nef increases the intrinsic infectivity of HIV-1 virions in a host-cell-dependent manner. Here we identify the host transmembrane protein SERINC5, and to a lesser extent SERINC3, as a potent inhibitor of HIV-1 particle infectivity that is counteracted by Nef. SERINC5 localizes to the plasma membrane, where it is efficiently incorporated into budding HIV-1 virions and impairs subsequent virion penetration of susceptible target cells. Nef redirects SERINC5 to a Rab7-positive endosomal compartment and thereby excludes it from HIV-1 particles. The ability to counteract SERINC5 was conserved in Nef encoded by diverse primate immunodeficiency viruses, as well as in the structurally unrelated glycosylated Gag from murine leukaemia virus. These examples of functional conservation and convergent evolution emphasize the fundamental importance of SERINC5 as a potent anti-retroviral factor. PMID:26416734

  10. Virions and intracellular nucleocapsids produced during mixed heterotypic influenza infection of MDCK cells

    SciTech Connect

    Sklyanskaya, E.I.; Varich, N.L.; Amvrosieva, T.V.; Kaverin, N.V.

    1985-02-01

    Phenotypically mixed virus yields, obtained by coinfection of MDCK cells with influenza A/WSN/33 and B/Lee/40 viruses, contained both A/WSN/33 and B/Lee/40 NP proteins, as revealed by polyacrylamide gel electrophoresis of the purified /sup 14/C-amino acids-labeled virus. Virions were lysed with 0.6 M KCl-Triton X-100 buffer, and nucleocapsids were immunoprecipitated with antibodies against NP protein of influenza A virus. Polyacrylamide gel electrophoresis of the immunoprecipitate revealed NP protein of A/WSN/33 but not of B/Lee/40 virus. However, in similar experiments with the lysates of doubly infected cells, the band of B/Lee/40 NP protein was revealed in the polyacrylamide gel electrophoresis patterns of the immunoprecipitates. In an attempt to analyze the RNA content of the immune complexes, the authors absorbed the lysates of doubly infected (/sup 3/H)uridine-labeled cells with protein A-containing Staphylococcus aureus covered with antibodies against the NP protein of influenza A virus. RNA extracted from the immune complexes contained genomic RNA segments of both A/WSN/33 and B/Lee/40 viruses. In control samples containing an artificial mixture of cell lysates separately infected with each virus, the analysis revealed homologous components (i.e., A/WSN/33 NP protein or RNA segments) in the immune complexes. The results suggest the presence of phenotypically mixed nucleocapsids in the cells doubly infected with influenza A and B viruses; in the course of the virion formation, the nucleocapsids lacking the heterologous NP protein are selected.

  11. Infection of a Single Cell Line with Distinct Strains of Human Cytomegalovirus Can Result in Large Variations in Virion Production and Facilitate Efficient Screening of Virus Protein Function

    PubMed Central

    Zavala, Anamaria G.; O'Dowd, John M.

    2015-01-01

    ABSTRACT Previously, we reported that the absence of the ataxia telangiectasia mutated (ATM) kinase, a critical DNA damage response (DDR) signaling component for double-strand breaks, caused no change in HCMV Towne virion production. Later, others reported decreased AD169 viral titers in the absence of ATM. To address this discrepancy, human foreskin fibroblasts (HFF) and three ATM− lines (GM02530, GM05823, and GM03395) were infected with both Towne and AD169. Two additional ATM− lines (GM02052 and GM03487) were infected with Towne. Remarkably, both previous studies' results were confirmed. However, the increased number of cell lines and infections with both lab-adapted strains confirmed that ATM was not necessary to produce wild-type-level titers in fibroblasts. Instead, interactions between individual virus strains and the cellular microenvironment of the individual ATM− line determined efficiency of virion production. Surprisingly, these two commonly used lab-adapted strains produced drastically different titers in one ATM− cell line, GM05823. The differences in titer suggested a rapid method for identifying genes involved in differential virion production. In silico comparison of the Towne and AD169 genomes determined a list of 28 probable candidates responsible for the difference. Using serial iterations of an experiment involving virion entry and input genome nuclear trafficking with a panel of related strains, we reduced this list to four (UL129, UL145, UL147, and UL148). As a proof of principle, reintroduction of UL148 largely rescued genome trafficking. Therefore, use of a battery of related strains offers an efficient method to narrow lists of candidate genes affecting various virus life cycle checkpoints. IMPORTANCE Human cytomegalovirus (HCMV) infection of multiple cell lines lacking ataxia telangiectasia mutated (ATM) protein produced wild-type levels of infectious virus. Interactions between virus strains and the microenvironment of individual

  12. Effect of the ionophore monensin on herpes simplex virus type 1-induced cell fusion, glycoprotein synthesis, and virion infectivity.

    PubMed

    Kousoulas, K G; Bzik, D J; Person, S

    1983-01-01

    The ionophore monensin inhibited the formation of mature, fully glycosylated glycoproteins gB, gC, and gD during herpes simplex virus type 1 infection of human embryonic lung cells. Underglycosylated forms, including the apparent high-mannose precursor forms of the major glycoproteins, appeared. Monensin inhibited virus-induced cell fusion. Infectious virions produced in the presence of monensin appeared to contain predominantly underglycosylated glycoproteins. PMID:6307921

  13. Nef Enhances Human Immunodeficiency Virus Type 1 Infectivity Resulting from Intervirion Fusion: Evidence Supporting a Role for Nef at the Virion Envelope

    PubMed Central

    Zhou, Jing; Aiken, Christopher

    2001-01-01

    The human immunodeficiency virus type 1 (HIV-1) accessory protein Nef stimulates viral infectivity by facilitating an early event in the HIV-1 life cycle. Although no structural or biochemical defects in Nef-defective HIV-1 particles have been demonstrated, the Nef protein is incorporated into HIV-1 particles. To localize the function of Nef within the virus particle, we developed a novel technology involving fusion of enveloped donor HIV-1 particles bearing core defects with envelope-defective target virions bearing HIV-1 receptors. Although neither virus alone was capable of infecting CD4+ target cells, the incubation of donor and target virions prior to addition to target cells resulted in infection. This effect, termed “virion transcomplementation,” required a functional Env protein on the donor virus and CD4 and an appropriate coreceptor on target virions. To provide evidence for intervirion fusion as the mechanism of complementation, experiments were performed using dual-enveloped HIV-1 particles bearing both HIV-1 and ecotropic murine leukemia virus (E-MLV) Env proteins as donor virions. Infection of CD4-negative target cells bearing E-MLV receptors was prevented by HIV-1 entry inhibitors when added before, but not after, incubation of donor and target virions prior to the addition to cells. When we used Nef+ and Nef− donor and target virions, Nef enhanced infection when present in donor virions. In contrast, no effect of Nef was detected when present in the target virus. These results reveal a potential mechanism for enhancing HIV-1 diversity in vivo through the rescue of defective viral genomes and provide a novel genetic system for the functional analysis of virion-associated proteins in HIV-1 infection. PMID:11390586

  14. HIV-1 dynamics in vivo: Virion clearance rate, infected cell life-span, and viral generation time

    SciTech Connect

    Perelson, A.S.; Neumann, A.U.; Markowitz, M.; Ho, D.D.; Leonard, J.M.

    1996-03-15

    A new mathematical model was used to analyze a detailed set of human immunodeficiency virus-type 1 (HIV-1) viral load data collected from five infected individuals after the administration of a potent inhibitor of HIV-1 protease. Productively infected cells were estimated to have, on average, a life-span of 2.2 days (half-life t{sub 1/2} = 1.6 days), and plasma virions were estimated to have, on average, a mean life-span of 0.3 days (t{sub 1/2} = 0.24 days). The estimated average total HIV-1 production was 10.3 x 10{sup 9}virions per day, which is substantially greater than previous minimum estimates. The results also suggest that the minimum duration of the HIV-1 life cycle in vivo is 1.2 days on average, and that the average HIV-1 generation time-defined as the time from release of a virion until it infects another cell and causes the release of a new generation of viral particles-is 2.6 days. These findings on viral dynamics provide not only a kinetic picture of HIV-1 pathogenesis, but also theoretical principles to guide the development of treatment strategies. 22 refs., 1 fig., 2 tabs.

  15. Early events in herpes simplex virus type 1 infection: photosensitivity of fluorescein isothiocyanate-treated virions.

    PubMed Central

    DeLuca, N; Bzik, D; Person, S; Snipes, W

    1981-01-01

    Herpes simplex virus type 1 is photosensitized by treatment with fluorescein isothiocyante (FITC). The inactivation of FITC-treated virions upon subsequent exposure to light is inhibited by the presence of sodium azide, suggesting the involvement of singlet oxygen in the process. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis revealed that treatment with FITC plus light induces crosslinks in viral envelope glycoproteins. Treatment of virions with high concentrations of FITC (50 micrograms/ml) plus light causes a reduction in the adsorption of the virus to monolayers of human embryonic lung cells. For lower concentrations of FITC (10 micrograms/ml) plus light, treated virions adsorb to the host cells, but remain sensitive to light until entry occurs. The loss of light sensitivity coincides with the development of resistance to antibodies. These results are most consistent with a mechanism of entry for herpes simplex virus involving fusion of the viral membrane with the plasma membrane of the host cell. Images PMID:6262783

  16. Early events in herpes simplex virus type 1 infection: photosensitivity of fluorescein isothiocyanate-treated virions.

    PubMed

    DeLuca, N; Bzik, D; Person, S; Snipes, W

    1981-02-01

    Herpes simplex virus type 1 is photosensitized by treatment with fluorescein isothiocyante (FITC). The inactivation of FITC-treated virions upon subsequent exposure to light is inhibited by the presence of sodium azide, suggesting the involvement of singlet oxygen in the process. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis revealed that treatment with FITC plus light induces crosslinks in viral envelope glycoproteins. Treatment of virions with high concentrations of FITC (50 micrograms/ml) plus light causes a reduction in the adsorption of the virus to monolayers of human embryonic lung cells. For lower concentrations of FITC (10 micrograms/ml) plus light, treated virions adsorb to the host cells, but remain sensitive to light until entry occurs. The loss of light sensitivity coincides with the development of resistance to antibodies. These results are most consistent with a mechanism of entry for herpes simplex virus involving fusion of the viral membrane with the plasma membrane of the host cell. PMID:6262783

  17. Early events in herpes simplex virus type 1 infection: photosensitivity of fluorescein isothiocyanate-treated virions

    SciTech Connect

    DeLuca, N.; Bzik, D.; Person, S.; Snipes, W.

    1981-02-01

    Herpes simplex virus type 1 is photosensitized by treatment with fluorescein isothiocyanate (FITC). The inactivation of FITC-treated virions upon subsequent exposure to light is inhibited by the presence of sodium azide, suggesting the involvement of singlet oxygen in the process. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis revealed that treatment with FITC plus light induces crosslinks in viral envelope glycoproteins. Treatment of virions with high concentrations of FITC (50 ..mu..g/ml) plus light causes a reduction in the adsorption of the virus to monolayers of human embryonic lung cells. For lower concentrations of FITC (10 ..mu..g/ml) plus light, treated virions adsorb to the host cells, but remain sensitive to light until entry occurs. The loss of light sensitivity coincides with the development of resistance to antibodies. These results are most consistent with a mechanism of entry for herpes simplex virus involving fusion of the viral membrane with the plasma membrane of the host cell.

  18. Mutations in the PPPY Motif of Vesicular Stomatitis Virus Matrix Protein Reduce Virus Budding by Inhibiting a Late Step in Virion Release

    PubMed Central

    Jayakar, Himangi R.; Murti, K. Gopal; Whitt, Michael A.

    2000-01-01

    The N terminus of the matrix (M) protein of vesicular stomatitis virus (VSV) and of other rhabdoviruses contains a highly conserved PPPY sequence (or PY motif) similar to the late (L) domains in the Gag proteins of some retroviruses. These L domains in retroviral Gag proteins are required for efficient release of virus particles. In this report, we show that mutations in the PPPY sequence of the VSV M protein reduce virus yield by blocking a late stage in virus budding. We also observed a delay in the ability of mutant viruses to cause inhibition of host gene expression compared to wild-type (WT) VSV. The effect of PY mutations on virus budding appears to be due to a block at a stage just prior to virion release, since electron microscopic examination of PPPA mutant-infected cells showed a large number of assembled virions at the plasma membrane trapped in the process of budding. Deletion of the glycoprotein (G) in addition to these mutations further reduced the virus yield to less than 1% of WT levels, and very few particles were assembled at the cell surface. This observation suggested that G protein aids in the initial stage of budding, presumably during the formation of the bud site. Overall, our results confirm that the PPPY sequence of the VSV M protein possesses L domain activity analogous to that of the retroviral Gag proteins. PMID:11024108

  19. Estimating the fraction of progeny virions that must incorporate APOBEC3G for suppression of productive HIV-1 infection

    SciTech Connect

    Thangavelu, Pulari U.; Gupta, Vipul; Dixit, Narendra M.

    2014-01-20

    The contest between the host factor APOBEC3G (A3G) and the HIV-1 protein Vif presents an attractive target of intervention. The extent to which the A3G–Vif interaction must be suppressed to tilt the balance in favor of A3G remains unknown. We employed stochastic simulations and mathematical modeling of the within-host dynamics and evolution of HIV-1 to estimate the fraction of progeny virions that must incorporate A3G to render productive infection unsustainable. Using three different approaches, we found consistently that a transition from sustained infection to suppression of productive infection occurred when the latter fraction exceeded ∼0.8. The transition was triggered by A3G-induced hypermutations that led to premature stop codons compromising viral production and was consistent with driving the basic reproductive number, R{sub 0}, below unity. The fraction identified may serve as a quantitative guideline for strategies targeting the A3G–Vif axis. - Highlights: • We perform simulations and mathematical modeling of the role of APOBEC3G in suppressing HIV-1 infection. • In three distinct ways, we estimate that when over 80% of progeny virions carry APOBEC3G, productive HIV-1 infection would be suppressed. • Our estimate of this critical fraction presents quantitative guidelines for strategies targeting the APOBEC3G–Vif axis.

  20. BST2/CD317 counteracts human coronavirus 229E productive infection by tethering virions at the cell surface

    SciTech Connect

    Wang, Shiu-Mei; Huang, Kuo-Jung; Wang, Chin-Tien

    2014-01-20

    Bone marrow stromal antigen 2 (BST2), an interferon-inducible antiviral factor, has been shown to block the release of various enveloped viruses from cells. It has also been identified as an innate immune system component. Most enveloped viruses subject to BST2 restriction bud at the plasma membrane. Here we report our findings that (a) the production of human coronavirus 229E (HCoV-229E) progeny viruses, whose budding occurs at the ER-Golgi intermediate compartment (ERGIC), markedly decreases in the presence of BST2; and (b) BST2 knockdown expression results in enhanced HCoV-229E virion production. Electron microscopy analyses indicate that HCoV-229E virions are tethered to cell surfaces or intracellular membranes by BST2. Our results suggest that BST2 exerts a broad blocking effect against enveloped virus release, regardless of whether budding occurs at the plasma membrane or intracellular compartments. - Highlights: • BST2 knockdown expression results in enhanced HCoV-229E egress. • HCoV-229E virions are tethered to cell surfaces or intracellular membranes by BST2. • HCoV-229E infection at high MOI can significantly downregulate HeLa BST2 and rescue HIV-1 egress.

  1. The virion N protein of infectious bronchitis virus is more phosphorylated than the N protein from infected cell lysates

    SciTech Connect

    Jayaram, Jyothi; Youn, Soonjeon; Collisson, Ellen W. . E-mail: ecollisson@cvm.tamu.edu

    2005-08-15

    Because phosphorylation of the infectious bronchitis virus (IBV) nucleocapsid protein (N) may regulate its multiple roles in viral replication, the dynamics of N phosphorylation were examined. {sup 32}P-orthophosphate labeling and Western blot analyses confirmed that N was the only viral protein that was phosphorylated. Pulse labeling with {sup 32}P-orthophosphate indicated that the IBV N protein was phosphorylated in the virion, as well as at all times during infection in either chicken embryo kidney cells or Vero cells. Pulse-chase analyses followed by immunoprecipitation of IBV N proteins using rabbit anti-IBV N polyclonal antibody demonstrated that the phosphate on the N protein was stable for at least 1 h. Simultaneous labeling with {sup 32}P-orthophosphate and {sup 3}H-leucine identified a 3.5-fold increase in the {sup 32}P:{sup 3}H counts per minute (cpm) ratio of N in the virion as compared to the {sup 32}P:{sup 3}H cpm ratio of N in the cell lysates from chicken embryo kidney cells, whereas in Vero cells the {sup 32}P:{sup 3}H cpm ratio of N from the virion was 10.5-fold greater than the {sup 32}P:{sup 3}H cpm ratio of N from the cell lysates. These studies are consistent with the phosphorylation of the IBV N playing a role in assembly or maturation of the viral particle.

  2. Murine Cytomegalovirus Virion-Associated Protein M45 Mediates Rapid NF-κB Activation after Infection

    PubMed Central

    Krause, Eva; de Graaf, Miranda; Fliss, Patricia M.; Dölken, Lars

    2014-01-01

    ABSTRACT Murine cytomegalovirus (MCMV) rapidly induces activation of nuclear factor κB (NF-κB) upon infection of host cells. After a transient phase of activation, the MCMV M45 protein blocks all canonical NF-κB-activating pathways by inducing the degradation of the gamma subunit of the inhibitor of κB kinase complex (IKKγ; commonly referred to as the NF-κB essential modulator [NEMO]). Here we show that the viral M45 protein also mediates rapid NF-κB activation immediately after infection. MCMV mutants lacking M45 or expressing C-terminally truncated M45 proteins induced neither NF-κB activation nor transcription of NF-κB-dependent genes within the first 3 h of infection. Rapid NF-κB activation was absent in MCMV-infected NEMO-deficient fibroblasts, indicating that activation occurs at or upstream of the IKK complex. NF-κB activation was strongly reduced in murine fibroblasts lacking receptor-interacting protein 1 (RIP1), a known M45-interacting protein, but was restored upon complementation with murine RIP1. However, the ability of M45 to interact with RIP1 and NEMO was not sufficient to induce NF-κB activation upon infection. In addition, incorporation of the M45 protein into virions was required. This was dependent on a C-terminal region of M45, which is not required for interaction with RIP1 and NEMO. We propose a model in which M45 delivered by viral particles activates NF-κB, presumably involving an interaction with RIP1 and NEMO. Later in infection, expression of M45 induces the degradation of NEMO and the shutdown of canonical NF-κB activation. IMPORTANCE Transcription factor NF-κB is an important regulator of innate and adaptive immunity. Its activation can be beneficial or detrimental for viral pathogens. Therefore, many viruses interfere with NF-κB signaling by stimulating or inhibiting the activation of this transcription factor. Cytomegaloviruses, opportunistic pathogens that cause lifelong infections in their hosts, activate NF

  3. Identification of regions of the Beet mild curly top virus (family Geminiviridae) capsid protein involved in systemic infection, virion formation and leafhopper transmission.

    PubMed

    Soto, Maria J; Chen, Li-Fang; Seo, Young-Su; Gilbertson, Robert L

    2005-10-25

    Plant viruses in the genus Curtovirus (family Geminiviridae) are vectored by the beet leafhopper (Circulifer tenellus) and cause curly top disease in a wide range of dicotyledonous plants. An infectious clone of an isolate of Beet mild curly top virus (BMCTV-[W4]), associated with an outbreak of curly top in pepper and tomato crops, was characterized and used to investigate the role of the capsid protein (CP) in viral biology and pathogenesis. Frameshift mutations were introduced into the overlapping CP and V2 genes, and a series of CP alanine scanning mutations were generated. All mutants replicated in tobacco protoplasts or systemically infected plants, consistent with these gene products not being required for viral DNA replication. The CP frameshift mutant and most C-terminal alanine scanning mutants did not systemically infect Nicotiana benthamiana plants or form detectable virions, and were not leafhopper-transmitted. In contrast, most N-terminal alanine scanning mutants systemically infected N. benthamiana and induced disease symptoms, formed virions and were leafhopper-transmissible; thus, these substitution mutations did not significantly alter the functional properties of this region. One N-terminal mutant (CP49-51) systemically infected N. benthamiana, but did not form detectable virions; whereas another (CP25-28) systemically infected N. benthamiana and formed virions, but was not insect-transmissible. These mutants may reveal regions involved in virus movement through the plant and/or leafhopper vector. Together, these results indicate an important role for virions in systemic infection (long-distance movement) and insect transmission, and strongly suggest that virions are the form in which BMCTV moves, long distance, in the phloem. PMID:16085227

  4. Use of RT-Defective HIV Virions: New Tool to Evaluate Specific Response in Chronic Asymptomatic HIV-Infected Individuals

    PubMed Central

    Arberas, Hodei; García-Pérez, Javier; García, Felipe; Bargalló, Manuel Enric; Maleno, María José; Gatell, José María; Mothe, Beatriz; Alcami, José; Sánchez-Palomino, Sonsoles; Plana, Montserrat

    2013-01-01

    Background Generation of new reagents that can be used to screen or monitor HIV-1-specific responses constituted an interesting field in the development of HIV vaccines to improve their efficacy. Methods We have evaluated the specific T cell response against different types of NL4-3 virions (including NL4-3 aldrithiol-2 treated, NL4-3/ΔRT and R5 envelopes: NL4-3/ΔRT/ΔEnv[AC10] and NL4-3/ΔRT/ΔEnv[Bal]) and against pools of overlapping peptides (15 mer) encompassing the HIV-1 Gag and Nef regions. Cryopreserved PBMC from a subset of 69 chronic asymptomatic HIV positive individuals have been employed using different techniques including IFN-γ ELISPOT assay, surface activation markers and intracellular cytokine staining (ICS) by flow cytometry. Results The differential response obtained against NL4-3 aldrithiol-2 treated and NL4-3/ΔRT virions (25% vs 55%, respectively) allow us to divide the population in three groups: “full-responders” (positive response against both viral particles), “partial-responders” (positive response only against NL4-3/ΔRT virions) and “non-responders” (negative responses). There was no difference between X4 and R5 envelopes. The magnitude of the total responses was higher against NL4-3/ΔRT and was positively correlated with gender and inverse correlated with viral load. On the contrary CD4+ T cell count was not associated with this response. In any case responses to the viruses tended to be lower in magnitude than those detected by the overlapping peptides tested. Finally we have found an increased frequency of HLA-B27 allele (23% vs 9%) and a significant reduction in some activation markers (CD69 and CD38) on T cells surface in responders vs non-responders individuals. Conclusions In summary these virions could be considered as alternative and useful reagents for screening HIV-1-specific T cell responses in HIV exposed uninfected people, HIV infected patients and to assess immunogenicity of new prototypes both in vitro and

  5. Differential segregation of nodaviral coat protein and RNA into progeny virions during mixed infection with FHV and NoV

    PubMed Central

    Gopal, Radhika; Venter, P. Arno; Schneemann, Anette

    2014-01-01

    Nodaviruses are icosahedral viruses with a bipartite, positive-sense RNA genome. The two RNAs are packaged into a single virion by a poorly understood mechanism. We chose two distantly related nodaviruses, Flock House virus and Nodamura virus, to explore formation of viral reassortants as a means to further understand genome recognition and encapsidation. In mixed infections, the viruses were incompatible at the level of RNA replication and their coat proteins segregated into separate populations of progeny particles. RNA packaging, on the other hand, was indiscriminate as all four viral RNAs were detectable in each progeny population. Consistent with the trans-encapsidation phenotype, fluorescence in situ hybridization of viral RNA revealed that the genomes of the two viruses co-localized throughout the cytoplasm. Our results imply that nodaviral RNAs lack rigorously defined packaging signals and that coencapsidation of the viral RNAs does not require a pair of cognate RNA1 and RNA2. PMID:24725955

  6. The use of Nanotrap particles technology in capturing HIV-1 virions and viral proteins from infected cells.

    PubMed

    Jaworski, Elizabeth; Saifuddin, Mohammed; Sampey, Gavin; Shafagati, Nazly; Van Duyne, Rachel; Iordanskiy, Sergey; Kehn-Hall, Kylene; Liotta, Lance; Petricoin, Emanuel; Young, Mary; Lepene, Benjamin; Kashanchi, Fatah

    2014-01-01

    HIV-1 infection results in a chronic but incurable illness since long-term HAART can keep the virus to an undetectable level. However, discontinuation of therapy rapidly increases viral burden. Moreover, patients under HAART frequently develop various metabolic disorders and HIV-associated neuronal disease. Today, the main challenge of HIV-1 research is the elimination of the residual virus in infected individuals. The current HIV-1 diagnostics are largely comprised of serological and nucleic acid based technologies. Our goal is to integrate the nanotrap technology into a standard research tool that will allow sensitive detection of HIV-1 infection. This study demonstrates that majority of HIV-1 virions in culture supernatants and Tat/Nef proteins spiked in culture medium can be captured by nanotrap particles. To determine the binding affinities of different baits, we incubated target molecules with nanotrap particles at room temperature. After short sequestration, materials were either eluted or remained attached to nanotrap particles prior to analysis. The unique affinity baits of nanotrap particles preferentially bound HIV-1 materials while excluded albumin. A high level capture of Tat or Tat peptide by NT082 and NT084 particles was measured by western blot (WB). Intracellular Nef protein was captured by NT080, while membrane-associated Nef was captured by NT086 and also detected by WB. Selective capture of HIV-1 particles by NT073 and NT086 was measured by reverse transcriptase assay, while capture of infectious HIV-1 by these nanoparticles was demonstrated by functional transactivation in TZM-bl cells. We also demonstrated specific capture of HIV-1 particles and exosomes-containing TAR-RNA in patients' serum by NT086 and NT082 particles, respectively, using specific qRT-PCR. Collectively, our data indicate that certain types of nanotrap particles selectively capture specific HIV-1 molecules, and we propose to use this technology as a platform to enhance HIV-1

  7. The Use of Nanotrap Particles Technology in Capturing HIV-1 Virions and Viral Proteins from Infected Cells

    PubMed Central

    Sampey, Gavin; Shafagati, Nazly; Van Duyne, Rachel; Iordanskiy, Sergey; Kehn-Hall, Kylene; Liotta, Lance; Petricoin, Emanuel; Young, Mary; Lepene, Benjamin; Kashanchi, Fatah

    2014-01-01

    HIV-1 infection results in a chronic but incurable illness since long-term HAART can keep the virus to an undetectable level. However, discontinuation of therapy rapidly increases viral burden. Moreover, patients under HAART frequently develop various metabolic disorders and HIV-associated neuronal disease. Today, the main challenge of HIV-1 research is the elimination of the residual virus in infected individuals. The current HIV-1 diagnostics are largely comprised of serological and nucleic acid based technologies. Our goal is to integrate the nanotrap technology into a standard research tool that will allow sensitive detection of HIV-1 infection. This study demonstrates that majority of HIV-1 virions in culture supernatants and Tat/Nef proteins spiked in culture medium can be captured by nanotrap particles. To determine the binding affinities of different baits, we incubated target molecules with nanotrap particles at room temperature. After short sequestration, materials were either eluted or remained attached to nanotrap particles prior to analysis. The unique affinity baits of nanotrap particles preferentially bound HIV-1 materials while excluded albumin. A high level capture of Tat or Tat peptide by NT082 and NT084 particles was measured by western blot (WB). Intracellular Nef protein was captured by NT080, while membrane-associated Nef was captured by NT086 and also detected by WB. Selective capture of HIV-1 particles by NT073 and NT086 was measured by reverse transcriptase assay, while capture of infectious HIV-1 by these nanoparticles was demonstrated by functional transactivation in TZM-bl cells. We also demonstrated specific capture of HIV-1 particles and exosomes-containing TAR-RNA in patients' serum by NT086 and NT082 particles, respectively, using specific qRT-PCR. Collectively, our data indicate that certain types of nanotrap particles selectively capture specific HIV-1 molecules, and we propose to use this technology as a platform to enhance HIV-1

  8. A conserved carboxy-terminal domain in the major tegument structural protein VP22 facilitates virion packaging of a chimeric protein during productive herpes simplex virus 1 infection

    SciTech Connect

    Schlegel, Elisabeth F.M.; Blaho, John A.

    2009-05-10

    Recombinant virus HSV-1(RF177) was previously generated to examine tegument protein VP22 function by inserting the GFP gene into the gene encoding VP22. During a detailed analysis of this virus, we discovered that RF177 produces a novel fusion protein between the last 15 amino acids of VP22 and GFP, termed GCT-VP22. Thus, the VP22 carboxy-terminal specific antibody 22-3 and two anti-GFP antibodies reacted with an approximately 28 kDa protein from RF177-infected Vero cells. GCT-VP22 was detected at 1 and 3 hpi. Examination of purified virions indicated that GCT-VP22 was incorporated into RF177 virus particles. These observations imply that at least a portion of the information required for virion targeting is located in this domain of VP22. Indirect immunofluorescence analyses showed that GCT-VP22 also localized to areas of marginalized chromatin during RF177 infection. These results indicate that the last fifteen amino acids of VP22 participate in virion targeting during HSV-1 infection.

  9. Reducing haemodialysis access infection rates.

    PubMed

    Dorman, Amanda; Dainton, Marissa

    Infections are the second most common cause of vascular access loss in the long-term haemodialysis patient, and recent years have seen an increase in healthcare-associated infections (HCAIs) associated with vascular access (Suhail, 2009). There have been a number of drivers including publication guidelines (Department of Health, 2006; 2007) and local protocols providing evidence-based recommendations that, when implemented, can reduce the risk of these infections. In England, the selection of bloodstream infections caused by methicillin resistant staphylococcus aureus (MRSA) as a significant clinical outcome has led to a vast amount of work in this area. Root cause analysis of individual infections (by the clinical teams when these occur) in many specialities identified areas where practice could be improved, including practice relating to vascular access within the renal setting. Manufacturers have also supported this work by focusing on developing products that are designed to reduce the likelihood of infections occurring. One product identified and used within the NHS is Chloraprep. PMID:21646994

  10. The vaccinia virus E8R gene product is required for formation of transcriptionally active virions.

    PubMed

    Kato, Sayuri E M; Condit, Richard C; Moussatché, Nissin

    2007-10-25

    Two vaccinia virus temperature-sensitive mutants were mapped to the E8R gene and subjected to phenotypic characterization. Dts23 contains a missense mutation in the coding region of E8R (L81F), and in Cts19 the initiating methionine codon is changed from ATG to ATA (M1I). The two ts mutants display normal patterns of gene expression and DNA replication during infection. The E8 protein is synthesized exclusively late during infection and packaged into virion cores Western blot analysis revealed that E8 synthesis is reduced in Dts23 infected cells at permissive (31 degrees C) and non-permissive temperature (39.7 degrees C) and absent in Cts19 infection under both conditions. Dts23 virions produced at 39.7 degrees C were indistinguishable in appearance from wt virions. Cts19 fails to produce identifiable viral structures when incubated at 39.7 degrees C. Purified Dts23 virions produced at 39.7 degrees C contain reduced amounts of E8 and have a high particle to infectivity ratio; purified Cts19 virions grown at 31 degrees C also show reduced infectivity and do not contain detectable E8. Dts23 grown at 39.7 degrees C could enter cells but failed to synthesize early mRNA or produce CPE. Soluble extracts from mutant virions were active in a promoter dependent in vitro transcription assay, however intact mutant cores were defective in transcription. We suggest that E8 plays a subtle role in virion core structure that impacts directly or indirectly on core transcription. PMID:17619043

  11. HIV-1 infection of T cells and macrophages are differentially modulated by virion-associated Hck: a Nef-dependent phenomenon.

    PubMed

    Cornall, Alyssa; Mak, Johnson; Greenway, Alison; Tachedjian, Gilda

    2013-09-01

    The proline repeat motif (PxxP) of Nef is required for interaction with the SH3 domains of macrophage-specific Src kinase Hck. However, the implication of this interaction for viral replication and infectivity in macrophages and T lymphocytes remains unclear. Experiments in HIV-1 infected macrophages confirmed the presence of a Nef:Hck complex which was dependent on the Nef proline repeat motif. The proline repeat motif of Nef also enhanced both HIV-1 infection and replication in macrophages, and was required for incorporation of Hck into viral particles. Unexpectedly, wild-type Hck inhibited infection of macrophages, but Hck was shown to enhance infection of primary T lymphocytes. These results indicate that the interaction between Nef and Hck is important for Nef-dependent modulation of viral infectivity. Hck-dependent enhancement of HIV-1 infection of T cells suggests that Nef-Hck interaction may contribute to the spread of HIV-1 infection from macrophages to T cells by modulating events in the producer cell, virion and target cell. PMID:24051604

  12. The Interferon-Inducible Protein Tetherin Inhibits Hepatitis B Virus Virion Secretion

    PubMed Central

    Yan, Ran; Zhao, Xuesen; Cai, Dawei; Liu, Yuanjie; Block, Timothy M.; Guo, Ju-Tao

    2015-01-01

    ABSTRACT Interferon alpha (IFN-α) is an approved medication for chronic hepatitis B therapy. Besides acting as an immunomodulator, IFN-α elicits a pleiotropic antiviral state in hepatitis B virus (HBV)-infected hepatocytes, but whether or not IFN-α impedes the late steps of the HBV life cycle, such as HBV secretion, remains elusive. Here we report that IFN-α treatment of HepAD38 cells with established HBV replication selectively reduced HBV virion release without altering intracellular viral replication or the secretion of HBV subviral particles and nonenveloped capsids. In search of the interferon-stimulated gene(s) that is responsible for the reduction of HBV virion release, we found that tetherin, a broad-spectrum antiviral transmembrane protein that inhibits the egress of a variety of enveloped viruses, was highly induced by IFN-α in HepAD38 cells and in primary human hepatocytes. We further demonstrated that the expression of full-length tetherin, but not the C-terminal glycosylphosphatidylinositol (GPI) anchor-truncated form, inhibited HBV virion egress from HepAD38 cells. In addition, GPI anchor-truncated tetherin exhibited a dominant-negative effect and was incorporated into the liberated virions. We also found colocalization of tetherin and HBV L protein at the intracellular multivesicular body, where the budding of HBV virions takes place. In line with this, electron microscopy demonstrated that HBV virions were tethered in the lumen of the cisterna membrane under tetherin expression. Finally, knockdown of tetherin or overexpression of dominant negative tetherin attenuated the IFN-α-mediated reduction of HBV virion release. Taken together, our study suggests that IFN-α inhibits HBV virion egress from hepatocytes through the induction of tetherin. IMPORTANCE Tetherin is a host restriction factor that blocks the egress of a variety of enveloped viruses through tethering the budding virions on the cell surface with its membrane anchor domains. Here we

  13. Baculovirus virions displaying infectious bursal disease virus VP2 protein protect chickens against infectious bursal disease virus infection.

    PubMed

    Xu, Xin-Gang; Tong, De-Wen; Wang, Zhi Sheng; Zhang, Qi; Li, Zhao-Cai; Zhang, Kuan; Li, Wei; Liu, Hung-Jen

    2011-06-01

    Infectious bursal disease (IBD) is an acute and contagious viral infection of young chickens caused by IBD virus (IBDV). The VP2 protein of IBDV is the only antigen for inducing neutralizing antibodies and protective immunity in the natural host. In the current study, we have succeeded in construction of one recombinant baculovirus BacSC-VP2 expressing His6-tagged VP2 with the baculovirus envelope protein gp64 transmembrane domain (TM) and cytoplasmic domain (CTD). The His6-tagged recombinant VP2 was expressed and anchored on the plasma membrane of Sf-9 cells, as examined by western blot and confocal microscopy. Immunogold electron microscopy demonstrated that the VP2 protein of IBDV was successfully displayed on the viral surface. Vaccination of chickens with the VP2-pseudotyped baculovirus vaccine (BacSC-VP2) elicited significantly higher levels of VP2-specific enzyme-linked immunosorbent assay antibodies and neutralizing antibodies than the control groups. IBDV-specific proliferation of lymphocytes was observed in chickens immunized with the recombinant BacSC-VP2. An in vivo challenge study of the recombinant baculovirus BacSC-VP2 showed effective protection against a very virulent (vv) IBDV infection in chickens. In addition, mortality and gross and histopathological findings in the bursa demonstrated the efficacy of the vaccine in reducing virulence of the disease. These results indicate that the recombinant baculovirus BacSC-VP2 can be a potential vaccine against IBDV infections. PMID:21793437

  14. Cleavage of the HPV16 Minor Capsid Protein L2 during Virion Morphogenesis Ablates the Requirement for Cellular Furin during De Novo Infection

    PubMed Central

    Cruz, Linda; Biryukov, Jennifer; Conway, Michael J.; Meyers, Craig

    2015-01-01

    Infections by high-risk human papillomaviruses (HPV) are the causative agents for the development of cervical cancer. As with other non-enveloped viruses, HPVs are taken up by the cell through endocytosis following primary attachment to the host cell. Through studies using recombinant pseudovirus particles (PsV), many host cellular proteins have been implicated in the process. The proprotein convertase furin has been demonstrated to cleave the minor capsid protein, L2, post-attachment to host cells and is required for infectious entry by HPV16 PsV. In contrast, using biochemical inhibition by a furin inhibitor and furin-negative cells, we show that tissue-derived HPV16 native virus (NV) initiates infection independent of cellular furin. We show that HPV16 L2 is cleaved during virion morphogenesis in differentiated tissue. In addition, HPV45 is also not dependent on cellular furin, but two other alpha papillomaviruses, HPV18 and HPV31, are dependent on the activity of cellular furin for infection. PMID:26569287

  15. Deficient incorporation of spike protein into virions contributes to the lack of infectivity following establishment of a persistent, non-productive infection in oligodendroglial cell culture by murine coronavirus

    SciTech Connect

    Liu Yin; Herbst, Werner; Cao Jianzhong; Zhang Xuming

    2011-01-05

    Infection of mouse oligodendrocytes with a recombinant mouse hepatitis virus (MHV) expressing a green fluorescence protein facilitated specific selection of virus-infected cells and subsequent establishment of persistence. Interestingly, while viral genomic RNAs persisted in infected cells over 14 subsequent passages with concomitant synthesis of viral subgenomic mRNAs and structural proteins, no infectious virus was isolated beyond passage 2. Further biochemical and electron microscopic analyses revealed that virions, while assembled, contained little spike in the envelope, indicating that lack of infectivity during persistence was likely due to deficiency in spike incorporation. This type of non-lytic, non-productive persistence in oligodendrocytes is unique among animal viruses and resembles MHV persistence previously observed in the mouse central nervous system. Thus, establishment of such a culture system that can recapitulate the in vivo phenomenon will provide a powerful approach for elucidating the mechanisms of coronavirus persistence in glial cells at the cellular and molecular levels.

  16. [Research Advances in Baculovirus Occlusion-derived Virions].

    PubMed

    Zhu, Shimao; Li, Hui

    2016-01-01

    Baculoviruses are a family of arthropod-specific viruses that produce two morphologically distinct types of virions (budded and occlusion-derived) in their lifecycle. Baculoviruses establish infection in the midgut of their host via the oral route: occlusion-derived virions have pivotal roles in these processes. This review summarizes the basic characteristics of baculoviruses, and discusses the composition and classification of baculovirus occlusion-derived virions. The latter focuses mainly on the evolution and role of multiple occlusion-derived virions in the lifecycle of baculoviruses. These achievements should aid understanding the evolution and infection mechanisms of baculoviruses. PMID:27295890

  17. Detection of L1, infectious virions and anti-L1 antibody in domestic rabbits infected with cottontail rabbit papillomavirus.

    PubMed

    Hu, Jiafen; Budgeon, Lynn R; Cladel, Nancy M; Culp, Timothy D; Balogh, Karla K; Christensen, Neil D

    2007-12-01

    Shope papillomavirus or cottontail rabbit papillomavirus (CRPV) is one of the first small DNA tumour viruses to be characterized. Although the natural host for CRPV is the cottontail rabbit (Sylvilagus floridanus), CRPV can infect domestic laboratory rabbits (Oryctolagus cuniculus) and induce tumour outgrowth and cancer development. In previous studies, investigators attempted to passage CRPV in domestic rabbits, but achieved very limited success, leading to the suggestion that CRPV infection in domestic rabbits was abortive. The persistence of specific anti-L1 antibody in sera from rabbits infected with either virus or viral DNA led us to revisit the questions as to whether L1 and infectious CRPV can be produced in domestic rabbit tissues. We detected various levels of L1 protein in most papillomas from CRPV-infected rabbits using recently developed monoclonal antibodies. Sensitive in vitro infectivity assays additionally confirmed that extracts from these papillomas were infectious. These studies demonstrated that the CRPV/New Zealand White rabbit model could be used as an in vivo model to study natural virus infection and viral life cycle of CRPV and not be limited to studies on abortive infections. PMID:18024897

  18. Reducing urinary tract infections in catheterised patients.

    PubMed

    Howe, Pam; Adams, John

    2015-01-20

    Urinary tract infections in catheterised patients continue to present a challenge in reducing healthcare-associated infection. In this article, an infection prevention and control team in one NHS trust reports on using audit results to focus attention on measures to reduce bacterial infections. Educational initiatives have an important role in reducing infection, but there is no single solution to the problem. Practice can be improved using a multi-targeted approach, peer review and clinical audit to allow for shared learning and experiences. These, along with informal education in the clinical area and more formal classroom lectures, can ultimately lead to improved patient outcomes. PMID:25585767

  19. Modulation of virion incorporation of Ebolavirus glycoprotein: effects on attachment, cellular entry and neutralization.

    PubMed

    Marzi, Andrea; Wegele, Anja; Pöhlmann, Stefan

    2006-09-01

    The filoviruses Ebolavirus (EBOV) and Marburgvirus (MARV) cause severe hemorrhagic fever in humans and are potential agents of biological warfare. The envelope glycoprotein (GP) of filoviruses mediates viral entry into cells and is an attractive target for therapeutic intervention and vaccine design. Here, we asked if the efficiency of virion incorporation of EBOV-GP impacts attachment and entry into target cells and modulates susceptibility to neutralizing antibodies. In order to control the level of EBOV-GP expression, we generated cell lines expressing the GPs of the four known EBOV subspecies in an inducible fashion. Regulated expression of GP on the cell surface allowed production of reporter viruses harboring different amounts of GP. A pronounced reduction of virion incorporation of EBOV-GP had relatively little effect on virion infectivity, suggesting that only a few copies of GP might be sufficient for efficient engagement of cellular receptors. In contrast, optimal interactions with cellular attachment factors like the DC-SIGN protein required incorporation of high amounts of GP. Antibody-mediated neutralization of virions bearing high amounts of GP was slightly more efficient than neutralization of virions harboring low amounts of GP, suggesting that the efficiency of GP incorporation into virions might modulate susceptibility to neutralizing antibodies. Finally, regulated expression of GP in permissive 293 cells did not reduce EBOV-GP-driven infection but diminished vesicular stomatitis virus GP (VSV-G) and amphotropic murine leukemia virus (A-MLV) GP mediated entry in a dose-dependent manner. Therefore, intracellular GP does not seem to downmodulate expression of its receptor(s) but might alter expression and/or function of molecules involved in VSV-G and A-MLV-GP-dependent entry. Our results suggest that the efficiency of virion incorporation of GP could impact EBOV attachment to target cells and might modulate control of viral spread by the humoral

  20. Antibodies of symptomatic human immunodeficiency virus type 1-infected individuals are directed to the V3 domain of noninfectious and not of infectious virions present in autologous serum.

    PubMed Central

    Schreiber, M; Petersen, H; Wachsmuth, C; Müller, H; Hufert, F T; Schmitz, H

    1994-01-01

    The present study was designed to determine the antibody specificity for the human immunodeficiency virus type 1 (HIV-1) V3 domains of infectious and noninfectious virions present in the serum of AIDS patients. To accomplish this, HIV-1 was isolated in the presence of autologous antibodies from the serum samples of six AIDS patients in HIV-1-negative donor peripheral blood mononuclear cells by short-term cultivation. The isolated virus, defined as the infectious cell-free virus (iCFV), was characterized by sequence analysis of the proviral DNA coding for the third hypervariable (V3) region of the external glycoprotein gp120. This was carried out by amplifying and cloning the V3 region. In all six cases studied, 20 randomly selected V3 clones derived from the proviral DNA of the iCFV, 20 clones from patient cell-free virus, and 20 clones from cell-integrated virus were sequenced to study the distribution and frequency of the intrapatient virus population. The number of major virus variants in the six patients ranged from three to nine. The various V3 sequences found in the AIDS patients showed the typical amino acid pattern of the syncytium-inducing and non-syncytium-inducing viral phenotypes characteristic for the late stage of infection. However, only one patient-specific iCFV variant was detected within the 20 V3 clones analyzed per virus isolation. For the six patients a total of 34 V3-loop variants, either iCFV or non-iCFV, was observed. All 34 V3-loop sequences were expressed as glutathione-S-transferase fusion proteins (V3-GST). The autologous antibody response to the V3-GST fusion proteins was studied by Western immunoblot analysis. A strong antibody response to almost all non-iCFV V3-GST proteins was found in the sera of the six patients. In contrast, the autologous antibody response to the six iCFV V3 loops was undetectable (in four patients) or very faint (in two patients) compared with that to the non-iCFV V3 loops. Five of the six iCFV loops showed

  1. Transcriptional analysis of the virion-sense genes of the geminivirus beet curly top virus.

    PubMed

    Frischmuth, S; Frischmuth, T; Latham, J R; Stanley, J

    1993-11-01

    The genome of the geminivirus beet curly top virus (BCTV) consists of a single circular DNA containing overlapping open reading frames (ORFs) located on both the virion-sense and complementary-sense DNA strands. To investigate the expression of these ORFs, RNA extracted from infected Nicotiana benthamiana and Beta vulgaris has been examined for the presence of viral transcripts. An abundant 1.1-kb virion-sense polyadenylated RNA and four complementary-sense polyadenylated RNAs of 1.7, 1.5, 1.3, and 0.7 kb have been identified by northern blot hybridization, confirming the bidirectional transcription strategy implied by the arrangement of ORFs. We previously demonstrated that two overlapping virion-sense ORFs are involved in coat protein synthesis (ORF V1) and viral single-stranded DNA accumulation (ORF V2). Mutants of a third virion-sense ORF (ORF V3), located upstream and overlapping ORFs V1 and V2, retain the ability to replicate efficiently in N. benthamiana leaf discs but produce an asymptomatic infection in N. benthamiana and B. vulgaris at low frequency, associated with reduced levels of viral DNA compared to wild-type infection. Our data support the recent suggestion that ORF V3 participates in virus movement. The 1.1 kb virion-sense RNA comprises a population of overlapping transcripts with 5' termini suitably positioned for the expression of ORFs V1, V2, and V3. The overlapping arrangement of the transcripts and juxtaposition of putative regulatory elements could provide a means for the temporal control of virion-sense gene expression. PMID:7692668

  2. Cell-to-Cell Transfer of HIV-1 via Virological Synapses Leads to Endosomal Virion Maturation that Activates Viral Membrane Fusion

    PubMed Central

    Dale, Benjamin M.; McNerney, Gregory P.; Thompson, Deanna L.; Hubner, Wolfgang; de los Reyes, Kevin; Chuang, Frank Y.S.; Huser, Thomas; Chen, Benjamin K.

    2012-01-01

    SUMMARY HIV-1 can infect T cells by cell-free virus or by direct virion transfer between cells through cell contact-induced structures called virological synapses (VS). During VS-mediated infection, virions accumulate within target cell endosomes. We show that after crossing the VS, the transferred virus undergoes both maturation and viral membrane fusion. Following VS transfer, viral membrane fusion occurs with delayed kinetics and transferred virions display reduced sensitivity to patient antisera compared to mature, cell-free virus. Furthermore, particle fusion requires that the transferred virions undergo proteolytic maturation within acceptor cell endosomes, which occurs over several hours. Rapid, live cell confocal microscopy demonstrated that viral fusion can occur in compartments that have moved away from the VS. Thus, HIV particle maturation activates viral fusion in target CD4+ T cell endosomes following transfer across the VS and may represent a pathway by which HIV evades antibody neutralization. PMID:22177560

  3. An Anti-Influenza Virus Antibody Inhibits Viral Infection by Reducing Nucleus Entry of Influenza Nucleoprotein

    PubMed Central

    Yoon, Aerin; Yi, Kye Sook; Chang, So Young; Kim, Sung Hwan; Song, Manki; Choi, Jung Ah; Bourgeois, Melissa; Hossain, M. Jaber; Chen, Li-Mei; Donis, Ruben O.; Kim, Hyori; Lee, Yujean; Hwang, Do Been; Min, Ji-Young; Chang, Shin Jae; Chung, Junho

    2015-01-01

    To date, four main mechanisms mediating inhibition of influenza infection by anti-hemagglutinin antibodies have been reported. Anti-globular-head-domain antibodies block either influenza virus receptor binding to the host cell or progeny virion release from the host cell. Anti-stem region antibodies hinder the membrane fusion process or induce antibody-dependent cytotoxicity to infected cells. In this study we identified a human monoclonal IgG1 antibody (CT302), which does not inhibit both the receptor binding and the membrane fusion process but efficiently reduced the nucleus entry of viral nucleoprotein suggesting a novel inhibition mechanism of viral infection by antibody. This antibody binds to the subtype-H3 hemagglutinin globular head domain of group-2 influenza viruses circulating throughout the population between 1997 and 2007. PMID:26512723

  4. An Anti-Influenza Virus Antibody Inhibits Viral Infection by Reducing Nucleus Entry of Influenza Nucleoprotein.

    PubMed

    Yoon, Aerin; Yi, Kye Sook; Chang, So Young; Kim, Sung Hwan; Song, Manki; Choi, Jung Ah; Bourgeois, Melissa; Hossain, M Jaber; Chen, Li-Mei; Donis, Ruben O; Kim, Hyori; Lee, Yujean; Hwang, Do Been; Min, Ji-Young; Chang, Shin Jae; Chung, Junho

    2015-01-01

    To date, four main mechanisms mediating inhibition of influenza infection by anti-hemagglutinin antibodies have been reported. Anti-globular-head-domain antibodies block either influenza virus receptor binding to the host cell or progeny virion release from the host cell. Anti-stem region antibodies hinder the membrane fusion process or induce antibody-dependent cytotoxicity to infected cells. In this study we identified a human monoclonal IgG1 antibody (CT302), which does not inhibit both the receptor binding and the membrane fusion process but efficiently reduced the nucleus entry of viral nucleoprotein suggesting a novel inhibition mechanism of viral infection by antibody. This antibody binds to the subtype-H3 hemagglutinin globular head domain of group-2 influenza viruses circulating throughout the population between 1997 and 2007. PMID:26512723

  5. Vaccine Reduces HPV Infections in Young Men

    Cancer.gov

    An international randomized clinical trial has shown that the vaccine Gardasil can reduce the incidence of anogenital human papillomavirus (HPV) infections in young men 16 to 26 years of age at the time of vaccination.

  6. The vaccinia virus E6 protein influences virion protein localization during virus assembly.

    PubMed

    Condit, Richard C; Moussatche, Nissin

    2015-08-01

    Vaccinia virus mutants in which expression of the virion core protein gene E6R is repressed are defective in virion morphogenesis. E6 deficient infections fail to properly package viroplasm into viral membranes, resulting in an accumulation of empty immature virions and large aggregates of viroplasm. We have used immunogold electron microscopy and immunofluorescence confocal microscopy to assess the intracellular localization of several virion structural proteins and enzymes during E6R mutant infections. We find that during E6R mutant infections virion membrane proteins and virion transcription enzymes maintain a normal localization within viral factories while several major core and lateral body proteins accumulate in aggregated virosomes. The results support a model in which vaccinia virions are assembled from at least three substructures, the membrane, the viroplasm and a "pre-nucleocapsid", and that the E6 protein is essential for maintaining proper localization of the seven-protein complex and the viroplasm during assembly. PMID:25863879

  7. Using PGD to reduce surgical infection risk.

    PubMed

    Archyangelio, Annesha; Shakhon, Amritpal

    Patients with spinal injuries are at increased risk of surgical site infection due to increased numbers of comorbidities and prolonged surgical procedures. This article describes the impact of a patient group direction that was used in a pre-operative assessment clinic to provide Staphylococcus aureus decolonisation to patients with a spinal injury who required prophylaxis. A post-implementation audit revealed that, in the main, staff and patients adhered to the direction, and infection rates were reduced. PMID:27089755

  8. Exogenous and endogenous hyaluronic acid reduces HIV infection of CD4+ T cells

    PubMed Central

    Li, Peilin; Fujimoto, Katsuya; Bourguingnon, Lilly; Yukl, Steven; Deeks, Steven; Wong, Joseph K

    2014-01-01

    Preventing mucosal transmission of HIV is critical to halting the HIV epidemic. Novel approaches to preventing mucosal transmission are needed. Hyaluronic acid (HA) is a major extracellular component of mucosa and the primary ligand for the cell surface receptor CD44. CD44 enhances HIV infection of CD4+ T cells, but the role of HA in this process is not clear. To study this, virions were generated with CD44 (HIVCD44) or without CD44 (HIVmock). Exogenous HA reduced HIV infection of unstimulated CD4+ T cells in a CD44-dependent manner. Conversely, hyaluronidase-mediated reduction of endogenous HA on the cell surface enhanced HIV binding to and infection of unstimulated CD4+ T cells. Exogenous HA treatment reduced activation of protein kinase C alpha via CD44 on CD4+ T cells during infection with HIVCD44. These results reveal new roles for HA during the interaction of HIV with CD4+ T cells that may be relevant to mucosal HIV transmission and could be exploitable as a future strategy to prevent HIV infection. PMID:24957217

  9. M2BP inhibits HIV-1 virion production in a vimentin filaments-dependent manner

    PubMed Central

    Wang, Qin; Zhang, Xiaolin; Han, Yuling; Wang, Xinlu; Gao, Guangxia

    2016-01-01

    M2BP (also called 90K) is an interferon-stimulated gene product that is upregulated in HIV-1 infection. A recent study revealed that M2BP reduces the infectivity of HIV-1 by inhibiting the processing of the viral envelope protein. Here we report that in addition to reducing viral infectivity, M2BP inhibits HIV-1 virion production. We provide evidence showing that M2BP inhibits HIV-1 Gag trafficking to the plasma membrane in a vimentin-dependent manner. When vimentin filaments were collapsed by treating cells with acrylamide or by overexpression of a dominant-negative mutant of vimentin, M2BP inhibition of HIV-1 virion production was significantly relieved. We further show that M2BP interacts with both HIV-1 Gag and vimentin and thereby mediates their interactions. We propose that M2BP traps HIV-1 Gag to vimentin filaments to inhibit the transportation of HIV-1 Gag to the plasma membrane. These findings uncover a novel mechanism by which a host antiviral factor inhibits HIV-1 virion production. PMID:27604950

  10. M2BP inhibits HIV-1 virion production in a vimentin filaments-dependent manner.

    PubMed

    Wang, Qin; Zhang, Xiaolin; Han, Yuling; Wang, Xinlu; Gao, Guangxia

    2016-01-01

    M2BP (also called 90K) is an interferon-stimulated gene product that is upregulated in HIV-1 infection. A recent study revealed that M2BP reduces the infectivity of HIV-1 by inhibiting the processing of the viral envelope protein. Here we report that in addition to reducing viral infectivity, M2BP inhibits HIV-1 virion production. We provide evidence showing that M2BP inhibits HIV-1 Gag trafficking to the plasma membrane in a vimentin-dependent manner. When vimentin filaments were collapsed by treating cells with acrylamide or by overexpression of a dominant-negative mutant of vimentin, M2BP inhibition of HIV-1 virion production was significantly relieved. We further show that M2BP interacts with both HIV-1 Gag and vimentin and thereby mediates their interactions. We propose that M2BP traps HIV-1 Gag to vimentin filaments to inhibit the transportation of HIV-1 Gag to the plasma membrane. These findings uncover a novel mechanism by which a host antiviral factor inhibits HIV-1 virion production. PMID:27604950

  11. Glutathione is required for efficient production of infectious picornavirus virions

    SciTech Connect

    Smith, Allen D. . E-mail: smitha@ba.ars.usda.gov; Dawson, Harry . E-mail: dawsonh@ba.ars.usda.gov

    2006-09-30

    Glutathione is an intracellular reducing agent that helps maintain the redox potential of the cell and is important for immune function. The drug L-buthionine sulfoximine (BSO) selectively inhibits glutathione synthesis. Glutathione has been reported to block replication of HIV, HSV-1, and influenza virus, whereas cells treated with BSO exhibit increased replication of Sendai virus. Pre-treatment of HeLa cell monolayers with BSO inhibited replication of CVB3, CVB4, and HRV14 with viral titers reduced by approximately 6, 5, and 3 log{sub 1}, respectively. The addition of glutathione ethyl ester, but not dithiothreitol or 2-mercaptoethanol, to the culture medium reversed the inhibitory effect of BSO. Viral RNA and protein synthesis were not inhibited by BSO treatment. Fractionation of lysates from CVB3-infected BSO-treated cells on cesium chloride and sucrose gradients revealed that empty capsids but not mature virions were being produced. The levels of the 5S and 14S assembly intermediates, however, were not affected by BSO treatment. These results demonstrate that glutathione is important for production of mature infectious picornavirus virions.

  12. LEDGINs inhibit late stage HIV-1 replication by modulating integrase multimerization in the virions

    PubMed Central

    2013-01-01

    Background LEDGINs are novel allosteric HIV integrase (IN) inhibitors that target the lens epithelium-derived growth factor (LEDGF)/p75 binding pocket of IN. They block HIV-1 integration by abrogating the interaction between LEDGF/p75 and IN as well as by allosterically inhibiting the catalytic activity of IN. Results Here we demonstrate that LEDGINs reduce the replication capacity of HIV particles produced in their presence. We systematically studied the molecular basis of this late effect of LEDGINs and demonstrate that HIV virions produced in their presence display a severe replication defect. Both the late effect and the previously described, early effect on integration contribute to LEDGIN antiviral activity as shown by time-of-addition, qPCR and infectivity assays. The late effect phenotype requires binding of LEDGINs to integrase without influencing proteolytic cleavage or production of viral particles. LEDGINs augment IN multimerization during virion assembly or in the released viral particles and severely hamper the infectivity of progeny virions. About 70% of the particles produced in LEDGIN-treated cells do not form a core or display aberrant empty cores with a mislocalized electron-dense ribonucleoprotein. The LEDGIN-treated virus displays defective reverse transcription and nuclear import steps in the target cells. The LEDGIN effect is possibly exerted at the level of the Pol precursor polyprotein. Conclusion Our results suggest that LEDGINs modulate IN multimerization in progeny virions and impair the formation of regular cores during the maturation step, resulting in a decreased infectivity of the viral particles in the target cells. LEDGINs thus profile as unique antivirals with combined early (integration) and late (IN assembly) effects on the HIV replication cycle. PMID:23721378

  13. Herpes simplex virus virion host shutoff function.

    PubMed Central

    Kwong, A D; Kruper, J A; Frenkel, N

    1988-01-01

    Herpes simplex virus (HSV) virions contain one or more functions which mediate the shutoff of host protein synthesis and the degradation of host mRNA. HSV type 1 (HSV-1) mutants deficient in the virion shutoff of host protein synthesis (vhs mutants) were isolated and were found to be defective in their ability to degrade host mRNA. Furthermore, it was found that viral mRNAs in cells infected with the vhs 1 mutant have a significantly longer functional half-life than viral mRNAs in wild-type virus-infected cells. In the present study we have mapped the vhs1 mutation affecting the virion shutoff of host protein synthesis to a 265-base-pair NruI-XmaIII fragment spanning map coordinates 0.604 to 0.606 of the HSV-1 genome. The mutation(s) affecting the functional half-lives of host mRNA as well as the alpha (immediate-early), beta (early), and gamma (late) viral mRNAs were also mapped within this 265-base-pair fragment. Thus, the shutoff of host protein synthesis is most likely mediated by the same function which decreases the half-life of viral mRNA. The shorter half-life of infected-cell mRNAs may allow a more rapid modulation of viral gene expression in response to changes in the transcription of viral genes. Interestingly, the vhs1 mutation of HSV-1 maps within a region which overlaps the Bg/II-N sequences of HSV-2 DNA shown previously to transform cells in culture. The possible relationship between the transformation and host shutoff functions are discussed. Images PMID:2828686

  14. Bacteriophage Infection of Model Metal Reducing Bacteria

    NASA Astrophysics Data System (ADS)

    Weber, K. A.; Bender, K. S.; Gandhi, K.; Coates, J. D.

    2008-12-01

    Microbially-mediated metal reduction plays a significant role controlling contaminant mobility in aqueous, soil, and sedimentary environments. From among environmentally relevant microorganisms mediating metal reduction, Geobacter spp. have been identified as predominant metal-reducing bacteria under acetate- oxidizing conditions. Due to the significance of these bacteria in environmental systems, it is necessary to understand factors influencing their metabolic physiology. Examination of the annotated finished genome sequence of G. sulfurreducens PCA, G. uraniumreducens Rf4, G. metallireduceans GS-15 as well as a draft genome sequence of Geobacter sp. FRC-32 have identified gene sequences of putative bacteriophage origin. Presence of these sequences indicates that these bacteria are susceptible to phage infection. Polymerase chain reaction (PCR) primer sets designed tested for the presence of 12 of 25 annotated phage-like sequences in G. sulfurreducens PCA and 9 of 17 phage-like sequences in FRC- 32. The following genes were successfully amplified in G. sulfurreducens PCA: prophage type transcription regulator, phage-induced endonuclease, phage tail sheath, 2 phage tail proteins, phage protein D, phage base plate protein, phage-related DNA polymerase, integrase, phage transcriptional regulator, and Cro-like transcription regulator. Nine of the following sequences were present in FRC-32: 4 separate phage- related proteins, phage-related tail component, viron core protein, phage Mu protein, phage base plate, and phage tail sheath. In addition to the bioinformatics evidence, incubation of G. sulfurreducens PCA with 1 μg mL-1 mytomycin C (mutagen stimulating prophage induction) during mid-log phase resulted in significant cell lysis relative to cultures that remained unamended. Cell lysis was concurrent with an increase in viral like particles enumerated using epifluorescent microscopy. In addition, samples collected following this lytic event (~44hours) were

  15. Bacteriophage virion-associated peptidoglycan hydrolases: potential new enzybiotics

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Virion-associated peptidoglycan hydrolases (VAPGH) are phage-encoded lytic enzymes that locally degrade the peptidoglycan (PG) of the bacterial cell wall during infection. Their action usually generates a small hole through which the phage tail crosses the cell envelope to inject the phage genetic m...

  16. M48U1 CD4 mimetic has a sustained inhibitory effect on cell-associated HIV-1 by attenuating virion infectivity through gp120 shedding

    PubMed Central

    2013-01-01

    Background HIV-1 infected cells can establish new infections by crossing the vaginal epithelia and subsequently producing virus in a milieu that avoids the high microbicide concentrations of the vaginal lumen. Findings To address this problem, here, we report that pretreatment of HIV-infected peripheral blood mononuclear cells (PBMCs) with a 27 amino acid CD4-mimetic, M48U1, causes dramatic and prolonged reduction of infectious virus output, due to its induction of gp120 shedding. Conclusions M48U1 may, therefore, be valuable for prophylaxis of mucosal HIV-1 transmission. PMID:23375046

  17. Reducing risk of mosquito-borne infections.

    PubMed

    2016-06-29

    Mosquitoes transmit a number of infections around the globe. Vaccines or chemoprophylaxis protect against few of these diseases, and current outbreaks of Zika and chikungunya viruses are causing significant concern. PMID:27353794

  18. Reduced Risk of Disease During Postsecondary Dengue Virus Infections

    PubMed Central

    Olkowski, Sandra; Forshey, Brett M.; Morrison, Amy C.; Rocha, Claudio; Vilcarromero, Stalin; Halsey, Eric S.; Kochel, Tadeusz J.; Scott, Thomas W.; Stoddard, Steven T.

    2013-01-01

    Background. Antibodies induced by infection with any 1 of 4 dengue virus (DENV) serotypes (DENV-1–4) may influence the clinical outcome of subsequent heterologous infections. To quantify potential cross-protective effects, we estimated disease risk as a function of DENV infection, using data from longitudinal studies performed from September 2006 through February 2011 in Iquitos, Peru, during periods of DENV-3 and DENV-4 transmission. Methods. DENV infections before and during the study period were determined by analysis of serial serum samples with virus neutralization tests. Third and fourth infections were classified as postsecondary infections. Dengue fever cases were detected by door-to-door surveillance for acute febrile illness. Results. Among susceptible participants, 39% (420/1077) and 53% (1595/2997) seroconverted to DENV-3 and DENV-4, respectively. Disease was detected in 7% of DENV-3 infections and 10% of DENV-4 infections. Disease during postsecondary infections was reduced by 93% for DENV-3 and 64% for DENV-4, compared with primary and secondary infections. Despite lower disease rates, postsecondary infections constituted a significant proportion of apparent infections (14% [for DENV-3 infections], 45% [for DENV-4 infections]). Conclusions. Preexisting heterotypic antibodies markedly reduced but did not eliminate the risk of disease in this study population. These results improve understanding of how preinfection history can be associated with dengue outcomes and DENV transmission dynamics. PMID:23776195

  19. Reducing Surgical Site Infections: A Review

    PubMed Central

    Reichman, David E; Greenberg, James A

    2009-01-01

    Infection at or near surgical incisions within 30 days of an operative procedure contributes substantially to surgical morbidity and mortality each year. The prevention of surgical site infections encompasses meticulous operative technique, timely administration of appropriate preoperative antibiotics, and a variety of preventive measures aimed at neutralizing the threat of bacterial, viral, and fungal contamination posed by operative staff, the operating room environment, and the patient’s endogenous skin flora. It is the latter aspect of contamination, and specifically mechanical methods of prevention, on which this review focuses. PMID:20111657

  20. Identification of Human Cytomegalovirus Genes Important for Biogenesis of the Cytoplasmic Virion Assembly Complex

    PubMed Central

    Das, Subhendu; Ortiz, Daniel A.; Gurczynski, Stephen J.; Khan, Fatin

    2014-01-01

    infected cells to build a factory for assembling new infectious particles (virions), the cytoplasmic virion assembly complex (cVAC). Here, we identified three HCMV genes (UL48, UL94, and UL103) as important contributors to cVAC development. In addition, we found that mutant viruses that express an unstable form of the UL103 protein have defects in cVAC development and production of infectious virions and produce small plaques and intracellular virions with aberrant appearances. Of these, only the reduced production of infectious virions is not eliminated by chemically stabilizing the protein. In addition to identifying new functions for these HCMV genes, this work is a necessary prelude to developing novel antivirals that would block cVAC development. PMID:24899189

  1. Reducing infections through nanotechnology and nanoparticles

    PubMed Central

    Taylor, Erik; Webster, Thomas J

    2011-01-01

    The expansion of bacterial antibiotic resistance is a growing problem today. When medical devices are inserted into the body, it becomes especially difficult for the body to clear robustly adherent antibiotic-resistant biofilm infections. In addition, concerns about the spread of bacterial genetic tolerance to antibiotics, such as that found in multiple drug-resistant Staphylococcus aureus (MRSA), have significantly increased of late. As a growing direction in biomaterial design, nanomaterials (materials with at least one dimension less than 100 nm) may potentially prevent bacterial functions that lead to infections. As a first step in this direction, various nanoparticles have been explored for improving bacteria and biofilm penetration, generating reactive oxygen species, and killing bacteria, potentially providing a novel method for fighting infections that is nondrug related. This review article will first examine in detail the mechanisms and applications of some of these nanoparticles, then follow with some recent material designs utilizing nanotechnology that are centered on fighting medical device infections. PMID:21796248

  2. The vaccinia virus E6 protein influences virion protein localization during virus assembly

    SciTech Connect

    Condit, Richard C. Moussatche, Nissin

    2015-08-15

    Vaccinia virus mutants in which expression of the virion core protein gene E6R is repressed are defective in virion morphogenesis. E6 deficient infections fail to properly package viroplasm into viral membranes, resulting in an accumulation of empty immature virions and large aggregates of viroplasm. We have used immunogold electron microscopy and immunofluorescence confocal microscopy to assess the intracellular localization of several virion structural proteins and enzymes during E6R mutant infections. We find that during E6R mutant infections virion membrane proteins and virion transcription enzymes maintain a normal localization within viral factories while several major core and lateral body proteins accumulate in aggregated virosomes. The results support a model in which vaccinia virions are assembled from at least three substructures, the membrane, the viroplasm and a “pre-nucleocapsid”, and that the E6 protein is essential for maintaining proper localization of the seven-protein complex and the viroplasm during assembly. - Highlights: • Mutation of E6 disrupts association of viral membranes with viral core proteins • Mutation of E6 does not perturb viral membrane biosynthesis • Mutation of E6 does not perturb localization of viral transcription enzymes • Mutation of E6 causes mis-localization and aggregation of viral core proteins • Vaccinia assembly uses three subassemblies: membranes, viroplasm, prenucleocapsid.

  3. GP3 is a structural component of the PRRSV type II (US) virion

    SciTech Connect

    Lima, M. de; Ansari, I.H.; Das, P.B.; Ku, B.J.; Martinez-Lobo, F.J.; Pattnaik, A.K.; Osorio, F.A.

    2009-07-20

    Glycoprotein 3 (GP3) is a highly glycosylated PRRSV envelope protein which has been reported as being present in the virions of PRRSV type I, while missing in the type II PRRSV (US) virions. We herein present evidence that GP3 is indeed incorporated in the virus particles of a North American strain of PRRSV (FL12), at a density that is consistent with the minor structural role assigned to GP3 in members of the Arterivirus genus. Two 15aa peptides corresponding to two different immunodominant linear epitopes of GP3 derived from the North American strain of PRRSV (FL12) were used as antigen to generate a rabbit monospecific antiserum to this protein. The specificity of this anti-GP3 antiserum was confirmed by radioimmunoprecipitation (RIP) assay using BHK-21 cells transfected with GP3 expressing plasmid, MARC-145 cells infected with FL12 PRRSV, as well as by confocal microscopy on PRRSV-infected MARC-145 cells. To test if GP3 is a structural component of the virion, {sup 35}S-labelled PRRSV virions were pelleted through a 30% sucrose cushion, followed by a second round of purification on a sucrose gradient (20-60%). Virions were detected in specific gradient fractions by radioactive counts and further confirmed by viral infectivity assay in MARC 145 cells. The GP3 was detected in gradient fractions containing purified virions by RIP using anti-GP3 antiserum. Predictably, the GP3 was less abundant in purified virions than other major structural envelope proteins such as GP5 and M. Further evidence of the presence of GP3 at the level of PRRSV FL12 envelope was obtained by immunogold staining of purified virions from the supernatant of infected cells with anti-GP3 antiserum. Taken together, these results indicate that GP3 is a minor structural component of the PRRSV type II (FL12 strain) virion, as had been previously described for PRRSV type I.

  4. HIV binding, penetration, and primary infection in human cervicovaginal tissue

    PubMed Central

    Maher, Diane; Wu, Xiaoyun; Schacker, Timothy; Horbul, Julie; Southern, Peter

    2005-01-01

    We have developed human cervicovaginal organ culture systems to examine the initiating events in HIV transmission after exposure to various sources of HIV infectivity, including semen. Newly infected cells were detected in the cervical submucosa 3–4 days after exposure to a primary HIV isolate. At earlier times, extensive and stable binding occurred when cervical surfaces were exposed to virions or seminal cells. Cervical mucus provided some protection for the endocervical surface, by physically trapping virions and seminal cells. Confocal microscopy combined with 3D surface reconstruction revealed that virions could both bind to the external surface of the cervical epithelium and actually penetrate beneath the epithelial surface. In quantitative assays, pretreatment with a blocking antibody directed against β1 integrin reduced HIV virion binding. Collectively, these results highlight a continuum of complex interactions that occurs when natural sources of HIV infectivity are deposited onto mucosal surfaces in the female reproductive tract. PMID:16061810

  5. Mutations in the catalytic core or the C-terminus of murine leukemia virus (MLV) integrase disrupt virion infectivity and exert diverse effects on reverse transcription

    SciTech Connect

    Steinrigl, Adolf; Nosek, Dagmara; Ertl, Reinhard; Guenzburg, Walter H.; Salmons, Brian; Klein, Dieter . E-mail: dieter.klein@vu-wien.ac.at

    2007-05-25

    Understanding of the structures and functions of the retroviral integrase (IN), a key enzyme in the viral replication cycle, is essential for developing antiretroviral treatments and facilitating the development of safer gene therapy vehicles. Thus, four MLV IN-mutants were constructed in the context of a retroviral vector system, harbouring either a substitution in the catalytic centre, deletions in the C-terminus, or combinations of both modifications. IN-mutants were tested for their performance in different stages of the viral replication cycle: RNA-packaging; RT-activity; transient and stable infection efficiency; dynamics of reverse transcription and nuclear entry. All mutant vectors packaged viral RNA with wild-type efficiencies and displayed only slight reductions in RT-activity. Deletion of either the IN C-terminus alone, or in addition to part of the catalytic domain exerted contrasting effects on intracellular viral DNA levels, implying that IN influences reverse transcription in more than one direction.

  6. Reducing periprosthetic joint infection: what really counts?

    PubMed Central

    SOLARINO, GIUSEPPE; ABATE, ANTONELLA; VICENTI, GIOVANNI; SPINARELLI, ANTONIO; PIAZZOLLA, ANDREA; MORETTI, BIAGIO

    2015-01-01

    Periprosthetic joint infection (PJI) remains one of the most challenging complications after joint arthroplasty. Despite improvements in surgical techniques and in the use of antibiotic prophylaxis, it remains a major cause of implant failure and need for revision. PJI is associated with both human host-related and bacterial agent-related factors that can interact in all the phases of the procedure (preoperative, intraoperative and postoperative). Prevention is the first strategy to implement in order to minimize this catastrophic complication. The present review focuses on the preoperative period, and on what to do once risk factors are fully understood and have been identified. PMID:26904527

  7. Virus factories of cauliflower mosaic virus are virion reservoirs that engage actively in vector transmission.

    PubMed

    Bak, Aurélie; Gargani, Daniel; Macia, Jean-Luc; Malouvet, Enrick; Vernerey, Marie-Stéphanie; Blanc, Stéphane; Drucker, Martin

    2013-11-01

    Cauliflower mosaic virus (CaMV) forms two types of inclusion bodies within infected plant cells: numerous virus factories, which are the sites for viral replication and virion assembly, and a single transmission body (TB), which is specialized for virus transmission by aphid vectors. The TB reacts within seconds to aphid feeding on the host plant by total disruption and redistribution of its principal component, the viral transmission helper protein P2, onto microtubules throughout the cell. At the same time, virions also associate with microtubules. This redistribution of P2 and virions facilitates transmission and is reversible; the TB reforms within minutes after vector departure. Although some virions are present in the TB before disruption, their subsequent massive accumulation on the microtubule network suggests that they also are released from virus factories. Using drug treatments, mutant viruses, and exogenous supply of viral components to infected protoplasts, we show that virions can rapidly exit virus factories and, once in the cytoplasm, accumulate together with the helper protein P2 on the microtubule network. Moreover, we show that during reversion of this phenomenon, virions from the microtubule network can either be incorporated into the reverted TB or return to the virus factories. Our results suggest that CaMV factories are dynamic structures that participate in vector transmission by controlled release and uptake of virions during TB reaction. PMID:24006440

  8. A virus capsid component mediates virion retention and transmission by its insect vector

    PubMed Central

    Chen, Angel Y. S.; Walker, Gregory P.; Carter, David; Ng, James C. K.

    2011-01-01

    Numerous pathogens of humans, animals, and plants are transmitted by specific arthropod vectors. However, understanding the mechanisms governing these pathogen–vector interactions is hampered, in part, by the lack of easy-to-use analytical tools. We investigated whitefly transmission of Lettuce infectious yellows virus (LIYV) by using a unique immunofluorescent localization approach in which we fed virions or recombinant virus capsid components to whiteflies, followed by feeding them antibodies to the virions or capsid components, respectively. Fluorescent signals, indicating the retention of virions, were localized in the anterior foregut or cibarium of a whitefly vector biotype but not within those of a whitefly nonvector biotype. Retention of virions in these locations strongly corresponded with the whitefly vector transmission of LIYV. When four recombinant LIYV capsid components were individually fed to whitefly vectors, significantly more whiteflies retained the recombinant minor coat protein (CPm). As demonstrated previously and in the present study, whitefly vectors failed to transmit virions preincubated with anti-CPm antibodies but transmitted virions preincubated with antibodies recognizing the major coat protein (CP). Correspondingly, the number of insects that specifically retained virions preincubated with anti-CPm antibodies were significantly reduced compared with those that specifically retained virions preincubated with anti-CP antibodies. Notably, a transmission-defective CPm mutant was deficient in specific virion retention, whereas the CPm-restored virus showed WT levels of specific virion retention and transmission. These data provide strong evidence that transmission of LIYV is determined by a CPm-mediated virion retention mechanism in the anterior foregut or cibarium of whitefly vectors. PMID:21930903

  9. Getting IN on Viral RNA Condensation and Virion Maturation.

    PubMed

    Freed, Eric O

    2016-08-25

    The retroviral enzyme integrase plays an essential role in the virus replication cycle by catalyzing the covalent insertion of newly synthesized viral DNA into the host cell chromosome early after infection. Now, Kessl et al. report a second function of integrase: binding to the viral RNA genome in virion particles late in the virus replication cycle to promote particle maturation. PMID:27565339

  10. A phenyl-thiadiazolylidene-amine derivative ejects zinc from retroviral nucleocapsid zinc fingers and inactivates HIV virions

    PubMed Central

    2012-01-01

    Background Sexual acquisition of the human immunodeficiency virus (HIV) through mucosal transmission may be prevented by using topically applied agents that block HIV transmission from one individual to another. Therefore, virucidal agents that inactivate HIV virions may be used as a component in topical microbicides. Results Here, we have identified 2-methyl-3-phenyl-2H-[1,2,4]thiadiazol-5-ylideneamine (WDO-217) as a low-molecular-weight molecule that inactivates HIV particles. Both HIV-1 and HIV-2 virions pretreated with this compound were unable to infect permissive cells. Moreover, WDO-217 was able to inhibit infections of a wide spectrum of wild-type and drug-resistant HIV-1, including clinical isolates, HIV-2 and SIV strains. Whereas the capture of virus by DC-SIGN was unaffected by the compound, it efficiently prevented the transmission of DC-SIGN-captured virus to CD4+ T-lymphocytes. Interestingly, exposure of virions to WDO-217 reduced the amount of virion-associated genomic RNA as measured by real-time RT-qPCR. Further mechanism-of-action studies demonstrated that WDO-217 efficiently ejects zinc from the zinc fingers of the retroviral nucleocapsid protein NCp7 and inhibits the cTAR destabilization properties of this protein. Importantly, WDO-217 was able to eject zinc from both zinc fingers, even when NCp7 was bound to oligonucleotides, while no covalent interaction between NCp7 and WDO-217 could be observed. Conclusion This compound is a new lead structure that can be used for the development of a new series of NCp7 zinc ejectors as candidate topical microbicide agents. PMID:23146561

  11. Spacecraft Environment May Reduce Resistance To Infection

    NASA Technical Reports Server (NTRS)

    Pierson, Duane L.; Ott, C. Mark; Castro, V. A.; Leal, Melanie; Mehta, Satish K.

    2006-01-01

    conditions. Data indicates that space flight is a unique stress environment that may produce stress-induced changes in the host-microbe relationship resulting in increased risk of infection.

  12. Nanoparticle-based flow virometry for the analysis of individual virions.

    PubMed

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

    2013-09-01

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

  13. Endemic infection reduces transmission potential of an epidemic parasite during co-infection.

    PubMed

    Randall, J; Cable, J; Guschina, I A; Harwood, J L; Lello, J

    2013-10-22

    Endemic, low-virulence parasitic infections are common in nature. Such infections may deplete host resources, which in turn could affect the reproduction of other parasites during co-infection. We aimed to determine whether the reproduction, and therefore transmission potential, of an epidemic parasite was limited by energy costs imposed on the host by an endemic infection. Total lipids, triacylglycerols (TAG) and polar lipids were measured in cockroaches (Blattella germanica) that were fed ad libitum, starved or infected with an endemic parasite, Gregarina blattarum. Reproductive output of an epidemic parasite, Steinernema carpocapsae, was then assessed by counting the number of infective stages emerging from these three host groups. We found both starvation and gregarine infection reduced cockroach lipids, mainly through depletion of TAG. Further, both starvation and G. blattarum infection resulted in reduced emergence of nematode transmission stages. This is, to our knowledge, the first study to demonstrate directly that host resource depletion caused by endemic infection could affect epidemic disease transmission. In view of the ubiquity of endemic infections in nature, future studies of epidemic transmission should take greater account of endemic co-infections. PMID:23966641

  14. Endemic infection reduces transmission potential of an epidemic parasite during co-infection

    PubMed Central

    Randall, J.; Cable, J.; Guschina, I. A.; Harwood, J. L.; Lello, J.

    2013-01-01

    Endemic, low-virulence parasitic infections are common in nature. Such infections may deplete host resources, which in turn could affect the reproduction of other parasites during co-infection. We aimed to determine whether the reproduction, and therefore transmission potential, of an epidemic parasite was limited by energy costs imposed on the host by an endemic infection. Total lipids, triacylglycerols (TAG) and polar lipids were measured in cockroaches (Blattella germanica) that were fed ad libitum, starved or infected with an endemic parasite, Gregarina blattarum. Reproductive output of an epidemic parasite, Steinernema carpocapsae, was then assessed by counting the number of infective stages emerging from these three host groups. We found both starvation and gregarine infection reduced cockroach lipids, mainly through depletion of TAG. Further, both starvation and G. blattarum infection resulted in reduced emergence of nematode transmission stages. This is, to our knowledge, the first study to demonstrate directly that host resource depletion caused by endemic infection could affect epidemic disease transmission. In view of the ubiquity of endemic infections in nature, future studies of epidemic transmission should take greater account of endemic co-infections. PMID:23966641

  15. Secondary metabolites in floral nectar reduce parasite infections in bumblebees

    PubMed Central

    Richardson, Leif L.; Adler, Lynn S.; Leonard, Anne S.; Andicoechea, Jonathan; Regan, Karly H.; Anthony, Winston E.; Manson, Jessamyn S.; Irwin, Rebecca E.

    2015-01-01

    The synthesis of secondary metabolites is a hallmark of plant defence against herbivores. These compounds may be detrimental to consumers, but can also protect herbivores against parasites. Floral nectar commonly contains secondary metabolites, but little is known about the impacts of nectar chemistry on pollinators, including bees. We hypothesized that nectar secondary metabolites could reduce bee parasite infection. We inoculated individual bumblebees with Crithidia bombi, an intestinal parasite, and tested effects of eight naturally occurring nectar chemicals on parasite population growth. Secondary metabolites strongly reduced parasite load, with significant effects of alkaloids, terpenoids and iridoid glycosides ranging from 61 to 81%. Using microcolonies, we also investigated costs and benefits of consuming anabasine, the compound with the strongest effect on parasites, in infected and uninfected bees. Anabasine increased time to egg laying, and Crithidia reduced bee survival. However, anabasine consumption did not mitigate the negative effects of Crithidia, and Crithidia infection did not alter anabasine consumption. Our novel results highlight that although secondary metabolites may not rescue survival in infected bees, they may play a vital role in mediating Crithidia transmission within and between colonies by reducing Crithidia infection intensities. PMID:25694627

  16. Secondary metabolites in floral nectar reduce parasite infections in bumblebees.

    PubMed

    Richardson, Leif L; Adler, Lynn S; Leonard, Anne S; Andicoechea, Jonathan; Regan, Karly H; Anthony, Winston E; Manson, Jessamyn S; Irwin, Rebecca E

    2015-03-22

    The synthesis of secondary metabolites is a hallmark of plant defence against herbivores. These compounds may be detrimental to consumers, but can also protect herbivores against parasites. Floral nectar commonly contains secondary metabolites, but little is known about the impacts of nectar chemistry on pollinators, including bees. We hypothesized that nectar secondary metabolites could reduce bee parasite infection. We inoculated individual bumblebees with Crithidia bombi, an intestinal parasite, and tested effects of eight naturally occurring nectar chemicals on parasite population growth. Secondary metabolites strongly reduced parasite load, with significant effects of alkaloids, terpenoids and iridoid glycosides ranging from 61 to 81%. Using microcolonies, we also investigated costs and benefits of consuming anabasine, the compound with the strongest effect on parasites, in infected and uninfected bees. Anabasine increased time to egg laying, and Crithidia reduced bee survival. However, anabasine consumption did not mitigate the negative effects of Crithidia, and Crithidia infection did not alter anabasine consumption. Our novel results highlight that although secondary metabolites may not rescue survival in infected bees, they may play a vital role in mediating Crithidia transmission within and between colonies by reducing Crithidia infection intensities. PMID:25694627

  17. Bacterial lipopolysaccharide binding enhances virion stability and promotes environmental fitness of an enteric virus

    PubMed Central

    Robinson, Christopher M.; Jesudhasan, Palmy R.; Pfeiffer, Julie K.

    2014-01-01

    Summary Enteric viruses, including poliovirus and reovirus, encounter a vast microbial community in the mammalian gastrointestinal tract, which has been shown to promote virus replication and pathogenesis. Investigating the underlying mechanisms, we find that poliovirus binds bacterial surface polysaccharides, which enhances virion stability and cell attachment by increasing binding to the viral receptor. Additionally, we identified a poliovirus mutant, VP1-T99K, with reduced lipopolysaccharide (LPS) binding. Although T99K and WT poliovirus cell attachment, replication and pathogenesis in mice are equivalent, following peroral inoculation of mice, VP1-T99K poliovirus was unstable in feces. Consequently, the ratio of mutant virus in feces is reduced following additional cycles of infection in mice. Thus, the mutant virus incurs a fitness cost when environmental stability is a factor. These data suggest that poliovirus binds bacterial surface polysaccharides, enhancing cell attachment and environmental stability, potentially promoting transmission to a new host. PMID:24439896

  18. Recruitment of a SAP18-HDAC1 complex into HIV-1 virions and its requirement for viral replication.

    PubMed

    Sorin, Masha; Cano, Jennifer; Das, Supratik; Mathew, Sheeba; Wu, Xuhong; Davies, Kelvin P; Shi, Xuanling; Cheng, S-W Grace; Ott, David; Kalpana, Ganjam V

    2009-06-01

    HIV-1 integrase (IN) is a virally encoded protein required for integration of viral cDNA into host chromosomes. INI1/hSNF5 is a component of the SWI/SNF complex that interacts with HIV-1 IN, is selectively incorporated into HIV-1 (but not other retroviral) virions, and modulates multiple steps, including particle production and infectivity. To gain further insight into the role of INI1 in HIV-1 replication, we screened for INI1-interacting proteins using the yeast two-hybrid system. We found that SAP18 (Sin3a associated protein 18 kD), a component of the Sin3a-HDAC1 complex, directly binds to INI1 in yeast, in vitro and in vivo. Interestingly, we found that IN also binds to SAP18 in vitro and in vivo. SAP18 and components of a Sin3A-HDAC1 complex were specifically incorporated into HIV-1 (but not SIV and HTLV-1) virions in an HIV-1 IN-dependent manner. Using a fluorescence-based assay, we found that HIV-1 (but not SIV) virion preparations harbour significant deacetylase activity, indicating the specific recruitment of catalytically active HDAC into the virions. To determine the requirement of virion-associated HDAC1 to HIV-1 replication, an inactive, transdominant negative mutant of HDAC1 (HDAC1(H141A)) was utilized. Incorporation of HDAC1(H141A) decreased the virion-associated histone deacetylase activity. Furthermore, incorporation of HDAC1(H141A) decreased the infectivity of HIV-1 (but not SIV) virions. The block in infectivity due to virion-associated HDAC1(H141A) occurred specifically at the early reverse transcription stage, while entry of the virions was unaffected. RNA-interference mediated knock-down of HDAC1 in producer cells resulted in decreased virion-associated HDAC1 activity and a reduction in infectivity of these virions. These studies indicate that HIV-1 IN and INI1/hSNF5 bind SAP18 and selectively recruit components of Sin3a-HDAC1 complex into HIV-1 virions. Furthermore, HIV-1 virion-associated HDAC1 is required for efficient early post

  19. Physicochemical Studies on L-Cell Virions

    PubMed Central

    Nichols, J. L.; Quade, Kristina; Luftig, R. B.

    1973-01-01

    The L-cell virion (LCV) has been purified from supernatant fluids of mouse L cells grown in suspension culture. The virion is similar to other RNA tumor viruses by several criteria: (i) the density of the virion is 1.16 g/cm3; (ii) the virion appears as a rounded membranous particle with an outer diameter of 146.7 ± 11.8 nm, and contains knobs (7-nm diameter) over its surface; (iii) 15 polypeptides (ranging in molecular weight from 7,000 to 110,000) are detectable after electrophoresis of virion protein in sodium dodecyl sulfate-polyacrylamide gels; (iv) three species of RNA can be isolated—high molecular weight (80 to 88s) (50%), 7s (35%) and 4s (15%); (v) heat denaturation of the high-molecular-weight RNA yields a heterogeneous population of molecules (20 to 35s) as well as a 7s and 4s species. Despite the general similarity to infectious RNA tumor viruses, LCV is apparently defective as evidenced by the fact that it does not induce tumors in animals or transform normal mouse cells in vitro (Kindig and Kirsten [17]). The defective nature of the LCV might be related to the fact that assays for DNA polymerase in the virion showed only a negligible activity when compared to Rous sarcoma virus. Images PMID:4120638

  20. The herpes simplex virus virion host shutoff function.

    PubMed Central

    Kwong, A D; Frenkel, N

    1989-01-01

    The virion host shutoff (vhs) function of herpes simplex virus (HSV) limits the expression of genes in the infected cells by destabilizing both host and viral mRNAs. vhs function mutants have been isolated which are defective in their ability to degrade host mRNA. Furthermore, the half-life of viral mRNAs is significantly longer in cells infected with the vhs-1 mutant virus than in cells infected with the wild-type (wt) virus. Recent data have shown that the vhs-1 mutation resides within the open reading frame UL41. We have analyzed the shutoff of host protein synthesis in cells infected with a mixture of the wt HSV-1 (KOS) and the vhs-1 mutant virus. The results of these experiments revealed that (i) the wt virus shutoff activity requires a threshold level of input virions per cell and (ii) the mutant vhs-1 virus protein can irreversibly block the wt virus shutoff activity. These results are consistent with a stoichiometric model in which the wt vhs protein interacts with a cellular factor which controls the half-life of cell mRNA. This wt virus interaction results in the destabilization of both host and viral mRNAs. In contrast, the mutant vhs function interacts with the cellular factor irreversibly, resulting in the increased half-life of both host and viral mRNAs. Images PMID:2552156

  1. Formation of virions is strictly required for turnip yellows virus long-distance movement in plants.

    PubMed

    Hipper, Clémence; Monsion, Baptiste; Bortolamiol-Bécet, Diane; Ziegler-Graff, Véronique; Brault, Véronique

    2014-02-01

    Viral genomic RNA of the Turnip yellows virus (TuYV; genus Polerovirus; family Luteoviridae) is protected in virions formed by the major capsid protein (CP) and the minor component, the readthrough (RT*) protein. Long-distance transport, used commonly by viruses to systemically infect host plants, occurs in phloem sieve elements and two viral forms of transport have been described: virions and ribonucleoprotein (RNP) complexes. With regard to poleroviruses, virions have always been presumed to be the long-distance transport form, but the potential role of RNP complexes has not been investigated. Here, we examined the requirement of virions for polerovirus systemic movement by analysing CP-targeted mutants that were unable to form viral particles. We confirmed that TuYV mutants that cannot encapsidate into virions are not able to reach systemic leaves. To completely discard the possibility that the introduced mutations in CP simply blocked the formation or the movement of RNP complexes, we tested in trans complementation of TuYV CP mutants by providing WT CP expressed in transgenic plants. WT CP was able to facilitate systemic movement of TuYV CP mutants and this observation was always correlated with the formation of virions. This demonstrated clearly that virus particles are essential for polerovirus systemic movement. PMID:24214396

  2. Mixed genotype transmission bodies and virions contribute to the maintenance of diversity in an insect virus

    PubMed Central

    Clavijo, Gabriel; Williams, Trevor; Muñoz, Delia; Caballero, Primitivo; López-Ferber, Miguel

    2010-01-01

    An insect nucleopolyhedrovirus naturally survives as a mixture of at least nine genotypes. Infection by multiple genotypes results in the production of virus occlusion bodies (OBs) with greater pathogenicity than those of any genotype alone. We tested the hypothesis that each OB contains a genotypically diverse population of virions. Few insects died following inoculation with an experimental two-genotype mixture at a dose of one OB per insect, but a high proportion of multiple infections were observed (50%), which differed significantly from the frequencies predicted by a non-associated transmission model in which genotypes are segregated into distinct OBs. By contrast, insects that consumed multiple OBs experienced higher mortality and infection frequencies did not differ significantly from those of the non-associated model. Inoculation with genotypically complex wild-type OBs indicated that genotypes tend to be transmitted in association, rather than as independent entities, irrespective of dose. To examine the hypothesis that virions may themselves be genotypically heterogeneous, cell culture plaques derived from individual virions were analysed to reveal that one-third of virions was of mixed genotype, irrespective of the genotypic composition of the OBs. We conclude that co-occlusion of genotypically distinct virions in each OB is an adaptive mechanism that favours the maintenance of virus diversity during insect-to-insect transmission. PMID:19939845

  3. The intracellular dynamics of hepatitis B virus (HBV) replication with reproduced virion "re-cycling".

    PubMed

    Nakabayashi, Jun

    2016-05-01

    Hepatitis B virus (HBV) is a causative agent of hepatitis. Clinical outcome of hepatitis type B depends on the viral titer observed in the peripheral blood of the patient. In the chronic hepatitis patient, production of HBV virion remains low level. On the other hand, the viral load prominently increases in fulminant hepatitis patient as compared with that in the chronic hepatitis patient. We previously proposed a mathematical model describing the intracellular dynamics of HBV replication. Our model clarified that there are two distinguishable replication patterns of HBV named "arrested" and "explosive" replication. In the arrested replication, the amount of virion newly reproduced from an infected cell remains low level, while the amount of virion extremely increases in the explosive replication. Viral load is drastically changed by slight alteration of expression ratio of 3.5kb RNA to 2.4kb mRNA of HBV. Though our model provided the switching mechanism determining the replication pattern of HBV, HBV dynamics is determined by not only the expression pattern of viral genes. In this study, "recycling" of HBV virion in the replication cycle is investigated as a new factor affecting the intracellular dynamics of HBV replication. A part of newly produced virion of HBV is reused as a core particle that is a resource of HBV replication. This recycling of HBV virion lowers the threshold for the explosive replication when waiting time for the next cycle of the replication is large. It is seemingly contradicting that prominent production of HBV is caused by large recycling rate and small release rate of HBV virion from infected cell to extracellular space. But the recycling of HBV virion can contribute to the positive feedback cycle of HBV replication for the explosive replication to accumulate the core particle as a resource of HBV replication in an infected cell. Accumulation of core particle in the infected cell can be risk factor for the exacerbation of hepatitis rather

  4. Preoperative skin disinfection methodologies for reducing prosthetic joint infections.

    PubMed

    Banerjee, Samik; Kapadia, Bhaveen H; Mont, Michael A

    2014-08-01

    Surgical site infections following lower extremity total joint arthroplasty procedures remain a substantial economic burden to the patient, the treating institution, and the health care system. The complexity of these surgical procedures creates the potential for various patient- or surgery-related risk factors for infection. Although there is much literature regarding the use of many preventative methods, a consensus regarding the true efficacy and application of such practices is generally not available. In this review, we reviewed the preoperative skin disinfection methodologies that have been used for reducing periprosthetic infections following lower extremity total joint arthroplasty. Currently, cumulative evidence suggests that preoperative chlorhexidine baths or chlorhexidine-impregnated wipes may reduce the colonization of pathogenic organisms on the skin. In addition, multiple showers or topical applications of chlorhexidine may lead to more substantial reduction in colony counts. Advanced preoperative whole-body cleaning with chlorhexidine-containing cloths rather than site-specific application may confer additional advantages. Further randomized controlled trials with carefully planned protocols and endpoints are needed to determine if this conclusively leads to reduction in the rate of surgical site infections. PMID:24622912

  5. Proteomics of HCV virions reveals an essential role for the nucleoporin Nup98 in virus morphogenesis.

    PubMed

    Lussignol, Marion; Kopp, Martina; Molloy, Kelly; Vizcay-Barrena, Gema; Fleck, Roland A; Dorner, Marcus; Bell, Kierstin L; Chait, Brian T; Rice, Charles M; Catanese, Maria Teresa

    2016-03-01

    Hepatitis C virus (HCV) is a unique enveloped virus that assembles as a hybrid lipoviral particle by tightly interacting with host lipoproteins. As a result, HCV virions display a characteristic low buoyant density and a deceiving coat, with host-derived apolipoproteins masking viral epitopes. We previously described methods to produce high-titer preparations of HCV particles with tagged envelope glycoproteins that enabled ultrastructural analysis of affinity-purified virions. Here, we performed proteomics studies of HCV isolated from culture media of infected hepatoma cells to define viral and host-encoded proteins associated with mature virions. Using two different affinity purification protocols, we detected four viral and 46 human cellular proteins specifically copurifying with extracellular HCV virions. We determined the C terminus of the mature capsid protein and reproducibly detected low levels of the viral nonstructural protein, NS3. Functional characterization of virion-associated host factors by RNAi identified cellular proteins with either proviral or antiviral roles. In particular, we discovered a novel interaction between HCV capsid protein and the nucleoporin Nup98 at cytosolic lipid droplets that is important for HCV propagation. These results provide the first comprehensive view to our knowledge of the protein composition of HCV and new insights into the complex virus-host interactions underlying HCV infection. PMID:26884193

  6. Conserved and host-specific features of influenza virion architecture

    PubMed Central

    Hutchinson, Edward C; Charles, Philip D; Hester, Svenja S; Thomas, Benjamin; Trudgian, David; Martínez-Alonso, Mónica; Fodor, Ervin

    2014-01-01

    Viruses use virions to spread between hosts, and virion composition is therefore the primary determinant of viral transmissibility and immunogenicity. However, the virions of many viruses are complex and pleomorphic, making them difficult to analyse in detail. Here we address this by identifying and quantifying virion proteins with mass spectrometry, producing a complete and quantified model of the hundreds of viral and host-encoded proteins that make up the pleomorphic virions of influenza viruses. We show that a conserved influenza virion architecture is maintained across diverse combinations of virus and host. This ‘core’ architecture, which includes substantial quantities of host proteins as well as the viral protein NS1, is elaborated with abundant host-dependent features. As a result, influenza virions produced by mammalian and avian hosts have distinct protein compositions. Finally we note that influenza virions share an underlying protein composition with exosomes, suggesting that influenza virions form by subverting microvesicle production. PMID:25226414

  7. Coxsackievirus B3 infection reduces female mouse fertility

    PubMed Central

    Shim, Hye Min; Hwang, Ji Young; Lee, Kyung Min; Kim, Yunhwa; Jeong, Daewon; Roh, Jaesook; Choi, Hyeonhae; Hwang, Jung Hye; Park, Hosun

    2015-01-01

    Previously we demonstrated coxsackievirus B3 (CVB3) infection during early gestation as a cause of pregnancy loss. Here, we investigated the impacts of CVB3 infection on female mouse fertility. Coxsackievirus-adenovirus receptor (CAR) expression and CVB3 replication in the ovary were evaluated by immunohistochemistry or reverse transcription-polymerase chain reaction (RT-PCR). CAR was highly expressed in granulosa cells (GCs) and CVB3 replicated in the ovary. Histological analysis showed a significant increase in the number of atretic follicles in the ovaries of CVB3-infected mice (CVBM). Estrous cycle evaluation demonstrated that a higher number of CVBM were in proestrus compared to mock mice (CVBM vs. mock; 61.5%, 28.5%, respectively). Estradiol concentration in GC culture supernatant and serum were measured by an enzyme-linked immunosorbent assay. Baseline and stimulated levels of estradiol in GC were decreased in CVBM, consistent with significantly reduced serum levels in these animals. In addition, aromatase transcript levels in GCs from CVBM were also decreased by 40% relative to the mock. Bone mineral density evaluated by micro-computed tomography was significantly decreased in the CVBM. Moreover, the fertility rate was also significantly decreased for the CVBM compared to the mock (CVBM vs. mock; 20%, 94.7%, respectively). This study suggests that CVB3 infection could interfere with reproduction by disturbing ovarian function and cyclic changes of the uterus. PMID:26062767

  8. Reducing bloodstream infection with a chlorhexidine gel IV dressing.

    PubMed

    Jeanes, Annette; Bitmead, James

    The use of vascular access devices (VAD) is common in healthcare provision but there is a significant risk of acquiring an infection. Central venous catheters (CVC) are associated with the highest risk of intravenous catheter-related bloodstream infection (CRBSI). 3M™ Tegaderm™ CHG IV dressing is a semi-permeable transparent adhesive dressing with an integrated gel pad containing chlorhexidine gluconate 2%. This product was reviewed by the National Institute for Health and Care Excellence (NICE) in 2015, recommending that Tegaderm CHG could be used for CVC and arterial line dressings in high-dependency and intensive-care settings. This article discusses issues around CRBSI, interventions to reduce the risk of CRBSI, and the use of Tegaderm CHG dressing. PMID:26496869

  9. Getting to Zero: Goal Commitment to Reduce Blood Stream Infections.

    PubMed

    McAlearney, Ann Scheck; Hefner, Jennifer L

    2016-08-01

    While preventing health care-associated infections (HAIs) can save lives and reduce health care costs, efforts designed to eliminate HAIs have had mixed results. Variability in contextual factors such as work culture and management practices has been suggested as a potential explanation for inconsistent results across organizations and interventions. We examine goal-setting as a factor contributing to program outcomes in eight hospitals focused on preventing central line-associated bloodstream infections (CLABSIs). We conducted qualitative case studies to compare higher- and lower-performing hospitals, and explored differences in contextual factors that might contribute to performance variation. We present a goal commitment framework that characterizes factors associated with successful CLABSI program outcomes. Across 194 key informant interviews, internal and external moderators and characteristics of the goal itself differentiated actors' goal commitment at higher- versus lower-performing hospitals. Our findings have implications for organizations struggling to prevent HAIs, as well as informing the broader goal commitment literature. PMID:26589674

  10. Modeling of Virion Collisions in Cervicovaginal Mucus Reveals Limits on Agglutination as the Protective Mechanism of Secretory Immunoglobulin A

    PubMed Central

    Chen, Alex; McKinley, Scott A.; Shi, Feng; Wang, Simi; Mucha, Peter J.; Harit, Dimple; Forest, M. Gregory; Lai, Samuel K.

    2015-01-01

    Secretory immunoglobulin A (sIgA), a dimeric antibody found in high quantities in the gastrointestinal mucosa, is broadly associated with mucosal immune protection. A distinguishing feature of sIgA is its ability to crosslink pathogens, thereby creating pathogen/sIgA aggregates that are too large to traverse the dense matrix of mucin fibers in mucus layers overlying epithelial cells and consequently reducing infectivity. Here, we use modeling to investigate this mechanism of “immune exclusion” based on sIgA-mediated agglutination, in particular the potential use of sIgA to agglutinate HIV in cervicovaginal mucus (CVM) and prevent HIV transmission. Utilizing reported data on HIV diffusion in CVM and semen, we simulate HIV collision kinetics in physiologically-thick mucus layers–a necessary first step for sIgA-induced aggregation. We find that even at the median HIV load in semen of acutely infected individuals possessing high viral titers, over 99% of HIV virions will penetrate CVM and reach the vaginal epithelium without colliding with another virion. These findings imply that agglutination is unlikely to be the dominant mechanism of sIgA-mediated protection against HIV or other sexually transmitted pathogens. Rather, we surmise that agglutination is most effective against pathogens either present at exceedingly high concentrations or that possess motility mechanisms other than Brownian diffusion that significantly enhance encounter rates. PMID:26132216

  11. A Small Molecule Inhibits Virion Attachment to Heparan Sulfate- or Sialic Acid-Containing Glycans

    PubMed Central

    Colpitts, Che C.

    2014-01-01

    ABSTRACT Primary attachment to cellular glycans is a critical entry step for most human viruses. Some viruses, such as herpes simplex virus type 1 (HSV-1) and hepatitis C virus (HCV), bind to heparan sulfate, whereas others, such as influenza A virus (IAV), bind to sialic acid. Receptor mimetics that interfere with these interactions are active against viruses that bind to either heparan sulfate or to sialic acid. However, no molecule that inhibits the attachment of viruses in both groups has yet been identified. Epigallocatechin gallate (EGCG), a green tea catechin, is active against many unrelated viruses, including several that bind to heparan sulfate or to sialic acid. We sought to identify the basis for the broad-spectrum activity of EGCG. Here, we show that EGCG inhibits the infectivity of a diverse group of enveloped and nonenveloped human viruses. EGCG acts directly on the virions, without affecting the fluidity or integrity of the virion envelopes. Instead, EGCG interacts with virion surface proteins to inhibit the attachment of HSV-1, HCV, IAV, vaccinia virus, adenovirus, reovirus, and vesicular stomatitis virus (VSV) virions. We further show that EGCG competes with heparan sulfate for binding of HSV-1 and HCV virions and with sialic acid for binding of IAV virions. Therefore, EGCG inhibits unrelated viruses by a common mechanism. Most importantly, we have identified EGCG as the first broad-spectrum attachment inhibitor. Our results open the possibility for the development of small molecule broad-spectrum antivirals targeting virion attachment. IMPORTANCE This study shows that it is possible to develop a small molecule antiviral or microbicide active against the two largest groups of human viruses: those that bind to glycosaminoglycans and those that bind to sialoglycans. This group includes the vast majority of human viruses, including herpes simplex viruses, cytomegalovirus, influenza virus, poxvirus, hepatitis C virus, HIV, and many others. PMID

  12. New molecular strategies for reducing implantable medical devices associated infections.

    PubMed

    Holban, Alina Maria; Gestal, Monica Cartelle; Grumezescu, Alexandru Mihai

    2014-01-01

    Due to the great prevalence of persistent and recurrent implanted device associated-infections novel and alternative therapeutic approaches are intensely investigated. For reducing complications and antibiotic resistance development, one major strategy is using natural or synthetic modulators for targeting microbial molecular pathways which are not related with cell multiplication and death, as Quorum Sensing, virulence and biofilm formation. The purpose of this review paper is to discuss the most recent in vitro approaches, investigating the efficiency of some novel antimicrobial products and the nano-technologic progress performed in order to increase their effect and stability. PMID:24606502

  13. Management strategies to reduce risk of postoperative infections

    PubMed Central

    Galor, Anat; Goldhardt, Raquel; Wellik, Sarah R.; Gregori, Ninel Z.; Flynn, Harry W.

    2013-01-01

    Postoperative infections, although rare, are still of great concern to the ophthalmologist. The incidence of post-cataract endophthalmitis is low, with a range of .28 per 1,000 to 2.99 per 1000. In addition to intraoperative considerations such as poor wound construction, vitreous loss, topical anesthesia, and prolonged surgical time, other risk factors include preoperative factors such as a diseased ocular surface and systemic immunosuppression. Potential methods of reducing risk of endophthalmitis after anterior segment surgery are discussed and available literature is summarized. PMID:24319649

  14. Putative site for the acquisition of human herpesvirus 6 virion tegument.

    PubMed Central

    Roffman, E; Albert, J P; Goff, J P; Frenkel, N

    1990-01-01

    The virion of human herpesvirus 6 (HHV-6) contains a very distinct tegument layer, occupying the space between the nucleocapsid and the virion envelope. Ultrastructural analyses of thymocytes infected with HHV-6 revealed the presence of intranuclear spherical compartments, approximately 1.5 microns in diameter, in which tegumentation seems to take place. These compartments, termed tegusomes, were bounded by two membranes and contained ribosomes, consistent with their derivation by cytoplasmic invagination into the nucleus. Capsids located within the nucleus outside the tegusomes were all naked, while those located in the cytoplasm were uniformly tegumented. In contrast, capsids present inside the tegusomes contains teguments of variable thicknesses. In addition, nucleocapsids were documented in the process of budding into the tegusomes. We thus suggest that the tegusomes represent a cellular site in which HHV-6 virions acquire their tegument. Images PMID:2173796

  15. Expression of the highly conserved vaccinia virus E6 protein is required for virion morphogenesis

    SciTech Connect

    Resch, Wolfgang; Weisberg, Andrea S.; Moss, Bernard

    2009-04-10

    The vaccinia virus E6R gene (VACVWR062) is conserved in all members of the poxvirus family and encodes a protein associated with the mature virion. We confirmed this association and provided evidence for an internal location. An inducible mutant that conditionally expresses E6 was constructed. In the absence of inducer, plaque formation and virus production were severely inhibited in several cell lines, whereas some replication occurred in others. This difference could be due to variation in the stringency of repression, since we could not isolate a stable deletion mutant even in the more 'permissive' cells. Under non-permissive conditions, viral late proteins were synthesized but processing of core proteins was inefficient, indicative of an assembly block. Transmission electron microscopy of sections of cells infected with the mutant in the absence of inducer revealed morphogenetic defects with crescents and empty immature virions adjacent to dense inclusions of viroplasm. Mature virions were infrequent and cores appeared to have lucent centers.

  16. Structure and Assembly of TP901-1 Virion Unveiled by Mutagenesis

    PubMed Central

    Douillard, François P.; Mahony, Jennifer; Cambillau, Christian; van Sinderen, Douwe

    2015-01-01

    Bacteriophages of the Siphoviridae family represent the most abundant viral morphology in the biosphere, yet many molecular aspects of their virion structure, assembly and associated functions remain to be unveiled. In this study, we present a comprehensive mutational and molecular analysis of the temperate Lactococcus lactis-infecting phage TP901-1. Fourteen mutations located within the structural module of TP901-1 were created; twelve mutations were designed to prevent full length translation of putative proteins by non-sense mutations, while two additional mutations caused aberrant protein production. Electron microscopy and Western blot analysis of mutant virion preparations, as well as in vitro assembly of phage mutant combinations, revealed the essential nature of many of the corresponding gene products and provided information on their biological function(s). Based on the information obtained, we propose a functional and assembly model of the TP901-1 Siphoviridae virion. PMID:26147978

  17. Culture-Independent Evaluation of Nonenveloped-Virus Infectivity Reduced by Free-Chlorine Disinfection

    PubMed Central

    Ohta, Takatomo; Nakamura, Arata; Nakagomi, Toyoko; Nakagomi, Osamu; Okabe, Satoshi

    2015-01-01

    The inability of molecular detection methods to distinguish disinfected virions from infectious ones has hampered the assessment of infectivity for enteric viruses caused by disinfection practices. In the present study, the reduction of infectivity of murine norovirus S7-PP3 and mengovirus vMC0, surrogates of human noroviruses and enteroviruses, respectively, caused by free-chlorine treatment was characterized culture independently by detecting carbonyl groups on viral capsid protein. The amount of carbonyls on viral capsid protein was evaluated by the proportion of biotinylated virions trapped by avidin-immobilized gel (percent adsorbed). This culture-independent approach demonstrated that the percent adsorbed was significantly correlated with the logarithm of the infectious titer of tested viruses. Taken together with the results of previous reports, the result obtained in this study indicates that the amount of carbonyls on viral capsid protein of four important families of waterborne pathogenic viruses, Astroviridae, Reoviridae, Caliciviridae, and Picornaviridae, is increased in proportion to the received oxidative stress of free chlorine. There was also a significant correlation between the percent adsorbed and the logarithm of the ratio of genome copy number to PFU, which enables estimation of the infectious titer of a subject virus by measuring values of the total genome copy number and the percent adsorbed. The proposed method is applicable when the validation of a 4-log reduction of viruses, a requirement in U.S. EPA guidelines for virus removal from water, is needed along with clear evidence of the oxidation of virus particles with chlorine-based disinfectants. PMID:25681178

  18. Predicting first traversal times for virions and nanoparticles in mucus with slowed diffusion.

    PubMed

    Erickson, Austen M; Henry, Bruce I; Murray, John M; Klasse, Per Johan; Angstmann, Christopher N

    2015-07-01

    Particle-tracking experiments focusing on virions or nanoparticles in mucus have measured mean-square displacements and reported diffusion coefficients that are orders of magnitude smaller than the diffusion coefficients of such particles in water. Accurate description of this subdiffusion is important to properly estimate the likelihood of virions traversing the mucus boundary layer and infecting cells in the epithelium. However, there are several candidate models for diffusion that can fit experimental measurements of mean-square displacements. We show that these models yield very different estimates for the time taken for subdiffusive virions to traverse through a mucus layer. We explain why fits of subdiffusive mean-square displacements to standard diffusion models may be misleading. Relevant to human immunodeficiency virus infection, using computational methods for fractional subdiffusion, we show that subdiffusion in normal acidic mucus provides a more effective barrier against infection than previously thought. By contrast, the neutralization of the mucus by alkaline semen, after sexual intercourse, allows virions to cross the mucus layer and reach the epithelium in a short timeframe. The computed barrier protection from fractional subdiffusion is some orders of magnitude greater than that derived by fitting standard models of diffusion to subdiffusive data. PMID:26153713

  19. Predicting First Traversal Times for Virions and Nanoparticles in Mucus with Slowed Diffusion

    PubMed Central

    Erickson, Austen M.; Henry, Bruce I.; Murray, John M.; Klasse, Per Johan; Angstmann, Christopher N.

    2015-01-01

    Particle-tracking experiments focusing on virions or nanoparticles in mucus have measured mean-square displacements and reported diffusion coefficients that are orders of magnitude smaller than the diffusion coefficients of such particles in water. Accurate description of this subdiffusion is important to properly estimate the likelihood of virions traversing the mucus boundary layer and infecting cells in the epithelium. However, there are several candidate models for diffusion that can fit experimental measurements of mean-square displacements. We show that these models yield very different estimates for the time taken for subdiffusive virions to traverse through a mucus layer. We explain why fits of subdiffusive mean-square displacements to standard diffusion models may be misleading. Relevant to human immunodeficiency virus infection, using computational methods for fractional subdiffusion, we show that subdiffusion in normal acidic mucus provides a more effective barrier against infection than previously thought. By contrast, the neutralization of the mucus by alkaline semen, after sexual intercourse, allows virions to cross the mucus layer and reach the epithelium in a short timeframe. The computed barrier protection from fractional subdiffusion is some orders of magnitude greater than that derived by fitting standard models of diffusion to subdiffusive data. PMID:26153713

  20. Genetic analysis of beet curly top virus: evidence for three virion sense genes involved in movement and regulation of single- and double-stranded DNA levels.

    PubMed

    Hormuzdi, S G; Bisaro, D M

    1993-04-01

    The monopartite DNA genome of beet curly top geminivirus (BCTV, strain Logan) contains four leftward, complementary sense open reading frames (ORFs) designated L1, L2, L3, and L4 and three rightward, virion sense ORFs designated R1, R2, and R3 (R1 encodes the coat protein). The R3 ORF has not been reported previously in the BCTV genome, and evidence for three functional virion sense genes on one genome component has not been presented before for any geminivirus. We investigated the functions of the virion sense ORFs by introducing mutations into each of them. We found that in Nicotiana benthamiana plants, BCTV genomes containing mutations in ORF R1 were not infectious, whereas an R3- mutant was very weakly infectious. The small proportion of plants infected by the R3- mutant remained asymptomatic and contained greatly reduced amounts of viral DNA. An R2- mutant was highly infectious but asymptomatic, and in infected plants it accumulated mostly the double-stranded DNA (dsDNA) replicative form in nearly wild-type amounts. All of the mutants replicated in tobacco protoplasts, although R1- and R2- mutants accumulated reduced amounts of genomic single-stranded DNA (ssDNA) relative to wild-type virus. In the case of R2- mutants, the reduction was large (approx. ninefold) and was accompanied by a similar increase in dsDNA levels. The results suggest that the R1 and R3 gene products are required for efficient movement of the virus in the infected plant, whereas the R2 gene product may be involved in the regulation of ssDNA vs dsDNA levels. PMID:8460493

  1. Reducing uncertainty in managing respiratory tract infections in primary care

    PubMed Central

    Stanton, Naomi; Francis, Nick A; Butler, Chris C

    2010-01-01

    Respiratory tract infections (RTIs) remain the commonest reason for acute consultations in primary care in resource-rich countries. Their spectrum and severity has changed from the time that antibiotics were discovered, largely from improvements in the socioeconomic determinants of health as well as vaccination. The benefits from antibiotic treatment for common RTIs have been shown to be largely overstated. Nevertheless, serious infections do occur. Currently, no clinical features or diagnostic test, alone or in combination, adequately determine diagnosis, aetiology, prognosis, or response to treatment. This narrative review focuses on emerging evidence aimed at helping clinicians reduce and manage uncertainty in treating RTIs. Consultation rate and prescribing rate trends are described, evidence of increasing rates of complications are discussed, and studies and the association with antibiotic prescribing are examined. Methods of improving diagnosis and identifying those patients who are at increased risk of complications from RTIs, using clinical scoring systems, biomarkers, and point of care tests are also discussed. The evidence for alternative management options for RTIs are summarised and the methods for changing public and clinicians' beliefs about antibiotics, including ways in which we can improve clinician–patient communication skills for management of RTIs, are described. PMID:21144191

  2. Major histocompatibility complex heterozygosity reduces fitness in experimentally infected mice.

    PubMed

    Ilmonen, Petteri; Penn, Dustin J; Damjanovich, Kristy; Morrison, Linda; Ghotbi, Laleh; Potts, Wayne K

    2007-08-01

    It is often suggested that heterozygosity at major histocompatibility complex (MHC) loci confers enhanced resistance to infectious diseases (heterozygote advantage, HA, hypothesis), and overdominant selection should contribute to the evolution of these highly polymorphic genes. The evidence for the HA hypothesis is mixed and mainly from laboratory studies on inbred congenic mice, leaving the importance of MHC heterozygosity for natural populations unclear. We tested the HA hypothesis by infecting mice, produced by crossbreeding congenic C57BL/10 with wild ones, with different strains of Salmonella, both in laboratory and in large population enclosures. In the laboratory, we found that MHC influenced resistance, despite interacting wild-derived background loci. Surprisingly, resistance was mostly recessive rather than dominant, unlike in most inbred mouse strains, and it was never overdominant. In the enclosures, heterozygotes did not show better resistance, survival, or reproductive success compared to homozygotes. On the contrary, infected heterozygous females produced significantly fewer pups than homozygotes. Our results show that MHC effects are not masked on an outbred genetic background, and that MHC heterozygosity provides no immunological benefits when resistance is recessive, and can actually reduce fitness. These findings challenge the HA hypothesis and emphasize the need for studies on wild, genetically diverse species. PMID:17603099

  3. Human pDCs preferentially sense enveloped hepatitis A virions.

    PubMed

    Feng, Zongdi; Li, You; McKnight, Kevin L; Hensley, Lucinda; Lanford, Robert E; Walker, Christopher M; Lemon, Stanley M

    2015-01-01

    Unlike other picornaviruses, hepatitis A virus (HAV) is cloaked in host membranes when released from cells, providing protection from neutralizing antibodies and facilitating spread in the liver. Acute HAV infection is typified by minimal type I IFN responses; therefore, we questioned whether plasmacytoid dendritic cells (pDCs), which produce IFN when activated, are capable of sensing enveloped virions (eHAV). Although concentrated nonenveloped virus failed to activate freshly isolated human pDCs, these cells produced substantial amounts of IFN-α via TLR7 signaling when cocultured with infected cells. pDCs required either close contact with infected cells or exposure to concentrated culture supernatants for IFN-α production. In isopycnic and rate-zonal gradients, pDC-activating material cosedimented with eHAV but not membrane-bound acetylcholinesterase, suggesting that eHAV, and not viral RNA exosomes, is responsible for IFN-α induction. pDC activation did not require virus replication and was associated with efficient eHAV uptake, which was facilitated by phosphatidylserine receptors on pDCs. In chimpanzees, pDCs were transiently recruited to the liver early in infection, during or shortly before maximal intrahepatic IFN-stimulated gene expression, but disappeared prior to inflammation onset. Our data reveal that, while membrane envelopment protects HAV against neutralizing antibody, it also facilitates an early but limited detection of HAV infection by pDCs. PMID:25415438

  4. Characterization of a ubiquitinated protein which is externally located in African swine fever virions.

    PubMed Central

    Hingamp, P M; Leyland, M L; Webb, J; Twigger, S; Mayer, R J; Dixon, L K

    1995-01-01

    An antiserum was raised against the African swine fever virus (ASFV)-encoded ubiquitin-conjugating enzyme (UBCv1) and used to demonstrate by Western blotting (immunoblotting) and immunofluorescence that the enzyme is present in purified extracellular virions, is expressed both early and late after infection of cells with ASFV, and is cytoplasmically located. Antiubiquitin serum was used to identify novel ubiquitin conjugates present during ASFV infections. This antiserum stained virus factories late after infection, suggesting that virion proteins may be ubiquitinated. This possibility was confirmed by Western blotting, which identified three major antiubiquitin-immunoreactive proteins with molecular masses of 5, 18, and 58 kDa in purified extracellular virions. The 18-kDa protein was solubilized from virions at relatively low concentrations of the detergent n-octyl-beta-D-glucopyranoside, indicating that it is externally located and is possibly in the virus capsid. The 18-kDa protein was purified, and N-terminal amino acid sequencing confirmed that the protein was ubiquitinated and was ASFV encoded. The ASFV gene encoding this protein (PIG1) was sequenced, and the encoded protein expressed in an Escherichia coli expression vector. Recombinant PIG1 was ubiquitinated in the presence of E. coli expressed UBCv1 in vitro. These results suggest that PIG1 may be a substrate for UBCv1. The predicted molecular masses of the PIG1 protein and recombinant ubiquitinated protein were larger than the 18-kDa molecular mass of the ubiquitinated protein present in virions. Therefore, during viral replication, a precursor protein may undergo limited proteolysis to generate the ubiquitinated 18-kDa protein. PMID:7853518

  5. Phenotypic characterization of temperature-sensitive mutants of vaccinia virus with mutations in a 135,000-Mr subunit of the virion-associated DNA-dependent RNA polymerase.

    PubMed Central

    Ensinger, M J

    1987-01-01

    The phenotypic defects of three temperature-sensitive (ts) mutants of vaccinia virus, the ts mutations of which were mapped to the gene for one of the high-molecular-weight subunits of the virion-associated DNA-dependent RNA polymerase, were characterized. Because the virion RNA polymerase is required for the initiation of the viral replication cycle, it has been predicted that this type of mutant is defective in viral DNA replication and the synthesis of early viral proteins at the nonpermissive temperature. However, all three mutants synthesized both DNA and early proteins, and two of the three synthesized late proteins as well. RNA synthesis in vitro by permeabilized mutant virions was not more ts than that by the wild type. Furthermore, only one of three RNA polymerase activities that was partially purified from virions assembled at the permissive temperature displayed altered biochemical properties in vitro that could be correlated with its ts mutation: the ts13 activity had reduced specific activity, increased temperature sensitivity, and increased thermolability under a variety of preincubation conditions. Although the partially purified polymerase activity of a second mutant, ts72, was also more thermolabile than the wild-type activity, the thermolability was shown to be the result of a second mutation within the RNA polymerase gene. These results suggest that the defects in these mutants affect the assembly of newly synthesized polymerase subunits into active enzyme or the incorporation of RNA polymerase into maturing virions; once synthesized at the permissive temperature, the mutant polymerases are able to function in the initiation of subsequent rounds of infection at the nonpermissive temperature. Images PMID:3573151

  6. Antibody-Mediated Internalization of Infectious HIV-1 Virions Differs among Antibody Isotypes and Subclasses.

    PubMed

    Tay, Matthew Zirui; Liu, Pinghuang; Williams, LaTonya D; McRaven, Michael D; Sawant, Sheetal; Gurley, Thaddeus C; Xu, Thomas T; Dennison, S Moses; Liao, Hua-Xin; Chenine, Agnès-Laurence; Alam, S Munir; Moody, M Anthony; Hope, Thomas J; Haynes, Barton F; Tomaras, Georgia D

    2016-08-01

    Emerging data support a role for antibody Fc-mediated antiviral activity in vaccine efficacy and in the control of HIV-1 replication by broadly neutralizing antibodies. Antibody-mediated virus internalization is an Fc-mediated function that may act at the portal of entry whereby effector cells may be triggered by pre-existing antibodies to prevent HIV-1 acquisition. Understanding the capacity of HIV-1 antibodies in mediating internalization of HIV-1 virions by primary monocytes is critical to understanding their full antiviral potency. Antibody isotypes/subclasses differ in functional profile, with consequences for their antiviral activity. For instance, in the RV144 vaccine trial that achieved partial efficacy, Env IgA correlated with increased risk of HIV-1 infection (i.e. decreased vaccine efficacy), whereas V1-V2 IgG3 correlated with decreased risk of HIV-1 infection (i.e. increased vaccine efficacy). Thus, understanding the different functional attributes of HIV-1 specific IgG1, IgG3 and IgA antibodies will help define the mechanisms of immune protection. Here, we utilized an in vitro flow cytometric method utilizing primary monocytes as phagocytes and infectious HIV-1 virions as targets to determine the capacity of Env IgA (IgA1, IgA2), IgG1 and IgG3 antibodies to mediate HIV-1 infectious virion internalization. Importantly, both broadly neutralizing antibodies (i.e. PG9, 2G12, CH31, VRC01 IgG) and non-broadly neutralizing antibodies (i.e. 7B2 mAb, mucosal HIV-1+ IgG) mediated internalization of HIV-1 virions. Furthermore, we found that Env IgG3 of multiple specificities (i.e. CD4bs, V1-V2 and gp41) mediated increased infectious virion internalization over Env IgG1 of the same specificity, while Env IgA mediated decreased infectious virion internalization compared to IgG1. These data demonstrate that antibody-mediated internalization of HIV-1 virions depends on antibody specificity and isotype. Evaluation of the phagocytic potency of vaccine

  7. AABB Committee Report: reducing transfusion-transmitted cytomegalovirus infections.

    PubMed

    Heddle, Nancy M; Boeckh, Michael; Grossman, Brenda; Jacobson, Jessica; Kleinman, Steven; Tobian, Aaron A R; Webert, Kathryn; Wong, Edward C C; Roback, John D

    2016-06-01

    Transfusion-transmitted cytomegalovirus (TT-CMV) is often asymptomatic, but certain patient populations, such as very low birth weight neonates, fetuses requiring intrauterine transfusion, pregnant women, patients with primary immunodeficiencies, transplant recipients, and patients receiving chemotherapy or transplantation for malignant disease, may be at risk of life-threatening CMV infection. It is unclear whether leukoreduction of cellular blood components is sufficient to reduce TT-CMV or whether CMV serological testing adds additional benefit to leukoreduction. The AABB CMV Prevention Work Group commissioned a systematic review to address these issues and subsequently develop clinical practice guidelines. However, the data were of poor quality, and no studies of significant size have been performed for over a decade. Rather than creating guidelines of questionable utility, the Work Group (with approval of the AABB Board of Directors) voted to prepare this Committee Report. There is wide variation in practices of using leukoreduced components alone or combining CMV-serology and leukoreduction to prevent TT-CMV for at-risk patients. Other approaches may also be feasible to prevent TT-CMV, including plasma nucleic acid testing, pathogen inactivation, and patient blood management programs to reduce the frequency of inappropriate transfusions. It is unlikely that future large-scale clinical trials will be performed to determine whether leukoreduction, CMV-serology, or a combination of both is superior. Consequently, alternative strategies including pragmatic randomized controlled trials, registries, and collaborations for electronic data merging, nontraditional approaches to inform evidence, or development of a systematic approach to inform expert opinion may help to address the issue of CMV-safe blood components. PMID:26968400

  8. Structural Lability of Barley Stripe Mosaic Virus Virions

    PubMed Central

    Semenyuk, Pavel I.; Abashkin, Dmitry A.; Kalinina, Natalya O.; Arutyunyan, Alexsandr M.; Solovyev, Andrey G.; Dobrov, Eugeny N.

    2013-01-01

    Virions of Barley stripe mosaic virus (BSMV) were neglected for more than thirty years after their basic properties were determined. In this paper, the physicochemical characteristics of BSMV virions and virion-derived viral capsid protein (CP) were analyzed, namely, the absorption and intrinsic fluorescence spectra, circular dichroism spectra, differential scanning calorimetry curves, and size distributions by dynamic laser light scattering. The structural properties of BSMV virions proved to be intermediate between those of Tobacco mosaic virus (TMV), a well-characterized virus with rigid rod-shaped virions, and flexuous filamentous plant viruses. The BSMV virions were found to be considerably more labile than expected from their rod-like morphology and a distant sequence relation of the BSMV and TMV CPs. The circular dichroism spectra of BSMV CP subunits incorporated into the virions, but not subunits of free CP, demonstrated a significant proportion of beta-structure elements, which were proposed to be localized mostly in the protein regions exposed on the virion outer surface. These beta-structure elements likely formed during virion assembly can comprise the N- and C-terminal protein regions unstructured in the non-virion CP and can mediate inter-subunit interactions. Based on computer-assisted structure modeling, a model for BSMV CP subunit structural fold compliant with the available experimental data was proposed. PMID:23613760

  9. Human Immunodeficiency Virus Type 1 Nef protein modulates the lipid composition of virions and host cell membrane microdomains

    PubMed Central

    Brügger, Britta; Krautkrämer, Ellen; Tibroni, Nadine; Munte, Claudia E; Rauch, Susanne; Leibrecht, Iris; Glass, Bärbel; Breuer, Sebastian; Geyer, Matthias; Kräusslich, Hans-Georg; Kalbitzer, Hans Robert; Wieland, Felix T; Fackler, Oliver T

    2007-01-01

    Background The Nef protein of Human Immunodeficiency Viruses optimizes viral spread in the infected host by manipulating cellular transport and signal transduction machineries. Nef also boosts the infectivity of HIV particles by an unknown mechanism. Recent studies suggested a correlation between the association of Nef with lipid raft microdomains and its positive effects on virion infectivity. Furthermore, the lipidome analysis of HIV-1 particles revealed a marked enrichment of classical raft lipids and thus identified HIV-1 virions as an example for naturally occurring membrane microdomains. Since Nef modulates the protein composition and function of membrane microdomains we tested here if Nef also has the propensity to alter microdomain lipid composition. Results Quantitative mass spectrometric lipidome analysis of highly purified HIV-1 particles revealed that the presence of Nef during virus production from T lymphocytes enforced their raft character via a significant reduction of polyunsaturated phosphatidylcholine species and a specific enrichment of sphingomyelin. In contrast, Nef did not significantly affect virion levels of phosphoglycerolipids or cholesterol. The observed alterations in virion lipid composition were insufficient to mediate Nef's effect on particle infectivity and Nef augmented virion infectivity independently of whether virus entry was targeted to or excluded from membrane microdomains. However, altered lipid compositions similar to those observed in virions were also detected in detergent-resistant membrane preparations of virus producing cells. Conclusion Nef alters not only the proteome but also the lipid composition of host cell microdomains. This novel activity represents a previously unrecognized mechanism by which Nef could manipulate HIV-1 target cells to facilitate virus propagation in vivo. PMID:17908312

  10. Autographa californica multiple nucleopolyhedrovirus ac66 is required for the efficient egress of nucleocapsids from the nucleus, general synthesis of preoccluded virions and occlusion body formation

    SciTech Connect

    Ke Jianhao Wang Jinwen; Deng Riqiang; Wang Xunzhang

    2008-05-10

    Although orf66 (ac66) of Autographa californica multiple nucleopolyhedrovirus (AcMNPV) is conserved in all sequenced lepidopteran baculovirus genomes, its function is not known. This paper describes generation of an ac66 knockout AcMNPV bacmid mutant and analyses of the influence of ac66 deletion on the virus replication in Sf-9 cells so as to determine the role of ac66 in the viral life cycle. Results indicated that budded virus (BV) yields were reduced over 99% in ac66-null mutant infected cells in comparison to that in wild-type virus infected cells. Optical microscopy revealed that occlusion body synthesis was significantly reduced in the ac66 knockout bacmid-transfected cells. In addition, ac66 deletion interrupted preoccluded virion synthesis. The mutant phenotype was rescued by an ac66 repair bacmid. On the other hand, real-time PCR analysis indicated that ac66 deletion did not affect the levels of viral DNA replication. Electron microscopy revealed that ac66 is not essential for nucleocapsid assembly, but for the efficient transport of nucleocapsids from the nucleus to the cytoplasm. These results suggested that ac66 plays an important role for the efficient exit of nucleocapsids from the nucleus to the cytoplasm for BV synthesis as well as for preoccluded virion and occlusion synthesis.

  11. Hepatitis C Virus Envelope Glycoprotein E1 Forms Trimers at the Surface of the Virion

    PubMed Central

    Falson, Pierre; Bartosch, Birke; Alsaleh, Khaled; Tews, Birke Andrea; Loquet, Antoine; Ciczora, Yann; Riva, Laura; Montigny, Cédric; Montpellier, Claire; Duverlie, Gilles; Pécheur, Eve-Isabelle; le Maire, Marc; Cosset, François-Loïc

    2015-01-01

    ABSTRACT In hepatitis C virus (HCV)-infected cells, the envelope glycoproteins E1 and E2 assemble as a heterodimer. To investigate potential changes in the oligomerization of virion-associated envelope proteins, we performed SDS-PAGE under reducing conditions but without thermal denaturation. This revealed the presence of SDS-resistant trimers of E1 in the context of cell-cultured HCV (HCVcc) as well as in the context of HCV pseudoparticles (HCVpp). The formation of E1 trimers was found to depend on the coexpression of E2. To further understand the origin of E1 trimer formation, we coexpressed in bacteria the transmembrane (TM) domains of E1 (TME1) and E2 (TME2) fused to reporter proteins and analyzed the fusion proteins by SDS-PAGE and Western blotting. As expected for strongly interacting TM domains, TME1–TME2 heterodimers resistant to SDS were observed. These analyses also revealed homodimers and homotrimers of TME1, indicating that such complexes are stable species. The N-terminal segment of TME1 exhibits a highly conserved GxxxG sequence, a motif that is well documented to be involved in intramembrane protein-protein interactions. Single or double mutations of the glycine residues (Gly354 and Gly358) in this motif markedly decreased or abrogated the formation of TME1 homotrimers in bacteria, as well as homotrimers of E1 in both HCVpp and HCVcc systems. A concomitant loss of infectivity was observed, indicating that the trimeric form of E1 is essential for virus infectivity. Taken together, these results indicate that E1E2 heterodimers form trimers on HCV particles, and they support the hypothesis that E1 could be a fusion protein. IMPORTANCE HCV glycoproteins E1 and E2 play an essential role in virus entry into liver cells as well as in virion morphogenesis. In infected cells, these two proteins form a complex in which E2 interacts with cellular receptors, whereas the function of E1 remains poorly understood. However, recent structural data suggest that E1

  12. Offset layered closure reduces deep wound infection in early-onset scoliosis surgery.

    PubMed

    Grzywna, Alexandra M; Miller, Patricia E; Glotzbecker, Michael P; Emans, John B

    2016-07-01

    Deep wound infection is a common complication in early-onset scoliosis (EOS) surgery. Soft tissue technique has received less attention as a means to reduce infection. A retrospective review of 1170 EOS surgeries (single surgeon, institution) investigated the impact of offset layered closure (OLC) and soft tissue awareness. The introduction of OLC reduced deep infection from 3.0% in 99 surgeries to 0.37% in 1071. Logistic regression confirmed that OLC led to significantly lower odds of infection (P=0.007). This deep infection rate (0.37%) is more typical of elective surgery, providing a more optimistic view of infection in EOS surgery than generally reported. PMID:27196268

  13. Natural malaria infection reduces starvation resistance of nutritionally stressed mosquitoes.

    PubMed

    Lalubin, Fabrice; Delédevant, Aline; Glaizot, Olivier; Christe, Philippe

    2014-07-01

    In disease ecology, there is growing evidence that environmental quality interacts with parasite and host to determine host susceptibility to an infection. Most studies of malaria parasites have focused on the infection costs incurred by the hosts, and few have investigated the costs on mosquito vectors. The interplay between the environment, the vector and the parasite has therefore mostly been ignored and often relied on unnatural or allopatric Plasmodium/vector associations. Here, we investigated the effects of natural avian malaria infection on both fecundity and survival of field-caught female Culex pipiens mosquitoes, individually maintained in laboratory conditions. We manipulated environmental quality by providing mosquitoes with different concentrations of glucose-feeding solution prior to submitting them to a starvation challenge. We used molecular-based methods to assess mosquitoes' infection status. We found that mosquitoes infected with Plasmodium had lower starvation resistance than uninfected ones only under low nutritional conditions. The effect of nutritional stress varied with time, with the difference of starvation resistance between optimally and suboptimally fed mosquitoes increasing from spring to summer, as shown by a significant interaction between diet treatment and months of capture. Infected and uninfected mosquitoes had similar clutch size, indicating no effect of infection on fecundity. Overall, this study suggests that avian malaria vectors may suffer Plasmodium infection costs in their natural habitat, under certain environmental conditions. This may have major implications for disease transmission in the wild. PMID:24286465

  14. The ns12.9 Accessory Protein of Human Coronavirus OC43 Is a Viroporin Involved in Virion Morphogenesis and Pathogenesis

    PubMed Central

    Zhang, Ronghua; Wang, Kai; Ping, Xianqiang; Yu, Wenjing

    2015-01-01

    ABSTRACT An accessory gene between the S and E gene loci is contained in all coronaviruses (CoVs), and its function has been studied in some coronaviruses. This gene locus in human coronavirus OC43 (HCoV-OC43) encodes the ns12.9 accessory protein; however, its function during viral infection remains unknown. Here, we engineered a recombinant mutant virus lacking the ns12.9 protein (HCoV-OC43-Δns12.9) to characterize the contributions of ns12.9 in HCoV-OC43 replication. The ns12.9 accessory protein is a transmembrane protein and forms ion channels in both Xenopus oocytes and yeast through homo-oligomerization, suggesting that ns12.9 is a newly recognized viroporin. HCoV-OC43-Δns12.9 presented at least 10-fold reduction of viral titer in vitro and in vivo. Intriguingly, exogenous ns12.9 and heterologous viroporins with ion channel activity could compensate for the production of HCoV-OC43-Δns12.9, indicating that the ion channel activity of ns12.9 plays a significant role in the production of infectious virions. Systematic dissection of single-cycle replication revealed that ns12.9 protein had no measurable effect on virus entry, subgenomic mRNA (sgmRNA) synthesis, and protein expression. Further characterization revealed that HCoV-OC43-Δns12.9 was less efficient in virion morphogenesis than recombinant wild-type virus (HCoV-OC43-WT). Moreover, reduced viral replication, inflammatory response, and virulence in HCoV-OC43-Δns12.9-infected mice were observed compared to the levels for HCoV-OC43-WT-infected mice. Taken together, our results demonstrated that the ns12.9 accessory protein functions as a viroporin and is involved in virion morphogenesis and the pathogenesis of HCoV-OC43 infection. IMPORTANCE HCoV-OC43 was isolated in the 1960s and is a major agent of the common cold. The functions of HCoV-OC43 structural proteins have been well studied, but few studies have focused on its accessory proteins. In the present study, we demonstrated that the ns12.9 protein

  15. Rhadinovirus Host Entry by Co-operative Infection

    PubMed Central

    May, Janet S.; Stevenson, Philip G.

    2015-01-01

    Rhadinoviruses establish chronic infections of clinical and economic importance. Several show respiratory transmission and cause lung pathologies. We used Murid Herpesvirus-4 (MuHV-4) to understand how rhadinovirus lung infection might work. A primary epithelial or B cell infection often is assumed. MuHV-4 targeted instead alveolar macrophages, and their depletion reduced markedly host entry. While host entry was efficient, alveolar macrophages lacked heparan - an important rhadinovirus binding target - and were infected poorly ex vivo. In situ analysis revealed that virions bound initially not to macrophages but to heparan+ type 1 alveolar epithelial cells (AECs). Although epithelial cell lines endocytose MuHV-4 readily in vitro, AECs did not. Rather bound virions were acquired by macrophages; epithelial infection occurred only later. Thus, host entry was co-operative - virion binding to epithelial cells licensed macrophage infection, and this in turn licensed AEC infection. An antibody block of epithelial cell binding failed to block host entry: opsonization provided merely another route to macrophages. By contrast an antibody block of membrane fusion was effective. Therefore co-operative infection extended viral tropism beyond the normal paradigm of a target cell infected readily in vitro; and macrophage involvement in host entry required neutralization to act down-stream of cell binding. PMID:25790477

  16. Characterization of the Determinants of NS2-3-Independent Virion Morphogenesis of Pestiviruses

    PubMed Central

    Klemens, O.; Dubrau, D.

    2015-01-01

    diarrhea virus, nonstructural protein NS2-3 is of critical importance to switch between these processes. While free NS3 is essential for RNA replication, uncleaved NS2-3, which accumulates over time in the infected cell, is required for virion morphogenesis. In contrast, the virion morphogenesis of the related hepatitis C virus is independent from uncleaved NS2-NS3. Here, we demonstrate that pestiviruses can adapt to virion morphogenesis in the absence of uncleaved NS2-3 by just two amino acid exchanges. While the mechanism behind this gain of function remains elusive, the fact that it can be achieved by such minor changes is in line with the assumption that an ancestral virus already used this mechanism but lost it in the course of adapting to a new host/infection strategy. PMID:26355097

  17. Multifunctional roles for the N-terminal basic motif of Alfalfa mosaic virus coat protein: nucleolar/cytoplasmic shuttling, modulation of RNA-binding activity, and virion formation.

    PubMed

    Herranz, Mari Carmen; Pallas, Vicente; Aparicio, Frederic

    2012-08-01

    In addition to virion formation, the coat protein (CP) of Alfalfa mosaic virus (AMV) is involved in the regulation of replication and translation of viral RNAs, and in cell-to-cell and systemic movement of the virus. An intriguing feature of the AMV CP is its nuclear and nucleolar accumulation. Here, we identify an N-terminal lysine-rich nucleolar localization signal (NoLS) in the AMV CP required to both enter the nucleus and accumulate in the nucleolus of infected cells, and a C-terminal leucine-rich domain which might function as a nuclear export signal. Moreover, we demonstrate that AMV CP interacts with importin-α, a component of the classical nuclear import pathway. A mutant AMV RNA 3 unable to target the nucleolus exhibited reduced plus-strand RNA synthesis and cell-to-cell spread. Moreover, virion formation and systemic movement were completely abolished in plants infected with this mutant. In vitro analysis demonstrated that specific lysine residues within the NoLS are also involved in modulating CP-RNA binding and CP dimerization, suggesting that the NoLS represents a multifunctional domain within the AMV CP. The observation that nuclear and nucleolar import signals mask RNA-binding properties of AMV CP, essential for viral replication and translation, supports a model in which viral expression is carefully modulated by a cytoplasmic/nuclear balance of CP accumulation. PMID:22746826

  18. UV-Sensitivity of Shiga Toxin-Converting Bacteriophage Virions Φ24B, 933W, P22, P27 and P32.

    PubMed

    Bloch, Sylwia; Nejman-Faleńczyk, Bożena; Topka, Gracja; Dydecka, Aleksandra; Licznerska, Katarzyna; Narajczyk, Magdalena; Necel, Agnieszka; Węgrzyn, Alicja; Węgrzyn, Grzegorz

    2015-09-01

    Shiga toxin-converting bacteriophages (Stx phages) are present as prophages in Shiga toxin-producing Escherichia coli (STEC) strains. Theses phages can be transmitted to previously non-pathogenic E. coli cells making them potential producers of Shiga toxins, as they bear genes for these toxins in their genomes. Therefore, sensitivity of Stx phage virions to various conditions is important in both natural processes of spreading of these viruses and potential prophylactic control of appearance of novel pathogenic E. coli strains. In this report we provide evidence that virions of Stx phages are significantly more sensitive to UV irradiation than bacteriophage λ. Following UV irradiation of Stx virions at the dose of 50 J/m², their infectivity dropped by 1-3 log10, depending on the kind of phage. Under these conditions, a considerable release of phage DNA from virions was observed, and electron microscopy analyses indicated a large proportion of partially damaged virions. Infection of E. coli cells with UV-irradiated Stx phages resulted in significantly decreased levels of expression of N and cro genes, crucial for lytic development. We conclude that inactivation of Stx virions caused by relatively low dose of UV light is due to damage of capsids that prevents effective infection of the host cells. PMID:26402701

  19. UV-Sensitivity of Shiga Toxin-Converting Bacteriophage Virions Φ24B, 933W, P22, P27 and P32

    PubMed Central

    Bloch, Sylwia; Nejman-Faleńczyk, Bożena; Topka, Gracja; Dydecka, Aleksandra; Licznerska, Katarzyna; Narajczyk, Magdalena; Necel, Agnieszka; Węgrzyn, Alicja; Węgrzyn, Grzegorz

    2015-01-01

    Shiga toxin-converting bacteriophages (Stx phages) are present as prophages in Shiga toxin-producing Escherichia coli (STEC) strains. Theses phages can be transmitted to previously non-pathogenic E. coli cells making them potential producers of Shiga toxins, as they bear genes for these toxins in their genomes. Therefore, sensitivity of Stx phage virions to various conditions is important in both natural processes of spreading of these viruses and potential prophylactic control of appearance of novel pathogenic E. coli strains. In this report we provide evidence that virions of Stx phages are significantly more sensitive to UV irradiation than bacteriophage λ. Following UV irradiation of Stx virions at the dose of 50 J/m2, their infectivity dropped by 1–3 log10, depending on the kind of phage. Under these conditions, a considerable release of phage DNA from virions was observed, and electron microscopy analyses indicated a large proportion of partially damaged virions. Infection of E. coli cells with UV-irradiated Stx phages resulted in significantly decreased levels of expression of N and cro genes, crucial for lytic development. We conclude that inactivation of Stx virions caused by relatively low dose of UV light is due to damage of capsids that prevents effective infection of the host cells. PMID:26402701

  20. Use of information to reduce the risk of OPP infection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This presentation reviewed maternal and non-maternal routes of exposure to ovine progressive pneumonia virus (OPPV), their relative impact on transmission, and factors that affect infection rates. Results of an experiment to investigate additive and dominance effects of the two most common haplotyp...

  1. Virion Glycoprotein-Mediated Immune Evasion by Human Cytomegalovirus: a Sticky Virus Makes a Slick Getaway.

    PubMed

    Gardner, Thomas J; Tortorella, Domenico

    2016-09-01

    The prototypic herpesvirus human cytomegalovirus (CMV) exhibits the extraordinary ability to establish latency and maintain a chronic infection throughout the life of its human host. This is even more remarkable considering the robust adaptive immune response elicited by infection and reactivation from latency. In addition to the ability of CMV to exist in a quiescent latent state, its persistence is enabled by a large repertoire of viral proteins that subvert immune defense mechanisms, such as NK cell activation and major histocompatibility complex antigen presentation, within the cell. However, dissemination outside the cell presents a unique existential challenge to the CMV virion, which is studded with antigenic glycoprotein complexes targeted by a potent neutralizing antibody response. The CMV virion envelope proteins, which are critical mediators of cell attachment and entry, possess various characteristics that can mitigate the humoral immune response and prevent viral clearance. Here we review the CMV glycoprotein complexes crucial for cell attachment and entry and propose inherent properties of these proteins involved in evading the CMV humoral immune response. These include viral glycoprotein polymorphism, epitope competition, Fc receptor-mediated endocytosis, glycan shielding, and cell-to-cell spread. The consequences of CMV virion glycoprotein-mediated immune evasion have a major impact on persistence of the virus in the population, and a comprehensive understanding of these evasion strategies will assist in designing effective CMV biologics and vaccines to limit CMV-associated disease. PMID:27307580

  2. Experimental evidence for proteins constituting virion components and particle morphogenesis of bacteriophage ZF40.

    PubMed

    Korol, Natalia; Van den Bossche, An; Romaniuk, Liudmyla; Noben, Jean-Paul; Lavigne, Rob; Tovkach, Fedor

    2016-03-01

    Bacteriophage ZF40 is the only currently available, temperate Myoviridae phage infecting the potato pathogen Pectobacterium carotovorum subsp. carotovorum. Despite its unusual tail morphology, its major tail sheath and tube proteins remained uncharacterized after the initial genome annotation. Using ESI tandem mass-spectrometry, 24 structural proteins of the ZF40 virion were identified, with a sequence coverage ranging between 15.8% and 87.8%. The putative function of 16 proteins could be elucidated based on secondary structure analysis and conservative domain searches. The experimental annotation of 35% of the encoded gene products within the structural region of the genome represents a complete view of the virion structure, which can serve as the basis for future structural analysis as a model phage. PMID:26887841

  3. Virion Structure of Israeli Acute Bee Paralysis Virus

    PubMed Central

    Mullapudi, Edukondalu; Přidal, Antonín; Pálková, Lenka; de Miranda, Joachim R.

    2016-01-01

    ABSTRACT The pollination services provided by the western honeybee (Apis mellifera) are critical for agricultural production and the diversity of wild flowering plants. However, honeybees suffer from environmental pollution, habitat loss, and pathogens, including viruses that can cause fatal diseases. Israeli acute bee paralysis virus (IAPV), from the family Dicistroviridae, has been shown to cause colony collapse disorder in the United States. Here, we present the IAPV virion structure determined to a resolution of 4.0 Å and the structure of a pentamer of capsid protein protomers at a resolution of 2.7 Å. IAPV has major capsid proteins VP1 and VP3 with noncanonical jellyroll β-barrel folds composed of only seven instead of eight β-strands, as is the rule for proteins of other viruses with the same fold. The maturation of dicistroviruses is connected to the cleavage of precursor capsid protein VP0 into subunits VP3 and VP4. We show that a putative catalytic site formed by the residues Asp-Asp-Phe of VP1 is optimally positioned to perform the cleavage. Furthermore, unlike many picornaviruses, IAPV does not contain a hydrophobic pocket in capsid protein VP1 that could be targeted by capsid-binding antiviral compounds. IMPORTANCE Honeybee pollination is required for agricultural production and to sustain the biodiversity of wild flora. However, honeybee populations in Europe and North America are under pressure from pathogens, including viruses that cause colony losses. Viruses from the family Dicistroviridae can cause honeybee infections that are lethal, not only to individual honeybees, but to whole colonies. Here, we present the virion structure of an Aparavirus, Israeli acute bee paralysis virus (IAPV), a member of a complex of closely related viruses that are distributed worldwide. IAPV exhibits unique structural features not observed in other picorna-like viruses. Capsid protein VP1 of IAPV does not contain a hydrophobic pocket, implying that capsid

  4. Foodborne disease prevention and broiler chickens with reduced Campylobacter infection.

    PubMed

    Bahrndorff, Simon; Rangstrup-Christensen, Lena; Nordentoft, Steen; Hald, Birthe

    2013-03-01

    Studies have suggested that flies play a linking role in the epidemiology of Campylobacter spp. in broiler chickens and that fly screens can reduce the prevalence of Campylobacter spp. We examined the year-round and long-term effects of fly screens in 10 broiler chicken houses (99 flocks) in Denmark. Prevalence of Campylobacter spp.-positive flocks was significantly reduced, from 41.4% during 2003-2005 (before fly screens) to 10.3% in 2006-2009 (with fly screens). In fly screen houses, Campylobacter spp. prevalence did not peak during the summer. Nationally, prevalence of Campylobacter spp.-positive flocks in Denmark could have been reduced by an estimated 77% during summer had fly screens been part of biosecurity practices. These results imply that fly screens might help reduce prevalence of campylobacteriosis among humans, which is closely linked to Campylobacter spp. prevalence among broiler chicken flocks. PMID:23628089

  5. The Sheep Tetherin Paralog oBST2B Blocks Envelope Glycoprotein Incorporation into Nascent Retroviral Virions

    PubMed Central

    Murphy, Lita; Varela, Mariana; Desloire, Sophie; Ftaich, Najate; Murgia, Claudio; Golder, Matthew; Neil, Stuart; Spencer, Thomas E.; Wootton, Sarah K.; Lavillette, Dimitri; Terzian, Christophe; Palmarini, Massimo

    2014-01-01

    ABSTRACT Bone marrow stromal cell antigen 2 (BST2) is a cellular restriction factor with a broad antiviral activity. In sheep, the BST2 gene is duplicated into two paralogs termed oBST2A and oBST2B. oBST2A impedes viral exit of the Jaagsiekte sheep retroviruses (JSRV), most probably by retaining virions at the cell membrane, similar to the “tethering” mechanism exerted by human BST2. In this study, we provide evidence that unlike oBST2A, oBST2B is limited to the Golgi apparatus and disrupts JSRV envelope (Env) trafficking by sequestering it. In turn, oBST2B leads to a reduction in Env incorporation into viral particles, which ultimately results in the release of virions that are less infectious. Furthermore, the activity of oBST2B does not seem to be restricted to retroviruses, as it also acts on vesicular stomatitis virus glycoproteins. Therefore, we suggest that oBST2B exerts antiviral activity using a mechanism distinct from the classical tethering restriction observed for oBST2A. IMPORTANCE BST2 is a powerful cellular restriction factor against a wide range of enveloped viruses. Sheep possess two paralogs of the BST2 gene called oBST2A and oBST2B. JSRV, the causative agent of a transmissible lung cancer of sheep, is known to be restricted by oBST2A. In this study, we show that unlike oBST2A, oBST2B impairs the normal cellular trafficking of JSRV envelope glycoproteins by sequestering them within the Golgi apparatus. We also show that oBST2B decreases the incorporation of envelope glycoprotein into JSRV viral particles, which in turn reduces virion infectivity. In conclusion, oBST2B exerts a novel antiviral activity that is distinct from those of BST2 proteins of other species. PMID:25339764

  6. Ebola virion attachment and entry into human macrophages profoundly effects early cellular gene expression.

    PubMed

    Wahl-Jensen, Victoria; Kurz, Sabine; Feldmann, Friedericke; Buehler, Lukas K; Kindrachuk, Jason; DeFilippis, Victor; da Silva Correia, Jean; Früh, Klaus; Kuhn, Jens H; Burton, Dennis R; Feldmann, Heinz

    2011-10-01

    Zaire ebolavirus (ZEBOV) infections are associated with high lethality in primates. ZEBOV primarily targets mononuclear phagocytes, which are activated upon infection and secrete mediators believed to trigger initial stages of pathogenesis. The characterization of the responses of target cells to ZEBOV infection may therefore not only further understanding of pathogenesis but also suggest possible points of therapeutic intervention. Gene expression profiles of primary human macrophages exposed to ZEBOV were determined using DNA microarrays and quantitative PCR to gain insight into the cellular response immediately after cell entry. Significant changes in mRNA concentrations encoding for 88 cellular proteins were observed. Most of these proteins have not yet been implicated in ZEBOV infection. Some, however, are inflammatory mediators known to be elevated during the acute phase of disease in the blood of ZEBOV-infected humans. Interestingly, the cellular response occurred within the first hour of Ebola virion exposure, i.e. prior to virus gene expression. This observation supports the hypothesis that virion binding or entry mediated by the spike glycoprotein (GP(1,2)) is the primary stimulus for an initial response. Indeed, ZEBOV virions, LPS, and virus-like particles consisting of only the ZEBOV matrix protein VP40 and GP(1,2) (VLP(VP40-GP)) triggered comparable responses in macrophages, including pro-inflammatory and pro-apoptotic signals. In contrast, VLP(VP40) (particles lacking GP(1,2)) caused an aberrant response. This suggests that GP(1,2) binding to macrophages plays an important role in the immediate cellular response. PMID:22028943

  7. Vaccinia Virus Extracellular Enveloped Virion Neutralization In Vitro and Protection In Vivo Depend on Complement▿

    PubMed Central

    Benhnia, Mohammed Rafii-El-Idrissi; McCausland, Megan M.; Moyron, Juan; Laudenslager, John; Granger, Steven; Rickert, Sandra; Koriazova, Lilia; Kubo, Ralph; Kato, Shinichiro; Crotty, Shane

    2009-01-01

    Antibody neutralization is an important component of protective immunity against vaccinia virus (VACV). Two distinct virion forms, mature virion and enveloped virion (MV and EV, respectively), possess separate functions and nonoverlapping immunological properties. In this study we examined the mechanics of EV neutralization, focusing on EV protein B5 (also called B5R). We show that neutralization of EV is predominantly complement dependent. From a panel of high-affinity anti-B5 monoclonal antibodies (MAbs), the only potent neutralizer in vitro (90% at 535 ng/ml) was an immunoglobulin G2a (IgG2a), and neutralization was complement mediated. This MAb was the most protective in vivo against lethal intranasal VACV challenge. Further studies demonstrated that in vivo depletion of complement caused a >50% loss of anti-B5 IgG2a protection, directly establishing the importance of complement for protection against the EV form. However, the mechanism of protection is not sterilizing immunity via elimination of the inoculum as the viral inoculum consisted of a purified MV form. The prevention of illness in vivo indicated rapid control of infection. We further demonstrate that antibody-mediated killing of VACV-infected cells expressing surface B5 is a second protective mechanism provided by complement-fixing anti-B5 IgG. Cell killing was very efficient, and this effector function was highly isotype specific. These results indicate that anti-B5 antibody-directed cell lysis via complement is a powerful mechanism for clearance of infected cells, keeping poxvirus-infected cells from being invisible to humoral immune responses. These findings highlight the importance of multiple mechanisms of antibody-mediated protection against VACV and point to key immunobiological differences between MVs and EVs that impact the outcome of infection. PMID:19019965

  8. A collaborative approach to reduce healthcare-associated infections.

    PubMed

    Su, Guizhen

    Healthcare-associated infections (HAIs) continue to be an ongoing issue for patients in acute hospital settings. Effectively preventing and controlling HAIs requires a collaborative approach compelling all healthcare staff to take up responsibilities and be involved. A surgical ward in an acute hospital aimed to implement comprehensive HAI prevention strategies by applying both Kotter's eight-step change model and the practice development principles into its current system. The project motivated staff to be involved and engaged in the assessment, implementation and evaluation of the project processes, and take ownership of the practice change. It focused on ensuring a clean environment, improving hand hygiene compliance, increasing staff's knowledge base regarding HAIs and enhancing active surveillance. The project achieved success in the reduction and prevention of HAIs as well as the development of a sustainable workplace culture. PMID:27281587

  9. How malaria merozoites reduce the deformability of infected RBC

    NASA Astrophysics Data System (ADS)

    Hosseini, Majid; Feng, James

    2011-11-01

    This talk presents a three-dimensional particle-based model for the red blood cell (RBC), and uses it to explore the changes in the deformability of RBC due to presence of malaria parasite. The cell membrane is represented by a set of discrete particles connected by nonlinear springs that represent shear and bending elasticity. The cytoplasm and the external liquid are modeled as homogeneous Newtonian fluids, and discretized by particles as in standard smoothed-particle-hydrodynamics models. The merozoite is modeled as an aggregate of particles constrained to rigid-body motion. The fluid flow and membrane deformation are computed, via the particle motion, by a two-step explicit scheme, with model parameters determined from experiments. The stretching of healthy and infected RBC by optical tweezers has been simulated to investigate the contribution of rigid merozoites to the decrease in deformability. Department of Mathematics, University of British Columbia, Vancouver, BC V6T 1Z2, Canada.

  10. The Human Cytomegalovirus-Specific UL1 Gene Encodes a Late-Phase Glycoprotein Incorporated in the Virion Envelope

    PubMed Central

    Shikhagaie, Medya; Mercé-Maldonado, Eva; Isern, Elena; Muntasell, Aura; Albà, M. Mar; López-Botet, Miguel; Hengel, Hartmut

    2012-01-01

    We have investigated the previously uncharacterized human cytomegalovirus (HCMV) UL1 open reading frame (ORF), a member of the rapidly evolving HCMV RL11 family. UL1 is HCMV specific; the absence of UL1 in chimpanzee cytomegalovirus (CCMV) and sequence analysis studies suggest that UL1 may have originated by the duplication of an ancestor gene from the RL11-TRL cluster (TRL11, TRL12, and TRL13). Sequence similarity searches against human immunoglobulin (Ig)-containing proteins revealed that HCMV pUL1 shows significant similarity to the cellular carcinoembryonic antigen-related (CEA) protein family N-terminal Ig domain, which is responsible for CEA ligand recognition. Northern blot analysis revealed that UL1 is transcribed during the late phase of the viral replication cycle in both fibroblast-adapted and endotheliotropic strains of HCMV. We characterized the protein encoded by hemagglutinin (HA)-tagged UL1 in the AD169-derived HB5 background. UL1 is expressed as a 224-amino-acid type I transmembrane glycoprotein which becomes detectable at 48 h postinfection. In infected human fibroblasts, pUL1 colocalized at the cytoplasmic site of virion assembly and secondary envelopment together with TGN-46, a marker for the trans-Golgi network, and viral structural proteins, including the envelope glycoprotein gB and the tegument phosphoprotein pp28. Furthermore, analyses of highly purified AD169 UL1-HA epitope-tagged virions revealed that pUL1 is a novel constituent of the HCMV envelope. Importantly, the deletion of UL1 in HCMV TB40/E resulted in reduced growth in a cell type-specific manner, suggesting that pUL1 may be implicated in regulating HCMV cell tropism. PMID:22345456

  11. The T7-Related Pseudomonas putida Phage ϕ15 Displays Virion-Associated Biofilm Degradation Properties

    PubMed Central

    Cornelissen, Anneleen; Ceyssens, Pieter-Jan; T'Syen, Jeroen; Van Praet, Helena; Noben, Jean-Paul; Shaburova, Olga V.; Krylov, Victor N.; Volckaert, Guido; Lavigne, Rob

    2011-01-01

    Formation of a protected biofilm environment is recognized as one of the major causes of the increasing antibiotic resistance development and emphasizes the need to develop alternative antibacterial strategies, like phage therapy. This study investigates the in vitro degradation of single-species Pseudomonas putida biofilms, PpG1 and RD5PR2, by the novel phage ϕ15, a ‘T7-like virus’ with a virion-associated exopolysaccharide (EPS) depolymerase. Phage ϕ15 forms plaques surrounded by growing opaque halo zones, indicative for EPS degradation, on seven out of 53 P. putida strains. The absence of haloes on infection resistant strains suggests that the EPS probably act as a primary bacterial receptor for phage infection. Independent of bacterial strain or biofilm age, a time and dose dependent response of ϕ15-mediated biofilm degradation was observed with generally a maximum biofilm degradation 8 h after addition of the higher phage doses (104 and 106 pfu) and resistance development after 24 h. Biofilm age, an in vivo very variable parameter, reduced markedly phage-mediated degradation of PpG1 biofilms, while degradation of RD5PR2 biofilms and ϕ15 amplification were unaffected. Killing of the planktonic culture occurred in parallel with but was always more pronounced than biofilm degradation, accentuating the need for evaluating phages for therapeutic purposes in biofilm conditions. EPS degrading activity of recombinantly expressed viral tail spike was confirmed by capsule staining. These data suggests that the addition of high initial titers of specifically selected phages with a proper EPS depolymerase are crucial criteria in the development of phage therapy. PMID:21526174

  12. Synaptic transmission and the susceptibility of HIV infection to anti-viral drugs

    NASA Astrophysics Data System (ADS)

    Komarova, Natalia L.; Levy, David N.; Wodarz, Dominik

    2013-07-01

    Cell-to-cell viral transmission via virological synapses has been argued to reduce susceptibility of the virus population to anti-viral drugs through multiple infection of cells, contributing to low-level viral persistence during therapy. Using a mathematical framework, we examine the role of synaptic transmission in treatment susceptibility. A key factor is the relative probability of individual virions to infect a cell during free-virus and synaptic transmission, a currently unknown quantity. If this infection probability is higher for free-virus transmission, then treatment susceptibility is lowest if one virus is transferred per synapse, and multiple infection of cells increases susceptibility. In the opposite case, treatment susceptibility is minimized for an intermediate number of virions transferred per synapse. Hence, multiple infection via synapses does not simply lower treatment susceptibility. Without further experimental investigations, one cannot conclude that synaptic transmission provides an additional mechanism for the virus to persist at low levels during anti-viral therapy.

  13. Mapping the Small RNA Content of Simian Immunodeficiency Virions (SIV)

    PubMed Central

    Brameier, Markus; Ibing, Wiebke; Höfer, Katharina; Montag, Judith; Stahl-Hennig, Christiane; Motzkus, Dirk

    2013-01-01

    Recent evidence indicates that regulatory small non-coding RNAs are not only components of eukaryotic cells and vesicles, but also reside within a number of different viruses including retroviral particles. Using ultra-deep sequencing we have comprehensively analyzed the content of simian immunodeficiency virions (SIV), which were compared to mock-control preparations. Our analysis revealed that more than 428,000 sequence reads matched the SIV mac239 genome sequence. Among these we could identify 12 virus-derived small RNAs (vsRNAs) that were highly abundant. Beside known retrovirus-enriched small RNAs, like 7SL-RNA, tRNALys3 and tRNALys isoacceptors, we also identified defined fragments derived from small ILF3/NF90-associated RNA snaR-A14, that were enriched more than 50 fold in SIV. We also found evidence that small nucleolar RNAs U2 and U12 were underrepresented in the SIV preparation, indicating that the relative number or the content of co-isolated exosomes was changed upon infection. Our comprehensive atlas of SIV-incorporated small RNAs provides a refined picture of the composition of retrovirions, which gives novel insights into viral packaging. PMID:24086438

  14. Mapping the small RNA content of simian immunodeficiency virions (SIV).

    PubMed

    Brameier, Markus; Ibing, Wiebke; Höfer, Katharina; Montag, Judith; Stahl-Hennig, Christiane; Motzkus, Dirk

    2013-01-01

    Recent evidence indicates that regulatory small non-coding RNAs are not only components of eukaryotic cells and vesicles, but also reside within a number of different viruses including retroviral particles. Using ultra-deep sequencing we have comprehensively analyzed the content of simian immunodeficiency virions (SIV), which were compared to mock-control preparations. Our analysis revealed that more than 428,000 sequence reads matched the SIV mac239 genome sequence. Among these we could identify 12 virus-derived small RNAs (vsRNAs) that were highly abundant. Beside known retrovirus-enriched small RNAs, like 7SL-RNA, tRNA(Lys3) and tRNA(Lys) isoacceptors, we also identified defined fragments derived from small ILF3/NF90-associated RNA snaR-A14, that were enriched more than 50 fold in SIV. We also found evidence that small nucleolar RNAs U2 and U12 were underrepresented in the SIV preparation, indicating that the relative number or the content of co-isolated exosomes was changed upon infection. Our comprehensive atlas of SIV-incorporated small RNAs provides a refined picture of the composition of retrovirions, which gives novel insights into viral packaging. PMID:24086438

  15. VP3 is crucial for the stability of Nora virus virions.

    PubMed

    Sadanandan, Sajna Anand; Ekström, Jens-Ola; Jonna, Venkateswara Rao; Hofer, Anders; Hultmark, Dan

    2016-09-01

    Nora virus is an enteric virus that causes persistent, non-pathological infection in Drosophila melanogaster. It replicates in the fly gut and is transmitted via the fecal-oral route. Nora virus has a single-stranded positive-sense RNA genome, which is translated in four open reading frames. Reading frame three encodes the VP3 protein, the structure and function of which we have investigated in this work. We have shown that VP3 is a trimer that has an α-helical secondary structure, with a functionally important coiled-coil domain. In order to identify the role of VP3 in the Nora virus life cycle, we constructed VP3-mutants using the cDNA clone of the virus. Our results show that VP3 does not have a role in the actual assembly of the virus particles, but virions that lack VP3 or harbor VP3 with a disrupted coiled coil domain are incapable of transmission via the fecal-oral route. Removing the region downstream of the putative coiled coil appears to have an effect on the fitness of the virus but does not hamper its replication or transmission. We also found that the VP3 protein and particularly the coiled coil domain are crucial for the stability of Nora virus virions when exposed to heat or proteases. Hence, we propose that VP3 is imperative to Nora virus virions as it confers stability to the viral capsid. PMID:27329665

  16. Proteomic Analysis of Mamestra Brassicae Nucleopolyhedrovirus Progeny Virions from Two Different Hosts

    PubMed Central

    Hou, Dianhai; Chen, Xi

    2016-01-01

    Mamestra brassicae nucleopolyhedrovirus (MabrNPV) has a wide host range replication in more than one insect species. In this study, a sequenced MabrNPV strain, MabrNPV-CTa, was used to perform proteomic analysis of both BVs and ODVs derived from two infected hosts: Helicoverpa armigera and Spodoptera exigua. A total of 82 and 39 viral proteins were identified in ODVs and BVs, respectively. And totally, 23 and 76 host proteins were identified as virion-associated with ODVs and BVs, respectively. The host proteins incorporated into the virus particles were mainly involved in cytoskeleton, signaling, vesicle trafficking, chaperone and metabolic systems. Some host proteins, such as actin, cyclophilin A and heat shock protein 70 would be important for viral replication. Several host proteins involved in immune response were also identified in BV, and a C-type lectin protein was firstly found to be associated with BV and its family members have been demonstrated to be involved in entry process of other viruses. This study facilitated the annotation of baculovirus genome, and would help us to understand baculovirus virion structure. Furthermore, the identification of host proteins associated with virions produced in vivo would facilitate investigations on the involvement of intriguing host proteins in virus replication. PMID:27058368

  17. Exocytosis of Alphaherpesvirus Virions, Light Particles, and Glycoproteins Uses Constitutive Secretory Mechanisms

    PubMed Central

    Hogue, Ian B.; Scherer, Julian

    2016-01-01

    ABSTRACT Many molecular and cell biological details of the alphaherpesvirus assembly and egress pathway remain unclear. Recently we developed a live-cell fluorescence microscopy assay of pseudorabies virus (PRV) exocytosis, based on total internal reflection fluorescence (TIRF) microscopy and a virus-encoded pH-sensitive fluorescent probe. Here, we use this assay to distinguish three classes of viral exocytosis in a nonpolarized cell type: (i) trafficking of viral glycoproteins to the plasma membrane, (ii) exocytosis of viral light particles, and (iii) exocytosis of virions. We find that viral glycoproteins traffic to the cell surface in association with constitutive secretory Rab GTPases and exhibit free diffusion into the plasma membrane after exocytosis. Similarly, both virions and light particles use these same constitutive secretory mechanisms for egress from infected cells. Furthermore, we show that viral light particles are distinct from cellular exosomes. Together, these observations shed light on viral glycoprotein trafficking steps that precede virus particle assembly and reinforce the idea that virions and light particles share a biogenesis and trafficking pathway. PMID:27273828

  18. Proteomic Analysis of Mamestra Brassicae Nucleopolyhedrovirus Progeny Virions from Two Different Hosts.

    PubMed

    Hou, Dianhai; Chen, Xi; Zhang, Lei-Ke

    2016-01-01

    Mamestra brassicae nucleopolyhedrovirus (MabrNPV) has a wide host range replication in more than one insect species. In this study, a sequenced MabrNPV strain, MabrNPV-CTa, was used to perform proteomic analysis of both BVs and ODVs derived from two infected hosts: Helicoverpa armigera and Spodoptera exigua. A total of 82 and 39 viral proteins were identified in ODVs and BVs, respectively. And totally, 23 and 76 host proteins were identified as virion-associated with ODVs and BVs, respectively. The host proteins incorporated into the virus particles were mainly involved in cytoskeleton, signaling, vesicle trafficking, chaperone and metabolic systems. Some host proteins, such as actin, cyclophilin A and heat shock protein 70 would be important for viral replication. Several host proteins involved in immune response were also identified in BV, and a C-type lectin protein was firstly found to be associated with BV and its family members have been demonstrated to be involved in entry process of other viruses. This study facilitated the annotation of baculovirus genome, and would help us to understand baculovirus virion structure. Furthermore, the identification of host proteins associated with virions produced in vivo would facilitate investigations on the involvement of intriguing host proteins in virus replication. PMID:27058368

  19. Virion and soluble antigens of japanese encephalitis virus.

    PubMed Central

    Eckels, K H; Hetrick, F M; Russell, P K

    1975-01-01

    Japanese encephalitis virions contain a 58 X 10-3-molecular-weight envelope glycoprotein antigen that can be solubilized with sodium lauryl sulfate and separated from other virion structural polypeptides and viral ribonucleic acid by gel filtration chromatography. The 58 X 10-3-molecular-weight envelope protein is the major antigen responsible for cross-reactivity of the virion in complement fixation tests with other closely related arboviruses. A naturally occurring soluble complement-fixing antigen is found in Japanese encephalitis mouse brain preparations after removal of particulate antigens. After partial purification by gel filtration and isoelectric focusing, the 53 X 10-3-molecular weight soluble complement-fixing antigen is more type specific than the Japanese encephalitis envelope antigen in complement fixation tests. Further, the Japanese encephalitis soluble complement-fixing antigen is stable to treatment with sodium lauryl sulfate and 2-mercaptoethanol, whereas virion complement-fixing antigens are unstable after this treatment. Images PMID:47312

  20. New operative technique to reduce surgeons' risk of HIV infection.

    PubMed

    Raahave, D; Bremmelgaard, A

    1991-06-01

    The surgical team is potentially at risk of acquiring human immunodeficiency virus (HIV) from the patient. Assuming that the probability of an accidental injury during surgery is 0.01 (P2), the prevalence of HIV is 0.01 (P3) and the seroconversion rate is 0.01 (P1), we have estimated the risk (actuarial model) for a surgeon as 0.2% per year, and 5.82% for 30 years of surgery. In view of this we have made changes in surgical technique to reduce the risk to the surgical team from splash or injury. The surgeon must handle tissue with instruments only and minimize the use of fingers. Whenever possible, sharp instruments should be replaced by a blunt type. The surgical nurse loads needles to the needle carrier using forceps. Sharp instruments are placed in a neutral zone on the nurse's stand so that the surgeon and the nurse never touch the same sharp instrument at the same time. Movements should be controlled, and instrument handling accompanied by eye contact. We consider that these changes will reduce the risk of accidental injuries and thereby the transmission of HIV during operations to a greater degree than knowledge of the patient's HIV status. PMID:1716276

  1. A simple method for measuring porcine circovirus 2 whole virion particles and standardizing vaccine formulation.

    PubMed

    Zanotti, Cinzia; Amadori, Massimo

    2015-03-01

    Porcine Circovirus 2 (PCV2) is involved in porcine circovirus-associated disease, that causes great economic losses to the livestock industry worldwide. Vaccination against PCV2 proved to be very effective in reducing disease occurrence and it is currently performed on a large scale. Starting from a previous model concerning Foot-and Mouth Disease Virus antigens, we developed a rapid and simple method to quantify PCV2 whole virion particles in inactivated vaccines. This procedure, based on sucrose gradient analysis and fluorometric evaluation of viral genomic content, allows for a better standardization of the antigen payload in vaccine batches. It also provides a valid indication of virion integrity. Most important, such a method can be applied to whole virion vaccines regardless of the production procedures, thus enabling meaningful comparisons on a common basis. In a future batch consistency approach to PCV2 vaccine manufacture, our procedure represents a valuable tool to improve in-process controls and to guarantee conformity of the final product with passmarks for approval. This might have important repercussions in terms of reduced usage of animals for vaccine batch release, in the framework of the current 3Rs policy. PMID:25687800

  2. Effectiveness and cost of failure mode and effects analysis methodology to reduce neurosurgical site infections.

    PubMed

    Hover, Alexander R; Sistrunk, William W; Cavagnol, Robert M; Scarrow, Alan; Finley, Phillip J; Kroencke, Audrey D; Walker, Judith L

    2014-01-01

    Mercy Hospital Springfield is a tertiary care facility with 32 000 discharges and 15 000 inpatient surgeries in 2011. From June 2009 through January 2011, a stable inpatient elective neurosurgery infection rate of 2.15% was observed. The failure mode and effects analysis (FMEA) methodology to reduce inpatient neurosurgery infections was utilized. Following FMEA implementation, overall elective neurosurgery infection rates were reduced to 1.51% and sustained through May 2012. Compared with baseline, the post-FMEA deep-space and organ infection rate was reduced by 41% (P = .052). Overall hospital inpatient clean surgery infection rates for the same time frame did not decrease to the same extent, suggesting a specific effect of the FMEA. The study team believes that the FMEA interventions resulted in 14 fewer expected infections, $270 270 in savings, a 168-day reduction in expected length of stay, and 22 fewer readmissions. Given the serious morbidity and cost of health care-associated infections, the study team concludes that FMEA implementation was clinically cost-effective. PMID:24101683

  3. Arginine reduces Cryptosporidium parvum infection in undernourished suckling mice involving both nitric oxide synthase and arginase

    PubMed Central

    Castro, Ibraim C.; Oliveira, Bruna B.; Slowikowski, Jacek J.; Coutinho, Bruna P.; Siqueira, Francisco Júlio W.S.; Costa, Lourrany B.; Sevilleja, Jesus Emmanuel; Almeida, Camila A.; Lima, Aldo A.M.; Warren, Cirle A.; Oriá, Reinaldo B.; Guerrant, Richard L.

    2011-01-01

    Objective This study investigated the role of L-arginine supplementation to undernourished and Cryptosporidium parvum-infected suckling mice. Methods The following regimens were initiated on the 4th day of life and given subcutaneously daily: either 200mM of L-arginine or PBS for the C. parvum-infected controls. L-arginine-treated mice were grouped to receive either 20mM of NG-nitroarginine-methyl-ester (L-NAME) or PBS. Infected mice received orally 106 excysted-C. parvum oocysts on day 6 and were euthanized on day 14th at the infection peak. Results L-arginine improved weight gain compared to the untreated infected controls. L-NAME profoundly impaired body weight gain as compared to all other groups. Cryptosporidiosis was associated with ileal crypt hyperplasia, villus blunting, and inflammation. L-arginine improved mucosal histology following infection. L-NAME abrogated these arginine-induced improvements. Infected control mice showed an intense arginase expression, which was even greater with L-NAME. L-arginine reduced parasite burden, an effect that was reversed by L-NAME. C. parvum infection increased urine NO3-/NO2- concentration when compared to uninfected controls, which was increased by L-arginine supplementation, an effect that was also reversed by L-NAME. Conclusion These findings show a protective role of L-arginine during C. parvum infection in undernourished mice with involvement of arginase I and nitric oxide synthase enzymatic actions. PMID:22261576

  4. Reducing the risk of infection associated with vascular access devices through nanotechnology: a perspective

    PubMed Central

    Zhang, Li; Keogh, Samantha; Rickard, Claire M

    2013-01-01

    Intravascular catheter-related infections are still a major problem in health care and are associated with significant morbidity, mortality, and additional cost. The formation of microbial biofilm on catheters makes these infections particularly complicated, as microbial cells that detach from the biofilm can lead to infection, and because these microorganisms are highly resistant to many antimicrobial agents; thus, catheter removal is often required to successfully treat infection. To reduce the risks of catheter-related infections, many strategies have been applied, such as improvements in aseptic insertion and post-insertion care practices, implantation techniques, and antibiotic coated or impregnated materials. However, despite significant advances in using these methods, it has not been possible to completely eradicate biofilm infections. Currently, nanotechnology approaches seem to be among the most promising for preventing biofilm formation and resultant catheter-related bloodstream infection (especially with multi-resistant bacterial strains). In this review, current knowledge about catheter technology and design, the mechanisms of catheter-related bloodstream infection, and the insertion and care practices performed by medical staff, are discussed, along with novel, achievable approaches to infection prevention, based on nanotechnology. PMID:24293997

  5. Antibiotic and Antiinflammatory Therapy Transiently Reduces Inflammation and Hypercoagulation in Acutely SIV-Infected Pigtailed Macaques

    PubMed Central

    Pandrea, Ivona; Xu, Cuiling; Stock, Jennifer L.; Frank, Daniel N.; Ma, Dongzhu; Policicchio, Benjamin B.; He, Tianyu; Kristoff, Jan; Cornell, Elaine; Haret-Richter, George S.; Trichel, Anita; Ribeiro, Ruy M.; Tracy, Russell; Wilson, Cara; Landay, Alan L.; Apetrei, Cristian

    2016-01-01

    Increased chronic immune activation and inflammation are hallmarks of HIV/SIV infection and are highly correlated with progression to AIDS and development of non-AIDS comorbidities, such as hypercoagulability and cardiovascular disease. Intestinal dysfunction resulting in microbial translocation has been proposed as a lead cause of systemic immune activation and hypercoagulability in HIV/SIV infection. Our goal was to assess the biological and clinical impact of a therapeutic strategy designed to reduce microbial translocation through reduction of the microbial content of the intestine (Rifaximin-RFX) and of gut inflammation (Sulfasalazine-SFZ). RFX is an intraluminal antibiotic that was successfully used in patients with hepatic encephalopathy. SFZ is an antiinflammatory drug successfully used in patients with mild to moderate inflammatory bowel disease. Both these clinical conditions are associated with increased microbial translocation, similar to HIV-infected patients. Treatment was administered for 90 days to five acutely SIV-infected pigtailed macaques (PTMs) starting at the time of infection; seven untreated SIVsab-infected PTMs were used as controls. RFX+SFZ were also administered for 90 days to three chronically SIVsab-infected PTMs. RFX+SFZ administration during acute SIVsab infection of PTMs resulted in: significantly lower microbial translocation, lower systemic immune activation, lower viral replication, better preservation of mucosal CD4+ T cells and significantly lower levels of hypercoagulation biomarkers. This effect was clear during the first 40 days of treatment and was lost during the last stages of treatment. Administration of RFX+SFZ to chronically SIVsab–infected PTMs had no discernible effect on infection. Our data thus indicate that early RFX+SFZ administration transiently improves the natural history of acute and postacute SIV infection, but has no effect during chronic infection. PMID:26764484

  6. Antibiotic and Antiinflammatory Therapy Transiently Reduces Inflammation and Hypercoagulation in Acutely SIV-Infected Pigtailed Macaques.

    PubMed

    Pandrea, Ivona; Xu, Cuiling; Stock, Jennifer L; Frank, Daniel N; Ma, Dongzhu; Policicchio, Benjamin B; He, Tianyu; Kristoff, Jan; Cornell, Elaine; Haret-Richter, George S; Trichel, Anita; Ribeiro, Ruy M; Tracy, Russell; Wilson, Cara; Landay, Alan L; Apetrei, Cristian

    2016-01-01

    Increased chronic immune activation and inflammation are hallmarks of HIV/SIV infection and are highly correlated with progression to AIDS and development of non-AIDS comorbidities, such as hypercoagulability and cardiovascular disease. Intestinal dysfunction resulting in microbial translocation has been proposed as a lead cause of systemic immune activation and hypercoagulability in HIV/SIV infection. Our goal was to assess the biological and clinical impact of a therapeutic strategy designed to reduce microbial translocation through reduction of the microbial content of the intestine (Rifaximin-RFX) and of gut inflammation (Sulfasalazine-SFZ). RFX is an intraluminal antibiotic that was successfully used in patients with hepatic encephalopathy. SFZ is an antiinflammatory drug successfully used in patients with mild to moderate inflammatory bowel disease. Both these clinical conditions are associated with increased microbial translocation, similar to HIV-infected patients. Treatment was administered for 90 days to five acutely SIV-infected pigtailed macaques (PTMs) starting at the time of infection; seven untreated SIVsab-infected PTMs were used as controls. RFX+SFZ were also administered for 90 days to three chronically SIVsab-infected PTMs. RFX+SFZ administration during acute SIVsab infection of PTMs resulted in: significantly lower microbial translocation, lower systemic immune activation, lower viral replication, better preservation of mucosal CD4+ T cells and significantly lower levels of hypercoagulation biomarkers. This effect was clear during the first 40 days of treatment and was lost during the last stages of treatment. Administration of RFX+SFZ to chronically SIVsab-infected PTMs had no discernible effect on infection. Our data thus indicate that early RFX+SFZ administration transiently improves the natural history of acute and postacute SIV infection, but has no effect during chronic infection. PMID:26764484

  7. Retromer Regulates HIV-1 Envelope Glycoprotein Trafficking and Incorporation into Virions

    PubMed Central

    Groppelli, Elisabetta; Jolly, Clare

    2014-01-01

    The envelope glycoprotein (Env) of the Human Immunodeficiency Virus Type-1 (HIV-1) is a critical determinant of viral infectivity, tropism and is the main target for humoral immunity; however, little is known about the cellular machinery that directs Env trafficking and its incorporation into nascent virions. Here we identify the mammalian retromer complex as a novel and important cellular factor regulating Env trafficking. Retromer mediates endosomal sorting and is most closely associated with endosome-to-Golgi transport. Consistent with this function, inactivating retromer using RNAi targeting the cargo selective trimer complex inhibited retrograde trafficking of endocytosed Env to the Golgi. Notably, in HIV-1 infected cells, inactivating retromer modulated plasma membrane expression of Env, along with Env incorporation into virions and particle infectivity. Mutagenesis studies coupled with coimmunoprecipitations revealed that retromer-mediated trafficking requires the Env cytoplasmic tail that we show binds directly to retromer components Vps35 and Vps26. Taken together these results provide novel insight into regulation of HIV-1 Env trafficking and infectious HIV-1 morphogenesis and show for the first time a role for retromer in the late-steps of viral replication and assembly of a virus. PMID:25393110

  8. The Plant Host Can Affect the Encapsidation of Brome Mosaic Virus (BMV) RNA; BMV Virions Are Surprisingly Heterogeneous

    PubMed Central

    Ni, Peng; Vaughan, Robert C.; Tragesser, Brady; Hoover, Haley; Kao, C. Cheng

    2013-01-01

    Brome mosaic virus (BMV) packages its genomic and subgenomic RNAs into three separate viral particles. BMV purified from barley, wheat and tobacco have distinct relative abundances of the encapsidated RNAs. We seek to identify the basis for the host-dependent differences in viral RNA encapsidation. Sequencing of the viral RNAs revealed recombination events in the 3′ untranslated region of RNA1 of BMV purified from barley and wheat, but not from tobacco. However, the relative amounts of the BMV RNAs that accumulated in barley and wheat are similar and RNA accumulation is not sufficient to account for the difference in RNA encapsidation. Virions purified from barley and wheat were found to differ in their isoelectric points, resistance to proteolysis, and contacts between the capsid residues and the RNA. Mass spectrometric analyses revealed that virions from the three hosts had different post-translational modifications that should impact the physiochemical properties of the virions. Another major source of variation in RNA encapsidation was due to the purification of BMV particles to homogeneity. Highly enriched BMV present in lysates had a surprising range of sizes, buoyant densities, and distinct relative amounts of encapsidated RNAs. These results show that the encapsidated BMV RNAs reflect a combination of host effects on the physiochemical properties of the viral capsids and the enrichment of a subset of virions. The previously unexpected heterogeneity in BMV should influence the timing of the infection and also the host innate immune responses. PMID:24036424

  9. A Rhesus Rhadinovirus Viral Interferon (IFN) Regulatory Factor Is Virion Associated and Inhibits the Early IFN Antiviral Response

    PubMed Central

    Morin, Gabriela; Robinson, Bridget A.; Rogers, Kelsey S.

    2015-01-01

    ABSTRACT The interferon (IFN) response is the earliest host immune response dedicated to combating viral infection. As such, viruses have evolved strategies to subvert this potent antiviral response. Two closely related gammaherpesviruses, Kaposi's sarcoma-associated herpesvirus (KSHV) and rhesus macaque rhadinovirus (RRV), are unique in that they express viral homologues to cellular interferon regulatory factors (IRFs), termed viral IRFs (vIRFs). Cellular IRFs are a family of transcription factors that are particularly important for the transcription of type I IFNs. Here, we demonstrate a strategy employed by RRV to ensure rapid inhibition of virus-induced type I IFN induction. We found that RRV vIRF R6, when expressed ectopically, interacts with a transcriptional coactivator, CREB-binding protein (CBP), in the nucleus. As a result, phosphorylated IRF3, an important transcriptional regulator in beta interferon (IFN-β) transcription, fails to effectively bind to the IFN-β promoter, thus inhibiting the activation of IFN-β genes. In addition, we found R6 within RRV virion particles via immunoelectron microscopy and, furthermore, that virion-associated R6 is capable of inhibiting the type I IFN response by preventing efficient binding of IRF3/CBP complexes to the IFN-β promoter in the context of infection. The work shown here is the first example of a vIRF being associated with either the KSHV or RRV virion. The presence of this immunomodulatory protein in the RRV virion provides the virus with an immediate mechanism to evade the host IFN response, thus enabling the virus to effectively establish an infection within the host. IMPORTANCE Kaposi's sarcoma-associated herpesvirus (KSHV) and the closely related rhesus macaque rhadinovirus (RRV) are the only viruses known to encode viral homologues to cellular interferon regulatory factors (IRFs), known as vIRFs. In KSHV, these proteins have been shown to play major roles in a variety of cellular processes and are

  10. A new Hydrocephalus Clinical Research Network protocol to reduce cerebrospinal fluid shunt infection.

    PubMed

    Kestle, John R W; Holubkov, Richard; Douglas Cochrane, D; Kulkarni, Abhaya V; Limbrick, David D; Luerssen, Thomas G; Jerry Oakes, W; Riva-Cambrin, Jay; Rozzelle, Curtis; Simon, Tamara D; Walker, Marion L; Wellons, John C; Browd, Samuel R; Drake, James M; Shannon, Chevis N; Tamber, Mandeep S; Whitehead, William E

    2016-04-01

    OBJECT In a previous report by the same research group (Kestle et al., 2011), compliance with an 11-step protocol was shown to reduce CSF shunt infection at Hydrocephalus Clinical Research Network (HCRN) centers (from 8.7% to 5.7%). Antibiotic-impregnated catheters (AICs) were not part of the protocol but were used off protocol by some surgeons. The authors therefore began using a new protocol that included AICs in an effort to reduce the infection rate further. METHODS The new protocol was implemented at HCRN centers on January 1, 2012, for all shunt procedures (excluding external ventricular drains [EVDs], ventricular reservoirs, and subgaleal shunts). Procedures performed up to September 30, 2013, were included (21 months). Compliance with the protocol and outcome events up to March 30, 2014, were recorded. The definition of infection was unchanged from the authors' previous report. RESULTS A total of 1935 procedures were performed on 1670 patients at 8 HCRN centers. The overall infection rate was 6.0% (95% CI 5.1%-7.2%). Procedure-specific infection rates varied (insertion 5.0%, revision 5.4%, insertion after EVD 8.3%, and insertion after treatment of infection 12.6%). Full compliance with the protocol occurred in 77% of procedures. The infection rate was 5.0% after compliant procedures and 8.7% after noncompliant procedures (p = 0.005). The infection rate when using this new protocol (6.0%, 95% CI 5.1%-7.2%) was similar to the infection rate observed using the authors' old protocol (5.7%, 95% CI 4.6%-7.0%). CONCLUSIONS CSF shunt procedures performed in compliance with a new infection prevention protocol at HCRN centers had a lower infection rate than noncompliant procedures. Implementation of the new protocol (including AICs) was associated with a 6.0% infection rate, similar to the infection rate of 5.7% from the authors' previously reported protocol. Based on the current data, the role of AICs compared with other infection prevention measures is unclear

  11. Drug treatment of malaria infections can reduce levels of protection transferred to offspring via maternal immunity

    PubMed Central

    Staszewski, Vincent; Reece, Sarah E.; O'Donnell, Aidan J.; Cunningham, Emma J. A.

    2012-01-01

    Maternally transferred immunity can have a fundamental effect on the ability of offspring to deal with infection. However, levels of antibodies in adults can vary both quantitatively and qualitatively between individuals and during the course of infection. How infection dynamics and their modification by drug treatment might affect the protection transferred to offspring remains poorly understood. Using the rodent malaria parasite Plasmodium chabaudi, we demonstrate that curing dams part way through infection prior to pregnancy can alter their immune response, with major consequences for offspring health and survival. In untreated maternal infections, maternally transferred protection suppressed parasitaemia and reduced pup mortality by 75 per cent compared with pups from naïve dams. However, when dams were treated with anti-malarial drugs, pups received fewer maternal antibodies, parasitaemia was only marginally suppressed, and mortality risk was 25 per cent higher than for pups from dams with full infections. We observed the same qualitative patterns across three different host strains and two parasite genotypes. This study reveals the role that within-host infection dynamics play in the fitness consequences of maternally transferred immunity. Furthermore, it highlights a potential trade-off between the health of mothers and offspring suggesting that anti-parasite treatment may significantly affect the outcome of infection in newborns. PMID:22357264

  12. Herpes simplex virus-infected cells contain a function(s) that destabilizes both host and viral mRNAs.

    PubMed Central

    Kwong, A D; Frenkel, N

    1987-01-01

    The herpes simplex virus virion contains a function that mediates the shutoff of host-protein synthesis and the degradation of host mRNA. Viral mutants affected in this function (vhs mutants) have previously been derived. Cells infected with these mutants exhibit a more stable synthesis of host as well as the immediate early (alpha)-viral proteins. We now show that a function associated with purified virions of the wild-type virus reduces the half-life of host and alpha mRNAs, whereas purified vhs-1 mutant virions lack this activity. The functional half-life of many early (beta)- and late (gamma)-viral mRNAs is also prolonged in mutant virus infections. These studies suggest that the wild-type virion brings into cells a function that indiscriminately reduces the half-life of both host and viral transcripts and that the early translational shutoff of the host is a consequence of this function. This function may facilitate rapid transitions in the expression of groups of genes that are transcriptionally turned on at different times after infection. Images PMID:3031658

  13. Cystic fibrosis-niche adaptation of Pseudomonas aeruginosa reduces virulence in multiple infection hosts.

    PubMed

    Lorè, Nicola Ivan; Cigana, Cristina; De Fino, Ida; Riva, Camilla; Juhas, Mario; Schwager, Stephan; Eberl, Leo; Bragonzi, Alessandra

    2012-01-01

    The opportunistic pathogen Pseudomonas aeruginosa is able to thrive in diverse ecological niches and to cause serious human infection. P. aeruginosa environmental strains are producing various virulence factors that are required for establishing acute infections in several host organisms; however, the P. aeruginosa phenotypic variants favour long-term persistence in the cystic fibrosis (CF) airways. Whether P. aeruginosa strains, which have adapted to the CF-niche, have lost their competitive fitness in the other environment remains to be investigated. In this paper, three P. aeruginosa clonal lineages, including early strains isolated at the onset of infection, and late strains, isolated after several years of chronic lung infection from patients with CF, were analysed in multi-host model systems of acute infection. P. aeruginosa early isolates caused lethality in the three non-mammalian hosts, namely Caenorhabditis elegans, Galleria mellonella, and Drosophila melanogaster, while late adapted clonal isolates were attenuated in acute virulence. When two different mouse genetic background strains, namely C57Bl/6NCrl and Balb/cAnNCrl, were used as acute infection models, early P. aeruginosa CF isolates were lethal, while late isolates exhibited reduced or abolished acute virulence. Severe histopathological lesions, including high leukocytes recruitment and bacterial load, were detected in the lungs of mice infected with P. aeruginosa CF early isolates, while late isolates were progressively cleared. In addition, systemic bacterial spread and invasion of epithelial cells, which were detected for P. aeruginosa CF early strains, were not observed with late strains. Our findings indicate that niche-specific selection in P. aeruginosa reduced its ability to cause acute infections across a broad range of hosts while maintaining the capacity for chronic infection in the CF host. PMID:22558188

  14. Cystic Fibrosis-Niche Adaptation of Pseudomonas aeruginosa Reduces Virulence in Multiple Infection Hosts

    PubMed Central

    De Fino, Ida; Riva, Camilla; Juhas, Mario; Schwager, Stephan; Eberl, Leo; Bragonzi, Alessandra

    2012-01-01

    The opportunistic pathogen Pseudomonas aeruginosa is able to thrive in diverse ecological niches and to cause serious human infection. P. aeruginosa environmental strains are producing various virulence factors that are required for establishing acute infections in several host organisms; however, the P. aeruginosa phenotypic variants favour long-term persistence in the cystic fibrosis (CF) airways. Whether P. aeruginosa strains, which have adapted to the CF-niche, have lost their competitive fitness in the other environment remains to be investigated. In this paper, three P. aeruginosa clonal lineages, including early strains isolated at the onset of infection, and late strains, isolated after several years of chronic lung infection from patients with CF, were analysed in multi-host model systems of acute infection. P. aeruginosa early isolates caused lethality in the three non-mammalian hosts, namely Caenorhabditis elegans, Galleria mellonella, and Drosophila melanogaster, while late adapted clonal isolates were attenuated in acute virulence. When two different mouse genetic background strains, namely C57Bl/6NCrl and Balb/cAnNCrl, were used as acute infection models, early P. aeruginosa CF isolates were lethal, while late isolates exhibited reduced or abolished acute virulence. Severe histopathological lesions, including high leukocytes recruitment and bacterial load, were detected in the lungs of mice infected with P. aeruginosa CF early isolates, while late isolates were progressively cleared. In addition, systemic bacterial spread and invasion of epithelial cells, which were detected for P. aeruginosa CF early strains, were not observed with late strains. Our findings indicate that niche-specific selection in P. aeruginosa reduced its ability to cause acute infections across a broad range of hosts while maintaining the capacity for chronic infection in the CF host. PMID:22558188

  15. Incorporation of Spike and Membrane Glycoproteins into Coronavirus Virions

    PubMed Central

    Ujike, Makoto; Taguchi, Fumihiro

    2015-01-01

    The envelopes of coronaviruses (CoVs) contain primarily three proteins; the two major glycoproteins spike (S) and membrane (M), and envelope (E), a non-glycosylated protein. Unlike other enveloped viruses, CoVs bud and assemble at the endoplasmic reticulum (ER)-Golgi intermediate compartment (ERGIC). For efficient virion assembly, these proteins must be targeted to the budding site and to interact with each other or the ribonucleoprotein. Thus, the efficient incorporation of viral envelope proteins into CoV virions depends on protein trafficking and protein–protein interactions near the ERGIC. The goal of this review is to summarize recent findings on the mechanism of incorporation of the M and S glycoproteins into the CoV virion, focusing on protein trafficking and protein–protein interactions. PMID:25855243

  16. Inhibition of the receptor-mediated virion attachment to a lipid membrane

    NASA Astrophysics Data System (ADS)

    Zhdanov, Vladimir P.

    2012-10-01

    The forefront of the anti-viral defence is sometimes aimed at virion attachment to a host membrane. This step or, more specifically, virion contacts with cellular membrane receptors (or, e.g., glycolipids) can be inhibited by antibodies (or specially chosen or designed compounds) via their association with virions. In this case, the full-scale attachment of virions to a host membrane occurs via a subtle interplay of the formation and rupture of multiple virion-inhibitor and virion-receptor bonds. We present a kinetic model describing this interplay and illustrating general trends in the process under consideration.

  17. Reducing the incidence of infection after caesarean section: implications of prophylaxis with antibiotics for hospital resources.

    PubMed Central

    Mugford, M.; Kingston, J.; Chalmers, I.

    1989-01-01

    OBJECTIVES--To estimate the cost effectiveness of giving prophylactic antibiotics routinely to reduce the incidence of wound infection after caesarean section. DESIGN--Estimation of cost effectiveness was based, firstly, on a retrospective overview of 58 controlled trials and, secondly, on evidence about costs derived from data and observations of practice. SETTING--Trials included in the overview were from obstetric units in several different countries, including the United Kingdom. The costing study was based on data referring to the John Radcliffe Maternity Hospital, Oxford. SUBJECTS--A total of 7777 women were included in the 58 controlled trials comparing the effects of giving routine prophylactic antibiotics at caesarean section with either treatment with a placebo or no treatment. Cost estimates were based on data on 486 women who had caesarean sections between January and September 1987. MAIN OUTCOME MEASURE--Cost effectiveness of prophylaxis with antibiotics. RESULTS--The odds of wound infection are likely to be reduced by between about 50 and 70% by giving antibiotics routinely at caesarean section. Forty one (8.4%) women who had caesarean section were coded by the Oxford obstetric data system as having developed wound infection. The additional average cost of hospital postnatal care for women with wound infection (compared with women who had had caesarean section and no wound infection) was estimated to be 716 pounds; introducing routine prophylaxis with antibiotics would reduce average costs of postnatal care by between 1300 pounds and 3900/100 pounds caesarean sections (at 1988 prices), depending on the cost of the antibiotic used and its effectiveness. CONCLUSIONS--The results suggest that giving antibiotics routinely at caesarean section will not only reduce rates of infection after caesarean section but also reduce costs. PMID:2511938

  18. Virus-producing cells determine the host protein profiles of HIV-1 virion cores

    PubMed Central

    2012-01-01

    Background Upon HIV entry into target cells, viral cores are released and rearranged into reverse transcription complexes (RTCs), which support reverse transcription and also protect and transport viral cDNA to the site of integration. RTCs are composed of viral and cellular proteins that originate from both target and producer cells, the latter entering the target cell within the viral core. However, the proteome of HIV-1 viral cores in the context of the type of producer cells has not yet been characterized. Results We examined the proteomic profiles of the cores purified from HIV-1 NL4-3 virions assembled in Sup-T1 cells (T lymphocytes), PMA and vitamin D3 activated THP1 (model of macrophages, mMΦ), and non-activated THP1 cells (model of monocytes, mMN) and assessed potential involvement of identified proteins in the early stages of infection using gene ontology information and data from genome-wide screens on proteins important for HIV-1 replication. We identified 202 cellular proteins incorporated in the viral cores (T cells: 125, mMΦ: 110, mMN: 90) with the overlap between these sets limited to 42 proteins. The groups of RNA binding (29), DNA binding (17), cytoskeleton (15), cytoskeleton regulation (21), chaperone (18), vesicular trafficking-associated (12) and ubiquitin-proteasome pathway-associated proteins (9) were most numerous. Cores of the virions from SupT1 cells contained twice as many RNA binding proteins as cores of THP1-derived virus, whereas cores of virions from mMΦ and mMN were enriched in components of cytoskeleton and vesicular transport machinery, most probably due to differences in virion assembly pathways between these cells. Spectra of chaperones, cytoskeletal proteins and ubiquitin-proteasome pathway components were similar between viral cores from different cell types, whereas DNA-binding and especially RNA-binding proteins were highly diverse. Western blot analysis showed that within the group of overlapping proteins, the level of

  19. Nef Stimulates Human Immunodeficiency Virus Type 1 Replication in Primary T Cells by Enhancing Virion-Associated gp120 Levels: Coreceptor-Dependent Requirement for Nef in Viral Replication†

    PubMed Central

    Lundquist, Christopher A.; Zhou, Jing; Aiken, Christopher

    2004-01-01

    The Nef protein enhances human immunodeficiency virus type 1 (HIV-1) replication through an unknown mechanism. We and others have previously reported that efficient HIV-1 replication in activated primary CD4+ T cells depends on the ability of Nef to downregulate CD4 from the cell surface. Here we demonstrate that Nef greatly enhances the infectivity of HIV-1 particles produced in primary T cells. Nef-defective HIV-1 particles contained significantly reduced quantities of gp120 on their surface; however, Nef did not affect the levels of virion-associated gp41, indicating that Nef indirectly stabilizes the association of gp120 with gp41. Surprisingly, Nef was not required for efficient replication of viruses that use CCR5 for entry, nor did Nef influence the infectivity or gp120 content of these virions. Nef also inhibited the incorporation of CD4 into HIV-1 particles released from primary T cells. We propose that Nef, by downregulating cell surface CD4, enhances HIV-1 replication by inhibiting CD4-induced dissociation of gp120 from gp41. The preferential requirement for Nef in the replication of X4-tropic HIV-1 suggests that the ability of Nef to downregulate CD4 may be most important at later stages of disease when X4-tropic viruses emerge. PMID:15163722

  20. The type I interferon response bridles rabies virus infection and reduces pathogenicity.

    PubMed

    Chopy, Damien; Detje, Claudia N; Lafage, Mireille; Kalinke, Ulrich; Lafon, Monique

    2011-08-01

    Rabies virus (RABV) is a neurotropic virus transmitted by the bite of an infected animal that triggers a fatal encephalomyelitis. During its migration in the nervous system (NS), RABV triggers an innate immune response, including a type I IFN response well known to limit viral infections. We showed that although the neuroinvasive RABV strain CVS-NIV dampens type I IFN signaling by inhibiting IRF3 phosphorylation and STAT2 translocation, an early and transient type I IFN response is still triggered in the infected neuronal cells and NS. This urged us to investigate the role of type I IFN on RABV infection. We showed that primary mouse neurons (DRGs) of type I IFN(α/β) receptor deficient mice (IFNAR(-/-) mice) were more susceptible to RABV than DRGs of WT mice. In addition, exogenous type I IFN is partially efficient in preventing and slowing down infection in human neuroblastoma cells. Intra-muscular inoculation of type I IFNAR deficient mice [IFNAR(-/-) mice and NesCre ((+/-)) IFNAR ((flox/flox)) mice lacking IFNAR in neural cells of neuroectodermal origin only] with RABV reveals that the type I IFN response limits RABV dissemination in the inoculated muscle, slows down invasion of the spinal cord, and delays mortality. Thus, the type I IFN which is still produced in the NS during RABV infection is efficient enough to reduce neuroinvasiveness and pathogenicity and partially protect the host from fatal infection. PMID:21805057

  1. Ultrafast Tracking of a Single Live Virion During the Invagination of a Cell Membrane.

    PubMed

    Pan, Yangang; Wang, Shaowen; Shan, Yuping; Zhang, Dinglin; Gao, Jing; Zhang, Min; Liu, Shuheng; Cai, Mingjun; Xu, Haijiao; Li, Guohui; Qin, Qiwei; Wang, Hongda

    2015-06-01

    The first step in most viral infections is the penetration of the cell membrane via endocytosis. However, the underlying mechanism of this important process has not been quantitatively characterized; for example, the velocity and force of a single virion during invagination remain unknown. Here, the endocytosis of a single live virion (Singapore grouper iridovirus, SGIV) through the apical membranes of a host cell is monitored by developing and using a novel ultrafast (at the microsecond level) tracking technique: force tracing. For the first time, these results unambiguously reveal that the maximum velocity during the cell entry of a single SGIV by membrane invagination is approximately 200 nm s(-1), the endocytic force is approximately 60.8 ± 18.5 pN, and the binding energy density increases with the engulfment depth. This report utilizing high temporospatial resolution (subnanometer and microsecond levels) approaches provides new insight into the dynamic process of viral infection via endocytosis and the mechanism of membrane invagination at the single-particle level. PMID:25689837

  2. Inhibitors of Endoplasmic Reticulum α-Glucosidases Potently Suppress Hepatitis C Virus Virion Assembly and Release▿

    PubMed Central

    Qu, Xiaowang; Pan, Xiaoben; Weidner, Jessica; Yu, Wenquan; Alonzi, Dominic; Xu, Xiaodong; Butters, Terry; Block, Timothy; Guo, Ju-Tao; Chang, Jinhong

    2011-01-01

    α-Glucosidases I and II are endoplasmic reticulum-resident enzymes that are essential for N-linked glycan processing and subsequent proper folding of glycoproteins. In this report, we first demonstrate that downregulation of the expression of α-glucosidase I, II, or both in Huh7.5 cells by small hairpin RNA technology inhibited the production of hepatitis C virus (HCV). In agreement with the essential role of α-glucosidases in HCV envelope glycoprotein processing and folding, treatment of HCV-infected cells with a panel of imino sugar derivatives, which are competitive inhibitors of α-glucosidases, did not affect intracellular HCV RNA replication and nonstructural protein expression but resulted in the inhibition of glycan processing and subsequent degradation of HCV E2 glycoprotein. As a consequence, HCV virion assembly and secretion were inhibited. In searching for imino sugars with better antiviral activity, we found that a novel imino sugar, PBDNJ0804, had a superior ability to inhibit HCV virion assembly and secretion. In summary, we demonstrated that glucosidases are important host factor-based antiviral targets for HCV infection. The low likelihood of drug-resistant virus emergence and potent antiviral efficacy of the novel glucosidase inhibitor hold promise for its development as a therapeutic agent for the treatment of chronic hepatitis C. PMID:21173177

  3. Current Evidence for the Use of Laminar Flow in Reducing Infection Rates in Total Joint Arthroplasty

    PubMed Central

    James, M; Khan, W.S; Nannaparaju, M.R; Bhamra, J.S; Morgan-Jones, R

    2015-01-01

    Since the introduction of laminar air flow in orthopaedic theatres by Sir John Charnley, it has widely become accepted as the standard during orthopaedic procedures such as joint arthroplasty. We present a review of available current literature for the use of laminar flow operating theatre ventilation during total joint arthroplasty and examines the effectiveness of laminar flow ventilated operating theatres in preventing post-operative wound infection. Results of our findings suggest that while bacterial and air particulate is reduced by laminar air flow systems, there is no conclusive effect on the reduction of post-operative wound infections following total joint arthroplasty. We conclude that a combination of strict aseptic technique, prophylactic antibiotics and good anaesthetic control during surgery remains crucial to reduce post-operative surgical infections. PMID:26587068

  4. The Flavonoid Isoliquiritigenin Reduces Lung Inflammation and Mouse Morbidity during Influenza Virus Infection

    PubMed Central

    Traboulsi, Hussein; Cloutier, Alexandre; Boyapelly, Kumaraswamy; Bonin, Marc-André; Marsault, Éric; Cantin, André M.

    2015-01-01

    The host response to influenza virus infection is characterized by an acute lung inflammatory response in which intense inflammatory cell recruitment, hypercytokinemia, and a high level of oxidative stress are present. The sum of these events contributes to the virus-induced lung damage that leads to high a level of morbidity and mortality in susceptible infected patients. In this context, we identified compounds that can simultaneously reduce the excessive inflammatory response and the viral replication as a strategy to treat influenza virus infection. We investigated the anti-inflammatory and antiviral potential activities of isoliquiritigenin (ILG). Interestingly, we demonstrated that ILG is a potent inhibitor of influenza virus replication in human bronchial epithelial cells (50% effective concentration [EC50] = 24.7 μM). In addition, our results showed that this molecule inhibits the expression of inflammatory cytokines induced after the infection of cells with influenza virus. We demonstrated that the anti-inflammatory activity of ILG in the context of influenza virus infection is dependent on the activation of the peroxisome proliferator-activated receptor gamma pathway. Interestingly, ILG phosphate (ILG-p)-treated mice displayed decreased lung inflammation as depicted by reduced cytokine gene expression and inflammatory cell recruitment. We also demonstrated that influenza virus-specific CD8+ effector T cell recruitment was reduced up to 60% in the lungs of mice treated with ILG-p (10 mg/kg) compared to that in saline-treated mice. Finally, we showed that administration of ILG-p reduced lung viral titers and morbidity of mice infected with the PR8/H1N1 virus. PMID:26248373

  5. Influenza virus pyrogenicity: central role of structural orientation of virion components and involvement of viral lipid and glycoproteins.

    PubMed

    Pickering, J M; Smith, H; Sweet, C

    1992-06-01

    Ultraviolet light-inactivated, non-infectious influenza virus is pyrogenic; virion components are probably responsible for this pyrogenicity. To try to identify the pyrogenic component, influenza virions were disrupted with either bromelain or sodium deoxycholate (DOC). Treatment of infectious virions with bromelain, under conditions that removed the surface glycoproteins (spikes), destroyed their pyrogenicity. The supernatant, containing non-aggregated and modified glycoproteins, was also non-pyrogenic. Disruption of virions with DOC considerably reduced pyrogenicity; however, some was retained by the sub-viral cores. Viral nucleoprotein and matrix protein, purified from the supernatant, were non-pyrogenic. Aggregated stellate clusters of surface glycoproteins separated on sucrose gradients were pyrogenic in half of numerous tests performed with different batches of material. Treatment of virus with ether resulted in complete loss of pyrogenicity. Liposomes made from extracted viral lipid were non-pyrogenic. In contrast, virosomes made from the viral lipid and the aggregated stellate clusters of surface glycoproteins were pyrogenic. Hence, optimum pyrogenicity depends upon the integrity of the virus particle, but haemagglutinin and/or neuraminidase appear essential, and lipid may be involved. PMID:1607857

  6. Reducing infection risk in implant-based breast-reconstruction surgery: challenges and solutions.

    PubMed

    Ooi, Adrian Sh; Song, David H

    2016-01-01

    Implant-based procedures are the most commonly performed method for postmastectomy breast reconstruction. While donor-site morbidity is low, these procedures are associated with a higher risk of reconstructive loss. Many of these are related to infection of the implant, which can lead to prolonged antibiotic treatment, undesired additional surgical procedures, and unsatisfactory results. This review combines a summary of the recent literature regarding implant-related breast-reconstruction infections and combines this with a practical approach to the patient and surgery aimed at reducing this risk. Prevention of infection begins with appropriate reconstructive choice based on an assessment and optimization of risk factors. These include patient and disease characteristics, such as smoking, obesity, large breast size, and immediate reconstructive procedures, as well as adjuvant therapy, such as radiotherapy and chemotherapy. For implant-based breast reconstruction, preoperative planning and organization is key to reducing infection. A logical and consistent intraoperative and postoperative surgical protocol, including appropriate antibiotic choice, mastectomy-pocket creation, implant handling, and considered acellular dermal matrix use contribute toward the reduction of breast-implant infections. PMID:27621667

  7. Interleukin-21 combined with ART reduces inflammation and viral reservoir in SIV-infected macaques

    PubMed Central

    Micci, Luca; Ryan, Emily S.; Fromentin, Rémi; Bosinger, Steven E.; Harper, Justin L.; He, Tianyu; Paganini, Sara; Easley, Kirk A.; Chahroudi, Ann; Benne, Clarisse; Gumber, Sanjeev; McGary, Colleen S.; Rogers, Kenneth A.; Deleage, Claire; Lucero, Carissa; Byrareddy, Siddappa N.; Apetrei, Cristian; Estes, Jacob D.; Lifson, Jeffrey D.; Piatak, Michael; Chomont, Nicolas; Villinger, Francois; Silvestri, Guido; Brenchley, Jason M.; Paiardini, Mirko

    2015-01-01

    Despite successful control of viremia, many HIV-infected individuals given antiretroviral therapy (ART) exhibit residual inflammation, which is associated with non–AIDS-related morbidity and mortality and may contribute to virus persistence during ART. Here, we investigated the effects of IL-21 administration on both inflammation and virus persistence in ART-treated, SIV-infected rhesus macaques (RMs). Compared with SIV-infected animals only given ART, SIV-infected RMs given both ART and IL-21 showed improved restoration of intestinal Th17 and Th22 cells and a more effective reduction of immune activation in blood and intestinal mucosa, with the latter maintained through 8 months after ART interruption. Additionally, IL-21, in combination with ART, was associated with reduced levels of SIV RNA in plasma and decreased CD4+ T cell levels harboring replication-competent virus during ART. At the latest experimental time points, which were up to 8 months after ART interruption, plasma viremia and cell-associated SIV DNA levels remained substantially lower than those before ART initiation in IL-21–treated animals but not in controls. Together, these data suggest that IL-21 supplementation of ART reduces residual inflammation and virus persistence in a relevant model of lentiviral disease and warrants further investigation as a potential intervention for HIV infection. PMID:26551680

  8. Reducing infection risk in implant-based breast-reconstruction surgery: challenges and solutions

    PubMed Central

    Ooi, Adrian SH; Song, David H

    2016-01-01

    Implant-based procedures are the most commonly performed method for postmastectomy breast reconstruction. While donor-site morbidity is low, these procedures are associated with a higher risk of reconstructive loss. Many of these are related to infection of the implant, which can lead to prolonged antibiotic treatment, undesired additional surgical procedures, and unsatisfactory results. This review combines a summary of the recent literature regarding implant-related breast-reconstruction infections and combines this with a practical approach to the patient and surgery aimed at reducing this risk. Prevention of infection begins with appropriate reconstructive choice based on an assessment and optimization of risk factors. These include patient and disease characteristics, such as smoking, obesity, large breast size, and immediate reconstructive procedures, as well as adjuvant therapy, such as radiotherapy and chemotherapy. For implant-based breast reconstruction, preoperative planning and organization is key to reducing infection. A logical and consistent intraoperative and postoperative surgical protocol, including appropriate antibiotic choice, mastectomy-pocket creation, implant handling, and considered acellular dermal matrix use contribute toward the reduction of breast-implant infections. PMID:27621667

  9. Using Reduced Personal Protective Equipment in an Endemically Infected Mouse Colony

    PubMed Central

    Baker, Samuel W; Prestia, Kevin A; Karolewski, Brian

    2014-01-01

    Personal protective equipment (PPE) frequently is used to reduce the risk of spreading adventitial diseases in rodent colonies. The PPE worn often reflects the historic practices of the research institution rather than published performance data. Standard PPE for a rodent facility typically consists of a disposable hair bonnet, gown, face mask, shoe covers, and gloves, which are donned on facility entry and removed on exiting. This study evaluated the effect of a reduced PPE protocol on disease spread within an endemically infected mouse colony. In the reduced protocol, only the parts of the wearer that came in direct contact with the mice or their environment were covered with PPE. To test the reduced PPE protocol, proven naïve mice were housed in a facility endemically infected with murine norovirus and mouse hepatitis virus for 12 wk. During that time, routine husbandry operations were conducted by using either the standard or reduced PPE protocols. All study mice remained free of virus antibody when reduced PPE was implemented. These results indicate that reduced PPE is adequate for disease containment when correct techniques for handling microisolation caging are used. Reducing the amount of PPE used in an animal facility affords considerable cost savings yet limits the risk of disease spread. PMID:24827569

  10. Filter-feeding bivalves can remove avian influenza viruses from water and reduce infectivity

    PubMed Central

    Faust, Christina; Stallknecht, David; Swayne, David; Brown, Justin

    2009-01-01

    Avian influenza (AI) viruses are believed to be transmitted within wild aquatic bird populations through an indirect faecal–oral route involving contaminated water. This study examined the influence of filter-feeding bivalves, Corbicula fluminea, on the infectivity of AI virus in water. Clams were placed into individual flasks with distilled water inoculated 1:100 with a low pathogenic (LP) AI virus (A/Mallard/MN/190/99 (H3N8)). Viral titres in water with clams were significantly lower at 24 and 48 h post-inoculation compared to LPAI-infected water without clams. To determine whether clams affected the infectivity of AI viruses, 18 wood ducks (Aix sponsa) were divided into test groups and inoculated with a variety of treatments of clam supernatants, whole clams and water exposed to a high pathogenic (HP) AI (A/whooper swan/Mongolia/244/05 (H5N1)). None of the wood ducks inoculated with HPAI-infected water that was filtered by clams or that was inoculated with or fed tissue from these clams exhibited morbidity or mortality. All wood ducks exposed to either HPAI-infected water without clams or the original viral inoculum died. These results indicate that filter-feeding bivalves can remove and reduce the infectivity of AI viruses in water and demonstrate the need to examine biotic environmental factors that can influence AI virus transmission. PMID:19656788

  11. Effects of Substituting Granulin or a Granulin-Polyhedrin Chimera for Polyhedrin on Virion Occlusion and Polyhedral Morphology in Autographa californica Multinucleocapsid Nuclear Polyhedrosis Virus

    PubMed Central

    Eason, Jane E.; Hice, Robert H.; Johnson, Jeffrey J.; Federici, Brian A.

    1998-01-01

    Substitution of granulin from the Trichoplusia ni granulosis virus (TnGV) for polyhedrin of the Autographa californica multinucleocapsid nuclear polyhedrosis virus (AcMNPV) yielded a few very large (2 to 5 μm) cuboidal inclusions in the cytoplasm and nucleus of infected cells. These polyhedra lacked the beveled edges characteristic of wild-type AcMNPV polyhedra, contained fractures, and occluded few virions. Placing a nuclear localization signal (KRKK) in granulin directed more granulin to the nucleus and resulted in more structurally uniform cuboidal inclusions in which no virions were observed. A granulin-polyhedrin chimera produced tetrahedral occlusions with more virions than granulin inclusions but many fewer than wild-type polyhedra. Despite the unusual structure of the granulin and granulin-polyhedrin inclusions, they interacted with AcMNPV p10 fibrillar structures and electron-dense spacers that are precursors of the polyhedral calyx. The change in inclusion shape obtained with the granulin-polyhedrin chimera demonstrates that the primary amino acid sequence affects occlusion body shape, but the large cuboidal inclusions formed by granulin indicate that the amino acid sequence is not the only determinant. The failure of granulin or the granulin-polyhedrin chimera to properly occlude AcMNPV virions suggests that specific interactions occur between polyhedrin and other viral proteins which facilitate normal virion occlusion and occlusion body assembly and shape in baculoviruses. PMID:9621097

  12. Vaccination with proteins involved in tick-pathogen interactions reduces vector infestations and pathogen infection.

    PubMed

    Merino, Octavio; Antunes, Sandra; Mosqueda, Juan; Moreno-Cid, Juan A; Pérez de la Lastra, José M; Rosario-Cruz, Rodrigo; Rodríguez, Sergio; Domingos, Ana; de la Fuente, José

    2013-12-01

    Tick-borne pathogens cause diseases that greatly impact animal health and production worldwide. The ultimate goal of tick vaccines is to protect against tick-borne diseases through the control of vector infestations and reducing pathogen infection and transmission. Tick genetic traits are involved in vector-pathogen interactions and some of these molecules such as Subolesin (SUB) have been shown to protect against vector infestations and pathogen infection. Based on these premises, herein we characterized the efficacy of cattle vaccination with tick proteins involved in vector-pathogen interactions, TROSPA, SILK, and Q38 for the control of cattle tick, Rhipicephalus (Boophilus) microplus infestations and infection with Anaplasma marginale and Babesia bigemina. SUB and adjuvant/saline placebo were used as positive and negative controls, respectively. The results showed that vaccination with Q38, SILK and SUB reduced tick infestations and oviposition with vaccine efficacies of 75% (Q38), 62% (SILK) and 60% (SUB) with respect to ticks fed on placebo control cattle. Vaccination with TROSPA did not have a significant effect on any of the tick parameters analyzed. The results also showed that vaccination with Q38, TROSPA and SUB reduced B. bigemina DNA levels in ticks while vaccination with SILK and SUB resulted in lower A. marginale DNA levels when compared to ticks fed on placebo control cattle. The positive correlation between antigen-specific antibody titers and reduction of tick infestations and pathogen infection strongly suggested that the effect of the vaccine was the result of the antibody response in vaccinated cattle. Vaccination and co-infection with A. marginale and B. bigemina also affected the expression of genes encoding for vaccine antigens in ticks fed on cattle. These results showed that vaccines using tick proteins involved in vector-pathogen interactions could be used for the dual control of tick infestations and pathogen infection. PMID:24084474

  13. Antigenic Properties of the Human Immunodeficiency Virus Envelope Glycoprotein Gp120 on Virions Bound to Target Cells

    PubMed Central

    Mengistu, Meron; Ray, Krishanu; Lewis, George K.; DeVico, Anthony L.

    2015-01-01

    The HIV-1 envelope glycoprotein, gp120, undergoes multiple molecular interactions and structural rearrangements during the course of host cell attachment and viral entry, which are being increasingly defined at the atomic level using isolated proteins. In comparison, antigenic markers of these dynamic changes are essentially unknown for single HIV-1 particles bound to target cells. Such markers should indicate how neutralizing and/or non-neutralizing antibodies might interdict infection by either blocking infection or sensitizing host cells for elimination by Fc-mediated effector function. Here we address this deficit by imaging fluorescently labeled CCR5-tropic HIV-1 pseudoviruses using confocal and superresolution microscopy to track the exposure of neutralizing and non-neutralizing epitopes as they appear on single HIV-1 particles bound to target cells. Epitope exposure was followed under conditions permissive or non-permissive for viral entry to delimit changes associated with virion binding from those associated with post-attachment events. We find that a previously unexpected array of gp120 epitopes is exposed rapidly upon target cell binding. This array comprises both neutralizing and non-neutralizing epitopes, the latter being hidden on free virions yet capable of serving as potent targets for Fc-mediated effector function. Under non-permissive conditions for viral entry, both neutralizing and non-neutralizing epitope exposures were relatively static over time for the majority of bound virions. Under entry-permissive conditions, epitope exposure patterns changed over time on subsets of virions that exhibited concurrent variations in virion contents. These studies reveal that bound virions are distinguished by a broad array of both neutralizing and non-neutralizing gp120 epitopes that potentially sensitize a freshly engaged target cell for destruction by Fc-mediated effector function and/or for direct neutralization at a post-binding step. The elucidation of

  14. So near and yet so far: harmonic radar reveals reduced homing ability of Nosema infected honeybees.

    PubMed

    Wolf, Stephan; McMahon, Dino P; Lim, Ka S; Pull, Christopher D; Clark, Suzanne J; Paxton, Robert J; Osborne, Juliet L

    2014-01-01

    Pathogens may gain a fitness advantage through manipulation of the behaviour of their hosts. Likewise, host behavioural changes can be a defence mechanism, counteracting the impact of pathogens on host fitness. We apply harmonic radar technology to characterize the impact of an emerging pathogen--Nosema ceranae (Microsporidia)--on honeybee (Apis mellifera) flight and orientation performance in the field. Honeybees are the most important commercial pollinators. Emerging diseases have been proposed to play a prominent role in colony decline, partly through sub-lethal behavioural manipulation of their hosts. We found that homing success was significantly reduced in diseased (65.8%) versus healthy foragers (92.5%). Although lost bees had significantly reduced continuous flight times and prolonged resting times, other flight characteristics and navigational abilities showed no significant difference between infected and non-infected bees. Our results suggest that infected bees express normal flight characteristics but are constrained in their homing ability, potentially compromising the colony by reducing its resource inputs, but also counteracting the intra-colony spread of infection. We provide the first high-resolution analysis of sub-lethal effects of an emerging disease on insect flight behaviour. The potential causes and the implications for both host and parasite are discussed. PMID:25098331

  15. So Near and Yet So Far: Harmonic Radar Reveals Reduced Homing Ability of Nosema Infected Honeybees

    PubMed Central

    Wolf, Stephan; McMahon, Dino P.; Lim, Ka S.; Pull, Christopher D.; Clark, Suzanne J.; Paxton, Robert J.; Osborne, Juliet L.

    2014-01-01

    Pathogens may gain a fitness advantage through manipulation of the behaviour of their hosts. Likewise, host behavioural changes can be a defence mechanism, counteracting the impact of pathogens on host fitness. We apply harmonic radar technology to characterize the impact of an emerging pathogen - Nosema ceranae (Microsporidia) - on honeybee (Apis mellifera) flight and orientation performance in the field. Honeybees are the most important commercial pollinators. Emerging diseases have been proposed to play a prominent role in colony decline, partly through sub-lethal behavioural manipulation of their hosts. We found that homing success was significantly reduced in diseased (65.8%) versus healthy foragers (92.5%). Although lost bees had significantly reduced continuous flight times and prolonged resting times, other flight characteristics and navigational abilities showed no significant difference between infected and non-infected bees. Our results suggest that infected bees express normal flight characteristics but are constrained in their homing ability, potentially compromising the colony by reducing its resource inputs, but also counteracting the intra-colony spread of infection. We provide the first high-resolution analysis of sub-lethal effects of an emerging disease on insect flight behaviour. The potential causes and the implications for both host and parasite are discussed. PMID:25098331

  16. Targeting α4β7 integrin reduces mucosal transmission of SIV and protects GALT from infection

    PubMed Central

    Byrareddy, Siddappa N.; Kallam, Brianne; Arthos, James; Cicala, Claudia; Nawaz, Fatima; Hiatt, Joseph; Kersh, Ellen N.; McNicholl, Janet M.; Hanson, Debra; Reimann, Keith A.; Brameier, Markus; Walter, Lutz; Rogers, Kenneth; Mayne, Ann E.; Dunbar, Paul; Villinger, Tara; Little, Dawn; Parslow, Tristram G.; Santangelo, Philip J.; Villinger, Francois; Fauci, Anthony S.; Ansari, Aftab A.

    2014-01-01

    α4β7 integrin expressing CD4+ T cells preferentially traffic to gut-associated lymphoid tissues (GALT) and play a key role in HIV/SIV pathogenesis. The administration of an anti-α4β7 monoclonal antibody during acute infection protects macaques from transmission following repeated low-dose intra-vaginal challenges with SIVmac251. In treated animals that became infected the GALT was significantly protected and CD4+ T–cell numbers were maintained. Thus, targeting α4β7 reduces mucosal transmission of SIV in macaques. PMID:25419708

  17. α1-Antitrypsin reduces rhinovirus infection in primary human airway epithelial cells exposed to cigarette smoke

    PubMed Central

    Berman, Reena; Jiang, Di; Wu, Qun; Chu, Hong Wei

    2016-01-01

    Human rhinovirus (HRV) infections target airway epithelium and are the leading cause of acute exacerbations of COPD. Cigarette smoke (CS) increases the severity of viral infections, but there is no effective therapy for HRV infection. We determined whether α1-antitrypsin (A1AT) reduces HRV-16 infection in CS-exposed primary human airway epithelial cells. Brushed bronchial epithelial cells from normal subjects and patients diagnosed with COPD were cultured at air–liquid interface to induce mucociliary differentiation. These cells were treated with A1AT or bovine serum albumin for 2 hours and then exposed to air or whole cigarette smoke (WCS) with or without HRV-16 (5×104 50% Tissue Culture Infective Dose [TCID50]/transwell) infection for 24 hours. WCS exposure significantly increased viral load by an average of fivefold and decreased the expression of antiviral genes interferon-λ1, OAS1, and MX1. When A1AT was added to WCS-exposed cells, viral load significantly decreased by an average of 29-fold. HRV-16 infection significantly increased HRV-16 receptor intercellular adhesion molecule-1 messenger RNA expression in air-exposed cells, which was decreased by A1AT. A1AT-mediated reduction of viral load was not accompanied by increased epithelial antiviral gene expression or by inhibiting the activity of 3C protease involved in viral replication or maturation. Our findings demonstrate that A1AT treatment prevents a WCS-induced increase in viral load and for the first time suggest a therapeutic effect of A1AT on HRV infection. PMID:27354786

  18. Antigenic Properties of the HIV Envelope on Virions in Solution

    PubMed Central

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

    2014-01-01

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

  19. Highly divergent strains of porcine reproductive and respiratory syndrome virus incorporate multiple isoforms of nonstructural protein 2 into virions.

    PubMed

    Kappes, Matthew A; Miller, Cathy L; Faaberg, Kay S

    2013-12-01

    Viral structural proteins form the critical intermediary between viral infection cycles within and between hosts, function to initiate entry, participate in immediate early viral replication steps, and are major targets for the host adaptive immune response. We report the identification of nonstructural protein 2 (nsp2) as a novel structural component of the porcine reproductive and respiratory syndrome virus (PRRSV) particle. A set of custom α-nsp2 antibodies targeting conserved epitopes within four distinct regions of nsp2 (the PLP2 protease domain [OTU], the hypervariable domain [HV], the putative transmembrane domain [TM], and the C-terminal region [C]) were obtained commercially and validated in PRRSV-infected cells. Highly purified cell-free virions of several PRRSV strains were isolated through multiple rounds of differential density gradient centrifugation and analyzed by immunoelectron microscopy (IEM) and Western blot assays using the α-nsp2 antibodies. Purified viral preparations were found to contain pleomorphic, predominantly spherical virions of uniform size (57.9 nm ± 8.1 nm diameter; n = 50), consistent with the expected size of PRRSV particles. Analysis by IEM indicated the presence of nsp2 associated with the viral particle of diverse strains of PRRSV. Western blot analysis confirmed the presence of nsp2 in purified viral samples and revealed that multiple nsp2 isoforms were associated with the virion. Finally, a recombinant PRRSV genome containing a myc-tagged nsp2 was used to generate purified virus, and these particles were also shown to harbor myc-tagged nsp2 isoforms. Together, these data identify nsp2 as a virion-associated structural PRRSV protein and reveal that nsp2 exists in or on viral particles as multiple isoforms. PMID:24089566

  20. Helicobacter pylori infection reduces disease severity in an experimental model of multiple sclerosis

    PubMed Central

    Cook, Katherine W.; Crooks, James; Hussain, Khiyam; O’Brien, Kate; Braitch, Manjit; Kareem, Huner; Constantinescu, Cris S.; Robinson, Karen; Gran, Bruno

    2015-01-01

    Recent research has demonstrated that infection with the bacterial pathogen Helicobacter pylori is less common amongst patients with multiple sclerosis (MS), an inflammatory demyelinating disease of the central nervous system (CNS). We aimed to compare the prevalence of H. pylori amongst MS patients and healthy controls, and also investigated the impact of this infection on an animal model for MS, experimental autoimmune encephalomyelitis (EAE). The H. pylori status of 71 MS patients and 42 healthy controls was determined by serology. Groups of C57BL/6 mice were infected with H. pylori, or given diluent alone as a placebo, prior to inducing EAE. Clinical scores were assessed for all mice, and spleens and spinal cord tissue were harvested. CD4+ T cell subsets were quantified by flow cytometry, and T cell proliferation assays were performed. In MS patients the seroprevalence of H. pylori was half that of healthy controls (p = 0.018). Over three independent experiments, prior H. pylori infection had a moderate effect in reducing the severity of EAE (p = 0.012). In line with this, the antigen-specific T cell proliferative responses of infected animals were significantly reduced (p = 0.001), and there was a fourfold reduction in the number of CD4+ cells in the CNS. CD4+ populations in both the CNS and the spleens of infected mice also contained greatly reduced proportions of IFNγ+, IL-17+, T-bet+, and RORγt+ cells, but the proportions of Foxp3+ cells were equivalent. There were no differences in the frequency of splenic CD4+cells expressing markers of apoptosis between infected and uninfected animals. H. pylori was less prevalent amongst MS patients. In mice, the infection exerted some protection against EAE, inhibiting both Th1 and Th17 responses. This could not be explained by the presence of increased numbers of Foxp3+ regulatory T cells, or T cell apoptosis. This is the first direct experimental evidence showing that H. pylori may provide protection against

  1. Cucumber Mosaic Virus as a carotenoid inhibitor reducing Phelipanche aegyptiaca infection in tobacco plants

    PubMed Central

    Ibdah, Mwafaq; Dubey, Neeraj Kumar; Eizenberg, Hanan; Dabour, Ziad; Abu-Nassar, Jacklin; Gal-On, Amit; Aly, Radi

    2014-01-01

    Cucumber Mosaic Virus (CMV) is a highly infectious cucumovirus, which infects more than 800 plant species and causes major diseases in greenhouse and field crops worldwide. Parasitic weeds such as Phelipanche aegyptiaca are a major constraint to the production of many crops in the world and the parasite's lifestyle makes control extremely difficult. The parasite seeds can germinate after conditioning and perceiving strigolactones secreted by the host roots. Strigolactones are rhizosphere signaling molecules in plants that are biosynthesized through carotenoid cleavage. In the present study we investigated the possibility of reducing β-carotene and then strigolactone production in the host roots by blocking carotenoid biosynthesis using CMV-infected tobacco. It was found that CMV downregulated the enzyme phytoene desaturase(PDS) and reduced significantly both carotenoid production and Phelipanche infection in tobacco host roots infected with both CMV and P. aegyptiaca. Based on our results (decrease of β-carotene and repression of PDS transcripts in tobacco roots), we hypothesized that the reduction of Phelipanche tubercles and shoots occurred due to an effect of CMV on secondary metabolite stimulators such as strigolacetones. Our study indicated that mass production of the host roots was not affected by CMV; however, most inflorescences of Phelipanche grown on CMV-infected tobacco developed abnormally (deformed shoots and short nodes). Carotenoid biosynthesis inhibitors such as CMV can be used to reduce the production of strigolactones, which will lead to decreased Phelipanche attachment. Interestingly, attenuated CMV strains may provide a safe means for enhancing crop resistance against parasitic weeds in a future plan. PMID:25482816

  2. Cucumber Mosaic Virus as a carotenoid inhibitor reducing Phelipanche aegyptiaca infection in tobacco plants.

    PubMed

    Ibdah, Mwafaq; Dubey, Neeraj Kumar; Eizenberg, Hanan; Dabour, Ziad; Abu-Nassar, Jacklin; Gal-On, Amit; Aly, Radi

    2014-01-01

    Cucumber Mosaic Virus (CMV) is a highly infectious cucumovirus, which infects more than 800 plant species and causes major diseases in greenhouse and field crops worldwide. Parasitic weeds such as Phelipanche aegyptiaca are a major constraint to the production of many crops in the world and the parasite's lifestyle makes control extremely difficult. The parasite seeds can germinate after conditioning and perceiving strigolactones secreted by the host roots. Strigolactones are rhizosphere signaling molecules in plants that are biosynthesized through carotenoid cleavage. In the present study we investigated the possibility of reducing β-carotene and then strigolactone production in the host roots by blocking carotenoid biosynthesis using CMV-infected tobacco. It was found that CMV downregulated the enzyme phytoene desaturase(PDS) and reduced significantly both carotenoid production and Phelipanche infection in tobacco host roots infected with both CMV and P. aegyptiaca. Based on our results (decrease of β-carotene and repression of PDS transcripts in tobacco roots), we hypothesized that the reduction of Phelipanche tubercles and shoots occurred due to an effect of CMV on secondary metabolite stimulators such as strigolacetones. Our study indicated that mass production of the host roots was not affected by CMV; however, most inflorescences of Phelipanche grown on CMV-infected tobacco developed abnormally (deformed shoots and short nodes). Carotenoid biosynthesis inhibitors such as CMV can be used to reduce the production of strigolactones, which will lead to decreased Phelipanche attachment. Interestingly, attenuated CMV strains may provide a safe means for enhancing crop resistance against parasitic weeds in a future plan. PMID:25482816

  3. Immunogenicity Studies of Bivalent Inactivated Virions of EV71/CVA16 Formulated with Submicron Emulsion Systems

    PubMed Central

    Lin, Chih-Wei; Lu, Tsung-Chun; Chow, Yen-Hung; Huang, Ming-Hsi

    2014-01-01

    We assessed two strategies for preparing candidate vaccines against hand, foot, and mouth disease (HFMD) caused mainly by infections of enterovirus (EV) 71 and coxsackievirus (CV) A16. We firstly design and optimize the potency of adjuvant combinations of emulsion-based delivery systems, using EV71 candidate vaccine as a model. We then perform immunogenicity studies in mice of EV71/CVA16 antigen combinations formulated with PELC/CpG. A single dose of inactivated EV71 virion (0.2 μg) emulsified in submicron particles was found (i) to induce potent antigen-specific neutralizing antibody responses and (ii) consistently to elicit broad antibody responses against EV71 neutralization epitopes. A single dose immunogenicity study of bivalent activated EV71/CVA16 virion formulated with either Alum or PELC/CpG adjuvant showed that CVA16 antigen failed to elicit CVA16 neutralizing antibody responses and did not affect EV71-specific neutralizing antibody responses. A boosting dose of emulsified EV71/CVA16 bivalent vaccine candidate was found to be necessary to achieve high seroconversion of CVA16-specific neutralizing antibody responses. The current results are important for the design and development of prophylactic vaccines against HFMD and other emerging infectious diseases. PMID:25006583

  4. Orsay virus utilizes ribosomal frameshifting to express a novel protein that is incorporated into virions

    SciTech Connect

    Jiang, Hongbing; Franz, Carl J.; Wu, Guang; Renshaw, Hilary; Zhao, Guoyan; Firth, Andrew E.; Wang, David

    2014-02-15

    Orsay virus is the first identified virus that is capable of naturally infecting Caenorhabditis elegans. Although it is most closely related to nodaviruses, Orsay virus differs from nodaviruses in its genome organization. In particular, the Orsay virus RNA2 segment encodes a putative novel protein of unknown function, termed delta, which is absent from all known nodaviruses. Here we present evidence that Orsay virus utilizes a ribosomal frameshifting strategy to express a novel fusion protein from the viral capsid (alpha) and delta ORFs. Moreover, the fusion protein was detected in purified virus fractions, demonstrating that it is most likely incorporated into Orsay virions. Furthermore, N-terminal sequencing of both the fusion protein and the capsid protein demonstrated that these proteins must be translated from a non-canonical initiation site. While the function of the alpha–delta fusion remains cryptic, these studies provide novel insights into the fundamental properties of this new clade of viruses. - Highlights: • Orsay virus encodes a novel fusion protein by a ribosomal frameshifting mechanism. • Orsay capsid and fusion protein is translated from a non-canonical initiation site. • The fusion protein is likely incorporated into Orsay virions.

  5. An Ensemble Method to Distinguish Bacteriophage Virion from Non-Virion Proteins Based on Protein Sequence Characteristics

    PubMed Central

    Zhang, Lina; Zhang, Chengjin; Gao, Rui; Yang, Runtao

    2015-01-01

    Bacteriophage virion proteins and non-virion proteins have distinct functions in biological processes, such as specificity determination for host bacteria, bacteriophage replication and transcription. Accurate identification of bacteriophage virion proteins from bacteriophage protein sequences is significant to understand the complex virulence mechanism in host bacteria and the influence of bacteriophages on the development of antibacterial drugs. In this study, an ensemble method for bacteriophage virion protein prediction from bacteriophage protein sequences is put forward with hybrid feature spaces incorporating CTD (composition, transition and distribution), bi-profile Bayes, PseAAC (pseudo-amino acid composition) and PSSM (position-specific scoring matrix). When performing on the training dataset 10-fold cross-validation, the presented method achieves a satisfactory prediction result with a sensitivity of 0.870, a specificity of 0.830, an accuracy of 0.850 and Matthew’s correlation coefficient (MCC) of 0.701, respectively. To evaluate the prediction performance objectively, an independent testing dataset is used to evaluate the proposed method. Encouragingly, our proposed method performs better than previous studies with a sensitivity of 0.853, a specificity of 0.815, an accuracy of 0.831 and MCC of 0.662 on the independent testing dataset. These results suggest that the proposed method can be a potential candidate for bacteriophage virion protein prediction, which may provide a useful tool to find novel antibacterial drugs and to understand the relationship between bacteriophage and host bacteria. For the convenience of the vast majority of experimental scientists, a user-friendly and publicly-accessible web-server for the proposed ensemble method is established. PMID:26370987

  6. Rifampin Reduces Concentrations of Trimethoprim and Sulfamethoxazole in Serum in Human Immunodeficiency Virus-Infected Patients

    PubMed Central

    Ribera, Esteban; Pou, Leonor; Fernandez-Sola, Antoni; Campos, Francisco; Lopez, Rosa M.; Ocaña, Imma; Ruiz, Isabel; Pahissa, Albert

    2001-01-01

    To determine whether rifampin reduces concentrations of trimethoprim (TMP) and sulfamethoxazole (SMX) in serum of human immunodeficiency virus (HIV)-infected persons, levels of these agents were determined by high-performance liquid chromatography before and after more than 12 days of standard antituberculosis treatment for 10 patients who had been taking one double-strength tablet of co-trimoxazole once daily for more than 1 month. Statistically significant, 47 and 23% decreases in TMP and SMX mean areas under the concentration-time curve from 0 to 24 h (AUC0–24), respectively, were observed after administration of rifampin. N-Acetyl-SMX profiles without and with rifampin were similar. The steady-state AUC0–24 metabolite/parent drug ratio increased by 32% with rifampin administration. Our study shows that rifampin reduces profiles of TMP and SMX in serum of HIV-infected patients. PMID:11600390

  7. Growing steers grazing high versus low endophyte (Neotyphodium coenophialum)-infected tall fescue have reduced serum enzymes increased hepatic glucogenic enzymes and reduced liver and carcass mass

    Technology Transfer Automated Retrieval System (TEKTRAN)

    It is well established that grazing Neotyphodium coenophialum-infected forages results in reduced weight gain and serum prolactin levels of cattle. The objective of this study was to determine the potential effects of toxic endophyte-infected tall fescue consumption on carcass characteristics, bloo...

  8. Enhancing Patient Safety by Reducing Healthcare-Associated Infections: The Role of Discovery and Dissemination

    PubMed Central

    2010-01-01

    Healthcare-associated infections (HAIs) take a major human toll on society and reduce public confidence in the healthcare system. The current convergence of scientific, public, and legislative interest in reducing rates of HAI can provide the necessary momentum to address and answer important questions in HAI research. This position paper outlines priorities for a national approach to HAIs: scrutinizing the science base, developing a prioritized research agenda, conducting studies that address the questions that have been identified, creating and deploying guidelines that are based on the outcomes of these studies, and then initiating new studies that assess the efficacy of the interventions. PMID:20038249

  9. Isolation of a herpes simplex virus type 1 mutant with a deletion in the virion host shutoff gene and identification of multiple forms of the vhs (UL41) polypeptide.

    PubMed Central

    Read, G S; Karr, B M; Knight, K

    1993-01-01

    The virion host shutoff (vhs) gene (UL41) of herpes simplex virus type 1 (HSV-1) encodes a virion component that induces degradation of host mRNAs and the shutoff of most host protein synthesis. Subsequently, the vhs protein accelerates the turnover of all kinetic classes of viral mRNA. To identify the vhs (UL41) polypeptide within infected cells and virions, antisera raised against a UL41-lacZ fusion protein were used to characterize the polypeptides encoded by wild-type HSV-1 and two mutants: vhs1, a previously characterized mutant that lacks detectable virion host shutoff activity, and vhs-delta Sma, a newly constructed mutant containing a deletion of 196 codons from UL41. Two forms of the vhs (UL41) polypeptide were identified in cells infected with the wild-type virus or vhs1. Wild-type HSV-1 produced a major 58-kDa polypeptide, as well as a less abundant 59.5-kDa form of the protein, while vhs1 produced 57- and 59-kDa polypeptides that were approximately equally abundant. Although for either virus, both forms of the protein were phosphorylated, they differed in the extent of phosphorylation. While both vhs polypeptides were found in infected cells, only the faster migrating, less phosphorylated form was incorporated into virions. vhs-delta Sma encoded a smaller, 31-kDa polypeptide which, although present in infected cells, was not incorporated into virions. The results identify multiple forms of the vhs (UL41) polypeptide and suggest that posttranslational processing affects its packaging into virions, as well as its ability to induce mRNA degradation. Images PMID:8230437

  10. Nosema ceranae Can Infect Honey Bee Larvae and Reduces Subsequent Adult Longevity.

    PubMed

    Eiri, Daren M; Suwannapong, Guntima; Endler, Matthew; Nieh, James C

    2015-01-01

    Nosema ceranae causes a widespread disease that reduces honey bee health but is only thought to infect adult honey bees, not larvae, a critical life stage. We reared honey bee (Apis mellifera) larvae in vitro and provide the first demonstration that N. ceranae can infect larvae and decrease subsequent adult longevity. We exposed three-day-old larvae to a single dose of 40,000 (40K), 10,000 (10K), zero (control), or 40K autoclaved (control) N. ceranae spores in larval food. Spores developed intracellularly in midgut cells at the pre-pupal stage (8 days after egg hatching) of 41% of bees exposed as larvae. We counted the number of N. ceranae spores in dissected bee midguts of pre-pupae and, in a separate group, upon adult death. Pre-pupae exposed to the 10K or 40K spore treatments as larvae had significantly elevated spore counts as compared to controls. Adults exposed as larvae had significantly elevated spore counts as compared to controls. Larval spore exposure decreased longevity: a 40K treatment decreased the age by which 75% of adult bees died by 28%. Unexpectedly, the low dose (10K) led to significantly greater infection (1.3 fold more spores and 1.5 fold more infected bees) than the high dose (40K) upon adult death. Differential immune activation may be involved if the higher dose triggered a stronger larval immune response that resulted in fewer adult spores but imposed a cost, reducing lifespan. The impact of N. ceranae on honey bee larval development and the larvae of naturally infected colonies therefore deserve further study. PMID:26018139

  11. Nosema ceranae Can Infect Honey Bee Larvae and Reduces Subsequent Adult Longevity

    PubMed Central

    Eiri, Daren M.; Suwannapong, Guntima; Endler, Matthew; Nieh, James C.

    2015-01-01

    Nosema ceranae causes a widespread disease that reduces honey bee health but is only thought to infect adult honey bees, not larvae, a critical life stage. We reared honey bee (Apis mellifera) larvae in vitro and provide the first demonstration that N. ceranae can infect larvae and decrease subsequent adult longevity. We exposed three-day-old larvae to a single dose of 40,000 (40K), 10,000 (10K), zero (control), or 40K autoclaved (control) N. ceranae spores in larval food. Spores developed intracellularly in midgut cells at the pre-pupal stage (8 days after egg hatching) of 41% of bees exposed as larvae. We counted the number of N. ceranae spores in dissected bee midguts of pre-pupae and, in a separate group, upon adult death. Pre-pupae exposed to the 10K or 40K spore treatments as larvae had significantly elevated spore counts as compared to controls. Adults exposed as larvae had significantly elevated spore counts as compared to controls. Larval spore exposure decreased longevity: a 40K treatment decreased the age by which 75% of adult bees died by 28%. Unexpectedly, the low dose (10K) led to significantly greater infection (1.3 fold more spores and 1.5 fold more infected bees) than the high dose (40K) upon adult death. Differential immune activation may be involved if the higher dose triggered a stronger larval immune response that resulted in fewer adult spores but imposed a cost, reducing lifespan. The impact of N. ceranae on honey bee larval development and the larvae of naturally infected colonies therefore deserve further study. PMID:26018139

  12. Moderate physical exercise reduces parasitaemia and protects colonic myenteric neurons in mice infected with Trypanosoma cruzi

    PubMed Central

    Moreira, Neide M; Santos, Franciele d N; Toledo, Max Jean d O; Moraes, Solange M F d; Araujo, Eduardo J d A; Sant'Ana, Debora d M G; Araujo, Silvana M d

    2013-01-01

    This study evaluated the influence of moderate physical exercise on the myenteric neurons in the colonic intestinal wall of mice that had been infected with Trypanosoma cruzi. Parasitology and immunological aspects of the mice were considered. Forty-day-old male Swiss mice were divided into four groups: Trained Infected (TI), Sedentary Infected (SI), Trained Control (TC), and Sedentary Control (SC). The TC and TI were subjected to a moderate physical exercise program on a treadmill for 8 weeks. Three days after finishing exercise, the TI and SI groups were inoculated with 1,300 blood trypomastigotes of the Y strain-T. cruzi. After 75 days of infection results were obtained. Kruskal-Wallis or Analyze of variance (Tukey post hoc test) at 5% level of significance was performed. Moderate physical exercise reduced both the parasite peak (day 8 of infection) and total parasitemia compared with the sedentary groups (P < 0.05). This activity also contributed to neuronal survival (P < 0.05). Exercise caused neuronal hypertrophy (P < 0.05) and an increase in the total thickness of the intestinal wall (P < 0.05). The TI group exhibited an increase in the number of intraepithelial lymphocytes (P > 0.05). In trained animals, the number of goblet cells was reduced compared with sedentary animals (P < 0.05). Physical exercise prevented the formation of inflammatory foci in the TI group (P < 0.05) and increased the synthesis of TNF-α (P < 0.05) and TGF-β (P > 0.05). The present results demonstrated the benefits of moderate physical exercise, and reaffirmed the possibility of that it may contribute to improving clinical treatment in Chagas' disease patients. PMID:24205797

  13. Virion Stability Is Important for the Circulative Transmission of Tomato Yellow Leaf Curl Sardinia Virus by Bemisia tabaci, but Virion Access to Salivary Glands Does Not Guarantee Transmissibility▿ †

    PubMed Central

    Caciagli, Piero; Medina Piles, Vicente; Marian, Daniele; Vecchiati, Manuela; Masenga, Vera; Mason, Giovanna; Falcioni, Tania; Noris, Emanuela

    2009-01-01

    The capsid protein (CP) of the monopartite begomovirus Tomato yellow leaf curl Sardinia virus (TYLCSV), family Geminiviridae, is indispensable for plant infection and vector transmission. A region between amino acids 129 and 152 is critical for virion assembly and insect transmissibility. Two previously described mutants, one with a double Q129P Q134H mutation (PNHD) and another with a further D152E change (PNHE), were found nontransmissible (NT). Another NT mutant with a single N130D change (QDQD) was retrieved from a new mutational analysis. In this study, these three NT mutants and the wild-type (wt) virus were compared in their relationships with the whitefly vector Bemisia tabaci and the nonvector Trialeurodes vaporariorum. Retention kinetics of NT mutants were analyzed by quantitative dot blot hybridization in whiteflies fed on infected plants. The QDQD mutant, whose virions appeared nongeminate following purification, was hardly detectable in either whitefly species at any sampling time. The PNHD mutant was acquired and circulated in both whitefly species for up to 10 days, like the wt virus, while PNHE circulated in B. tabaci only. Using immunogold labeling, both PNHD and PNHE CPs were detected in B. tabaci salivary glands (SGs) like the wt virus, while no labeling was found in any whitefly tissue with the QDQD mutant. Significant inhibition of transmission of the wt virus was observed after prior feeding of the insects on plants infected with the PNHE mutant, but not on plants infected with the other mutants. Virion stability and ability to cross the SG barrier are necessary for TYLCSV transmission, but interactions with molecular components inside the SGs are also critical for transmissibility. PMID:19321611

  14. Oseltamivir reduces hippocampal abnormal EEG activities after a virus infection (influenza) in isoflurane-anesthetized rats

    PubMed Central

    Cissé, Youssouf; Inoue, Isao; Kido, Hiroshi

    2012-01-01

    Background Oseltamivir phosphate (OP, Tamiflu®) is a widely used drug in the treatment of influenza with fever. However, case reports have associated OP intake with sudden abnormal behaviors. In rats infected by the influenza A virus (IAV), the electroencephalogram (EEG) displayed abnormal high-voltage amplitudes with spikes and theta oscillations at a core temperature of 39.9°C to 41°C. Until now, there has been no information describing the effect of OP on intact brain hippocampal activity of IAV-infected animals during hyperthermia. Objective The aim of the present study was to examine the effect of OP on abnormal EEG activities in the hippocampus using the rat model of influenza-associated encephalopathy. Methods Male Wistar rats aged 3 to 4 weeks were used for the study. Influenza A/WSN/33 strain (1 × 105 plaque forming unit in PBS, 60 µL) was applied intranasally to the rats. To characterize OP effects on the IAV-infected rats, EEG activity was studied more particularly in isoflurane-anesthetized IAV-infected rats during hyperthermia. Results We found that the hippocampal EEG of the OP-administered (10 mg/kg) IAV-infected rats showed significant reduction of the high-voltage amplitudes and spikes, but the theta oscillations, which had been observed only at >40°C in OP non-administered rats, appeared at 38°C core temperature. Atropine (30 mg/kg) blocked the theta oscillations. Conclusion Our data suggest that OP efficiently reduces the abnormal EEG activities after IAV infection during hyperthermia. However, OP administration may stimulate ACh release in rats at normal core temperature.

  15. Reduced central line infection rates in children with leukemia following caregiver training

    PubMed Central

    Lo Vecchio, Andrea; Schaffzin, Joshua K.; Ruberto, Eliana; Caiazzo, Maria Angela; Saggiomo, Loredana; Mambretti, Daniela; Russo, Danila; Crispo, Sara; Continisio, Grazia Isabella; Dello Iacovo, Rossano; Poggi, Vincenzo; Guarino, Alfredo

    2016-01-01

    Abstract Infections are a leading cause of morbidity and mortality in children with acute leukemia. Central-line (CL) devices increase this population's risk of serious infections. Within the context of a quality improvement (QI) project, we tested the effect of caregiver education on CL management on the CL-associated bloodstream infection (CLABSI) rate among children with acute leukemia seen at a large referral center in Italy. The intervention consisted of 9 in-person sessions for education and practice using mannequins and children. One hundred and twenty caregivers agreed to participate in the initiative. One hundred and five (87.5%) completed the training, 5 (4.1%) withdrew after the first session, and 10 (8.3%) withdrew during practical sessions. After educational intervention, the overall CLABSI rate was reduced by 46% (from 6.86 to 3.70/1000 CL-days). CLABSI rate was lower in children whose caregivers completed the training (1.74/1000 CL-days, 95% CI 0.43–6.94) compared with those who did not receive any training (12.2/1000 CL-days, 95% CI 7.08–21.0, P < 0.05) or were in-training (3.96/1000 CL-days, 95% CI 1.98–7.91) at the time of infection. Caregiver training in CL management, applied within a multifaceted QI approach, reduced the rate of CLABSI in children with acute leukemia. Specific training and active involvement of caregivers in CL management may be effective to reduce CLABSI in high-risk children. PMID:27336888

  16. Involvement of the N-Terminal Deubiquitinating Protease Domain of Human Cytomegalovirus UL48 Tegument Protein in Autoubiquitination, Virion Stability, and Virus Entry

    PubMed Central

    Kim, Young-Eui; Oh, Se Eun; Kwon, Ki Mun; Lee, Chan Hee

    2016-01-01

    ABSTRACT Human cytomegalovirus (HCMV) protein pUL48 is closely associated with the capsid and has a deubiquitinating protease (DUB) activity in its N-terminal region. Although this DUB activity moderately increases virus replication in cultured fibroblast cells, the requirements of the N-terminal region of pUL48 in the viral replication cycle are not fully understood. In this study, we characterized the recombinant viruses encoding UL48(ΔDUB/NLS), which lacks the DUB domain and the adjacent nuclear localization signal (NLS), UL48(ΔDUB), which lacks only the DUB, and UL48(Δ360–1200), which lacks the internal region (amino acids 360 to 1200) downstream of the DUB/NLS. While ΔDUB/NLS and Δ360–1200 mutant viruses did not grow in fibroblasts, the ΔDUB virus replicated to titers 100-fold lower than those for wild-type virus and showed substantially reduced viral gene expression at low multiplicities of infection. The DUB domain contained ubiquitination sites, and DUB activity reduced its own proteasomal degradation in trans. Deletion of the DUB domain did not affect the nuclear and cytoplasmic localization of pUL48, whereas the internal region (360–1200) was necessary for cytoplasmic distribution. In coimmunoprecipitation assays, pUL48 interacted with three tegument proteins (pUL47, pUL45, and pUL88) and two capsid proteins (pUL77 and pUL85) but the DUB domain contributed to only pUL85 binding. Furthermore, we found that the ΔDUB virus showed reduced virion stability and less efficiently delivered its genome into the cell than the wild-type virus. Collectively, our results demonstrate that the N-terminal DUB domain of pUL48 contributes to efficient viral growth by regulating its own stability and promoting virion stabilization and virus entry. IMPORTANCE HCMV pUL48 and its herpesvirus homologs play key roles in virus entry, regulation of immune signaling pathways, and virion assembly. The N terminus of pUL48 contains the DUB domain, which is well conserved

  17. Vaccinia virions deficient in transcription enzymes lack a nucleocapsid

    SciTech Connect

    McFadden, Baron D.H.; Moussatche, Nissin; Kelley, Karen; Kang, Byung-Ho; Condit, Richard C.

    2012-12-05

    The poxvirus virion contains an inner tubular nucleocapsid structure. The nucleocapsid is apparently labile to conventional electron microscopy fixation procedures and has therefore been largely ignored for decades. Advancements in electron microscopy sample preparation, notably high pressure freezing, better preserve the nucleocapsid structure. Using high pressure freezing and electron microscopy, we have compared the virion structures of wt virus and mutant viruses known to be deficient in packaging of viral transcription enzymes. We show that the mutant viruses lack a defined nucleocapsid. These results support the hypothesis that the nucleocapsid contains the viral DNA genome complexed with viral transcription enzymes and structural proteins. The studies open the door to further investigation of the composition and ultrastructure of the poxvirus nucleocapsid.

  18. Vaccinia Virions Deficient in Transcription Enzymes Lack a Nucleocapsid

    PubMed Central

    McFadden, Baron D.H.; Moussatche, Nissin; Kelley, Karen; Kang, Byung-Ho; Condit, Richard C.

    2012-01-01

    The poxvirus virion contains an inner tubular nucleocapsid structure. The nucleocapsid is apparently labile to conventional electron microscopy fixation procedures and has therefore been largely ignored for decades. Advancements in electron microscopy sample preparation, notably high pressure freezing, better preserve the nucleocapsid structure. Using high pressure freezing and electron microscopy, we have compared the virion structures of wt virus and mutant viruses known to be deficient in packaging of viral transcription enzymes. We show that the mutant viruses lack a defined nucleocapsid. These results support the hypothesis that the nucleocapsid contains the viral DNA genome complexed with viral transcription enzymes and structural proteins. The studies open the door to further investigation of the composition and ultrastructure of the poxvirus nucleocapsid. PMID:22944110

  19. Reduced Infectivity in Cattle for an Outer Membrane Protein Mutant of Anaplasma marginale

    PubMed Central

    Brayton, Kelly A.; Magunda, Forgivemore; Munderloh, Ulrike G.; Kelley, Karen L.; Barbet, Anthony F.

    2015-01-01

    Anaplasma marginale is the causative agent of anaplasmosis in cattle. Transposon mutagenesis of this pathogen using the Himar1 system resulted in the isolation of an omp10 operon insertional mutant referred to as the omp10::himar1 mutant. The work presented here evaluated if this mutant had morphological and/or growth rate defects compared to wild-type A. marginale. Results showed that the morphology, developmental cycle, and growth in tick and mammalian cell cultures are similar for the mutant and the wild type. Tick transmission experiments established that tick infection levels with the mutant were similar to those with wild-type A. marginale and that infected ticks successfully infected cattle. However, this mutant exhibited reduced infectivity and growth in cattle. The possibility of transforming A. marginale by transposon mutagenesis coupled with in vitro and in vivo assessment of altered phenotypes can aid in the identification of genes associated with virulence. The isolation of deliberately attenuated organisms that can be evaluated in their natural biological system is an important advance for the rational design of vaccines against this species. PMID:25595772

  20. Extraintestinal Helminth Infection Reduces the Development of Colitis-Associated Tumorigenesis

    PubMed Central

    León-Cabrera, Sonia; Callejas, Blanca E.; Ledesma-Soto, Yadira; Coronel, Jossimar; Pérez-Plasencia, Carlos; Gutiérrez-Cirlos, Emma B.; Ávila-Moreno, Federico; Rodríguez-Sosa, Miriam; Hernández-Pando, Rogelio; Marquina-Castillo, Brenda; Chirino, Yolanda I.; Terrazas, Luis I.

    2014-01-01

    Colitis-associated colorectal cancer (CAC) is one of the most common cancers and is closely related to chronic or deregulated inflammation. Helminthic infections can modulate inflammatory responses in some diseases, but their immunomodulatory role during cancer development remains completely unknown. We have analyzed the role of Taenia crassiceps-induced anti-inflammatory response in determining the outcome of CAC. We show that extraintestinal T. crassiceps infection in CAC mice inhibited colonic inflammatory responses and tumor formation and prevented goblet cell loss. There was also increased expression of IL-4 and alternatively activated macrophages markers in colonic tissue and negative immunomodulation of pro-inflammatory cytokine expression. In addition, T. crassiceps infection prevented the upregulation of β-catenin and CXCR2 expression observed in the CAC mice, which are both markers associated with CAC-tumorigenesis, and reduced the numbers of circulating and colonic CD11b+Ly6ChiCCR2+ monocytes. Thus, immunomodulatory activities induced by helminth infections may have a role in the progression of CAC. PMID:25210492

  1. Generation of transmissible hepatitis C virions from a molecular clone in chimpanzees.

    PubMed

    Hong, Z; Beaudet-Miller, M; Lanford, R E; Guerra, B; Wright-Minogue, J; Skelton, A; Baroudy, B M; Reyes, G R; Lau, J Y

    1999-03-30

    Multiple alignments of hepatitis C virus (HCV) polyproteins from six different genotypes identified a total of 22 nonconsensus mutations in a clone derived from the Hutchinson (H77) isolate. These mutations, collectively, may have contributed to the failure in generating a "functionally correct" or "infectious" clone in earlier attempts. A consensus clone was constructed after systematic repair of these mutations, which yielded infectious virions in a chimpanzee after direct intrahepatic inoculation of in vitro transcribed RNAs. This RNA-infected chimpanzee has developed hepatitis and remained HCV positive for more than 11 months. To further verify this RNA-derived infectivity, a second naive chimpanzee was injected intravenously with serum collected from the first chimpanzee. Infectivity analysis of the second chimpanzee demonstrated that the HCV infection was successfully transmitted, which validated unequivocally the infectivity of our repaired molecular clone. Amino acid sequence comparisons revealed that our repaired infectious clone had 4 mismatches with the isogenic clone reported by Kolykhalov et al. (1997, Science 277, 570-574) and 8 mismatches with that reported by Yanagi et al. (1997, Proc. Natl. Acad. Sci. USA 94, 8738-8743). At the RNA level, more mismatches (43 and 67, respectively) were identified; most of them were synonymous substitutions. Further comparisons with 16 isolates from different genotypes demonstrated that our repaired clone shares greater consensus than the reported isogenic clones. This approach of generating infectious HCV RNA validates the importance of amino acid sequence consensus in relation to the biology of HCV. PMID:10087224

  2. Platelet-rich plasma inside the sternotomy wound reduces the incidence of sternal wound infections.

    PubMed

    Serraino, Giuseppe F; Dominijanni, Andrea; Jiritano, Federica; Rossi, Michele; Cuda, Aldo; Caroleo, Santo; Brescia, Adalgisa; Renzulli, Attilio

    2015-06-01

    Despite the large choice of wide-spectrum antibiotic therapy, deep sternal wound infection (DSWI) following cardiac surgery is a life-threatening complication worldwide. This study evaluated that the use of platelet-rich plasma (PRP) applied inside the sternotomy wound would reduce the effect of sternal wound infections, both superficial and deep. Between January 2007 and January 2012, 1093 consecutive patients underwent cardiac surgery through median sternotomy. Patients were divided into two groups. Group B, the study group, included those who received the PRP applied inside the sternotomy wound before closure. Group A, the control group, included patients who received a median sternotomy but without the application of PRP. Antibiotic prophylaxis remained unchanged across the study and between the two groups. Occurrence of DSWI was significantly higher in group A than in group B [10 of 671 (1·5%) versus 1 of 422 (0·20%), P = 0·043]. Also, superficial sternal wound infections (SSWIs) were significantly higher in group A than in group B [19 of 671 (2·8%) versus 2 of 422 (0·5%), P = 0·006]. The use of PRP can significantly reduce the occurrence of DSWI and SSWI in cardiac surgery. PMID:23692143

  3. Reducing the risk of HIV infection among South African sex workers: socioeconomic and gender barriers.

    PubMed Central

    Karim, Q A; Karim, S S; Soldan, K; Zondi, M

    1995-01-01

    OBJECTIVES. The social context within which women engaged in sex work at a popular truck stop in South Africa are placed at risk of human immunodeficiency virus (HIV) infection and the factors that influence their ability to reduce their risk were assessed. METHODS. Using qualitative and quantitative techniques, an elected sex worker from within the group collected all data. RESULTS. Given the various pressing needs for basic survival, the risk of HIV infection is viewed as one more burden imposed on these women by their lack of social, legal, and economic power. Violence, or the threat thereof, plays an important role in their disempowerment. In the few instances in which sex workers were able to insist on condom use, it resulted in a decrease in earnings, loss of clients, and physical abuse. CONCLUSIONS. Recommendations to reduce the sex workers' risk for HIV infection include negotiation and communication skills to enable them to persuade their clients to use condoms; development of strategies through which they can maximally use their group strength to facilitate unified action; and accessibility of protective methods they can use and control, such as intravaginal microbicides. PMID:7485664

  4. Ultraviolet-ozone treatment reduces levels of disease-associated prion protein and prion infectivity

    USGS Publications Warehouse

    Johnson, C.J.; Gilbert, P.; McKenzie, D.; Pedersen, J.A.; Aiken, Judd M.

    2009-01-01

    Background. Transmissible spongiform encephalopathies (TSEs) are a group of fatal neurodegenerative diseases caused by novel infectious agents referred to as prions. Prions appear to be composed primarily, if not exclusively, of a misfolded isoform of the cellular prion protein. TSE infectivity is remarkably stable and can resist many aggressive decontamination procedures, increasing human, livestock and wildlife exposure to TSEs. Findings. We tested the hypothesis that UV-ozone treatment reduces levels of the pathogenic prion protein and inactivates the infectious agent. We found that UV-ozone treatment decreased the carbon and prion protein content in infected brain homogenate to levels undetectable by dry-ashing carbon analysis or immunoblotting, respectively. After 8 weeks of ashing, UV-ozone treatment reduced the infectious titer of treated material by a factor of at least 105. A small amount of infectivity, however, persisted despite UV-ozone treatment. When bound to either montmorillonite clay or quartz surfaces, PrPTSE was still susceptible to degradation by UV-ozone. Conclusion. Our findings strongly suggest that UV-ozone treatment can degrade pathogenic prion protein and inactivate prions, even when the agent is associated with surfaces. Using larger UV-ozone doses or combining UV-ozone treatment with other decontaminant methods may allow the sterilization of TSE-contaminated materials. ?? 2009 Aiken et al; licensee BioMed Central Ltd.

  5. Protein Composition of the Vaccinia Virus Mature Virion

    SciTech Connect

    Resch, Wolfgang; Hixson, Kim K.; Moore, Ronald J.; Lipton, Mary S.; Moss, Bernard

    2007-02-05

    The protein content of vaccinia virus mature virions, purified by rate zonal and isopycnic centrifugation and solubilized by SDS or a solution of urea and thiourea, was determined by the accurate mass and time tag technology which uses both tandem mass spectrometry and Fourier transform-ion cyclotron resonance mass spectrometry to detect tryptic peptides separated by high-resolution liquid chromatography. Eighty vaccinia virus-encoded proteins representing 37% of the 218 genes annotated in the complete genome sequence were detected in at least three analyses. Ten proteins accounted for approximately 80% of the mass, while the least abundant proteins made up 1% or less of the mass. Thirteen identified proteins were not previously reported as components of virions. On the other hand, 8 previously described virion proteins were not detected here, presumably due to technical reasons including small size and hydrophobicity. In addition to vaccinia virus-encoded proteins, 24 host proteins omitting isoforms were detected. The most abundant of these were cytoskeletal proteins, heat shock proteins, and proteins involved in translation.

  6. Multiscale Computer Simulation of the Immature HIV-1 Virion

    PubMed Central

    Ayton, Gary S.; Voth, Gregory A.

    2010-01-01

    Multiscale computer simulations, employing a combination of experimental data and coarse-graining methods, are used to explore the structure of the immature HIV-1 virion. A coarse-grained (CG) representation is developed for the virion membrane shell and Gag polypeptides using molecular level information. Building on the results from electron cryotomography experiments, the simulations under certain conditions reveal the existence of an incomplete p6 hexameric lattice formed from hexameric bundles of the Gag CA domains. In particular, the formation and stability of the immature Gag lattice at the CG level requires enhanced interfacial interactions of the CA protein C-terminal domains (CTDs). An exact mapping of the CG representation back to the molecular level then allows for detailed atomistic molecular dynamics studies to confirm the existence of these enhanced CACTD interactions and to probe their possible origin. The multiscale simulations further provide insight into potential CACTD mutations that may disrupt or modify the Gag immature lattice assembly process in the immature HIV-1 virion. PMID:21044572

  7. PPARγ Agonists as an Anti-Inflammatory Treatment Inhibiting Rotavirus Infection of Small Intestinal Villi

    PubMed Central

    Gómez, Dory; Muñoz, Natalia; Guerrero, Rafael; Acosta, Orlando; Guerrero, Carlos A.

    2016-01-01

    Rotavirus infection has been reported to induce an inflammatory response in the host cell accompanied by the increased expression or activation of some cellular molecules including ROS, NF-κB, and COX-2. PPARγ stimulation and N-acetylcysteine (NAC) treatment have been found to interfere with viral infections including rotavirus infection. Small intestinal villi isolated from in vivo infected mice with rotavirus ECwt were analyzed for the percentage of ECwt-infected cells, the presence of rotavirus antigens, and infectious virion yield following treatment with pioglitazone. Isolated villi were also infected in vitro and treated with PPARγ agonists (PGZ, TZD, RGZ, DHA, and ALA), all-trans retinoic acid (ATRA), and NAC. After treatments, the expression of cellular proteins including PPARγ, NF-κB, PDI, Hsc70, and COX-2 was analyzed using immunochemistry, ELISA, immunofluorescence, and Western blotting. The results showed that rotavirus infection led to an increased accumulation of the cellular proteins studied and ROS. The virus infection-induced accumulation of the cellular proteins studied and ROS was reduced upon pioglitazone treatment, causing also a concomitant reduction of the infectious virion yield. We hypothesized that rotavirus infection is benefiting from the induction of a host cell proinflammatory response and that the interference of the inflammatory pathways involved leads to decreased infection. PMID:27382365

  8. Reducing infection in chronic leg ulcers with an activated carbon cloth dressing.

    PubMed

    Murphy, Nina

    2016-06-23

    Zorflex is a new type of antimicrobial dressing composed of 100% activated carbon cloth. It attracts and binds bacteria to its surface, enabling them to be safely removed at dressing change. It has no reported toxic effects and can be used on either a short-or long-term basis. This article describes 4 case studies in which patients with recalcitrant chronic venous leg ulcers that were prone to recurrent infection were treated with the activated carbon cloth dressing. All of the wounds had failed to respond to antimicrobial dressings containing silver, iodine or polyhexamethylene biguanide (PHMB), and were heavily exuding and painful. In all cases, the signs of infection reduced significantly within 4 weeks, resulting in good patient outcomes. PMID:27345081

  9. Immunotherapy of HIV-infected patients with Gc protein-derived macrophage activating factor (GcMAF).

    PubMed

    Yamamoto, Nobuto; Ushijima, Naofumi; Koga, Yoshihiko

    2009-01-01

    Serum Gc protein (known as vitamin D3-binding protein) is the precursor for the principal macrophage activating factor (MAF). The MAF precursor activity of serum Gc protein of HIV-infected patients was lost or reduced because Gc protein is deglycosylated by alpha-N-acetylgalactosaminidase (Nagalase) secreted from HIV-infected cells. Therefore, macrophages of HIV-infected patients having deglycosylated Gc protein cannot be activated, leading to immunosuppression. Since Nagalase is the intrinsic component of the envelope protein gp120, serum Nagalase activity is the sum of enzyme activities carried by both HIV virions and envelope proteins. These Nagalase carriers were already complexed with anti-HIV immunoglobulin G (IgG) but retained Nagalase activity that is required for infectivity. Stepwise treatment of purified Gc protein with immobilized beta-galactosidase and sialidase generated the most potent macrophage activating factor (termed GcMAF), which produces no side effects in humans. Macrophages activated by administration of 100 ng GcMAF develop a large amount of Fc-receptors as well as an enormous variation of receptors that recognize IgG-bound and unbound HIV virions. Since latently HIV-infected cells are unstable and constantly release HIV virions, the activated macrophages rapidly intercept the released HIV virions to prevent reinfection resulting in exhaustion of infected cells. After less than 18 weekly administrations of 100 ng GcMAF for nonanemic patients, they exhibited low serum Nagalase activities equivalent to healthy controls, indicating eradication of HIV-infection, which was also confirmed by no infectious center formation by provirus inducing agent-treated patient PBMCs. No recurrence occurred and their healthy CD + cell counts were maintained for 7 years. PMID:19031451

  10. Identification of Novel Antipoxviral Agents: Mitoxantrone Inhibits Vaccinia Virus Replication by Blocking Virion Assembly▿

    PubMed Central

    Deng, Liang; Dai, Peihong; Ciro, Anthony; Smee, Donald F.; Djaballah, Hakim; Shuman, Stewart

    2007-01-01

    The bioterror threat of a smallpox outbreak in an unvaccinated population has mobilized efforts to develop new antipoxviral agents. By screening a library of known drugs, we identified 13 compounds that inhibited vaccinia virus replication at noncytotoxic doses. The anticancer drug mitoxantrone is unique among the inhibitors identified in that it has no apparent impact on viral gene expression. Rather, it blocks processing of viral structural proteins and assembly of mature progeny virions. The isolation of mitoxantrone-resistant vaccinia strains underscores that a viral protein is the likely target of the drug. Whole-genome sequencing of mitoxantrone-resistant viruses pinpointed missense mutations in the N-terminal domain of vaccinia DNA ligase. Despite its favorable activity in cell culture, mitoxantrone administered intraperitoneally at the maximum tolerated dose failed to protect mice against a lethal intranasal infection with vaccinia virus. PMID:17928345

  11. Moloney murine sarcoma virions synthesize full-genome-length double-stranded DNA in vitro.

    PubMed Central

    Benz, E W; Dina, D

    1979-01-01

    Moloney murine sarcoma virus (MSV) virions incubated under optimal conditions were shown to support extensive synthesis of double-stranded DNA. The major product, a 5950-base-pair (6-kilobase-pair DNA) double-stranded DNA, was characterized by cleavage with restriction endonucleases and shown to contain a 600-nucleotide-long direct repeat at both ends of the MSV genome. Linear DNA molecules made in vivo shortly after infection were compared to the linear double-stranded DNA synthesized in vitro. The restriction maps of both viral DNA products were indistinguishable. The 600-base-pair repeat results in a progeny DNA molecule that is longer than the parental MSV genomic RNA. The generation of this repeat must involve a mechanism that allows the viral reverse transcriptase (RNA-dependent DNA nucleotidyltransferase) to copy 5'- and 3'-terminal genomic (+) strand sequences twice. Images PMID:291003

  12. Efficacy of Clonostachys rosea and Duddingtonia flagrans in Reducing the Haemonchus contortus Infective Larvae

    PubMed Central

    da Silva, Manoel Eduardo; Braga, Fabio Ribeiro; de Gives, Pedro Mendoza; Uriostegui, Miguel Angel Mercado; Reyes, Manuela; Soares, Filippe Elias de Freitas; de Carvalho, Lorendane Millena; Rodrigues, Francielle Bosi; de Araújo, Jackson Victor

    2015-01-01

    The biocontrol is proven effective in reducing in vitro and in situ free-living stages of major gastrointestinal helminths, allowing progress in reducing losses by parasitism, maximizing production, and productivity. This study aimed at evaluating the predatory activity of fungal isolates of Duddingtonia flagrans and Clonostachys rosea species and its association on infective larvae (L3) of H. contortus in microplots formed by grasses and maintained in a protected environment. All groups were added with 10 mL of an aqueous suspension with 618 H. contortus L3 approximately. Group 1 was used as control and only received the infective larvae. Groups 2 and 3 received D. flagrans chlamydospores and C. rosea conidia at doses of 5 × 106. Group 4 received the combination of 5 × 106 D. flagrans chlamydospores + 5 × 106 C. rosea conidia. D. flagrans and C. rosea showed nematicidal effectiveness reducing by 91.5 and 88.9%, respectively, the population of H. contortus L3. However, when used in combination efficiency decreased to 74.5% predation of H. contortus L3. These results demonstrate the need for further studies to determine the existence of additive effects, synergistic or antagonistic, between these species. PMID:26504809

  13. Thioridazine in PLGA nanoparticles reduces toxicity and improves rifampicin therapy against mycobacterial infection in zebrafish.

    PubMed

    Vibe, Carina Beatrice; Fenaroli, Federico; Pires, David; Wilson, Steven Ray; Bogoeva, Vanya; Kalluru, Raja; Speth, Martin; Anes, Elsa; Griffiths, Gareth; Hildahl, Jon

    2016-08-01

    Encapsulating antibiotics such as rifampicin in biodegradable nanoparticles provides several advantages compared to free drug administration, including reduced dosing due to localized targeting and sustained release. Consequently, these characteristics reduce systemic drug toxicity. However, new nanoformulations need to be tested in complex biological systems to fully characterize their potential for improved drug therapy. Tuberculosis, caused by infection with the bacterium Mycobacterium tuberculosis, requires lengthy and expensive treatment, and incomplete therapy contributes to an increasing incidence of drug resistance. Recent evidence suggests that standard therapy may be improved by combining antibiotics with bacterial efflux pump inhibitors, such as thioridazine. However, this drug is difficult to use clinically due to its toxicity. Here, we encapsulated thioridazine in poly(lactic-co-glycolic) acid nanoparticles and tested them alone and in combination with rifampicin nanoparticles, or free rifampicin in macrophages and in a zebrafish model of tuberculosis. Whereas free thioridazine was highly toxic in both cells and zebrafish embryos, after encapsulation in nanoparticles no toxicity was detected. When combined with rifampicin nanoparticles, the nanoparticles loaded with thioridazine gave a modest increase in killing of both Mycobacterium bovis BCG and M. tuberculosis in macrophages. In the zebrafish, the thioridazine nanoparticles showed a significant therapeutic effect in combination with rifampicin by enhancing embryo survival and reducing mycobacterial infection. Our results show that the zebrafish embryo is a highly sensitive indicator of drug toxicity and that thioridazine nanoparticle therapy can improve the antibacterial effect of rifampicin in vivo. PMID:26573343

  14. Small-Molecule Inhibition of Human Immunodeficiency Virus Type 1 Replication by Specific Targeting of the Final Step of Virion Maturation

    PubMed Central

    Zhou, Jing; Yuan, Xiong; Dismuke, David; Forshey, Brett M.; Lundquist, Christopher; Lee, Kuo-Hsiung; Aiken, Christopher; Chen, Chin Ho

    2004-01-01

    Despite the effectiveness of currently available human immunodeficiency virus type 1 (HIV-1) therapies, a continuing need exists for new drugs to treat HIV-1 infection. We investigated the mechanism by which 3-O-{3′,3′-dimethylsuccinyl}-betulinic acid (DSB) inhibits HIV-1 replication. DSB functions at a late stage of the virus life cycle but does not inhibit the HIV-1 protease in vitro or interfere with virus assembly or release. DSB specifically delays the cleavage of Gag between the capsid (CA) and p2, resulting in delayed formation of the mature viral core and reduced HIV-1 infectivity. Replication of simian immunodeficiency virus (SIV) was resistant to DSB; however, a chimeric SIV carrying CA-p2 sequences from HIV-1 was inhibited by the drug, indicating that susceptibility to DSB maps to the CA-p2 region of the HIV-1 Gag protein. A single point mutation at the CA-p2 cleavage site of HIV-1 conferred strong resistance to DSB, confirming the target of the drug. HIV-1 strains that are resistant to a variety of protease inhibitors were sensitive to DSB. These findings indicate that DSB specifically protects the CA-p2 cleavage site from processing by the viral protease during virion maturation, thereby revealing a novel mechanism for pharmacologic inhibition of HIV-1 replication. PMID:14694123

  15. High Dietary Folate in Mice Alters Immune Response and Reduces Survival after Malarial Infection

    PubMed Central

    Meadows, Danielle N.; Bahous, Renata H.; Best, Ana F.; Rozen, Rima

    2015-01-01

    Malaria is a significant global health issue, with nearly 200 million cases in 2013 alone. Parasites obtain folate from the host or synthesize it de novo. Folate consumption has increased in many populations, prompting concerns regarding potential deleterious consequences of higher intake. The impact of high dietary folate on the host’s immune function and response to malaria has not been examined. Our goal was to determine whether high dietary folate would affect response to malarial infection in a murine model of cerebral malaria. Mice were fed control diets (CD, recommended folate level for rodents) or folic acid-supplemented diets (FASD, 10x recommended level) for 5 weeks before infection with Plasmodium berghei ANKA. Survival, parasitemia, numbers of immune cells and other infection parameters were assessed. FASD mice had reduced survival (p<0.01, Cox proportional hazards) and higher parasitemia (p< 0.01, joint model of parasitemia and survival) compared with CD mice. FASD mice had lower numbers of splenocytes, total T cells, and lower numbers of specific T and NK cell sub-populations, compared with CD mice (p<0.05, linear mixed effects). Increased brain TNFα immunoreactive protein (p<0.01, t-test) and increased liver Abca1 mRNA (p<0.01, t-test), a modulator of TNFα, were observed in FASD mice; these variables correlated positively (rs = 0.63, p = 0.01). Bcl-xl/Bak mRNA was increased in liver of FASD mice (p<0.01, t-test), suggesting reduced apoptotic potential. We conclude that high dietary folate increases parasite replication, disturbs the immune response and reduces resistance to malaria in mice. These findings have relevance for malaria-endemic regions, when considering anti-folate anti-malarials, food fortification or vitamin supplementation programs. PMID:26599510

  16. BCG Vaccination Reduces Risk of Tuberculosis Infection in Vaccinated Badgers and Unvaccinated Badger Cubs

    PubMed Central

    Carter, Stephen P.; Chambers, Mark A.; Rushton, Stephen P.; Shirley, Mark D. F.; Schuchert, Pia; Pietravalle, Stéphane; Murray, Alistair; Rogers, Fiona; Gettinby, George; Smith, Graham C.; Delahay, Richard J.; Hewinson, R. Glyn; McDonald, Robbie A.

    2012-01-01

    Wildlife is a global source of endemic and emerging infectious diseases. The control of tuberculosis (TB) in cattle in Britain and Ireland is hindered by persistent infection in wild badgers (Meles meles). Vaccination with Bacillus Calmette-Guérin (BCG) has been shown to reduce the severity and progression of experimentally induced TB in captive badgers. Analysis of data from a four-year clinical field study, conducted at the social group level, suggested a similar, direct protective effect of BCG in a wild badger population. Here we present new evidence from the same study identifying both a direct beneficial effect of vaccination in individual badgers and an indirect protective effect in unvaccinated cubs. We show that intramuscular injection of BCG reduced by 76% (Odds ratio = 0.24, 95% confidence interval (CI) 0.11–0.52) the risk of free-living vaccinated individuals testing positive to a diagnostic test combination to detect progressive infection. A more sensitive panel of tests for the detection of infection per se identified a reduction of 54% (Odds ratio = 0.46, 95% CI 0.26–0.88) in the risk of a positive result following vaccination. In addition, we show the risk of unvaccinated badger cubs, but not adults, testing positive to an even more sensitive panel of diagnostic tests decreased significantly as the proportion of vaccinated individuals in their social group increased (Odds ratio = 0.08, 95% CI 0.01–0.76; P = 0.03). When more than a third of their social group had been vaccinated, the risk to unvaccinated cubs was reduced by 79% (Odds ratio = 0.21, 95% CI 0.05–0.81; P = 0.02). PMID:23251352

  17. Enterovirus 71 Virion-Associated Galectin-1 Facilitates Viral Replication and Stability

    PubMed Central

    Lee, Pei-Huan; Liu, Chia-Ming; Ho, Tzong-Shiann; Tsai, Yi-Che; Lin, Chi-Cheng; Wang, Ya-Fang; Chen, Yuh-Ling; Yu, Chun-Keung; Wang, Shih-Min; Liu, Ching-Chuan; Shiau, Ai-Li; Lei, Huan-Yao; Chang, Chih-Peng

    2015-01-01

    Enterovirus 71 (EV71) infection causes a myriad of diseases from mild hand-foot-and-mouth disease or herpangina to fatal brain stem encephalitis complicated with pulmonary edema. Several severe EV71 endemics have occurred in Asia-Pacific region, including Taiwan, and have become a serious threat to children’s health. EV71 infection is initiated by the attachment of the virion to the target cell surface. Although this process relies primarily upon interaction between viruses and cell surface receptors, soluble factors may also influence the binding of EV71 to host cells.Galectin-1 has been reported to participate in several virus infections, but is not addressed in EV71. In this study, we found that the serum levels of galectin-1 in EV71-infected children were higher than those in non-infected people. In EV71 infected cells, galectin-1 was found to be associated with the EV71 VP1 and VP3 via carbohydrate residues and subsequently released and bound to another cell surface along with the virus. EV71 propagated from galectin-1 knockdown SK-N-SH cells exhibited lower infectivity in cultured cells and less pathogenicity in mice than the virus propagated from parental cells. In addition, this galectin-1-free EV71 virus was sensitive to high temperature and lost its viability after long-term storage, which could be restored following supplement of recombinant galectin-1. Taken together, our findings uncover a new role of galectin-1 in facilitating EV71 virus infection. PMID:25706563

  18. Enterovirus 71 virion-associated galectin-1 facilitates viral replication and stability.

    PubMed

    Lee, Pei-Huan; Liu, Chia-Ming; Ho, Tzong-Shiann; Tsai, Yi-Che; Lin, Chi-Cheng; Wang, Ya-Fang; Chen, Yuh-Ling; Yu, Chun-Keung; Wang, Shih-Min; Liu, Ching-Chuan; Shiau, Ai-Li; Lei, Huan-Yao; Chang, Chih-Peng

    2015-01-01

    Enterovirus 71 (EV71) infection causes a myriad of diseases from mild hand-foot-and-mouth disease or herpangina to fatal brain stem encephalitis complicated with pulmonary edema. Several severe EV71 endemics have occurred in Asia-Pacific region, including Taiwan, and have become a serious threat to children's health. EV71 infection is initiated by the attachment of the virion to the target cell surface. Although this process relies primarily upon interaction between viruses and cell surface receptors, soluble factors may also influence the binding of EV71 to host cells. Galectin-1 has been reported to participate in several virus infections, but is not addressed in EV71. In this study, we found that the serum levels of galectin-1 in EV71-infected children were higher than those in non-infected people. In EV71 infected cells, galectin-1 was found to be associated with the EV71 VP1 and VP3 via carbohydrate residues and subsequently released and bound to another cell surface along with the virus. EV71 propagated from galectin-1 knockdown SK-N-SH cells exhibited lower infectivity in cultured cells and less pathogenicity in mice than the virus propagated from parental cells. In addition, this galectin-1-free EV71 virus was sensitive to high temperature and lost its viability after long-term storage, which could be restored following supplement of recombinant galectin-1. Taken together, our findings uncover a new role of galectin-1 in facilitating EV71 virus infection. PMID:25706563

  19. Defining the level of human immunodeficiency virus type 1 (HIV-1) protease activity required for HIV-1 particle maturation and infectivity.

    PubMed Central

    Rosé, J R; Babé, L M; Craik, C S

    1995-01-01

    The human immunodeficiency virus type 1 (HIV-1) protease is the enzyme required for processing of the Gag and Gag-Pol polyproteins to yield mature, infectious virions. Although the complete absence of proteolytic activity prevents maturation, the level of activity sufficient for maturation and subsequent infectivity has not been determined. Amino acid substitutions that reduce catalytic activity without affecting substrate recognition have been engineered into the active site of the HIV-1 protease. The catalytic efficiency (kcat) of the HIV-1 protease is decreased 4-fold when threonine 26 is replaced by serine (T26S) and approximately 50-fold when alanine 28 is replaced by serine (A28S). Genes containing these mutations were cloned into a proviral vector for analysis of their effects on virion maturation and infectivity. The results show that virions containing the T26S protease variant, in which only 25% of the protease is active, are very similar to wild-type virions, although slight reductions in infectivity are observed. Virions containing the A28S protease variant are not infectious, even though a limited amount of polyprotein processing does occur. There appears to be a linear correlation between the level of protease activity and particle infectivity. Our observations suggest that a threshold of protease activity exists between a 4-fold and 50-fold reduction, below which processing is insufficient to yield infectious particles. Our data also suggest that a reduction of protease activity by 50-fold or greater is sufficient to prevent the formation of infectious particles. PMID:7535864

  20. Secondary Defense Chemicals in Milkweed Reduce Parasite Infection in Monarch Butterflies, Danaus plexippus.

    PubMed

    Gowler, Camden D; Leon, Kristoffer E; Hunter, Mark D; de Roode, Jacobus C

    2015-06-01

    In tri-trophic systems, herbivores may benefit from their host plants in fighting parasitic infections. Plants can provide parasite resistance in two contrasting ways: either directly, by interfering with the parasite, or indirectly, by increasing herbivore immunity or health. In monarch butterflies, the larval diet of milkweed strongly influences the fitness of a common protozoan parasite. Toxic secondary plant chemicals known as cardenolides correlate strongly with parasite resistance of the host, with greater cardenolide concentrations in the larval diet leading to lower parasite growth. However, milkweed cardenolides may covary with other indices of plant quality including nutrients, and a direct experimental link between cardenolides and parasite performance has not been established. To determine if the anti-parasitic activity of milkweeds is indeed due to secondary chemicals, as opposed to nutrition, we supplemented the diet of infected and uninfected monarch larvae with milkweed latex, which contains cardenolides but no nutrients. Across three experiments, increased dietary cardenolide concentrations reduced parasite growth in infected monarchs, which consequently had longer lifespans. However, uninfected monarchs showed no differences in lifespan across treatments, confirming that cardenolide-containing latex does not increase general health. Our results suggest that cardenolides are a driving force behind plant-derived resistance in this system. PMID:25953502

  1. Carotenoid-dependent coloration of male American kestrels predicts ability to reduce parasitic infections

    NASA Astrophysics Data System (ADS)

    Dawson, Russell D.; Bortolotti, Gary R.

    2006-12-01

    The signaling function of sexually selected traits, such as carotenoid-dependent avian plumage coloration, has received a great deal of recent attention especially with respect to parasitism and immunocompetence. We argue that parasite-mediated models of sexual selection may have an implicit temporal component that many researchers have ignored. For example, previous studies have demonstrated that carotenoid-dependent traits can signal past parasite exposure, current levels of parasitism, or the ability of individuals to manage parasitic infections in the future. We examined repeated measures of carotenoid-dependent skin color and blood parasitism in American kestrels ( Falco sparverius) to distinguish whether coloration might signal current parasitism or the potential to deal with infections in the future. We found no evidence that coloration was related to current levels of parasitism in either sex. However, coloration of males significantly predicted their response to parasitism; males with bright orange coloration during prelaying, when mate choice is occurring, were more likely than dull yellow males to reduce their levels of infection by the time incubation began. Coloration during prelaying may advertise a male’s health later in the breeding season. For kestrels, the ability to predict future health would be highly beneficial given the male’s role in providing food to his mate and offspring. Coloration of females was not a significant predictor of parasitism in the future, and we provide several possible explanations for this result.

  2. Improved cell survival by the reduction of immediate-early gene expression in replication-defective mutants of herpes simplex virus type 1 but not by mutation of the virion host shutoff function.

    PubMed Central

    Johnson, P A; Wang, M J; Friedmann, T

    1994-01-01

    42-VP16 double mutant, in 1850 delta 42, showed reduced toxicity only at low multiplicities of infection. To test the role of the virion host shutoff function as an additional candidate to influence virus-induced CPE, we have introduced a large insertion mutation into the virion host shutoff gene of an IE 3 deletion mutant and the double mutant 14H delta 3.(ABSTRACT TRUNCATED AT 400 WORDS) Images PMID:8083974

  3. Reducing fungal infections and testing tag loss in juvenile Pacific lampreys implanted with passive integrated transponders.

    USGS Publications Warehouse

    Christiansen, H.E.; Gee, L.P.; Mesa, M.G.

    2012-01-01

    Pacific lamprey Entosphenus tridentatus are facing severe population declines, yet little is known about juvenile lamprey passage, life history, or adult return rates because until now, these small fish could not be tagged for unique identification of live individuals. Previously, we developed a simple and effective method for tagging juvenile lampreys with passive integrated transponder (PIT) tags and showed that tagging per se did not affect survival. Mortality in tagged and untagged control fish, however, was frequently associated with fungal infection. In this study, we addressed two outstanding issues related to handling and tagging juvenile lampreys. First, we tried to mitigate freshwater fungal infections by reducing irritation and stress from anesthesia and by treating tagged fish briefly with a prophylactic immediately after tagging. We tested four anesthetics at three concentrations each and determined that 100 mg/L MS-222 and 60 mg/L BENZOAK® (benzocaine) were the most effective for anesthetizing juvenile lampreys to handleable while minimizing irritation. We also showed that fish anesthetized with BENZOAK® may have lower rates of fungal infection than those anesthetized with MS-222 or AQUI-S® 20E (eugenol). When fish anesthetized with MS-222 or BENZOAK® were given a 30 min prophylactic treatment with Stress Coat®, hydrogen peroxide, or salt immediately after tagging, few fish presented with fungal infections. However, untreated, tagged control fish also showed few fungal infections, making it difficult to determine if the prophylactic treatments were successful. The second question we addressed was whether activity would increase tag loss in PIT-tagged lampreys. We found that active swimming did not cause tag loss if fish were first held for 20–24 h after tagging. Therefore, we recommend anesthesia with MS-222 or BENZOAK® and then tagging with a 20–24 h recovery period followed by immediate release. If field studies show that lampreys are not

  4. The UL13 and US3 Protein Kinases of Herpes Simplex Virus 1 Cooperate to Promote the Assembly and Release of Mature, Infectious Virions

    PubMed Central

    Gershburg, Svetlana; Geltz, Joshua; Peterson, Karin E.; Halford, William P.; Gershburg, Edward

    2015-01-01

    Herpes simplex virus type 1 (HSV-1) encodes two bona fide serine/threonine protein kinases, the US3 and UL13 gene products. HSV-1 ΔUS3 mutants replicate with wild-type efficiency in cultured cells, and HSV-1 ΔUL13 mutants exhibit <10-fold reduction in infectious viral titers. Given these modest phenotypes, it remains unclear how the US3 and UL13 protein kinases contribute to HSV-1 replication. In the current study, we designed a panel of HSV-1 mutants, in which portions of UL13 and US3 genes were replaced by expression cassettes encoding mCherry protein or green fluorescent protein (GFP), respectively, and analyzed DNA replication, protein expression, and spread of these mutants in several cell types. Loss of US3 function alone had largely negligible effect on viral DNA accumulation, gene expression, virion release, and spread. Loss of UL13 function alone also had no appreciable effects on viral DNA levels. However, loss of UL13 function did result in a measurable decrease in the steady-state levels of two viral glycoproteins (gC and gD), release of total and infectious virions, and viral spread. Disruption of both genes did not affect the accumulation of viral DNA, but resulted in further reduction in gC and gD steady-state levels, and attenuation of viral spread and infectious virion release. These data show that the UL13 kinase plays an important role in the late phase of HSV-1 infection, likely by affecting virion assembly and/or release. Moreover, the data suggest that the combined activities of the US3 and UL13 protein kinases are critical to the efficient assembly and release of infectious virions from HSV-1-infected cells. PMID:26115119

  5. The Impact of Implementation of Bundle to Reduce Catheter-Related Bloodstream Infection Rates

    PubMed Central

    Menegueti, Mayra Goncalves; Ardison, Kym Marcel Martins; Bellissimo-Rodrigues, Fernando; Gaspar, Gilberto Gambero; Martins-Filho, Olindo Assis; Puga, Marcelo Lourencini; Laus, Ana Maria; Basile-Filho, Anibal; Auxiliadora-Martins, Maria

    2015-01-01

    Background The aim of the study was to investigate how control bundles reduce the rate of central venous catheter-associated bloodstream infections (CVC-BSIs) rates in critically ill patients. Methods This is a prospective before-and-after study designed to evaluate whether a set of control measures (bundle) can help prevent CVC-BSI. The bundles included a checklist that aimed to correct practices related to CVC insertion, manipulation, and maintenance based on guidelines of the Center for Disease Control and Prevention (CDC). Results We examined 123 checklists before and 155 checklists after implementation of the training program. Compared with the pre-intervention period, CVC-BSI rates decreased. Hand hygiene techniques were used correctly. CVC-BSI incidence was 9.3 and 5.1 per 1,000 catheter-days before and after the training program, respectively. Conclusions The implementation of a bundle and training program effectively reduces CVC-BSI rates. PMID:26491498

  6. Yoga lifestyle intervention reduces blood pressure in HIV-infected adults with cardiovascular disease risk factors

    PubMed Central

    Cade, Todd; Reeds, Dominic N.; Mondy, Kristin E.; Overton, Turner; Grassino, Joseph; Tucker, Shawn; Bopp, Coco; Laciny, Erin; Hubert, Sara; Lassa-Claxton, Sherry; Yarasheski, Kevin E.

    2009-01-01

    People living with human immunodeficiency virus infection (HIV) are at increased risk for developing cardiovascular disease (CVD). Safe and effective interventions for lowering CVD risk in HIV are high priorities. Objective We conducted a prospective, randomized, controlled study to evaluate whether a yoga lifestyle intervention improves CVD risk factors, virologic or immunologic status, or quality of life in HIV-infected adults more than in a matched control group. Methods Sixty HIV-infected adults with mild-moderate CVD risk were assigned to 20 wks of supervised yoga practice or standard of care treatment. Baseline and week 20 measures were; 2hr-oral glucose tolerance test with insulin monitoring, body composition, fasting serum lipid/lipoprotein profile, resting blood pressures, CD4+ T-cell number and plasma HIV RNA, and the Medical Outcomes Study SF-36 health-related quality of life inventory. Results Resting systolic and diastolic blood pressures were reduced more (p=0.04) in the yoga group (−5±2 and −3±1 mmHg) than in the standard of care group (+1±2 and +2±2 mmHg), despite no greater reduction in body weight, fat mass, proatherogenic lipids, or improvements in glucose tolerance or overall quality of life after yoga. Immune and virologic status was not adversely affected. Conclusion Among traditional lifestyle modifications, yoga is a low cost, simple to administer, non-pharmacological, popular behavioral intervention that can lower blood pressure in pre-hypertensive HIV-infected adults with mild-moderate CVD risk factors. PMID:20059570

  7. Prophylactic, therapeutic and neutralizing effects of zinc oxide tetrapod structures against herpes simplex virus type-2 infection

    PubMed Central

    Antoine, Thessicar; Mishra, Yogendra K.; Trigilio, James; Tiwari, Vaibhav; Adelung, Rainer; Shukla, Deepak

    2012-01-01

    The attachment of Herpes simplex virus type-2 (HSV-2) to a target cell requires ionic interactions between negatively charged cell surface co-receptor heparan sulfate (HS) and positively charged residues on viral envelop glycoproteins, gB and gC. Effective blocking of this first step of HSV-2 pathogenesis demonstrates significant prophylactic effects against the viral disease; any in vitro therapeutic effects of blocking this interaction, however, are not clear. Here, we provide new evidence that zinc oxide tetrapod micro-nanostructures synthesized by flame transport approach significantly block HSV-2 entry into target cells and, in addition, demonstrate the potential to stop the spread of the virus among already infected cells. The zinc oxide tetrapods (ZnOTs) also exhibit the ability to neutralize HSV-2 virions. Natural target cells such as human vaginal epithelial and HeLa cells showed highly reduced infectivity when infected with HSV-2 virions that were pre-incubated with the ZnOTs. The mechanism behind the ability of ZnOTs to prevent, neutralize or reduce HSV-2 infection relies on their ability to bind the HSV-2 virions. We used fluorescently labeled ZnOTs and GFP-expressing HSV-2 virions to demonstrate the binding of the ZnOTs with HSV-2. We also show that the binding and hence, the anti-viral effects of ZnOTs can be enhanced by illuminating the ZnOTs with UV light. Our results provide new insights into the anti-HSV-2 effects of ZnOT and rationalize their development as a HSV-2 trapping agent for the prevention and/or treatment of infection. The observed results also demonstrate that blocking HSV-2 attachment can have prophylactic as well as therapeutic applications. PMID:23047013

  8. Propofol Increases Host Susceptibility to Microbial Infection by Reducing Subpopulations of Mature Immune Effector Cells at Sites of Infection

    PubMed Central

    Visvabharathy, Lavanya; Xayarath, Bobbi; Weinberg, Guy; Shilling, Rebecca A.; Freitag, Nancy E.

    2015-01-01

    Anesthetics are known to modulate host immune responses, but separating the variables of surgery from anesthesia when analyzing hospital acquired infections is often difficult. Here, the bacterial pathogen Listeria monocytogenes (Lm) was used to assess the impact of the common anesthetic propofol on host susceptibility to infection. Brief sedation of mice with physiologically relevant concentrations of propofol increased bacterial burdens in target organs by more than 10,000-fold relative to infected control animals. The adverse effects of propofol sedation on immune clearance of Lm persisted after recovery from sedation, as animals given the drug remained susceptible to infection for days following anesthesia. In contrast to propofol, sedation with alternative anesthetics such as ketamine/xylazine or pentobarbital did not increase susceptibility to systemic Lm infection. Propofol altered systemic cytokine and chemokine expression during infection, and prevented effective bacterial clearance by inhibiting the recruitment and/or activity of immune effector cells at sites of infection. Propofol exposure induced a marked reduction in marginal zone macrophages in the spleens of Lm infected mice, resulting in bacterial dissemination into deep tissue. Propofol also significantly increased mouse kidney abscess formation following infection with the common nosocomial pathogen Staphylococcus aureus. Taken together, these data indicate that even brief exposure to propofol severely compromises host resistance to microbial infection for days after recovery from sedation. PMID:26381144

  9. Reduced itraconazole concentration and durations are successful in treating Batrachochytrium dendrobatidis infection in amphibians.

    PubMed

    Brannelly, Laura A

    2014-01-01

    Amphibians are experiencing the greatest decline of any vertebrate class and a leading cause of these declines is a fungal pathogen, Batrachochytrium dendrobatidis (Bd), which causes the disease chytridiomycosis. Captive assurance colonies are important worldwide for threatened amphibian species and may be the only lifeline for those in critical threat of extinction. Maintaining disease free colonies is a priority of captive managers, yet safe and effective treatments for all species and across life stages have not been identified. The most widely used chemotherapeutic treatment is itraconazole, although the dosage commonly used can be harmful to some individuals and species. We performed a clinical treatment trial to assess whether a lower and safer but effective dose of itraconazole could be found to cure Bd infections. We found that by reducing the treatment concentration from 0.01-0.0025% and reducing the treatment duration from 11-6 days of 5 min baths, frogs could be cured of Bd infection with fewer side effects and less treatment-associated mortality. PMID:24686573

  10. Abortive replication of Bombyx mori nucleopolyhedrovirus in Sf9 and High Five cells: Defective nuclear transport of the virions

    SciTech Connect

    Katou, Yasuhiro; Ikeda, Motoko; Kobayashi, Michihiro . E-mail: michihir@agr.nagoya-u.ac.jp

    2006-04-10

    Despite close genetic relationship, Bombyx mori nucleopolyhedrovirus (BmNPV) and Autographa californica multicapsid NPV (AcMNPV) display a distinct host range property. Here, BmNPV replication was examined in Sf9 and High Five cells that were nonproductive for BmNPV infection but supported high titers of AcMNPV replication. Recombinant BmNPV, vBm/gfp/lac, containing bm-ie1 promoter-driven egfp showed that few Sf9 and High Five cells infected with vBm/gfp/lac expressed EGFP, while large proportion of EGFP-expressing cells was observed when transfected with vBm/gfp/lac DNA. Immunocytochemical analysis showed that BmNPV was not imported into the nucleus of these two cell lines, while recombinant BmNPV, vBm{delta}64/ac-gp64 possessing AcMNPV gp64 was imported into the nucleus, yielding progeny virions in High Five cells, but not Sf9 cells. These results indicate that the defective nuclear import of infected virions due to insufficient BmNPV GP64 function is involved in the restricted BmNPV replication in Sf9 and High Five cells.

  11. N-terminal basic amino acid residues of Beet black scorch virus capsid protein play a critical role in virion assembly and systemic movement

    PubMed Central

    2013-01-01

    Background Beet black scorch virus (BBSV) is a small single-stranded, positive-sense RNA plant virus belonging to the genus Necrovirus, family Tombusviridae. Its capsid protein (CP) contains a 13 amino acid long basic region at the N-terminus, rich in arginine and lysine residues, which is thought to interact with viral RNA to initiate virion assembly. Results In the current study, a series of BBSV mutants containing amino acid substitutions as well as deletions within the N-terminal region were generated and examined for their effects on viral RNA replication, virion assembly, and long distance spread in protoplasts and whole host plants of BBSV. The RNA-binding activities of the mutated CPs were also evaluated in vitro. These experiments allowed us to identify two key basic amino acid residues in this region that are responsible for initiating virus assembly through RNA-binding. Proper assembly of BBSV particles is in turn needed for efficient viral systemic movement. Conclusions We have identified two basic amino acid residues near the N-terminus of the BBSV CP that bind viral RNA with high affinity to initiate virion assembly. We further provide evidence showing that systemic spread of BBSV in infected plants requires intact virions. This study represents the first in-depth investigation of the role of basic amino acid residues within the N-terminus of a necroviral CP. PMID:23786675

  12. An Aptamer against the Matrix Binding Domain on the Hepatitis B Virus Capsid Impairs Virion Formation

    PubMed Central

    Orabi, Ahmed; Bieringer, Maria; Geerlof, Arie

    2015-01-01

    ABSTRACT The hepatitis B virus (HBV) particle is an icosahedral nucleocapsid surrounded by a lipid envelope containing viral surface proteins. A small domain (matrix domain [MD]) in the large surface protein L and a narrow region (matrix binding domain [MBD]) including isoleucine 126 on the capsid surface have been mapped, in which point mutations such as core I126A specifically blocked nucleocapsid envelopment. It is possible that the two domains interact with each other during virion morphogenesis. By the systematic evolution of ligands by exponential enrichment (SELEX) method, we evolved DNA aptamers from an oligonucleotide library binding to purified recombinant capsids but not binding to the corresponding I126A mutant capsids. Aptamers bound to capsids were separated from unbound molecules by filtration. After 13 rounds of selections and amplifications, 16 different aptamers were found among 73 clones. The four most frequent aptamers represented more than 50% of the clones. The main aptamer, AO-01 (13 clones, 18%), showed the lowest dissociation constant (Kd) of 180 ± 82 nM for capsid binding among the four molecules. Its Kd for I126A capsids was 1,306 ± 503 nM. Cotransfection of Huh7 cells with AO-01 and an HBV genomic construct resulted in 47% inhibition of virion production at 3 days posttransfection, but there was no inhibition by cotransfection of an aptamer with a random sequence. The half-life of AO-01 in cells was 2 h, which might explain the incomplete inhibition. The results support the importance of the MBD for nucleocapsid envelopment. Inhibiting the MD-MBD interaction with a low-molecular-weight substance might represent a new approach for an antiviral therapy. IMPORTANCE Approximately 240 million people are persistently infected with HBV. To date, antiviral therapies depend on a single target, the viral reverse transcriptase. Future additional targets could be viral protein-protein interactions. We selected a 55-base-long single-stranded DNA

  13. Capillarity-induced disassembly of virions in carbon nanotubes

    NASA Astrophysics Data System (ADS)

    Fan, Xiaobin; Barclay, J. Elaine; Peng, Wenchao; Li, Yang; Li, Xianyu; Zhang, Guoliang; Evans, David J.; Zhang, Fengbao

    2008-04-01

    Studying the transport and fate of viruses through nanochannels is of great importance. By using the nanochannel of a carbon nanotube (CNT) as an ideal model, we evaluated the possibility of capillarity-induced viral transport through a closely fitting nanochannel and explored the mechanisms involved. It is shown both experimentally and theoretically that Cowpea mosaic virus can enter CNTs by capillarity. However, when introduced into a nanotube the protein capsid may disassemble. During the initial capillary filling stage, anomalous needle-shaped high pressure exists in the centre of the nanotube's entrance. This high pressure, combining with the significant negative pressure within the nanotube, may account for the disassembly of the virions.

  14. A gradient-free method for the purification of infective dengue virus for protein-level investigations.

    PubMed

    Jensen, Stephanie M; Nguyen, Celina T; Jewett, John C

    2016-09-01

    Dengue virus (DENV) is a mosquito-transmitted flavivirus that infects approximately 100 million people annually. Multi-day protocols for purification of DENV reduce the infective titer due to viral sensitivity to both temperature and pH. Herein we describe a 5-h protocol for the purification of all DENV serotypes, utilizing traditional gradient-free ultracentrifugation followed by selective virion precipitation. This protocol allows for the separation of DENV from contaminating proteins - including intact C6/36 densovirus, for the production of infective virus at high concentration for protein-level analysis. PMID:27265428

  15. An intraoperative irrigation regimen to reduce the surgical site infection rate following adolescent idiopathic scoliosis surgery.

    PubMed

    Herwijnen, B van; Evans, N R; Dare, C J; Davies, E M

    2016-05-01

    Introduction The aim of this study was to compare the efficacy of a gentamicin antibiotic intraoperative irrigation regimen (regimen A) with a povidone-iodine intraoperative irrigation regimen (regimen B) and to evaluate the ability of adjunctive local vancomycin powder (regimen C) to reduce the surgical site infection (SSI) rate following idiopathic scoliosis correction. Methods This was a retrospective, single centre, two-surgeon cohort study of paediatric scoliosis procedures involving 118 patients under the age of 18 years who underwent correction for idiopathic scoliosis over a period of 42 months. Patients' baseline characteristics, pseudarthrosis and rates of SSI were compared. Results Baseline characteristics were comparable in all three groups, with the exception of sex distribution. Over a quarter (27%) of patients with regimen B were male compared with 13% and 6% for regimens A and C respectively. Patients were mostly followed up for a minimum of 12 months. The SSI rate for both superficial and deep infections was higher with regimen A (26.7%) than with regimens B and C (7.0% and 6.3% respectively). The SSI rates for regimens B and C were comparable. No patients developed complications related to vancomycin toxicity, metalwork failure or pseudarthrosis. Conclusions Wound irrigation with a povidone-iodine solution reduces SSIs following adolescent idiopathic scoliosis surgery. The direct application of vancomycin powder to the wound is safe but does not reduce the SSI rate further in low risk patients. Additional studies are needed to elucidate whether it is effective at higher doses and in high risk patient groups. PMID:27087324

  16. Stoichiometry of the antiviral protein APOBEC3G in HIV-1 virions.

    PubMed

    Xu, Hongzhan; Chertova, Elena; Chen, Jianbo; Ott, David E; Roser, James D; Hu, Wei-Shau; Pathak, Vinay K

    2007-04-10

    A host cytidine deaminase, APOBEC3G (A3G), inhibits replication of human immunodeficiency virus type 1 (HIV-1) by incorporating into virions in the absence of the virally encoded Vif protein (Deltavif virions), at least in part by causing G-to-A hypermutation. To gain insight into the antiretroviral function of A3G, we determined the quantities of A3G molecules that are incorporated in Deltavif virions. We combined three experimental approaches-reversed-phase high-pressure liquid chromatography (HPLC), scintillation proximity assay (SPA), and quantitative immunoblotting-to determine the molar ratio of A3G to HIV-1 capsid protein in Deltavif virions. Our studies revealed that the amount of the A3G incorporated into Deltavif virions was proportional to the level of its expression in the viral producing cells, and the ratio of the A3G to Gag in the Deltavif virions produced from activated human peripheral blood mononuclear cells (PBMC) was approximately 1:439. Based on previous estimates of the stoichiometry of HIV-1 Gag in virions (1400-5000), we conclude that approximately 7 (+/-4) molecules of A3G are incorporated into Deltavif virions produced from human PBMCs. These results indicate that virion incorporation of only a few molecules of A3G is sufficient to inhibit HIV-1 replication. PMID:17126871

  17. A review of health system infection control measures in developing countries: what can be learned to reduce maternal mortality

    PubMed Central

    2011-01-01

    A functional health system is a necessary part of efforts to achieve maternal mortality reduction in developing countries. Puerperal sepsis is an infection contracted during childbirth and one of the commonest causes of maternal mortality in developing countries, despite the discovery of antibiotics over eighty years ago. Infections can be contracted during childbirth either in the community or in health facilities. Some developing countries have recently experienced increased use of health facilities for labour and delivery care and there is a possibility that this trend could lead to rising rates of puerperal sepsis. Drug and technological developments need to be combined with effective health system interventions to reduce infections, including puerperal sepsis. This article reviews health system infection control measures pertinent to labour and delivery units in developing country health facilities. Organisational improvements, training, surveillance and continuous quality improvement initiatives, used alone or in combination have been shown to decrease infection rates in some clinical settings. There is limited evidence available on effective infection control measures during labour and delivery and from low resource settings. A health systems approach is necessary to reduce maternal mortality and the occurrence of infections resulting from childbirth. Organisational and behavioural change underpins the success of infection control interventions. A global, targeted initiative could raise awareness of the need for improved infection control measures during childbirth. PMID:21595872

  18. Immune evasion or avoidance: fungal skin infection linked to reduced defence peptides in Australian green-eyed treefrogs, Litoria serrata.

    PubMed

    Woodhams, Douglas C; Bell, Sara C; Kenyon, Nicole; Alford, Ross A; Rollins-Smith, Louise A

    2012-12-01

    Many parasites and pathogens suppress host immunity to maintain infection or initiate disease. On the skin of many amphibians, defensive peptides are active against the fungus Batrachochytrium dendrobatidis (Bd), the causative agent of the emerging infectious disease chytridiomycosis. We tested the hypothesis that infection with the fungus may be linked to lower levels of defensive peptides. We sampled both ambient (or constitutive) skin peptides on the ventral surface immediately upon capture, and stored skin peptides induced from granular glands by norepinephrine administration of Australian green-eyed treefrogs, Litoria serrata. Upon capture, uninfected frogs expressed an array of antimicrobial peptides on their ventral surface, whereas infected frogs had reduced skin peptide expression. Expression of ambient skin peptides differed with infection status, and antimicrobial peptides maculatin 1.1 and 2.1 were on average three times lower on infected frogs. However, the repertoire of skin peptides stored in granular glands did not differ with infection status; on average equal quantities were recovered from infected and from uninfected frogs. Our results could have at least two causes: (1) frogs with reduced peptide expression are more likely to become infected; (2) Bd infection interferes with defence peptides by inhibiting release or causing selective degradation of peptides on the skin surface. Immune evasion therefore may contribute to the pathogenesis of chytridiomycosis and a mechanistic understanding of this fungal strategy may lead to improved methods of disease control. PMID:23245614

  19. Down-regulated Th17 Responses Are Associated with Reduced Gastritis in Helicobacter pylori-infected Children

    PubMed Central

    Bimczok, Diane; Shaffer, Carrie L.; Cover, Timothy L.; Venegas, Alejandro; Salazar, Maria G.; Smythies, Lesley E.; Harris, Paul R.; Smith, Phillip D.

    2013-01-01

    Helicobacter pylori induces less gastric inflammation in children than adults. Here we investigated whether this reduced inflammation involves dysregulated Th17 responses. H. pylori-infected children and adults in Santiago, Chile had similar levels of H. pylori colonization, proportions of bacteria containing cagA and s1/s2 vacA markers of virulence and strain genotypes (predominantly hpEurope), but the children had significantly reduced levels of gastric inflammation and neutrophil infiltration. The reduced neutrophil accumulation in infected children was accompanied by significantly fewer gastric Th17 cells and significantly lower levels of IL-17-specific mRNA and protein compared to infected adults. The gastric mucosa of H. pylori-infected children also contained higher numbers of IL-10+ cells and increased levels of both IL-10 and Foxp3 mRNA compared to that of infected adults. Thus, reduced gastric inflammation, including diminished neutrophil accumulation, in H. pylori-infected children compared with infected adults is likely due to down-regulated gastric Th17/IL-17 responses as a consequence of enhanced mucosal regulatory T cell activity in the children. PMID:23299619

  20. Localization of the Houdinisome (Ejection Proteins) inside the Bacteriophage P22 Virion by Bubblegram Imaging

    PubMed Central

    Wu, Weimin; Leavitt, Justin C.; Cheng, Naiqian; Gilcrease, Eddie B.; Motwani, Tina; Teschke, Carolyn M.; Casjens, Sherwood R.

    2016-01-01

    ABSTRACT The P22 capsid is a T=7 icosahedrally symmetric protein shell with a portal protein dodecamer at one 5-fold vertex. Extending outwards from that vertex is a short tail, and putatively extending inwards is a 15-nm-long α-helical barrel formed by the C-terminal domains of portal protein subunits. In addition to the densely packed genome, the capsid contains three “ejection proteins” (E-proteins [gp7, gp16, and gp20]) destined to exit from the tightly sealed capsid during the process of DNA delivery into target cells. We estimated their copy numbers by quantitative SDS-PAGE as approximately 12 molecules per virion of gp16 and gp7 and 30 copies of gp20. To localize them, we used bubblegram imaging, an adaptation of cryo-electron microscopy in which gaseous bubbles induced in proteins by prolonged irradiation are used to map the proteins’ locations. We applied this technique to wild-type P22, a triple mutant lacking all three E-proteins, and three mutants each lacking one E-protein. We conclude that all three E-proteins are loosely clustered around the portal axis, in the region displaced radially inwards from the portal crown. The bubblegram data imply that approximately half of the α-helical barrel seen in the portal crystal structure is disordered in the mature virion, and parts of the disordered region present binding sites for E-proteins. Thus positioned, the E-proteins are strategically placed to pass down the shortened barrel and through the portal ring and the tail, as they exit from the capsid during an infection. PMID:27507825

  1. Expression of EBV-encoded oncogenes and EBV-like virions in multiple canine tumors.

    PubMed

    Chiu, Hung-Chuan; Chow, Kuan-Chih; Fan, Yi-Hsin; Chang, Shih-Chieh; Chiou, Shiow-Her; Chiang, Shu-Fen; Chiou, Che-Hao; Wu, Guo-Hua; Yang, Hsiu-Ching; Ho, Shu-Peng; Chen, Yuh-Kun; Lee, Wei-Cheng; Sun, H Sunny

    2013-04-12

    Epstein-Barr virus (EBV) is a ubiquitous human oncovirus. Previous studies by us and others have indicated that pet dogs frequently encounter EBV or EBV-related viral infection. In this study, we explored whether EBV is involved in canine malignancies in dogs. EBV-specific BamHI W sequence was detected by polymerase chain reaction (PCR) in 10 of 12 canine tumor specimens, including 8 of 10 oral tumors. Using reverse transcription-PCR, gene expressions of latent membrane protein 1 (LMP 1) and BamHI H rightward reading frame 1 (BHRF1) were identified in 8 and 7 of 12 specimens, respectively. A novel LMP1 variant, T0905, was predominant in 5 canine tumor specimens and found to exist in EBV positive human BC-2 cells. Another LMP1 variant, T0902, was similar to human tumor variant JB7. The BHRF1 sequence identified from these canine tumors was identical to that of the B95-8 viral strain. LMP1 protein and EBV-encoded RNA (EBER) were detected by immunohistochemistry and fluorescent in situ hybridization, respectively, in several tumors, particularly in tumor nests of oral amelanotic melanomas. Furthermore, EBV-like virions adopting a herpesvirus egress pathway were detected in a canthal fibroblastic osteosarcoma and an oral amelanotic melanoma. In conclusion, we report the expressions of BHRF1 transcript (a viral anti-apoptotic protein), LMP1 (a viral oncoprotein) transcript and protein, EBER (a viral oncogenic RNA), and EBV-like virions in multiple canine tumors. The identity of BHRF1 and the resemblance of LMP1 variants between canine and human tumors indicate either a close evolutionary relationship between canine and human EBV, or the possibility of zoonotic transmission. PMID:23380461

  2. Influenza virion transcriptase: synthesis in vitro of large, polyadenylic acid-containing complementary RNA.

    PubMed Central

    Plotch, S J; Krug, R M

    1977-01-01

    The influenza virion transcriptase is capable of synthesizing in vitro complementary RNA (cRNA) that is similar in several characteristics to the cRNA synthesized in the infected cell, which is the viral mRNA. Most of the in vitro cRNA is large (approximately 2.5 X 10(5) to 10(6) daltons), similar in size to in vivo cRNA. The in vitro transcripts initiate in adenosine (A) or guanosine (G) at the 5' end, as also appears to be the case with in vivo cRNA (R.M. Krug et al., 1976). The in vitro transcripts contain covalently linked polyadenylate [poly(A)] sequences, which are longer and more heterogeneous than the poly(A) sequences found on in vivo cRNA. The synthesis in vitro of cRNA with these characteristics requires both the proper divalent cation, Mg2+, and a specific dinulceside monophosphage (DNMP), ApG or GpG. These DNMPs stimulate cRNA synthesis about 100-fold in the presence of Mg2+ and act as primers to initiate RNA chains, as demonstrated by the fact that the 5'-phosphorylated derivatives of these DNMP's, 32pApG or 32pGpG, are incroporated at the 5' end of the product RNA. The RNA synthesized in vitro differs from in vivo cRNA in that neither capping nor methylation of the in vitro transcripts has been detected. The virion does contain a methylase activity, as shown by its ability to methylate exogenous methyl-deficient Escherichia coli tRNA. PMID:833924

  3. Translation and processing of mouse hepatitis virus virion RNA in a cell-free system

    SciTech Connect

    Denison, M.R.; Perlman, S.

    1986-10-01

    The first event after infection with mouse hepatitis virus strain A59 (MHV-A59) is presumed to be the synthesis of an RNA-dependent RNA polymerase from the input genomic RNA. The synthesis and processing of this putative ploymerase protein was studied in a cell-free translation system utilizing 60S RNA from MHV-A59 virions. The polypeptide products of this reaction included two major species of 220 and 28 kilodaltons. Kinetics experiments indicated that both p220 and p28 appeared after 60 min of incubation and that protein p28 was synthesized initially as the N-terminal portion of a larger precursor protein. When the cell-free translation products were labeled with N-formyl(/sup 35/S)methionyl-tRNA/sub i/, p28 was the predominant radioactive product, confirming its N-terminal location within a precursor protein. Translation in the presence of the protease inhibitors leupeptin and ZnCl/sub 2/ resulted in the disappearance of p28 and p220 and the appearance of a new protein, p250. This product, which approached the maximal size predicted for a protein synthesized from genomic RNA, was not routinely detected in the absence of inhibitors even under conditions which optimized the translation reaction for elongation of proteins. Subsequent chelation of ZnCl/sub 2/ resulted in the partial cleavage of the precursor protein and the reappearance of p28. One-dimensional peptide mapping with Staphylococcus aureus V-8 protease confirmed the precursor-product relationship of p250 and p28. The results show that MHV virion RNA, like many other viral RNAs, is translated into a large polyprotein, which is cleaved soon after synthesis into smaller, presumably functional proteins. This is in marked contrast to the synthesis of other MHV proteins, in which minimal proteolytic processing occurs.

  4. Hospital-wide multidisciplinary, multimodal intervention programme to reduce central venous catheter-associated bloodstream infection.

    PubMed

    Zingg, Walter; Cartier, Vanessa; Inan, Cigdem; Touveneau, Sylvie; Theriault, Michel; Gayet-Ageron, Angèle; Clergue, François; Pittet, Didier; Walder, Bernhard

    2014-01-01

    Central line-associated bloodstream infection (CLABSI) is the major complication of central venous catheters (CVC). The aim of the study was to test the effectiveness of a hospital-wide strategy on CLABSI reduction. Between 2008 and 2011, all CVCs were observed individually and hospital-wide at a large university-affiliated, tertiary care hospital. CVC insertion training started from the 3rd quarter and a total of 146 physicians employed or newly entering the hospital were trained in simulator workshops. CVC care started from quarter 7 and a total of 1274 nurses were trained by their supervisors using a web-based, modular, e-learning programme. The study included 3952 patients with 6353 CVCs accumulating 61,366 catheter-days. Hospital-wide, 106 patients had 114 CLABSIs with a cumulative incidence of 1.79 infections per 100 catheters. We observed a significant quarterly reduction of the incidence density (incidence rate ratios [95% confidence interval]: 0.92 [0.88-0.96]; P<0.001) after adjusting for multiple confounders. The incidence densities (n/1000 catheter-days) in the first and last study year were 2.3/1000 and 0.7/1000 hospital-wide, 1.7/1000 and 0.4/1000 in the intensive care units, and 2.7/1000 and 0.9/1000 in non-intensive care settings, respectively. Median time-to-infection was 15 days (Interquartile range, 8-22). Our findings suggest that clinically relevant reduction of hospital-wide CLABSI was reached with a comprehensive, multidisciplinary and multimodal quality improvement programme including aspects of behavioural change and key principles of good implementation practice. This is one of the first multimodal, multidisciplinary, hospital-wide training strategies successfully reducing CLABSI. PMID:24714418

  5. Reduced Immunocompetent B Cells and Increased Secondary Infection in Elderly Patients With Severe Sepsis.

    PubMed

    Suzuki, Kodai; Inoue, Shigeaki; Kametani, Yoshie; Komori, Yukako; Chiba, Sayuri; Sato, Takehito; Inokuchi, Sadaki; Ogura, Shinji

    2016-09-01

    Lymphocyte exhaustion was recently recognized as a mechanism of immunosuppression in sepsis. While B cells are known to play pivotal roles in bacterial infection and sepsis, changes in B-cell-mediated humoral immunity have not been evaluated in critically ill septic patients. We aimed to investigate changes in humoral immunity caused by defective B-cell function during severe sepsis. Thirty-three severe sepsis patients and 44 healthy subjects were prospectively enrolled. Blood was collected from patients within 72 h of and 8 to 11 h after sepsis onset to measure B-cell subtypes, serum immunoglobulin M concentration, and CpG-B oligodeoxynucleotide-induced immunoglobulin M (IgM) production ex vivo. Participants were divided into two age groups: adults (18-64 years) and elderly (≥65 years). The fraction of CD21 exhausted B cells in acute sepsis patients (3.18%) was higher than that observed in healthy donors (0.77%, respectively, P <0.01). Significantly, serum IgM in elderly septic patients (≥65 years) was negatively correlated with acute physiology and chronic health evaluation II score (r = -0.57, P <0.05). Consistently, in B cells stimulated ex vivo, both aging and sepsis induced significant reductions in supernatant IgM (P <0.01). This finding was clinically relevant, as elderly patients with decreased IgM production might be more susceptible to infection by Gram-negative bacteria and fungi. Reduced immunocompetent B cells may be related to increased secondary infection after sepsis, especially in the elderly. Finally, impaired humoral immunity with increased CD21 exhausted B cells and insufficient immunoglobulin M production may be a critical immunological change in sepsis. PMID:27172158

  6. Prednisolone reduces experimental arthritis, and inflammatory tissue destruction in SCID mice infected with Borrelia burgdorferi.

    PubMed

    Hurtenbach, U; Böggemeyer, E; Stehle, T; Museteanu, C; Del Pozo, E; Simon, M M

    1996-05-01

    Glucocorticosteroids (GC) are widely used as anti-inflammatory agents. The effects of Prednisolone on the development of Borrelia (B.) burgdorferi-induced clinical arthritis and organ inflammation was studied in severe combined immunodeficiency (SCID) mice. The drug was administered orally at a dose of 3, 10 and 30 mg/kg, starting shortly before experimental infection of the mice. A dose dependent inhibition of arthritic joint swelling was observed. Full protection was obtained with 30 mg/kg until 21 days after infection, subsequently, mild joint swelling developed but progression and severity of the disease was considerably less than in the other treated as well as in the untreated mice. Inhibition of clinical arthritis coincided with reduction of inflammatory cell infiltration in the joints, liver and muscle. Prednisolone was ineffective when application was initiated after arthritis was fully developed, i.e., 22 days after infection. Since the activated endothelium plays a critical role in development of inflammatory lesions, the expression of the cellular adhesion molecules (CAMs) E-selectin, P-selectin, ICAM-1 and VCAM-1 was determined in vitro using the bEnd3 endothelial cell line. Stimulation with a sonicated B. burgdorferi preparation in the presence of the water-soluble compound Prednisolone-21-hemisuccinate considerably reduced expression of ICAM-1, and marginally also of E-selectin, whereas the level of P-selectin and VCAM-1 remained unaltered. Thus, downregulation of ICAM-1 might be a critical factor in Prednisolone-mediated inhibition of B. burgdorferi-induced inflammation; the flare up of the disease after the initial protection indicates that additional therapy, e.g. with antibiotics, is necessary. PMID:8933206

  7. Loss of Glycosaminoglycan Receptor Binding after Mosquito Cell Passage Reduces Chikungunya Virus Infectivity

    PubMed Central

    Acharya, Dhiraj; Paul, Amber M.; Anderson, John F.; Huang, Faqing; Bai, Fengwei

    2015-01-01

    Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that can cause fever and chronic arthritis in humans. CHIKV that is generated in mosquito or mammalian cells differs in glycosylation patterns of viral proteins, which may affect its replication and virulence. Herein, we compare replication, pathogenicity, and receptor binding of CHIKV generated in Vero cells (mammal) or C6/36 cells (mosquito) through a single passage. We demonstrate that mosquito cell-derived CHIKV (CHIKVmos) has slower replication than mammalian cell-derived CHIKV (CHIKVvero), when tested in both human and murine cell lines. Consistent with this, CHIKVmos infection in both cell lines produce less cytopathic effects and reduced antiviral responses. In addition, infection in mice show that CHIKVmos produces a lower level of viremia and less severe footpad swelling when compared with CHIKVvero. Interestingly, CHIKVmos has impaired ability to bind to glycosaminoglycan (GAG) receptors on mammalian cells. However, sequencing analysis shows that this impairment is not due to a mutation in the CHIKV E2 gene, which encodes for the viral receptor binding protein. Moreover, CHIKVmos progenies can regain GAG receptor binding capability and can replicate similarly to CHIKVvero after a single passage in mammalian cells. Furthermore, CHIKVvero and CHIKVmos no longer differ in replication when N-glycosylation of viral proteins was inhibited by growing these viruses in the presence of tunicamycin. Collectively, these results suggest that N-glycosylation of viral proteins within mosquito cells can result in loss of GAG receptor binding capability of CHIKV and reduction of its infectivity in mammalian cells. PMID:26484530

  8. Antibodies elicited by yeast glycoproteins recognize HIV-1 virions and potently neutralize virions with high mannose N-glycans.

    PubMed

    Zhang, Hong; Fu, Hu; Luallen, Robert J; Liu, Bingfen; Lee, Fang-Hua; Doms, Robert W; Geng, Yu

    2015-09-22

    The glycan shield on the human immunodeficiency virus 1 (HIV-1) envelope (Env) glycoprotein has drawn attention as a target for HIV-1 vaccine design given that an increasing number of potent and broadly neutralizing antibodies (bNAbs) recognize epitopes entirely or partially comprised of high mannose type N-linked glycans. In an attempt to generate immunogens that target the glycan shield of HIV-1, we previously engineered a triple mutant (TM) strain of Saccharomyces cerevisiae that results in exclusive presentation of high mannose type N-glycans, and identified five TM yeast glycoproteins that support strong binding of 2G12, a bNAb that targets a cluster of high mannose glycans on the gp120 subunit of Env. Here, we further analyzed the antigenicity and immunogenicity of these proteins in inducing anti-HIV responses. Our study demonstrated that the 2G12-reactive TM yeast glycoproteins efficiently bound to recently identified bNAbs including PGT125-130 and PGT135 that recognize high mannose glycan-dependent epitopes. Immunization of rabbits with a single TM yeast glycoprotein (Gp38 or Pst1), when conjugated to a promiscuous T-cell epitope peptide and coadministered with a Toll-like receptor 2 agonist, induced glycan-specific HIV-1 Env cross-reactive antibodies. The immune sera bound to both synthetic mannose oligosaccharides and gp120 proteins from a broad range of HIV-1 strains. The purified antibodies recognized and captured virions that contain both complex- and high mannose-type of N-glycans, and potently neutralized virions from different HIV-1 clades but only when the virions were enforced to retain high mannose N-glycans. This study provides insights into the elicitation of anti-carbohydrate, HIV-1 Env-cross reactive antibodies with a heterologous glycoprotein and may have applications in the design and administration of immunogens that target the viral glycan shield for development of an effective HIV-1 vaccine. PMID:26277072

  9. Review of Subcutaneous Wound Drainage in Reducing Surgical Site Infections after Laparotomy

    PubMed Central

    Manzoor, B.; Heywood, N.; Sharma, A.

    2015-01-01

    Purpose. Surgical site infections (SSIs) remain a significant problem after laparotomies. The aim of this review was to assess the evidence on the efficacy of subcutaneous wound drainage in reducing SSI. Methods. MEDLINE database was searched. Studies were identified and screened according to criteria to determine their eligibility for meta-analysis. Meta-analysis was performed using the Mantel-Haenszel method and a fixed effects model. Results. Eleven studies were included with two thousand eight hundred and sixty-four patients. One thousand four hundred and fifty patients were in the control group and one thousand four hundred and fourteen patients were in the drain group. Wound drainage in all patients shows no statistically significant benefit in reducing SSI incidence. Use of drainage in high risk patients, contaminated wound types, and obese patients appears beneficial. Conclusion. Using subcutaneous wound drainage after laparotomy in all patients is unnecessary as it does not reduce SSI risk. Similarly, there seems to be no benefit in using it in clean and clean contaminated wounds. However, there may be benefit in using drains in patients who are at high risk, including patients who are obese and/or have contaminated wound types. A well designed trial is needed which examines these factors. PMID:26783556

  10. Reducing the risk of surgical site infections: does chlorhexidine gluconate provide a risk reduction benefit?

    PubMed

    Edmiston, Charles E; Bruden, Benjamin; Rucinski, Maria C; Henen, Cindy; Graham, Mary Beth; Lewis, Brian L

    2013-05-01

    Chlorhexidine gluconate (CHG) has been available as a topical antiseptic for over 50 years, having broad clinical application throughout the health care environment. Evidence-based clinical studies have shown chlorhexidine gluconate to be a safe and effective perioperative skin-prepping agent. Renewed interest has emerged for use of the antiseptic bath/shower to reduce the microbial skin burden prior to hospital admission. Recent clinical studies have documented that multiple applications of 2% or 4% CHG using a standardized protocol results in high skin surface concentrations sufficient to inhibit/kill skin colonizing flora, including methicillin-resistant Staphylococcus aureus. A new focus for the use of CHG in surgical patients involves irrigation of the wound prior to closure with 0.05% CHG followed by saline rinse. Recent laboratory studies suggest that, following a 1-minute exposure, 0.05% CHG produces a >5-log reduction against selective health care-associated pathogens and reduces microbial adherence to the surface of implantable biomedical devices. General, orthopedic, cardiothoracic, and obstetrical surgical studies have documented the safety of selective CHG formulations in elective surgical procedures. The following discussion will address both the evidence-based literature and preliminary findings suggesting that CHG has a broad and safe range of applications when used as an adjunctive interventional strategy for reducing the risk of postoperative surgical site infections (SSI). PMID:23622749