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Sample records for human cytomegalovirus nuclear

  1. A Role for Myosin Va in Human Cytomegalovirus Nuclear Egress.

    Science.gov (United States)

    Wilkie, Adrian R; Sharma, Mayuri; Pesola, Jean M; Ericsson, Maria; Fernandez, Rosio; Coen, Donald M

    2018-03-15

    Herpesviruses replicate and package their genomes into capsids in replication compartments within the nuclear interior. Capsids then move to the inner nuclear membrane for envelopment and release into the cytoplasm in a process called nuclear egress. We previously found that nuclear F-actin is induced upon infection with the betaherpesvirus human cytomegalovirus (HCMV) and is important for nuclear egress and capsid localization away from replication compartment-like inclusions toward the nuclear rim. Despite these and related findings, it has not been shown that any specific motor protein is involved in herpesvirus nuclear egress. In this study, we have investigated whether the host motor protein, myosin Va, could be fulfilling this role. Using immunofluorescence microscopy and coimmunoprecipitation, we observed associations between a nuclear population of myosin Va and the viral major capsid protein, with both concentrating at the periphery of replication compartments. Immunoelectron microscopy showed that nearly 40% of assembled nuclear capsids associate with myosin Va. We also found that myosin Va and major capsid protein colocalize with nuclear F-actin. Importantly, antagonism of myosin Va with RNA interference or a dominant negative mutant revealed that myosin Va is important for the efficient production of infectious virus, capsid accumulation in the cytoplasm, and capsid localization away from replication compartment-like inclusions toward the nuclear rim. Our results lead us to suggest a working model whereby human cytomegalovirus capsids associate with myosin Va for movement from replication compartments to the nuclear periphery during nuclear egress. IMPORTANCE Little is known regarding how newly assembled and packaged herpesvirus capsids move from the nuclear interior to the periphery during nuclear egress. While it has been proposed that an actomyosin-based mechanism facilitates intranuclear movement of alphaherpesvirus capsids, a functional role for

  2. A Role for Nuclear F-Actin Induction in Human Cytomegalovirus Nuclear Egress

    Science.gov (United States)

    Wilkie, Adrian R.; Lawler, Jessica L.

    2016-01-01

    ABSTRACT Herpesviruses, which include important pathogens, remodel the host cell nucleus to facilitate infection. This remodeling includes the formation of structures called replication compartments (RCs) in which herpesviruses replicate their DNA. During infection with the betaherpesvirus, human cytomegalovirus (HCMV), viral DNA synthesis occurs at the periphery of RCs within the nuclear interior, after which assembled capsids must reach the inner nuclear membrane (INM) for translocation to the cytoplasm (nuclear egress). The processes that facilitate movement of HCMV capsids to the INM during nuclear egress are unknown. Although an actin-based mechanism of alphaherpesvirus capsid trafficking to the INM has been proposed, it is controversial. Here, using a fluorescently-tagged, nucleus-localized actin-binding peptide, we show that HCMV, but not herpes simplex virus 1, strongly induced nuclear actin filaments (F-actin) in human fibroblasts. Based on studies using UV inactivation and inhibitors, this induction depended on viral gene expression. Interestingly, by 24 h postinfection, nuclear F-actin formed thicker structures that appeared by super-resolution microscopy to be bundles of filaments. Later in infection, nuclear F-actin primarily localized along the RC periphery and between the RC periphery and the nuclear rim. Importantly, a drug that depolymerized nuclear F-actin caused defects in production of infectious virus, capsid accumulation in the cytoplasm, and capsid localization near the nuclear rim, without decreasing capsid accumulation in the nucleus. Thus, our results suggest that for at least one herpesvirus, nuclear F-actin promotes capsid movement to the nuclear periphery and nuclear egress. We discuss our results in terms of competing models for these processes. PMID:27555312

  3. Human Cytomegalovirus Nuclear Capsids Associate with the Core Nuclear Egress Complex and the Viral Protein Kinase pUL97.

    Science.gov (United States)

    Milbradt, Jens; Sonntag, Eric; Wagner, Sabrina; Strojan, Hanife; Wangen, Christina; Lenac Rovis, Tihana; Lisnic, Berislav; Jonjic, Stipan; Sticht, Heinrich; Britt, William J; Schlötzer-Schrehardt, Ursula; Marschall, Manfred

    2018-01-13

    The nuclear phase of herpesvirus replication is regulated through the formation of regulatory multi-component protein complexes. Viral genomic replication is followed by nuclear capsid assembly, DNA encapsidation and nuclear egress. The latter has been studied intensely pointing to the formation of a viral core nuclear egress complex (NEC) that recruits a multimeric assembly of viral and cellular factors for the reorganization of the nuclear envelope. To date, the mechanism of the association of human cytomegalovirus (HCMV) capsids with the NEC, which in turn initiates the specific steps of nuclear capsid budding, remains undefined. Here, we provide electron microscopy-based data demonstrating the association of both nuclear capsids and NEC proteins at nuclear lamina budding sites. Specifically, immunogold labelling of the core NEC constituent pUL53 and NEC-associated viral kinase pUL97 suggested an intranuclear NEC-capsid interaction. Staining patterns with phospho-specific lamin A/C antibodies are compatible with earlier postulates of targeted capsid egress at lamina-depleted areas. Important data were provided by co-immunoprecipitation and in vitro kinase analyses using lysates from HCMV-infected cells, nuclear fractions, or infectious virions. Data strongly suggest that nuclear capsids interact with pUL53 and pUL97. Combined, the findings support a refined concept of HCMV nuclear trafficking and NEC-capsid interaction.

  4. Protein kinases responsible for the phosphorylation of the nuclear egress core complex of human cytomegalovirus.

    Science.gov (United States)

    Sonntag, Eric; Milbradt, Jens; Svrlanska, Adriana; Strojan, Hanife; Häge, Sigrun; Kraut, Alexandra; Hesse, Anne-Marie; Amin, Bushra; Sonnewald, Uwe; Couté, Yohann; Marschall, Manfred

    2017-10-01

    Nuclear egress of herpesvirus capsids is mediated by a multi-component nuclear egress complex (NEC) assembled by a heterodimer of two essential viral core egress proteins. In the case of human cytomegalovirus (HCMV), this core NEC is defined by the interaction between the membrane-anchored pUL50 and its nuclear cofactor, pUL53. NEC protein phosphorylation is considered to be an important regulatory step, so this study focused on the respective role of viral and cellular protein kinases. Multiply phosphorylated pUL50 varieties were detected by Western blot and Phos-tag analyses as resulting from both viral and cellular kinase activities. In vitro kinase analyses demonstrated that pUL50 is a substrate of both PKCα and CDK1, while pUL53 can also be moderately phosphorylated by CDK1. The use of kinase inhibitors further illustrated the importance of distinct kinases for core NEC phosphorylation. Importantly, mass spectrometry-based proteomic analyses identified five major and nine minor sites of pUL50 phosphorylation. The functional relevance of core NEC phosphorylation was confirmed by various experimental settings, including kinase knock-down/knock-out and confocal imaging, in which it was found that (i) HCMV core NEC proteins are not phosphorylated solely by viral pUL97, but also by cellular kinases; (ii) both PKC and CDK1 phosphorylation are detectable for pUL50; (iii) no impact of PKC phosphorylation on NEC functionality has been identified so far; (iv) nonetheless, CDK1-specific phosphorylation appears to be required for functional core NEC interaction. In summary, our findings provide the first evidence that the HCMV core NEC is phosphorylated by cellular kinases, and that the complex pattern of NEC phosphorylation has functional relevance.

  5. Nuclear domain 10 components upregulated via interferon during human cytomegalovirus infection potently regulate viral infection.

    Science.gov (United States)

    Ashley, Caroline L; Glass, Mandy S; Abendroth, Allison; McSharry, Brian P; Slobedman, Barry

    2017-07-01

    Human cytomegalovirus (HCMV) is a ubiquitous betaherpesvirus that causes life-threatening disease in immunocompromised and immunonaïve individuals. Type I interferons (IFNs) are crucial molecules in the innate immune response to HCMV and are also known to upregulate several components of the interchromosomal multiprotein aggregates collectively referred to as nuclear domain 10 (ND10). In the context of herpesvirus infection, ND10 components are known to restrict gene expression. This raises the question as to whether key ND10 components (PML, Sp100 and hDaxx) act as anti-viral IFN-stimulated genes (ISGs) during HCMV infection. In this study, analysis of ND10 component transcription during HCMV infection demonstrated that PML and Sp100 were significantly upregulated whilst hDaxx expression remained unchanged. In cells engineered to block the production of, or response to, type I IFNs, upregulation of PML and Sp100 was not detected during HCMV infection. Furthermore, pre-treatment with an IFN-β neutralizing antibody inhibited upregulation of PML and Sp100 during both infection and treatment with HCMV-infected cell supernatant. The significance of ND10 components functioning as anti-viral ISGs during HCMV infection was determined through knockdown of PML, Sp100 and hDaxx. ND10 knockdown cells were significantly more permissive to HCMV infection, as previously described but, in contrast to control cells, could support HCMV plaque formation following IFN-β pre-treatment. This ability of HCMV to overcome the potently anti-viral effects of IFN-β in ND10 expression deficient cells provides evidence that ND10 component upregulation is a key mediator of the anti-viral activity of IFN-β.

  6. Human Cytomegalovirus Persistence

    OpenAIRE

    Goodrum, Felicia; Caviness, Katie; Zagallo, Patricia

    2012-01-01

    Viral persistence is the rule following infection with all herpesviruses. The β-herpesvirus, human cytomegalovirus (HCMV), persists through chronic and latent states of infection. Both the chronic and latent states of infection contribute to HCMV persistence and to the high HCMV seroprevalence worldwide. The chronic infection is poorly defined molecularly, but clinically manifests as low-level virus shedding over extended periods of time and often in the absence of symptoms. Latency requires ...

  7. [Infection by human cytomegalovirus].

    Science.gov (United States)

    Sanbonmatsu Gámez, Sara; Ruiz, Mercedes Pérez; Navarro Marí, José María

    2014-02-01

    Prevalence of human cytomegalovirus infection is very high worldwide. Following primary infection, the virus remains latent, being able to cause recurrences either by reinfection with a new strain or by reactivation of the replication of the latent virus. The most severe disease is seen in congenital infection and in immunosuppressed patients, in whom the virus act as an opportunistic pathogen. Serological techniques are the methods of choice in primary infection and to determine the immune status against CMV in organ donor and receptor. Although well-standardized studies are lacking, the recent commercial availability of methods that measure cellular immune response are promising to predict the risk of CMV disease in immunosuppressed individuals. Molecular assays, that have gradually been substituting viral culture and/or antigen detection, are the most widely used methods for the diagnosis and control of CMV infection. Copyright © 2014 Elsevier España, S.L. All rights reserved.

  8. Human cytomegalovirus persistence.

    Science.gov (United States)

    Goodrum, Felicia; Caviness, Katie; Zagallo, Patricia

    2012-05-01

    Viral persistence is the rule following infection with all herpesviruses. The β-herpesvirus, human cytomegalovirus (HCMV), persists through chronic and latent states of infection. Both of these states of infection contribute to HCMV persistence and to the high HCMV seroprevalence worldwide. The chronic infection is poorly defined molecularly, but clinically manifests as low-level virus shedding over extended periods of time and often in the absence of symptoms. Latency requires long-term maintenance of viral genomes in a reversibly quiescent state in the immunocompetent host. In this review, we focus on recent advances in the biology of HCMV persistence, particularly with respect to the latent mode of persistence. Latently infected individuals harbour HCMV genomes in haematopoietic cells and maintain large subsets of HCMV-specific T-cells. In the last few years, impressive advances have been made in understanding virus-host interactions important to HCMV infection, many of which will profoundly impact HCMV persistence. We discuss these advances and their known or potential impact on viral latency. As herpesviruses are met with similar challenges in achieving latency and often employ conserved strategies to persist, we discuss current and future directions of HCMV persistence in the context of the greater body of knowledge regarding α- and γ-herpesviruses persistence. © 2012 Blackwell Publishing Ltd.

  9. The human cytomegalovirus nuclear egress complex unites multiple functions: Recruitment of effectors, nuclear envelope rearrangement, and docking to nuclear capsids.

    Science.gov (United States)

    Marschall, Manfred; Muller, Yves A; Diewald, Benedikt; Sticht, Heinrich; Milbradt, Jens

    2017-07-01

    Nuclear replication represents a common hallmark of herpesviruses achieved by a number of sequentially unrolled regulatory processes. A rate-limiting step is provided by nucleo-cytoplasmic capsid export, for which a defined multiregulatory protein complex, namely, the nuclear egress complex (NEC), is assembled comprising both viral and cellular components. The NEC regulates at least 3 aspects of herpesviral nuclear replication: (1) multimeric recruitment of NEC-associated effector proteins, (2) reorganization of the nuclear lamina and membranes, and (3) the docking to nuclear capsids. Here, we review published data and own experimental work that characterizes the NEC of HCMV and other herpesviruses. A systematic review of information on nuclear egress of HCMV compared to selected alpha-, beta-, and gamma-herpesviruses: proteomics-based approaches, high-resolution imaging techniques, and functional investigations. A large number of reports on herpesviral NECs have been published during the last two decades, focusing on protein-protein interactions, nuclear localization, regulatory phosphorylation, and functional validation. The emerging picture provides an illustrated example of well-balanced and sophisticated protein networking in virus-host interaction. Current evidence refined the view about herpesviral NECs. Datasets published for HCMV, murine CMV, herpes simplex virus, and Epstein-Barr virus illustrate the marked functional consistency in the way herpesviruses achieve nuclear egress. However, this compares with only limited sequence conservation of core NEC proteins and a structural conservation restricted to individual domains. The translational use of this information might help to define a novel antiviral strategy on the basis of NEC-directed small molecules. Copyright © 2017 John Wiley & Sons, Ltd.

  10. Human cytomegalovirus proteins PP65 and IEP72 are targeted to distinct compartments in nuclei and nuclear matrices of infected human embryo fibroblasts.

    Science.gov (United States)

    Arcangeletti, M C; De Conto, F; Ferraglia, F; Pinardi, F; Gatti, R; Orlandini, G; Calderaro, A; Motta, F; Medici, M C; Martinelli, M; Valcavi, P; Razin, S V; Chezzi, C; Dettori, G

    2003-12-01

    The cellular distribution of the human cytomegalovirus (HCMV)-specific UL83 phosphoprotein (pp65) and UL123 immediate-early protein (IEp72) in lytically infected human embryo fibroblasts was studied by means of indirect immunofluorescence and confocal microscopy. Both proteins were found to have a nuclear localization, but they were concentrated in different compartments within the nuclei. The pp65 was located predominantly in the nucleoli; this was already evident with the parental viral protein, which was targeted to the above nuclear compartment very soon after infection. The nucleolar localization of pp65 was also observed at later stages of the HCMV infectious cycle. After chromatin extraction (in the so-called in situ nuclear matrices), a significant portion of the pp65 remained associated with nucleoli within the first hour after infection, then gradually redistributed in a perinucleolar area, as well as throughout the nucleus, with a granular pattern. A quite different distribution was observed for IEp72 at very early stages after infection of human embryo fibroblasts with HCMV; indeed, this viral protein was found in bright foci, clearly observable in both non-extracted nuclei and in nuclear matrices. At later stages of infection, IEp72 became almost homogeneously distributed within the whole nucleus, while the foci increased in size and were more evenly spread; in several infected cells some of them lay within nucleoli. This peculiar nuclear distribution of IEp72 was preserved in nuclear matrices as well. The entire set of data is discussed in terms of the necessity of integration for HCMV-specific products into the pre-existing nuclear architecture, with the possibility of subsequent adaptation of nuclear compartments to fit the needs of the HCMV replicative cycle. Copyright 2003 Wiley-Liss, Inc.

  11. Human Cytomegalovirus and Autoimmune Disease

    Science.gov (United States)

    2014-01-01

    Human cytomegalovirus (HCMV) represents a prototypic pathogenic member of the β-subgroup of the herpesvirus family. A range of HCMV features like its lytic replication in multiple tissues, the lifelong persistence through periods of latency and intermitting reactivation, the extraordinary large proteome, and extensive manipulation of adaptive and innate immunity make HCMV a high profile candidate for involvement in autoimmune disorders. We surveyed the available literature for reports on HCMV association with onset or exacerbation of autoimmune disease. A causative linkage between HCMV and systemic lupus erythematosus (SLE), systemic sclerosis (SSc), diabetes mellitus type 1, and rheumatoid arthritis (RA) is suggested by the literature. However, a clear association of HCMV seroprevalence and disease could not be established, leaving the question open whether HCMV could play a coresponsible role for onset of disease. For convincing conclusions population-based prospective studies must be performed in the future. Specific immunopathogenic mechanisms by which HCMV could contribute to the course of autoimmune disease have been suggested, for example, molecular mimicry by UL94 in SSc and UL83/pp65 in SLE patients, as well as aggravation of joint inflammation by induction and expansion of CD4+/CD28− T-cells in RA patients. Further studies are needed to validate these findings and to lay the grounds for targeted therapeutic intervention. PMID:24967373

  12. Human Cytomegalovirus and Autoimmune Disease

    Directory of Open Access Journals (Sweden)

    Anne Halenius

    2014-01-01

    Full Text Available Human cytomegalovirus (HCMV represents a prototypic pathogenic member of the β-subgroup of the herpesvirus family. A range of HCMV features like its lytic replication in multiple tissues, the lifelong persistence through periods of latency and intermitting reactivation, the extraordinary large proteome, and extensive manipulation of adaptive and innate immunity make HCMV a high profile candidate for involvement in autoimmune disorders. We surveyed the available literature for reports on HCMV association with onset or exacerbation of autoimmune disease. A causative linkage between HCMV and systemic lupus erythematosus (SLE, systemic sclerosis (SSc, diabetes mellitus type 1, and rheumatoid arthritis (RA is suggested by the literature. However, a clear association of HCMV seroprevalence and disease could not be established, leaving the question open whether HCMV could play a coresponsible role for onset of disease. For convincing conclusions population-based prospective studies must be performed in the future. Specific immunopathogenic mechanisms by which HCMV could contribute to the course of autoimmune disease have been suggested, for example, molecular mimicry by UL94 in SSc and UL83/pp65 in SLE patients, as well as aggravation of joint inflammation by induction and expansion of CD4+/CD28− T-cells in RA patients. Further studies are needed to validate these findings and to lay the grounds for targeted therapeutic intervention.

  13. The Human Cytomegalovirus IE1 Protein Antagonizes PML Nuclear Body-Mediated Intrinsic Immunity via the Inhibition of PML De Novo SUMOylation.

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    Schilling, Eva-Maria; Scherer, Myriam; Reuter, Nina; Schweininger, Johannes; Muller, Yves A; Stamminger, Thomas

    2017-02-15

    PML nuclear bodies (NBs) are accumulations of cellular proteins embedded in a scaffold-like structure built by SUMO-modified PML/TRIM19. PML and other NB proteins act as cellular restriction factors against human cytomegalovirus (HCMV); however, this intrinsic defense is counteracted by the immediate early protein 1 (IE1) of HCMV. IE1 directly interacts with the PML coiled-coil domain via its globular core region and disrupts NB foci by inducing a loss of PML SUMOylation. Here, we demonstrate that IE1 acts via abrogating the de novo SUMOylation of PML. In order to overcome reversible SUMOylation dynamics, we made use of a cell-based assay that combines inducible IE1 expression with a SUMO mutant resistant to SUMO proteases. Interestingly, we observed that IE1 expression did not affect preSUMOylated PML; however, it clearly prevented de novo SUMO conjugation. Consistent results were obtained by in vitro SUMOylation assays, demonstrating that IE1 alone is sufficient for this effect. Furthermore, IE1 acts in a selective manner, since K160 was identified as the main target lysine. This is strengthened by the fact that IE1 also prevents As2O3-mediated hyperSUMOylation of K160, thereby blocking PML degradation. Since IE1 did not interfere with coiled-coil-mediated PML dimerization, we propose that IE1 affects PML autoSUMOylation either by directly abrogating PML E3 ligase function or by preventing access to SUMO sites. Thus, our data suggest a novel mechanism for how a viral protein counteracts a cellular restriction factor by selectively preventing the de novo SUMOylation at specific lysine residues without affecting global protein SUMOylation. The human cytomegalovirus IE1 protein acts as an important antagonist of a cellular restriction mechanism that is mediated by subnuclear structures termed PML nuclear bodies. This function of IE1 is required for efficient viral replication and thus constitutes a potential target for antiviral strategies. In this paper, we further

  14. Human Cytomegalovirus UL97 Kinase Activity Is Required for the Hyperphosphorylation of Retinoblastoma Protein and Inhibits the Formation of Nuclear Aggresomes

    Energy Technology Data Exchange (ETDEWEB)

    Prichard, Mark N.; Sztul, Elizabeth; Daily, Shannon L.; Perry, Amie L.; Frederick, Samuel L.; Gill, Rachel B.; Hartline, Caroll B.; Streblow, Daniel N.; Varnum, Susan M.; Smith, Richard D.; Kern, Earl R.

    2008-05-01

    Cells infected with human cytomegalovirus in the absence of UL97 kinase activity produce large nuclear aggregates that sequester considerable quantities of viral proteins. A transient expression assay suggested that pp71 and IE1 were also involved in this process, and this suggestion was significant, since both proteins have been reported to interact with components of promyelocytic leukemia (PML) bodies (ND10) and also interact functionally with retinoblastoma pocket proteins (RB). PML bodies have been linked to the formation of nuclear aggresomes, and colocalization studies suggested that viral proteins were recruited to these structures and that UL97 kinase activity inhibited their formation. Proteins associated with PML bodies were examined by Western blot analysis, and pUL97 appeared to specifically affect the phosphorylation of RB in a kinasedependent manner. Three consensus RB binding motifs were identified in the UL97 kinase, and recombinant viruses were constructed in which each was mutated to assess a potential role in the phosphorylation of RB and the inhibition of nuclear aggresome formation. The mutation of either the conserved LxCxE RB binding moti for the lysine required for kinase activity impaired the ability of the virus to stabilize and phosphorylate RB. We concluded from these studies that both UL97 kinase activity and the LxCxE RB binding motif are required for the phosphorylation and stabilization of RB in infected cells and that this effect can be antagonized by the antiviral drug maribavir. These data also suggest a potential link between RB function and the formation of aggresomes.

  15. Overlapping transcription structure of human cytomegalovirus ...

    Indian Academy of Sciences (India)

    2013-01-21

    Jan 21, 2013 ... Transcription of human cytomegalovirus UL/b′ region has been studied extensively for some genes. In this study, transcripts of the UL140 and UL141, two of the UL/b′ genes, were identified in late RNAs of three HCMV isolates using Northern blot hybridization, cDNA library screening and RACE-PCR.

  16. A novel polyclonal antibody against human cytomegalovirus ...

    African Journals Online (AJOL)

    There is no vaccine available for human cytomegalovirus (HCMV) infection and the treatment is very limited; therefore, it is still an important cause of morbidity and occasional mortality in transplant recipients. An HCMV peptide was selected based on the GenBank sequence M60929. The peptide was conjugated with ...

  17. Overlapping transcription structure of human cytomegalovirus ...

    Indian Academy of Sciences (India)

    Transcription of human cytomegalovirus UL/b′ region has been studied extensively for some genes. In this study, transcripts of the UL140 and UL141, two of the UL/b′ genes, were identified in late RNAs of three HCMV isolates using Northern blot hybridization, cDNA library screening and RACE-PCR. At least three ...

  18. Cytomegalovirus.

    Science.gov (United States)

    Griffiths, Paul; Lumley, Sheila

    2014-12-01

    To consider new treatment options for cytomegalovirus (CMV) infection, review recent trials, and anticipate their use in clinical practice, focussing on bone marrow transplantation, congenital infection, and intervention during pregnancy. Three double-blind randomized placebo-controlled phase 2 proof-of-concept studies have each identified a novel antiviral drug with activity against CMV infection in bone marrow transplant patients. One of these (brincidofovir) inhibits the DNA polymerase that is the target of the currently licensed drug ganciclovir. Another new drug (maribavir) inhibits a protein kinase which, coincidentally, is the enzyme responsible for activating ganciclovir through phosphorylation. The third drug (letermovir) inhibits the terminase enzyme complex responsible for packaging unit length DNA into assembling virions.In addition, in a double-blind randomized placebo-controlled trial in neonates with symptomatic congenital CMV infection, a 6-month course of valganciclovir was superior to the standard 6-week course of the same drug. In pregnant women with primary CMV infection, administration of hyperimmune immunoglobulin did not significantly reduce transmission of CMV across the placenta. The ability to diagnose CMV infections reliably in different clinical settings through application of molecular laboratory methods has ushered in new ways of evaluating potential new treatments for CMV. Several of these may help control the diseases caused by this important human pathogen.

  19. Human Cytomegalovirus Latency: Approaching the Gordian Knot.

    Science.gov (United States)

    Goodrum, Felicia

    2016-09-29

    Herpesviruses have evolved exquisite virus-host interactions that co-opt or evade a number of host pathways to enable the viruses to persist. Persistence of human cytomegalovirus (CMV), the prototypical betaherpesvirus, is particularly complex in the host organism. Depending on host physiology and the cell types infected, CMV persistence comprises latent, chronic, and productive states that may occur concurrently. Viral latency is a central strategy by which herpesviruses ensure their lifelong persistence. Although much remains to be defined about the virus-host interactions important to CMV latency, it is clear that checkpoints composed of viral and cellular factors exist to either maintain a latent state or initiate productive replication in response to host cues. CMV offers a rich platform for defining the virus-host interactions and understanding the host biology important to viral latency. This review describes current understanding of the virus-host interactions that contribute to viral latency and reactivation.

  20. Cytomegalovirus (CMV) infection

    Science.gov (United States)

    CMV mononucleosis; Cytomegalovirus; CMV; Human cytomegalovirus; HCMV ... Crumpacker CS. Cytomegalovirus (CMV). In: Bennett JE, Dolin R, Blaser MJ, eds. Mandell, Douglas, and Bennett's Principles and Practice of Infectious ...

  1. Glucocorticoids facilitate the transcription from the human cytomegalovirus major immediate early promoter in glucocorticoid receptor- and nuclear factor-I-like protein-dependent manner

    Energy Technology Data Exchange (ETDEWEB)

    Inoue-Toyoda, Maki [Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575 (Japan); Kato, Kohsuke [Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575 (Japan); Faculty of Medicine, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575 (Japan); Nagata, Kyosuke, E-mail: knagata@md.tsukuba.ac.jp [University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575 (Japan); Yoshikawa, Hiroyuki [Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575 (Japan); Faculty of Medicine, University of Tsukuba, 1-1-1 Tennodai, Tsukuba 305-8575 (Japan)

    2015-02-27

    Human cytomegalovirus (HCMV) is a common and usually asymptomatic virus agent in healthy individuals. Initiation of HCMV productive infection depends on expression of the major immediate early (MIE) genes. The transcription of HCMV MIE genes is regulated by a diverse set of transcription factors. It was previously reported that productive HCMV infection is triggered probably by elevation of the plasma hydroxycorticoid level. However, it is poorly understood whether the transcription of MIE genes is directly regulated by glucocorticoid. Here, we found that the dexamethasone (DEX), a synthetic glucocorticoid, facilitates the transcription of HCMV MIE genes through the MIE promoter and enhancer in a glucocorticoid receptor (GR)-dependent manner. By competitive EMSA and reporter assays, we revealed that an NF-I like protein is involved in DEX-mediated transcriptional activation of the MIE promoter. Thus, this study supports a notion that the increased level of hydroxycorticoid in the third trimester of pregnancy reactivates HCMV virus production from the latent state. - Highlights: • DEX facilitates the transcription from the HCMV MIE promoter. • GR is involved in DEX-dependent transcription from the HCMV MIE promoter. • A 17 bp repeat is responsible for the HCMV MIE promoter activation by DEX. • An NF-I-like protein is involved in the HCMV MIE promoter activation by DEX.

  2. Human Cytomegalovirus Manipulation of Latently Infected Cells

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    Sinclair, John H.; Reeves, Matthew B.

    2013-01-01

    Primary infection with human cytomegalovirus (HCMV) results in the establishment of a lifelong infection of the host which is aided by the ability of HCMV to undergo a latent infection. One site of HCMV latency in vivo is in haematopoietic progenitor cells, resident in the bone marrow, with genome carriage and reactivation being restricted to the cells of the myeloid lineage. Until recently, HCMV latency has been considered to be relatively quiescent with the virus being maintained essentially as a “silent partner” until conditions are met that trigger reactivation. However, advances in techniques to study global changes in gene expression have begun to show that HCMV latency is a highly active process which involves expression of specific latency-associated viral gene products which orchestrate major changes in the latently infected cell. These changes are argued to help maintain latent infection and to modulate the cellular environment to the benefit of latent virus. In this review, we will discuss these new findings and how they impact not only on our understanding of the biology of HCMV latency but also how they could provide tantalising glimpses into mechanisms that could become targets for the clearance of latent HCMV. PMID:24284875

  3. 76 FR 69743 - The Development and Evaluation of Human Cytomegalovirus Vaccines; Public Workshop

    Science.gov (United States)

    2011-11-09

    ... HUMAN SERVICES Food and Drug Administration The Development and Evaluation of Human Cytomegalovirus... Development and Evaluation of Human Cytomegalovirus Vaccines.'' The purpose of the public workshop is to identify and discuss key issues related to the development and evaluation of human cytomegalovirus (HCMV...

  4. Peptide inhibition of human cytomegalovirus infection

    Directory of Open Access Journals (Sweden)

    Morris Cindy A

    2011-02-01

    Full Text Available Abstract Background Human cytomegalovirus (HCMV is the most prevalent congenital viral infection in the United States and Europe causing significant morbidity and mortality to both mother and child. HCMV is also an opportunistic pathogen in immunocompromised individuals, including human immunodeficiency virus (HIV- infected patients with AIDS, and solid organ and allogeneic stem cell transplantation recipients. Current treatments for HCMV-associated diseases are insufficient due to the emergence of drug-induced resistance and cytotoxicity, necessitating novel approaches to limit HCMV infection. The aim of this study was to develop therapeutic peptides targeting glycoprotein B (gB, a major glycoprotein of HCMV that is highly conserved across the Herpesviridae family, that specifically inhibit fusion of the viral envelope with the host cell membrane preventing HCMV entry and infection. Results Using the Wimley-White Interfacial Hydrophobicity Scale (WWIHS, several regions within gB were identified that display a high potential to interact with lipid bilayers of cell membranes and hydrophobic surfaces within proteins. The ability of synthetic peptides analogous to WWIHS-positive sequences of HCMV gB to inhibit viral infectivity was evaluated. Human foreskin fibroblasts (HFF were infected with the Towne-GFP strain of HCMV (0.5 MOI, preincubated with peptides at a range of concentrations (78 nm to 100 μM, and GFP-positive cells were visualized 48 hours post-infection by fluorescence microscopy and analyzed quantitatively by flow cytometry. Peptides that inhibited HCMV infection demonstrated different inhibitory concentration curves indicating that each peptide possesses distinct biophysical properties. Peptide 174-200 showed 80% inhibition of viral infection at a concentration of 100 μM, and 51% and 62% inhibition at concentrations of 5 μM and 2.5 μM, respectively. Peptide 233-263 inhibited infection by 97% and 92% at concentrations of 100

  5. Intravitreal ganciclovir in the management of non-AIDS-related human cytomegalovirus retinitis

    NARCIS (Netherlands)

    Langner-Wegscheider, Beate J.; ten Dam-van Loon, Ninette; Mura, Marco; Faridpooya, Koorosh; de Smet, Marc D.

    2010-01-01

    To report and evaluate intravitreal ganciclovir injections in non-AIDS patients with human cytomegalovirus (HCMV) retinitis. Retrospective chart review. Two SLE patients and one patient post chemotherapy for a non Hodgkin's lymphoma presented with myelosuppression and persistent cytomegalovirus

  6. Cytomegalovirus infection of the human placenta: an immunocytochemical study.

    Science.gov (United States)

    Mühlemann, K; Miller, R K; Metlay, L; Menegus, M A

    1992-11-01

    In congenital cytomegalovirus (CMV) infection histologic evaluation of the placenta is often unrevealing. In the present study immunocytochemistry to CMV immediate early and early nuclear antigens was used to characterize placental involvement in six cases of symptomatic intrauterine CMV infection. Histologic examination had demonstrated diagnostic viral inclusions in one placenta and non-specific villitis in another. However, immunocytochemistry revealed CMV infection in five of the six placentas, including three with no pathologic changes on routine histologic evaluation. Infected cells were located primarily in the villous stroma. In one case immunoperoxidase staining showed infection in the syncytiotrophoblast. Infected endothelial cells were demonstrated by double staining for CMV and factor VIII antigen. No double-stained cells were seen in tissue sections stained for CMV immediate early nuclear antigen or the human macrophage-associated CD68 antigen, which is expressed in Hofbauer cells. In conclusion, specific immunoperoxidase staining was more sensitive for demonstrating placental CMV infection than was histologic examination and it aided in the characterization of infected cells.

  7. Human Cytomegalovirus-Encoded Receptor US28 Is Expressed in Renal Allografts and Facilitates Viral Spreading In Vitro

    NARCIS (Netherlands)

    Lollinga, Wouter T; de Wit, Raymond H; Rahbar, Afsar; Vasse, Gwenda F; Davoudi, Belghis; Diepstra, Arjan; Riezebos-Brilman, Annelies; Harmsen, Martin C; Hillebrands, Jan-Luuk; Söderberg-Naucler, Cecilia; van Son, Willem J; Smit, Martine J; Sanders, Jan-Stephan; van den Born, Jacob

    BACKGROUND: Renal transplantation is the preferred treatment for patients with end-stage renal disease. Human cytomegalovirus (HCMV) activation is associated with decreased renal graft function and survival. Human cytomegalovirus encodes several immune modulatory proteins, including the G

  8. Growth in Agarose of Human Cells Infected with Cytomegalovirus

    Science.gov (United States)

    Lang, David J.; Montagnier, Luc; Latarjet, Raymond

    1974-01-01

    After infection by human cytomegalovirus (CMV), human diploid fibroblasts could grow in agarose medium for several generations. Clones of infected cells grew for weeks, although in every case they ultimately underwent lysis owing to the cytopathic effect of the virus. Virus was inoculated at high dilution and after UV irradiation in an effort to derive cells infected with noninfectious defective particles still capable of inducing cell stimulation. Dilute or irradiated virus occasionally yielded large colonies of replicating cells, although permanent transformation was not observed. One clone derived from UV-CMV-infected cells was passaged four times before undergoing lysis. During these passages the cells exhibited alterations in morphology and orientation. Images PMID:4367907

  9. Dynamics of the cellular metabolome during human cytomegalovirus infection.

    Directory of Open Access Journals (Sweden)

    Joshua Munger

    2006-12-01

    Full Text Available Viral replication requires energy and macromolecular precursors derived from the metabolic network of the host cell. Despite this reliance, the effect of viral infection on host cell metabolic composition remains poorly understood. Here we applied liquid chromatography-tandem mass spectrometry to measure the levels of 63 different intracellular metabolites at multiple times after human cytomegalovirus (HCMV infection of human fibroblasts. Parallel microarray analysis provided complementary data on transcriptional regulation of metabolic pathways. As the infection progressed, the levels of metabolites involved in glycolysis, the citric acid cycle, and pyrimidine nucleotide biosynthesis markedly increased. HCMV-induced transcriptional upregulation of specific glycolytic and citric acid cycle enzymes mirrored the increases in metabolite levels. The peak levels of numerous metabolites during infection far exceeded those observed during normal fibroblast growth or quiescence, demonstrating that HCMV markedly disrupts cellular metabolic homeostasis and institutes its own specific metabolic program.

  10. Role of Human Cytomegalovirus Tegument Proteins in Virion Assembly

    Science.gov (United States)

    Smith, Rebecca Marie; Kosuri, Srivenkat; Kerry, Julie Anne

    2014-01-01

    Like other herpesviruses, human cytomegalovirus (HCMV) contains a unique proteinaceous layer between the virion envelope and capsid, termed the tegument. Upon infection, the contents of the tegument layer are delivered to the host cell, along with the capsid and the viral genome, where they facilitate the initial stages of virus replication. The tegument proteins also play important roles in virion assembly and this dual nature makes them attractive potential targets for antiviral therapies. While our knowledge regarding tegument protein function during the initiation of infection has been the subject of intense study, their roles in assembly are much less well understood. In this review, we will focus on recent studies that highlight the functions of HCMV tegument proteins during assembly, and pose key questions for further investigation. PMID:24509811

  11. Human cytomegalovirus-mediated immunomodulation: Effects on glioblastoma progression.

    Science.gov (United States)

    Foster, Haidn; Ulasov, Ilya V; Cobbs, Charles S

    2017-08-01

    The presence of human cytomegalovirus (HCMV) and glioblastoma multiforme (GBM), first established in 2002, has developed into an area of considerable interest and controversy. Numerous studies have found evidence of possible HCMV infection of GBM tumor cells as well as myriad onco- and immunomodulatory properties exhibited by HCMV antigens and transcripts, while recent reports have failed to detect HCMV particles in GBM and question the virus' role in tumor progression. This review highlights the known immunomodulatory properties of HCMV, independent of GBM infection status, that help drive the virus from peripheral blood into the vital tissues and subsequently dampen local immune response, assisting GBM tumors in evading immune surveillance and contributing to the disease's poor prognosis. Emerging antiviral approaches to treating GBM, including antiviral drugs and immunotherapies directed against HCMV, are also examined. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Modulation of the NFκb Signalling Pathway by Human Cytomegalovirus

    Science.gov (United States)

    Hancock, Meaghan H; Nelson, Jay A

    2017-01-01

    Many viruses trigger innate and adaptive immune responses and must circumvent the negative consequences to successfully establish infection in their hosts. Human Cytomegalovirus (HCMV) is no exception, and devotes a significant portion of its coding capacity to genes involved in immune evasion. Activation of the NFκB signalling pathway by viral binding and entry results in induction of antiviral and pro-inflammatory genes that have significant negative effects on HCMV infection. However, NFκB signalling stimulates transcription from the Major Immediate Early Promoter (MIEP) and pro-inflammatory signalling is crucial for cellular differentiation and viral reactivation from latency. Accordingly, HCMV encodes proteins that act to both stimulate and inhibit the NFκB signalling pathway. In this Review we will highlight the complex interactions between HCMV and NFκB, discussing the known agonists and antagonists encoded by the virus and suggest why manipulation of the pathway may be critical for both lytic and latent infections. PMID:29082387

  13. Human antibody technology and the development of antibodies against cytomegalovirus.

    Science.gov (United States)

    Ohlin, Mats; Söderberg-Nauclér, Cecilia

    2015-10-01

    Cytomegalovirus (CMV) is a virus that causes chronic infections in a large set of the population. It may cause severe disease in immunocompromised individuals, is linked to immunosenescence and implied to play an important role in the pathogenesis of cardiovascular diseases and cancer. Modulation of the immune system's abilities to manage the virus represent a highly viable therapeutic option and passive immunotherapy with polyclonal antibody preparations is already in clinical use. Defined monoclonal antibodies offer many advantages over polyclonal antibodies purified from serum. Human CMV-specific monoclonal antibodies have consequently been thoroughly investigated with respect to their potential in the treatment of diseases caused by CMV. Recent advances in human antibody technology have substantially expanded the breadth of antibodies for such applications. This review summarizes the fundamental basis for treating CMV disease by use of antibodies, the basic technologies to be used to develop such antibodies, and relevant human antibody specificities available to target this virus. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. BST2/Tetherin enhances entry of human cytomegalovirus.

    Directory of Open Access Journals (Sweden)

    Kasinath Viswanathan

    2011-11-01

    Full Text Available Interferon-induced BST2/Tetherin prevents budding of vpu-deficient HIV-1 by tethering mature viral particles to the plasma membrane. BST2 also inhibits release of other enveloped viruses including Ebola virus and Kaposi's sarcoma associated herpesvirus (KSHV, indicating that BST2 is a broadly acting antiviral host protein. Unexpectedly however, recovery of human cytomegalovirus (HCMV from supernatants of BST2-expressing human fibroblasts was increased rather than decreased. Furthermore, BST2 seemed to enhance viral entry into cells since more virion proteins were released into BST2-expressing cells and subsequent viral gene expression was elevated. A significant increase in viral entry was also observed upon induction of endogenous BST2 during differentiation of the pro-monocytic cell line THP-1. Moreover, treatment of primary human monocytes with siRNA to BST2 reduced HCMV infection, suggesting that BST2 facilitates entry of HCMV into cells expressing high levels of BST2 either constitutively or in response to exogenous stimuli. Since BST2 is present in HCMV particles we propose that HCMV entry is enhanced via a reverse-tethering mechanism with BST2 in the viral envelope interacting with BST2 in the target cell membrane. Our data suggest that HCMV not only counteracts the well-established function of BST2 as inhibitor of viral egress but also employs this anti-viral protein to gain entry into BST2-expressing hematopoietic cells, a process that might play a role in hematogenous dissemination of HCMV.

  15. Absence of human cytomegalovirus infection in childhood brain tumors.

    Science.gov (United States)

    Sardi, Iacopo; Lucchesi, Maurizio; Becciani, Sabrina; Facchini, Ludovica; Guidi, Milena; Buccoliero, Anna Maria; Moriondo, Maria; Baroni, Gianna; Stival, Alessia; Farina, Silvia; Genitori, Lorenzo; de Martino, Maurizio

    2015-01-01

    Human cytomegalovirus (HCMV) is a common human pathogen which induces different clinical manifestations related to the age and the immune conditions of the host. HCMV infection seems to be involved in the pathogenesis of adult glioblastomas. The aim of our study was to detect the presence of HCMV in high grade gliomas and other pediatric brain tumors. This hypothesis might have important therapeutic implications, offering a new target for adjuvant therapies. Among 106 pediatric patients affected by CNS tumors we selected 27 patients with a positive HCMV serology. The serological analysis revealed 7 patients with positive HCMV IGG (≥14 U/mL), whom had also a high HCMV IgG avidity, suggesting a more than 6 months-dated infection. Furthermore, HCMV IGM were positive (≥22 U/mL) in 20 patients. Molecular and immunohistochemical analyses were performed in all the 27 samples. Despite a positive HCMV serology, confirmed by ELISA, no viral DNA was shown at the PCR analysis in the patients' neoplastic cells. At immunohistochemistry, no expression of HCMV antigens was observed in tumoral cells. Our results are in agreement with recent results in adults which did not evidence the presence of HCMV genome in glioblastoma lesions. We did not find any correlation between HCMV infection and pediatric CNS tumors.

  16. Positive Expression of Human Cytomegalovirus Phosphoprotein 65 in Atherosclerosis

    Directory of Open Access Journals (Sweden)

    Zhe Wang

    2016-01-01

    Full Text Available Previous studies showed that human cytomegalovirus (HCMV is associated with atherosclerosis. However, local vascular atherosclerosis related HCMV infection and protein expression remain unclear. This study aimed to assess the relationship between HCMV infection and atherosclerosis. Formalin-fixed, paraffin-embedded peripheral artery specimens were obtained from 15 patients with atherosclerosis undergoing vascular surgery from 2008 to 2010 at Zhongnan Hospital, Wuhan University. Pathological analyses were carried out after hematoxylin and eosin (H&E and Masson trichrome staining. In situ hybridization and immunohistochemistry with two different monoclonal antibodies were employed to detect HCMV nucleic acids and proteins, respectively. H&E and Masson trichrome staining showed homogeneous extracellular matrix in femoral artery, while smooth muscle fibers were interlaced with collagen fibers; in carotid artery, inflammatory cell infiltration, foam cell vascular change, cholesterol crystals, and layered collagen fibers were observed. In situ hybridization showed no expression of HCMV nucleic acids in all 15 cases. Immunohistochemical staining for protein immediate-early protein (IE1 72 was negative in all cases, while phosphoprotein 65 (pp65 expression was detected in 14 cases. A high rate of positive pp65 signals was found in patients with atherosclerosis, suggesting that local HCMV infection may be associated with the pathogenesis of atherosclerosis. Further studies on this relationship are warranted.

  17. Positive Expression of Human Cytomegalovirus Phosphoprotein 65 in Atherosclerosis.

    Science.gov (United States)

    Wang, Zhe; Cai, Jun; Zhang, Mingming; Wang, Xiaojing; Chi, Hongjie; Feng, Haijun; Yang, Xinchun

    2016-01-01

    Previous studies showed that human cytomegalovirus (HCMV) is associated with atherosclerosis. However, local vascular atherosclerosis related HCMV infection and protein expression remain unclear. This study aimed to assess the relationship between HCMV infection and atherosclerosis. Formalin-fixed, paraffin-embedded peripheral artery specimens were obtained from 15 patients with atherosclerosis undergoing vascular surgery from 2008 to 2010 at Zhongnan Hospital, Wuhan University. Pathological analyses were carried out after hematoxylin and eosin (H&E) and Masson trichrome staining. In situ hybridization and immunohistochemistry with two different monoclonal antibodies were employed to detect HCMV nucleic acids and proteins, respectively. H&E and Masson trichrome staining showed homogeneous extracellular matrix in femoral artery, while smooth muscle fibers were interlaced with collagen fibers; in carotid artery, inflammatory cell infiltration, foam cell vascular change, cholesterol crystals, and layered collagen fibers were observed. In situ hybridization showed no expression of HCMV nucleic acids in all 15 cases. Immunohistochemical staining for protein immediate-early protein (IE1 72) was negative in all cases, while phosphoprotein 65 (pp65) expression was detected in 14 cases. A high rate of positive pp65 signals was found in patients with atherosclerosis, suggesting that local HCMV infection may be associated with the pathogenesis of atherosclerosis. Further studies on this relationship are warranted.

  18. The Human Cytomegalovirus UL44 C Clamp Wraps around DNA

    Science.gov (United States)

    Komazin-Meredith, Gloria; Petrella, Robert J.; Santos, Webster L.; Filman, David J.; Hogle, James M.; Verdine, Gregory L.; Karplus, Martin; Coen, Donald M.

    2010-01-01

    Summary Processivity factors tether the catalytic subunits of DNA polymerases to DNA so that continuous synthesis of long DNA strands is possible. The human cytomegalovirus DNA polymerase subunit UL44 forms a C clamp-shaped dimer intermediate in structure between monomeric herpes simplex virus UL42, which binds DNA directly via a basic surface, and the trimeric sliding clamp PCNA, which encircles DNA. To investigate how UL44 interacts with DNA, calculations were performed in which a 12 bp DNA oligonucleotide was docked to UL44. The calculations suggested that UL44 encircles DNA, which interacts with basic residues both within the cavity of the C clamp and in flexible loops of UL44 that complete the “circle.” The results of mutational and crosslinking studies were consistent with this model. Thus, UL44 is a “hybrid” of UL42 and PCNA: its structure is intermediate between the two and its mode of interaction with DNA has elements of both. PMID:18682223

  19. Resistance to antivirals in human cytomegalovirus: mechanisms and clinical significance.

    Science.gov (United States)

    Pérez, J L

    1997-09-01

    Long term therapies needed for managing human cytomegalovirus (HCMV) infections in immunosupressed patients provided the background for the emergence of the resistance to antivirals active against HCMV. In addition, laboratory selected mutants have also been readily achieved. Both clinical and laboratory resistant strains share the same determinants of resistance. Ganciclovir resistance may be due to a few mutations in the HCMV UL97 gene and/or viral DNA pol gene, the former being responsible for about 70% of clinical resistant isolates. Among them, V464, V594, S595 and F595 are the most frequent mutations. Because of their less extensive clinical use, much less is known about resistance to foscarnet and cidofovir (formerly, HPMPC) but in both cases, it has been associated to mutations in the DNA pol. Ganciclovir resistant strains showing DNA pol mutations are cross-resistant to cidofovir and their corresponding IC50 are normally higher than those from strains harboring only mutations at the UL97 gene. To date, foscarnet resistance seems to be independent of both ganciclovir and cidofovir resistance.

  20. The oncogenic potential of human cytomegalovirus and breast cancer.

    Directory of Open Access Journals (Sweden)

    Georges eHerbein

    2014-08-01

    Full Text Available Breast cancer is among the leading causes of cancer-related death among women. The vast majority of breast cancers are carcinomas that originate from cells lining the milk-forming ducts of the mammary gland. Numerous articles indicate that breast tumors exhibit diverse phenotypes depending on their distinct physiopathological signatures, clinical courses and therapeutic possibilities. The human cytomegalovirus (HCMV is a multifaceted highly host specific betaherpesvirus that is regarded as asymptomatic or mildly pathogenic virus in immunocompetent host. HCMV may cause serious in utero infections as well as acute and chronic complications in immunocompromised individual. The involvement of HCMV in late inflammatory complications underscores its possible role in inflammatory diseases and cancer. HCMV targets a variety of cell types in vivo, including macrophages, epithelial cells, endothelial cells, fibroblasts, stromal cells, neuronal cells, smooth muscle cells, and hepatocytes. HCMV can be detected in the milk after delivery and thereby HCMV could spread to adjacent mammary epithelial cells. HCMV also infects macrophages and induces an atypical M1/M2 phenotype, close to the tumor associated macrophage phenotype, which is associated with the release of cytokines involved in cancer initiation or promotion and breast cancer of poor prognosis. HCMV antigens and DNA have been detected in tissue biopsies of breast cancers and elevation in serum HCMV IgG antibody levels has been reported to precede the development of breast cancer in some women. In this review, we will discuss the potential role of HCMV in the initiation and progression of breast cancer.

  1. Functional annotation of human cytomegalovirus gene products: an update

    Directory of Open Access Journals (Sweden)

    Ellen eVan Damme

    2014-05-01

    Full Text Available Human Cytomegalovirus is an opportunistic double-stranded DNA virus with one of the largest viral genomes known. The 235kB genome is divided in a unique long (UL and a unique short (US region which are flanked by terminal and internal repeats. The expression of HCMV genes is highly complex and involves the production of protein coding transcripts, polyadenylated long non-coding RNAs, polyadenylated anti-sense transcripts and a variety of non-polyadenylated RNAs such as microRNAs. Although the function of many of these transcripts is unknown, they are suggested play a direct or regulatory role in the delicately orchestrated processes that ensure HCMV replication and life-long persistence. This review focuses on annotating the complete viral genome based on three sources of information. First, previous reviews were used as a template for the functional keywords to ensure continuity; second, the Uniprot database was used to further enrich the functional database; and finally, the literature was manually curated for novel functions of HCMV gene products. Novel discoveries were discussed in light of the viral life cycle. This functional annotation highlights still poorly understood regions of the genome but most importantly it can give insight in functional clusters and/or may be helpful in the analysis of transcriptomics and proteomics studies.

  2. Inhibition of Human Cytomegalovirus pUL89 Terminase Subunit Blocks Virus Replication and Genome Cleavage.

    Science.gov (United States)

    Wang, Yan; Mao, Lili; Kankanala, Jayakanth; Wang, Zhengqiang; Geraghty, Robert J

    2017-02-01

    The human cytomegalovirus terminase complex cleaves concatemeric genomic DNA into unit lengths during genome packaging and particle assembly. This process is an attractive drug target because cleavage of concatemeric DNA is not required in mammalian cell DNA replication, indicating that drugs targeting the terminase complex could be safe and selective. One component of the human cytomegalovirus terminase complex, pUL89, provides the endonucleolytic activity for genome cleavage, and the domain responsible is reported to have an RNase H-like fold. We hypothesize that the pUL89 endonuclease activity is inhibited by known RNase H inhibitors. Using a novel enzyme-linked immunosorbent assay (ELISA) format as a screening assay, we found that a hydroxypyridonecarboxylic acid compound, previously reported to be an inhibitor of human immunodeficiency virus RNase H, inhibited pUL89 endonuclease activity at low-micromolar concentrations. Further characterization revealed that this pUL89 endonuclease inhibitor blocked human cytomegalovirus replication at a relatively late time point, similarly to other reported terminase complex inhibitors. Importantly, this inhibitor also prevented the cleavage of viral genomic DNA in infected cells. Taken together, these results substantiate our pharmacophore hypothesis and validate our ligand-based approach toward identifying novel inhibitors of pUL89 endonuclease. Human cytomegalovirus infection in individuals lacking a fully functioning immune system, such as newborns and transplant patients, can have severe and debilitating consequences. The U.S. Food and Drug Administration-approved anti-human cytomegalovirus drugs mainly target the viral polymerase, and resistance to these drugs has appeared. Therefore, anti-human cytomegalovirus drugs from novel targets are needed for use instead of, or in combination with, current polymerase inhibitors. pUL89 is a viral ATPase and endonuclease and is an attractive target for anti-human cytomegalovirus

  3. Crystal Structure of the Human Cytomegalovirus Glycoprotein B.

    Directory of Open Access Journals (Sweden)

    Heidi G Burke

    2015-10-01

    Full Text Available Human cytomegalovirus (HCMV, a dsDNA, enveloped virus, is a ubiquitous pathogen that establishes lifelong latent infections and caused disease in persons with compromised immune systems, e.g., organ transplant recipients or AIDS patients. HCMV is also a leading cause of congenital viral infections in newborns. Entry of HCMV into cells requires the conserved glycoprotein B (gB, thought to function as a fusogen and reported to bind signaling receptors. gB also elicits a strong immune response in humans and induces the production of neutralizing antibodies although most anti-gB Abs are non-neutralizing. Here, we report the crystal structure of the HCMV gB ectodomain determined to 3.6-Å resolution, which is the first atomic-level structure of any betaherpesvirus glycoprotein. The structure of HCMV gB resembles the postfusion structures of HSV-1 and EBV homologs, establishing it as a new member of the class III viral fusogens. Despite structural similarities, each gB has a unique domain arrangement, demonstrating structural plasticity of gB that may accommodate virus-specific functional requirements. The structure illustrates how extensive glycosylation of the gB ectodomain influences antibody recognition. Antigenic sites that elicit neutralizing antibodies are more heavily glycosylated than those that elicit non-neutralizing antibodies, which suggest that HCMV gB uses glycans to shield neutralizing epitopes while exposing non-neutralizing epitopes. This glycosylation pattern may have evolved to direct the immune response towards generation of non-neutralizing antibodies thus helping HCMV to avoid clearance. HCMV gB structure provides a starting point for elucidation of its antigenic and immunogenic properties and aid in the design of recombinant vaccines and monoclonal antibody therapies.

  4. Crystal structure of human cytomegalovirus IL-10 bound to soluble human IL-10R1

    OpenAIRE

    Jones, Brandi C.; Logsdon, Naomi J.; Josephson, Kristopher; Cook, Jennifer; Barry, Peter A.; Walter, Mark R.

    2002-01-01

    Human IL-10 (hIL-10) modulates critical immune and inflammatory responses by way of interactions with its high- (IL-10R1) and low-affinity (IL-10R2) cell surface receptors. Human cytomegalovirus exploits the IL-10 signaling pathway by expressing a functional viral IL-10 homolog (cmvIL-10), which shares only 27% sequence identity with hIL-10 yet signals through IL-10R1 and IL-10R2. To define the molecular basis of this virus–host interaction, we determined the 2.7-Å crysta...

  5. Letermovir Treatment of Human Cytomegalovirus Infection Antiinfective Agent

    OpenAIRE

    Verghese, Priya S.; Schleiss, Mark R.

    2013-01-01

    Novel therapies are urgently needed for the management of cytomegalovirus (CMV) disease in high-risk patients. Currently licensed agents target the viral DNA polymerase, and although they are effective, they are fraught with toxicities to patients. Moreover, emergence of antiviral resistance is an increasing problem, particularly for patients on long-term suppressive therapy. A new agent, letermovir (AIC246), shows great promise for the management of CMV infection. Advantages include its good...

  6. Infection of human endothelium in vitro by cytomegalovirus causes enhanced expression of purinergic receptors : A potential virus escape mechanism?

    NARCIS (Netherlands)

    Zandberg, Mariet; van Son, Willem J.; Harmsen, Martin C.; Bakker, Winston W.

    2007-01-01

    Background. Human cytomegalovirus (CMV) uses different strategies to escape from human host defense reactions. Previously we have observed that infection of endothelial cells with CMV in vitro leads to enhanced activity of endothelial ectonucleotidases. These ectoenzymes are responsible for

  7. Complete Genome Sequence of a Human Cytomegalovirus Strain AD169 Bacterial Artificial Chromosome Clone

    OpenAIRE

    Ostermann, Eleonore; Spohn, Michael; Indenbirken, Daniela; Brune, Wolfram

    2016-01-01

    The complete sequence of the human cytomegalovirus strain AD169 (variant ATCC) cloned as a bacterial artificial chromosome (AD169-BAC, also known as HB15 or pHB15) was determined. The viral genome has a length of 230,290?bp and shows 52 nucleotide differences compared to a previously sequenced AD169varATCC clone.

  8. Partial functional complementation between human and mouse cytomegalovirus chemokine receptor homologues

    DEFF Research Database (Denmark)

    Farrell, Helen E; Abraham, Alexander M; Cardin, Rhonda D

    2011-01-01

    The human cytomegalovirus (CMV) proteins US28 and UL33 are homologous to chemokine receptors (CKRs). Knockout of the mouse CMV M33 protein (UL33 homologue) results in substantial attenuation of salivary gland infection/replication and reduced efficiency of reactivation from tissue explants. M33...

  9. Murine cytomegalovirus (CMV) M33 and human CMV US28 receptors exhibit similar constitutive signaling activities

    DEFF Research Database (Denmark)

    Waldhoer, Maria; Kledal, Thomas N; Farrell, Helen

    2002-01-01

    Cellular infection by cytomegalovirus (CMV) is associated with very early G-protein-mediated signal transduction and reprogramming of gene expression. Here we investigated the involvement of human CMV (HCMV)-encoded US27, US28, and UL33 receptors as well as murine CMV-encoded M33 transmembrane (7TM...

  10. Relationship between cytomegalovirus infection and procoagulant changes in human immunodeficiency virus-infected patients

    NARCIS (Netherlands)

    Mulder, R.; Tichelaar, Y. I. G. V.; Sprenger, H. G.; Mulder, A. B.; Lijfering, W. M.

    P>Cytomegalovirus is associated with hypercoagulability, and is reported to increase the risk of venous thrombosis in human immunodeficiency virus (HIV)-infected patients. Progression to AIDS, however, is also associated with hypercoagulability and venous thrombosis, and may result in more

  11. Human cytomegalovirus-encoded miR-US4-1 promotes cell ...

    Indian Academy of Sciences (India)

    2016-04-05

    Apr 5, 2016 ... Human cytomegalovirus (HCMV) can cause congenital diseases and opportunistic infections in immunocompromised individuals. Its functional proteins and microRNAs (miRNAs) facilitate efficient viral propagation by altering host cell behaviour. Identification of functional target genes of miRNAs is an ...

  12. Human cytomegalovirus induced pseudotumor of upper gastrointestinal tract mucosa: effects of long-term chronic disease?

    Science.gov (United States)

    Reggiani Bonetti, Luca; Barresi, Valeria; Bertani, Angela; Maccio, Livia; Palmiere, Cristian

    2015-06-01

    Human cytomegalovirus-induced lesions resembling malignancies have been described in the gastrointestinal tract and include ulcerated or exophytic large masses. The aim of this study was to review the cases registered in the databases of two academic hospitals and formulate a hypothesis concerning the pathogenic mechanisms responsible for cytomegalovirus-induced pseudotumor development. All the diagnoses of human cytomegalovirus infections of the upper gastrointestinal tract recorded from 1991 to 2013 were reviewed. Cases of mucosal alterations misdiagnosed endoscopically as malignancies were selected. Large ulcers occurring in the stomach (three cases) and an irregular exophytic mass at the gastro-jejunal anastomosis were misdiagnosed endoscopically as malignancies (4 cases out of 53). Histologically, all lesions reflected hyperplastic mucosal changes with a prevalence of epithelial and stroma infected cells, without signs of cell atypia. The hypothesis presented is that the development of human cytomegalovirus-induced pseudotumors may be the morphological expression of chronic mucosa damage underlying long-term infection. © 2015 Wiley Periodicals, Inc.

  13. Human Cytomegalovirus and Epstein-Barr Virus Genotypes in Apical Periodontitis Lesions.

    Science.gov (United States)

    Jakovljevic, Aleksandar; Andric, Miroslav; Knezevic, Aleksandra; Soldatovic, Ivan; Nikolic, Nadja; Karalic, Danijela; Milasin, Jelena

    2015-11-01

    Different genotypes of human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV) possess specific pathogenic abilities because of various interactions with the host's immune system and differences in cell tropism. The aim of this study was to determine the distribution of HCMV and EBV genotypes in apical periodontitis lesions in relation to their clinical and histopathologic features. One hundred samples of apical periodontitis lesions and 25 control samples (healthy pulp tissue) were collected. The presence of HCMV glycoprotein B (gB) and EBV nuclear antigen-2 genotypes was analyzed by nested polymerase chain reaction and restriction fragment length polymorphisms analysis. EBV and HCMV were detected in apical periodontitis lesions at significantly higher frequencies than in healthy pulp controls (P = .020 and P = .020, respectively). HCMV gB type II was significantly more frequent compared with gB type I in the examined groups (P = .036). No HCMV gB type III or IV products were found. In both periapical lesions and controls, EBV-1 occurred more often compared with EBV-2 (P = .001). Dual EBV and HCMV coinfection was more frequently detected in large-size periapical lesions (P = .038). Both HCMV and EBV are associated with inflammatory processes of periapical bone destruction. HCMV gB type II and EBV-1 are the most prevalent genotypes in apical periodontitis lesions. Copyright © 2015 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  14. The eIF4AIII RNA helicase is a critical determinant of human cytomegalovirus replication

    Energy Technology Data Exchange (ETDEWEB)

    Ziehr, Ben; Lenarcic, Erik; Cecil, Chad; Moorman, Nathaniel J., E-mail: nmoorman@med.unc.edu

    2016-02-15

    Human cytomegalovirus (HCMV) was recently shown to encode a large number of spliced mRNAs. While the nuclear export of unspliced viral transcripts has been extensively studied, the role of host mRNA export factors in HCMV mRNA trafficking remains poorly defined. We found that the eIF4AIII RNA helicase, a component of the exon junction complex, was necessary for efficient virus replication. Depletion of eIF4AIII limited viral DNA accumulation, export of viral mRNAs from the nucleus, and the production of progeny virus. However eIF4AIII was dispensable for the association of viral transcripts with ribosomes. We found that pateamine A, a natural compound that inhibits both eIF4AI/II and eIF4AIII, has potent antiviral activity and inhibits HCMV replication throughout the virus lytic cycle. Our results demonstrate that eIF4AIII is required for efficient HCMV replication, and suggest that eIF4A family helicases may be a new class of targets for the development of host-directed antiviral therapeutics. - Highlights: • The host eIF4AIII RNA helicase is required for efficient HCMV replication. • Depleting eIF4AIII inhibited the nuclear export of HCMV mRNAs. • HCMV mRNAs did not require eIF4AIII to associate with polyribosomes. • The eIF4A family helicases may be new targets for host-directed antiviral drugs.

  15. Letermovir Treatment of Human Cytomegalovirus Infection Antiinfective Agent.

    Science.gov (United States)

    Verghese, Priya S; Schleiss, Mark R

    2013-05-01

    Novel therapies are urgently needed for the management of cytomegalovirus (CMV) disease in high-risk patients. Currently licensed agents target the viral DNA polymerase, and although they are effective, they are fraught with toxicities to patients. Moreover, emergence of antiviral resistance is an increasing problem, particularly for patients on long-term suppressive therapy. A new agent, letermovir (AIC246), shows great promise for the management of CMV infection. Advantages include its good oral bioavailability, its lack of toxicity, and the apparent absence of drug-drug interactions. Letermovir has a novel mechanism of action, exerting its antiviral effect by interfering with the viral pUL56 gene product and in the process disrupting the viral terminase complex. This agent demonstrates substantial promise as an alternative to more toxic antivirals in patients at high risk for CMV disease, particularly in the transplantation setting.

  16. Cytoskeleton involvement during human cytomegalovirus replicative cycle in human embryo fibroblasts.

    Science.gov (United States)

    Arcangeletti, M C; Pinardi, F; Medici, M C; Pilotti, E; De Conto, F; Ferraglia, F; Landini, M P; Chezzi, C; Dettori, G

    2000-07-01

    Several studies indicate that viruses can induce different cytoskeletal modifications. The present investigation examines the possible involvement of human embryo fibroblast cytoskeleton in the replication of human cytomegalovirus (HCMV). Significant cytoskeletal modifications occur in infected cells; specifically, microfilament depolymerization is observed very early during the HCMV replicative cycle, whilst microtubules and intermediate filaments do not undergo any change for longer times after infection. Our data focus, in particular, on microfilament depolymerization, which starts within the first hour of the replicative cycle, and on the significance of this event, as a CMV-induced mechanism to modify the post-transcriptional regulation of cellular gene expression for its own benefit. Among the possible mechanisms exploited by HCMV to induce microfilament modifications, one might involve the cellular ADP-ribosylation activity, which is increased by HCMV very early in the infectious cycle. Experiments carried out on HCMV-infected cells, in the presence of ADP-ribosylation inhibitors, seem to confirm this hypothesis.

  17. Receptor expression and responsiveness of human peripheral blood mononuclear cells to a human cytomegalovirus encoded CC chemokine.

    Science.gov (United States)

    Zheng, Qi; Xu, Jun; Gao, Huihui; Tao, Ran; Li, Wei; Shang, Shiqiang; Gu, Weizhong

    2015-01-01

    Human cytomegalovirus is a ubiquitous pathogen that infects the majority of the world's population. After long period of time co-evolving with human being, this pathogen has developed several strategies to evade host immune surveillance. One of the major trick is encoding homologous to those of the host organism or stealing host cellular genes that have significant functions in immune system. To date, we have found several viral immune analogous which include G protein coupled receptor, class I major histocompatibility complex and chemokine. Chemokine is a small group of molecules which is defined by the presence of four cysteines in highly conserved region. The four kinds of chemokines (C, CC, CXC, and CX3C) are classified based on the arrangement of 1 or 2 N-terminal cysteine residues. UL128 protein is one of the analogous that encoded by human cytomegalovirus that has similar amino acid sequences to the human CC chemokine. It has been proved to be one of the essential particles that involved in human cytomegalovirus entry into epithelial/endothelial cells as well as macrophages. It is also the target of potent neutralizing antibodies in human cytomegalovirus-seropositive individuals. We had demonstrated the chemotactic trait of UL128 protein in our previous study. Recombinant UL128 in vitro has the ability to attract monocytes to the infection region and enhances peripheral blood mononuclear cell proliferation by activating the MAPK/ERK signaling pathway. However, the way that this viral encoded chemokine interacting with peripheral blood mononuclear cells and the detailed mechanism that involving the virus entry into host cells keeps unknown. Here we performed in vitro investigation into the effects of UL128 protein on peripheral blood mononuclear cell's activation and receptor binding, which may help us further understand the immunomodulatory function of UL128 protein as well as human cytomegalovirus diffusion mechanism. Copyright © 2015 Elsevier Editora Ltda

  18. Cytomegalovirus retinitis

    Science.gov (United States)

    ... sharing features on this page, please enable JavaScript. Cytomegalovirus (CMV) retinitis is a viral infection of the ... need treatment to prevent its return. Alternative Names Cytomegalovirus retinitis Images Eye CMV retinitis CMV (cytomegalovirus) References ...

  19. Complete Genome Sequence of a Human Cytomegalovirus Strain AD169 Bacterial Artificial Chromosome Clone.

    Science.gov (United States)

    Ostermann, Eleonore; Spohn, Michael; Indenbirken, Daniela; Brune, Wolfram

    2016-03-31

    The complete sequence of the human cytomegalovirus strain AD169 (variant ATCC) cloned as a bacterial artificial chromosome (AD169-BAC, also known as HB15 or pHB15) was determined. The viral genome has a length of 230,290 bp and shows 52 nucleotide differences compared to a previously sequenced AD169varATCC clone. Copyright © 2016 Ostermann et al.

  20. Geno- and phenotypic characterization of human cytomegalovirus mutants selected in vitro after letermovir (AIC246) exposure.

    Science.gov (United States)

    Goldner, Thomas; Hempel, Christine; Ruebsamen-Schaeff, Helga; Zimmermann, Holger; Lischka, Peter

    2014-01-01

    Letermovir is a novel antiviral compound currently in clinical development for the prevention of human cytomegalovirus (HCMV) infections. In contrast to all currently approved anti-HCMV drugs that target the viral DNA polymerase, letermovir acts via a distinct mode of action involving the viral terminase subunit pUL56. To extend our understanding of potential letermovir resistance mechanisms, we used marker transfer to characterize mutations identified in letermovir-resistant HCMV variants that were selected in cell culture.

  1. Geno- and Phenotypic Characterization of Human Cytomegalovirus Mutants Selected In Vitro after Letermovir (AIC246) Exposure

    OpenAIRE

    Goldner, Thomas; Hempel, Christine; Ruebsamen-Schaeff, Helga; Zimmermann, Holger; Lischka, Peter

    2014-01-01

    Letermovir is a novel antiviral compound currently in clinical development for the prevention of human cytomegalovirus (HCMV) infections. In contrast to all currently approved anti-HCMV drugs that target the viral DNA polymerase, letermovir acts via a distinct mode of action involving the viral terminase subunit pUL56. To extend our understanding of potential letermovir resistance mechanisms, we used marker transfer to characterize mutations identified in letermovir-resistant HCMV variants th...

  2. The Human Cytomegalovirus Major Immediate-Early Proteins as Antagonists of Intrinsic and Innate Antiviral Host Responses

    Directory of Open Access Journals (Sweden)

    Michael Nevels

    2009-11-01

    Full Text Available The major immediate-early (IE gene of human cytomegalovirus (CMV is believed to have a decisive role in acute infection and its activity is an important indicator of viral reactivation from latency. Although a variety of gene products are expressed from this region, the 72-kDa IE1 and the 86-kDa IE2 nuclear phosphoproteins are the most abundant and important. Both proteins have long been recognized as promiscuous transcriptional regulators. More recently, a critical role of the IE1 and IE2 proteins in counteracting nonadaptive host cell defense mechanisms has been revealed. In this review we will briefly summarize the available literature on IE1- and IE2-dependent mechanisms contributing to CMV evasion from intrinsic and innate immune responses.

  3. A Phase 1 Study of 4 Live, Recombinant Human Cytomegalovirus Towne/Toledo Chimera Vaccines in Cytomegalovirus-Seronegative Men.

    Science.gov (United States)

    Adler, Stuart P; Manganello, Anne-Marie; Lee, Ronzo; McVoy, Michael A; Nixon, Daniel E; Plotkin, Stanley; Mocarski, Edward; Cox, Josephine H; Fast, Patricia E; Nesterenko, Pavlo A; Murray, Susan E; Hill, Ann B; Kemble, George

    2016-11-01

    Human cytomegalovirus (HCMV) infection causes disease in newborns and transplant recipients. A HCMV vaccine (Towne) protects transplant recipients.  The genomes of Towne and the nonattenuated Toledo strain were recombined, yielding 4 Towne/Toledo chimera vaccines. Each of 36 HCMV-seronegative men received 1 subcutaneous dose of 10, 100, or 1000 plaque-forming units (PFU) in cohorts of 3. Safety and immunogenicity were evaluated over 12 weeks after immunization and for 52 weeks for those who seroconverted.  There were no serious local or systemic reactions. No subject had HCMV in urine or saliva. For chimera 3, none of 9 subjects seroconverted. For chimera 1, 1 of 9 seroconverted (the seroconverter received 100 PFU). For chimera 2, 3 subjects seroconverted (1 received 100 PFU, and 2 received 1000 PFU). For chimera 4, 7 subjects seroconverted (1 received 10 PFU, 3 received 100 PFU, and 3 received 1000 PFU). All 11 seroconverters developed low but detectable levels of neutralizing activity. CD4+ T-cell responses were detectable in 1 subject (who received 100 PFU of chimera 4). Seven subjects receiving chimera 2 or 4 had detectable CD8+ T-cell responses to IE1; 3 responded to 1-2 additional antigens.  The Towne/Toledo chimera vaccine candidates were well tolerated and were not excreted. Additional human trials of chimeras 2 and 4 are appropriate.  NCT01195571. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  4. Modulation of the Host Environment by Human Cytomegalovirus with Viral Interleukin 10 in Peripheral Blood.

    Science.gov (United States)

    Young, Vivian P; Mariano, Margarette C; Tu, Carolyn C; Allaire, Kathryn M; Avdic, Selmir; Slobedman, Barry; Spencer, Juliet V

    2017-03-15

    Human cytomegalovirus (HCMV) is a herpesvirus with both lytic and latent life cycles. Human cytomegalovirus encodes 2 viral cytokines that are orthologs of human cellular interleukin 10 (cIL-10). Both cytomegalovirus interleukin 10 (cmvIL-10) and Latency-associated cytomegalovirus interleukin 10 (LAcmvIL-10) (collectively vIL-10) are expressed during lytic infection and cause immunosuppressive effects that impede virus clearance. LAcmvIL-10 is also expressed during latent infection of myeloid progenitor cells and monocytes and facilitates persistence. Here, we investigated whether vIL-10 could be detected during natural infection. Plasma from healthy blood donors was tested by enzyme-linked immunosorbent assay for anti-HCMV immunoglobulin G and immunoglobulin M and for cIL-10 and vIL-10 levels using a novel vIL-10 assay that detects cmvIL-10 and LAcmvIL-10, with no cross-reactivity to cIL-10. vIL-10 was evident in HCMV+ donors (n = 19 of 26), at levels ranging 31-547 pg/mL. By comparison, cIL-10 was detected at lower levels ranging 3-69 pg/mL. There was a strong correlation between vIL-10 and cIL-10 levels (P = .01). Antibodies against vIL-10 were also detected and neutralized vIL-10 activity. vIL-10 was detected in peripheral blood of healthy blood donors. These findings suggest that vIL-10 may play a key role in sensing or modifying the host environment during latency and, therefore, may be a potential target for intervention strategies.

  5. Human Cytomegalovirus Intrahost Evolution – A New Avenue for Understanding and Controlling Herpesvirus Infections

    Science.gov (United States)

    Renzette, Nicholas; Gibson, Laura; Jensen, Jeffrey D.; Kowalik, Timothy F.

    2014-01-01

    Human cytomegalovirus (HCMV) is exquisitely adapted to the human host, and much research has focused on its evolution over long timescales spanning millennia. Here, we review recent data exploring the evolution of the virus on much shorter timescales, on the order of days or months. We describe the intrahost genetic diversity of the virus isolated from humans, and how this diversity contributes to HCMV spatiotemporal evolution. We propose mechanisms to explain the high levels of intrahost diversity and discuss how this new information may shed light on HCMV infection and pathogenesis. PMID:25154343

  6. Human induced pluripotent stem cell-derived models to investigate human cytomegalovirus infection in neural cells.

    Directory of Open Access Journals (Sweden)

    Leonardo D'Aiuto

    Full Text Available Human cytomegalovirus (HCMV infection is one of the leading prenatal causes of congenital mental retardation and deformities world-wide. Access to cultured human neuronal lineages, necessary to understand the species specific pathogenic effects of HCMV, has been limited by difficulties in sustaining primary human neuronal cultures. Human induced pluripotent stem (iPS cells now provide an opportunity for such research. We derived iPS cells from human adult fibroblasts and induced neural lineages to investigate their susceptibility to infection with HCMV strain Ad169. Analysis of iPS cells, iPS-derived neural stem cells (NSCs, neural progenitor cells (NPCs and neurons suggests that (i iPS cells are not permissive to HCMV infection, i.e., they do not permit a full viral replication cycle; (ii Neural stem cells have impaired differentiation when infected by HCMV; (iii NPCs are fully permissive for HCMV infection; altered expression of genes related to neural metabolism or neuronal differentiation is also observed; (iv most iPS-derived neurons are not permissive to HCMV infection; and (v infected neurons have impaired calcium influx in response to glutamate.

  7. Saturated very long chain fatty acids are required for the production of infectious human cytomegalovirus progeny.

    Directory of Open Access Journals (Sweden)

    Emre Koyuncu

    Full Text Available Human cytomegalovirus hijacks host cell metabolism, increasing the flux of carbon from glucose to malonyl-CoA, the committed precursor to fatty acid synthesis and elongation. Inhibition of acetyl-CoA carboxylase blocks the production of progeny virus. To probe further the role of fatty acid metabolism during infection, we performed an siRNA screen to identify host cell metabolic enzymes needed for the production of infectious cytomegalovirus progeny. The screen predicted that multiple long chain acyl-CoA synthetases and fatty acid elongases are needed during infection, and the levels of RNAs encoding several of these enzymes were upregulated by the virus. Roles for acyl-CoA synthetases and elongases during infection were confirmed by using small molecule antagonists. Consistent with a role for these enzymes, mass spectrometry-based fatty acid analysis with ¹³C-labeling revealed that malonyl-CoA is consumed by elongases to produce very long chain fatty acids, generating an approximately 8-fold increase in C26-C34 fatty acid tails in infected cells. The virion envelope was yet further enriched in C26-C34 saturated fatty acids, and elongase inhibitors caused the production of virions with lower levels of these fatty acids and markedly reduced infectivity. These results reveal a dependence of cytomegalovirus on very long chain fatty acid metabolism.

  8. Probable neuroimmunological link between Toxoplasma and cytomegalovirus infections and personality changes in the human host

    Directory of Open Access Journals (Sweden)

    Roubalová Kateřina

    2005-07-01

    Full Text Available Abstract Background Recently, a negative association between Toxoplasma-infection and novelty seeking was reported. The authors suggested that changes of personality trait were caused by manipulation activity of the parasite, aimed at increasing the probability of transmission of the parasite from an intermediate to a definitive host. They also suggested that low novelty seeking indicated an increased level of the neurotransmitter dopamine in the brain of infected subjects, a phenomenon already observed in experimentally infected rodents. However, the changes in personality can also be just a byproduct of any neurotropic infection. Moreover, the association between a personality trait and the toxoplasmosis can even be caused by an independent correlation of both the probability of Toxoplasma-infection and the personality trait with the third factor, namely with the size of living place of a subject. To test these two alternative hypotheses, we studied the influence of another neurotropic pathogen, the cytomegalovirus, on the personality of infected subjects, and reanalyzed the original data after the effect of the potential confounder, the size of living place, was controlled. Methods In the case-control study, 533 conscripts were tested for toxoplasmosis and presence of anti-cytomegalovirus antibodies and their novelty seeking was examined with Cloninger's TCI questionnaire. Possible association between the two infections and TCI dimensions was analyzed. Results The decrease of novelty seeking is associated also with cytomegalovirus infection. After the size of living place was controlled, the effect of toxoplasmosis on novelty seeking increased. Significant difference in novelty seeking was observed only in the largest city, Prague. Conclusion Toxoplasma and cytomegalovirus probably induce a decrease of novelty seeking. As the cytomegalovirus spreads in population by direct contact (not by predation as with Toxoplasma, the observed changes are

  9. Detection of Human Cytomegalovirus and Epstein-Barr Virus in Coronary Atherosclerotic Tissue

    Science.gov (United States)

    Imbronito, Ana Vitória; Marcelino, Silvia Linardi; Grande, Sabrina Rosa; Nunes, Fabio Daumas; Romito, Giuseppe Alexandre

    2010-01-01

    Previous studies indicated that patients with atherosclerosis are predominantly infected by human cytomegalovirus (HCMV), but rarely infected by type 1 Epstein-Barr virus (EBV-1). In this study, atheromas of 30 patients who underwent aortocoronary bypass surgery with coronary endartherectomy were tested for the presence of these two viruses. HCMV occurred in 93.3% of the samples and EBV-1 was present in 50% of them. Concurrent presence of both pathogens was detected in 43.3% of the samples. PMID:24031529

  10. Human Cytomegalovirus Strategies to Maintain and Promote mRNA Translation.

    Science.gov (United States)

    Vincent, Heather A; Ziehr, Benjamin; Moorman, Nathaniel J

    2016-04-13

    mRNA translation requires the ordered assembly of translation initiation factors and ribosomal subunits on a transcript. Host signaling pathways regulate each step in this process to match levels of protein synthesis to environmental cues. In response to infection, cells activate multiple defenses that limit viral protein synthesis, which viruses must counteract to successfully replicate. Human cytomegalovirus (HCMV) inhibits host defenses that limit viral protein expression and manipulates host signaling pathways to promote the expression of both host and viral proteins necessary for virus replication. Here we review key regulatory steps in mRNA translation, and the strategies used by HCMV to maintain protein synthesis in infected cells.

  11. Cytomegalovirus infection presenting as acute periodontal infection in a patient infected with the human immunodeficiency virus.

    Science.gov (United States)

    Dodd, C L; Winkler, J R; Heinic, G S; Daniels, T E; Yee, K; Greenspan, D

    1993-04-01

    During childhood, many people acquire primary infection with cytomegalovirus (CMV), one of the herpes viruses. If they later become immunosuppressed, such as occurs with human immunodeficiency virus (HIV) infection, CMV is likely to become reactivated. Severe disease caused by CMV is life-threatening in the HIV-infected population. CMV retinitis, gastritis, colitis, pneumonia, encephalitis and hepatitis have all been reported, but oral lesions due to infection with CMV are rarely reported. We report a case of oral CMV infection which at first was clinically indistinguishable from HIV-associated periodontal disease.

  12. Sequestration of human cytomegalovirus by human renal and mammary epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Twite, Nicolas [Institute for Medical Immunology, Université Libre de Bruxelles, Rue A. Bolland 8, B-6041 Charleroi (Belgium); Andrei, Graciela [Laboratory of Virology and Chemotherapy, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven (Belgium); Kummert, Caroline [ImmuneHealth, Rue A. Bolland 8, B-6041 Charleroi (Belgium); Donner, Catherine [Department of Obstetrics and Gynecology, Erasme Hospital, Route de Lennik 808, 1070 Brussels (Belgium); Perez-Morga, David [Laboratory of Molecular Parasitology, Institut de Biologie et Médecine Moléculaires, Université Libre de Bruxelles, Gosselies (Belgium); De Vos, Rita [Pathology Department, U.Z. Leuven, Minderbroedersstraat 12, Leuven (Belgium); Snoeck, Robert, E-mail: Robert.Snoeck@Rega.kuleuven.be [Laboratory of Virology and Chemotherapy, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven (Belgium); Marchant, Arnaud, E-mail: arnaud.marchant@ulb.ac.be [Institute for Medical Immunology, Université Libre de Bruxelles, Rue A. Bolland 8, B-6041 Charleroi (Belgium); ImmuneHealth, Rue A. Bolland 8, B-6041 Charleroi (Belgium)

    2014-07-15

    Urine and breast milk represent the main routes of human cytomegalovirus (HCMV) transmission but the contribution of renal and mammary epithelial cells to viral excretion remains unclear. We observed that kidney and mammary epithelial cells were permissive to HCMV infection and expressed immediate early, early and late antigens within 72 h of infection. During the first 24 h after infection, high titers of infectious virus were measured associated to the cells and in culture supernatants, independently of de novo synthesis of virus progeny. This phenomenon was not observed in HCMV-infected fibroblasts and suggested the sequestration and the release of HCMV by epithelial cells. This hypothesis was supported by confocal and electron microscopy analyses. The sequestration and progressive release of HCMV by kidney and mammary epithelial cells may play an important role in the excretion of the virus in urine and breast milk and may thereby contribute to HCMV transmission. - Highlights: • Primary renal and mammary epithelial cells are permissive to HCMV infection. • HCMV is sequestered by epithelial cells and this phenomenon does not require viral replication. • HCMV sequestration by epithelial cells is reduced by antibodies and IFN-γ.

  13. The Carboxy Terminal Region of the Human Cytomegalovirus Immediate Early 1 (IE1 Protein Disrupts Type II Inteferon Signaling

    Directory of Open Access Journals (Sweden)

    Bindu Raghavan

    2014-04-01

    Full Text Available Interferons (IFNs activate the first lines of defense against viruses, and promote innate and adaptive immune responses to viruses. We report that the immediate early 1 (IE1 protein of human cytomegalovirus (HCMV disrupts signaling by IFNγ. The carboxyl-terminal region of IE1 is required for this function. We found no defect in the initial events in IFNγ signaling or in nuclear accumulation of signal transducer and activator of transcription 1 (STAT1 in IE1-expressing cells. Moreover, we did not observe an association between disruption of IFNγ signaling and nuclear domain 10 (ND10 disruption. However, there is reduced binding of STAT1 homodimers to target gamma activated sequence (GAS elements in the presence of IE1. Co-immunoprecipitation studies failed to support a direct interaction between IE1 and STAT1, although these studies revealed that the C-terminal region of IE1 was required for interaction with STAT2. Together, these results indicate that IE1 disrupts IFNγ signaling by interfering with signaling events in the nucleus through a novel mechanism.

  14. Identification of common mechanisms by which human and mouse cytomegalovirus seven-transmembrane receptor homologues contribute to in vivo phenotypes in a mouse model

    DEFF Research Database (Denmark)

    Farrell, Helen E; Abraham, Alexander M; Cardin, Rhonda D

    2013-01-01

    The mouse cytomegalovirus chemokine receptor homologue (CKR) M33 is required for salivary gland tropism and efficient reactivation from latency, phenotypes partially rescued by the human cytomegalovirus CKR US28. Herein, we demonstrate that complementation of salivary gland tropism is mediated...

  15. Human cytomegalovirus UL145 gene is highly conserved among ...

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Biosciences; Volume 32; Issue 6 ... Articles Volume 32 Issue 6 September 2007 pp 1111-1118 ... strains, irrespective of whether the strains come from patients with different manifestations, from different areas of the world, or were passaged or not in human embryonic lung fibroblast (HELF) cells.

  16. Human cytomegalovirus antigens in malignant gliomas as targets for adoptive cellular therapy

    Directory of Open Access Journals (Sweden)

    Daniel eLandi

    2014-11-01

    Full Text Available Malignant gliomas are the most common primary brain tumor in adults, with over 12,000 new cases diagnosed in the United States each year. Over the last decade, investigators have reliably identified human cytomegalovirus (HCMV proteins, nucleic acids, and virions in most high-grade gliomas, including glioblastoma (GBM. This discovery is significant because human cytomegalovirus gene products can be targeted by immune-based therapies.In this review, we describe the current level of understanding regarding the presence and role in pathogenesis of HCMV in GBM. We describe our success detecting and expanding HCMV-specific cytotoxic T lymphocytes to kill GBM cells and explain how these cells can be used as a platform for enhanced cellular therapies. We discuss alternative approaches that capitalize on HCMV infection to treat patients with HCMV-positive tumors. Adoptive cellular therapy for HCMV-positive GBM has been tried in a small number of patients with some benefit, but we reason why, to date, these approaches generally fail to generate long-term remission or cure. We conjecture how cellular therapy for GBM can be improved and describe the barriers that must be overcome to cure these patients.

  17. A Hsp40 chaperone protein interacts with and modulates the cellular distribution of the primase protein of human cytomegalovirus.

    Directory of Open Access Journals (Sweden)

    Yonggang Pei

    Full Text Available Genomic DNA replication is a universal and essential process for all herpesvirus including human cytomegalovirus (HCMV. HCMV UL70 protein, which is believed to encode the primase activity of the viral DNA replication machinery and is highly conserved among herpesviruses, needs to be localized in the nucleus, the site of viral DNA synthesis. No host factors that facilitate the nuclear import of UL70 have been reported. In this study, we provided the first direct evidence that UL70 specifically interacts with a highly conserved and ubiquitously expressed member of the heat shock protein Hsp40/DNAJ family, DNAJB6, which is expressed as two isoforms, a and b, as a result of alternative splicing. The interaction of UL70 with a common region of DNAJB6a and b was identified by both a two hybrid screen in yeast and coimmunoprecipitation in human cells. In transfected cells, UL70 was primarily co-localized with DNAJB6a in the nuclei and with DNAJB6b in the cytoplasm, respectively. The nuclear import of UL70 was increased in cells in which DNAJB6a was up-regulated or DNAJB6b was down-regulated, and was reduced in cells in which DNAJB6a was down-regulated or DNAJB6b was up-regulated. Furthermore, the level of viral DNA synthesis and progeny production was increased in cells in which DNAJB6a was up-regulated or DNAJB6b was down-regulated, and was reduced in cells in which DNAJB6a was down-regulated or DNAJB6b was up-regulated. Thus, DNAJB6a and b appear to enhance the nuclear import and cytoplasmic accumulation of UL70, respectively. Our results also suggest that the relative expression levels of DNAJB6 isoforms may play a key role in regulating the cellular localization of UL70, leading to modulation of HCMV DNA synthesis and lytic infection.

  18. Infection and upregulation of proinflammatory cytokines in human brain vascular pericytes by human cytomegalovirus

    Directory of Open Access Journals (Sweden)

    Alcendor Donald J

    2012-05-01

    Full Text Available Abstract Background Congenital human cytomegalovirus (HCMV infections can result in CNS abnormalities in newborn babies including vision loss, mental retardation, motor deficits, seizures, and hearing loss. Brain pericytes play an essential role in the development and function of the blood–brain barrier yet their unique role in HCMV dissemination and neuropathlogy has not been reported. Methods Primary human brain vascular pericytes were exposed to a primary clinical isolate of HCMV designated ‘SBCMV’. Infectivity was analyzed by microscopy, immunofluorescence, Western blot, and qRT-PCR. Microarrays were performed to identify proinflammatory cytokines upregulated after SBCMV exposure, and the results validated by real-time quantitative polymerase chain reaction (qPCR methodology. In situ cytokine expression of pericytes after exposure to HCMV was examined by ELISA and in vivo evidence of HCMV infection of brain pericytes was shown by dual-labeled immunohistochemistry. Results HCMV-infected human brain vascular pericytes as evidenced by several markers. Using a clinical isolate of HCMV (SBCMV, microscopy of infected pericytes showed virion production and typical cytomegalic cytopathology. This finding was confirmed by the expression of major immediate early and late virion proteins and by the presence of HCMV mRNA. Brain pericytes were fully permissive for CMV lytic replication after 72 to 96 hours in culture compared to human astrocytes or human brain microvascular endothelial cells (BMVEC. However, temporal transcriptional expression of pp65 virion protein after SBCMV infection was lower than that seen with the HCMV Towne laboratory strain. Using RT-PCR and dual-labeled immunofluorescence, proinflammatory cytokines CXCL8/IL-8, CXCL11/ITAC, and CCL5/Rantes were upregulated in SBCMV-infected cells, as were tumor necrosis factor-alpha (TNF-alpha, interleukin-1 beta (IL-1beta, and interleukin-6 (IL-6. Pericytes exposed to SBCMV elicited

  19. Infection and upregulation of proinflammatory cytokines in human brain vascular pericytes by human cytomegalovirus

    Science.gov (United States)

    2012-01-01

    Background Congenital human cytomegalovirus (HCMV) infections can result in CNS abnormalities in newborn babies including vision loss, mental retardation, motor deficits, seizures, and hearing loss. Brain pericytes play an essential role in the development and function of the blood–brain barrier yet their unique role in HCMV dissemination and neuropathlogy has not been reported. Methods Primary human brain vascular pericytes were exposed to a primary clinical isolate of HCMV designated ‘SBCMV’. Infectivity was analyzed by microscopy, immunofluorescence, Western blot, and qRT-PCR. Microarrays were performed to identify proinflammatory cytokines upregulated after SBCMV exposure, and the results validated by real-time quantitative polymerase chain reaction (qPCR) methodology. In situ cytokine expression of pericytes after exposure to HCMV was examined by ELISA and in vivo evidence of HCMV infection of brain pericytes was shown by dual-labeled immunohistochemistry. Results HCMV-infected human brain vascular pericytes as evidenced by several markers. Using a clinical isolate of HCMV (SBCMV), microscopy of infected pericytes showed virion production and typical cytomegalic cytopathology. This finding was confirmed by the expression of major immediate early and late virion proteins and by the presence of HCMV mRNA. Brain pericytes were fully permissive for CMV lytic replication after 72 to 96 hours in culture compared to human astrocytes or human brain microvascular endothelial cells (BMVEC). However, temporal transcriptional expression of pp65 virion protein after SBCMV infection was lower than that seen with the HCMV Towne laboratory strain. Using RT-PCR and dual-labeled immunofluorescence, proinflammatory cytokines CXCL8/IL-8, CXCL11/ITAC, and CCL5/Rantes were upregulated in SBCMV-infected cells, as were tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1beta), and interleukin-6 (IL-6). Pericytes exposed to SBCMV elicited higher levels of IL-6

  20. Human cytomegalovirus IE1 downregulates Hes1 in neural progenitor cells as a potential E3 ubiquitin ligase.

    Directory of Open Access Journals (Sweden)

    Xi-Juan Liu

    2017-07-01

    Full Text Available Congenital human cytomegalovirus (HCMV infection is the leading cause of neurological disabilities in children worldwide, but the mechanisms underlying these disorders are far from well-defined. HCMV infection has been shown to dysregulate the Notch signaling pathway in human neural progenitor cells (NPCs. As an important downstream effector of Notch signaling, the transcriptional regulator Hairy and Enhancer of Split 1 (Hes1 is essential for governing NPC fate and fetal brain development. In the present study, we report that HCMV infection downregulates Hes1 protein levels in infected NPCs. The HCMV 72-kDa immediate-early 1 protein (IE1 is involved in Hes1 degradation by assembling a ubiquitination complex and promoting Hes1 ubiquitination as a potential E3 ubiquitin ligase, followed by proteasomal degradation of Hes1. Sp100A, an important component of PML nuclear bodies, is identified to be another target of IE1-mediated ubiquitination. A C-terminal acidic region in IE1, spanning amino acids 451 to 475, is required for IE1/Hes1 physical interaction and IE1-mediated Hes1 ubiquitination, but is dispensable for IE1/Sp100A interaction and ubiquitination. Our study suggests a novel mechanism linking downregulation of Hes1 protein to neurodevelopmental disorders caused by HCMV infection. Our findings also complement the current knowledge of herpesviruses by identifying IE1 as the first potential HCMV-encoded E3 ubiquitin ligase.

  1. Human cytomegalovirus IE1 downregulates Hes1 in neural progenitor cells as a potential E3 ubiquitin ligase.

    Science.gov (United States)

    Liu, Xi-Juan; Yang, Bo; Huang, Sheng-Nan; Wu, Cong-Cong; Li, Xiao-Jun; Cheng, Shuang; Jiang, Xuan; Hu, Fei; Ming, Ying-Zi; Nevels, Michael; Britt, William J; Rayner, Simon; Tang, Qiyi; Zeng, Wen-Bo; Zhao, Fei; Luo, Min-Hua

    2017-07-01

    Congenital human cytomegalovirus (HCMV) infection is the leading cause of neurological disabilities in children worldwide, but the mechanisms underlying these disorders are far from well-defined. HCMV infection has been shown to dysregulate the Notch signaling pathway in human neural progenitor cells (NPCs). As an important downstream effector of Notch signaling, the transcriptional regulator Hairy and Enhancer of Split 1 (Hes1) is essential for governing NPC fate and fetal brain development. In the present study, we report that HCMV infection downregulates Hes1 protein levels in infected NPCs. The HCMV 72-kDa immediate-early 1 protein (IE1) is involved in Hes1 degradation by assembling a ubiquitination complex and promoting Hes1 ubiquitination as a potential E3 ubiquitin ligase, followed by proteasomal degradation of Hes1. Sp100A, an important component of PML nuclear bodies, is identified to be another target of IE1-mediated ubiquitination. A C-terminal acidic region in IE1, spanning amino acids 451 to 475, is required for IE1/Hes1 physical interaction and IE1-mediated Hes1 ubiquitination, but is dispensable for IE1/Sp100A interaction and ubiquitination. Our study suggests a novel mechanism linking downregulation of Hes1 protein to neurodevelopmental disorders caused by HCMV infection. Our findings also complement the current knowledge of herpesviruses by identifying IE1 as the first potential HCMV-encoded E3 ubiquitin ligase.

  2. Global Mapping of O-Glycosylation of Varicella Zoster Virus, Human Cytomegalovirus, and Epstein-Barr Virus

    DEFF Research Database (Denmark)

    Bagdonaite, Ieva; Nordén, Rickard; Joshi, Hiren J

    2016-01-01

    of the herpesvirus family: varicella zoster virus, human cytomegalovirus, and Epstein-Barr virus. We identified a large number of O-glycosites distributed on most envelope proteins in all viruses and further demonstrated conserved patterns of O-glycans on distinct homologous proteins. Because glycosylation is highly...

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

    NARCIS (Netherlands)

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

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

  4. Effect of compounds with antibacterial activities in human milk on respiratory syncytial virus and cytomegalovirus in vitro.

    Science.gov (United States)

    Portelli, J; Gordon, A; May, J T

    1998-11-01

    The effect of some antibacterial compounds present in human milk were tested for antiviral activity against respiratory syncytial virus, Semliki Forest virus and cytomegalovirus. These included the gangliosides GM1, GM2 and GM3, sialyl-lactose, lactoferrin and chondroitin sulphate A, B and C, which were all tested for their ability to inhibit the viruses in cell culture. Of the compounds tested, only the ganglioside GM2, chondroitin sulphate B and lactoferrin inhibited the absorption and growth of respiratory syncytial virus in cell culture, and none inhibited the growth of Semliki Forest virus, indicating that lipid antiviral activity was not associated with any of the gangliosides. While the concentrations of these two compounds required to inhibit respiratory syncytial virus were in excess of those present in human milk, sialyl-lactose concentrations similar to those present in human milk increased the growth of cytomegalovirus. Lactoferrin was confirmed as inhibiting both respiratory syncytial virus and cytomegalovirus growth in culture even when used at lower concentrations than those present in human milk. The antiviral activities of GM2, chondroitin sulphate B and lactoferrin were tested when added to an infant formula. Lactoferrin continued to have antiviral activity against cytomegalovirus, but a lower activity against respiratory syncytial virus; ganglioside GM2 and chondroitin sulphate B still maintained antiviral activity against respiratory syncytial virus.

  5. The human cytomegalovirus US28 protein is located in endocytic vesicles and undergoes constitutive endocytosis and recycling

    DEFF Research Database (Denmark)

    Fraile-Ramos, A; Kledal, T N; Pelchen-Matthews, A

    2001-01-01

    Genes encoding chemokine receptor-like proteins have been found in herpes and poxviruses and implicated in viral pathogenesis. Here we describe the cellular distribution and trafficking of a human cytomegalovirus (HCMV) chemokine receptor encoded by the US28 gene, after transient and stable...

  6. Relationship between TLR4 signalling alterations and effective human cytomegalovirus infection

    Directory of Open Access Journals (Sweden)

    Germini D.

    2014-09-01

    Full Text Available Toll-like receptors (TLR, the main class of immune-sensor molecules triggering the innate immunity pathways, are known to be involved in the infection of different RNA and DNA viruses, including herpesviruses. Human cytomegalovirus (HCMV is a widespread human beta-herpesvirus that infects 80–90 % of the world’s population and it can cause severe and even fatal diseases in immunocompromised patients and it is also responsible for birth defects as a consequence of congenital infection. Aim of this review is to discuss the existing data regarding the role of TLRs in HCMV concentrating mainly on TLR4. A better understanding in this relationship could be exploited for the development of efficient early diagnosis methodologies and anti viral therapies.

  7. Synergistic effects by combination of ganciclovir and tricin on human cytomegalovirus replication in vitro.

    Science.gov (United States)

    Yamada, Rie; Suda, Hideki; Sadanari, Hidetaka; Matsubara, Keiko; Tuchida, Yuuzo; Murayama, Tsugiya

    2016-01-01

    It has been demonstrated as the first report that combination treatment with ganciclovir (GCV) and tricin (4',5,7-trihydroxy-3',5' -dimethoxyflavone), a derivative of Sasa albo-marginata, after human cytomegalovirus (HCMV) infection has synergistic effects on both infectious virus production and HCMV DNA synthesis in the human embryonic fibroblast cell line MRC-5. In this paper, we examined the anti-HCMV effects of GCV plus various concentrations of tricin, and tricin plus various concentrations of GCV in MRC-5 cells. We found that expression of the HCMV UL54 gene was significantly inhibited by combination of GCV with tricin when compared with GCV mono-treatment. These results suggest that tricin is a novel compound for combination therapy with GCV against HCMV replication. In addition, reduced-dose combination therapy may provide a direction for treatment in patients with HCMV infection while reducing drug toxicity. Copyright © 2015 Elsevier B.V. All rights reserved.

  8. Modulation of human mesenchymal stem cell immunogenicity through forced expression of human cytomegalovirus us proteins.

    Directory of Open Access Journals (Sweden)

    Melisa A Soland

    Full Text Available BACKGROUND: Mesenchymal stem cells (MSC are promising candidates for cell therapy, as they migrate to areas of injury, differentiate into a broad range of specialized cells, and have immunomodulatory properties. However, MSC are not invisible to the recipient's immune system, and upon in vivo administration, allogeneic MSC are able to trigger immune responses, resulting in rejection of the transplanted cells, precluding their full therapeutic potential. Human cytomegalovirus (HCMV has developed several strategies to evade cytotoxic T lymphocyte (CTL and Natural Killer (NK cell recognition. Our goal is to exploit HCMV immunological evasion strategies to reduce MSC immunogenicity. METHODOLOGY/PRINCIPAL FINDINGS: We genetically engineered human MSC to express HCMV proteins known to downregulate HLA-I expression, and investigated whether modified MSC were protected from CTL and NK attack. Flow cytometric analysis showed that amongst the US proteins tested, US6 and US11 efficiently reduced MSC HLA-I expression, and mixed lymphocyte reaction demonstrated a corresponding decrease in human and sheep mononuclear cell proliferation. NK killing assays showed that the decrease in HLA-I expression did not result in increased NK cytotoxicity, and that at certain NK∶MSC ratios, US11 conferred protection from NK cytotoxic effects. Transplantation of MSC-US6 or MSC-US11 into pre-immune fetal sheep resulted in increased liver engraftment when compared to control MSC, as demonstrated by qPCR and immunofluorescence analyses. CONCLUSIONS AND SIGNIFICANCE: These data demonstrate that engineering MSC to express US6 and US11 can be used as a means of decreasing recognition of MSC by the immune system, allowing higher levels of engraftment in an allogeneic transplantation setting. Since one of the major factors responsible for the failure of allogeneic-donor MSC to engraft is the mismatch of HLA-I molecules between the donor and the recipient, MSC-US6 and MSC-US11

  9. Structural basis for the recognition of human cytomegalovirus glycoprotein B by a neutralizing human antibody.

    Directory of Open Access Journals (Sweden)

    Nadja Spindler

    2014-10-01

    Full Text Available Human cytomegalovirus (HCMV infections are life-threating to people with a compromised or immature immune system. Upon adhesion, fusion of the virus envelope with the host cell is initiated. In this step, the viral glycoprotein gB is considered to represent the major fusogen. Here, we present for the first time structural data on the binding of an anti-herpes virus antibody and describe the atomic interactions between the antigenic domain Dom-II of HCMV gB and the Fab fragment of the human antibody SM5-1. The crystal structure shows that SM5-1 binds Dom-II almost exclusively via only two CDRs, namely light chain CDR L1 and a 22-residue-long heavy chain CDR H3. Two contiguous segments of Dom-II are targeted by SM5-1, and the combining site includes a hydrophobic pocket on the Dom-II surface that is only partially filled by CDR H3 residues. SM5-1 belongs to a series of sequence-homologous anti-HCMV gB monoclonal antibodies that were isolated from the same donor at a single time point and that represent different maturation states. Analysis of amino acid substitutions in these antibodies in combination with molecular dynamics simulations show that key contributors to the picomolar affinity of SM5-1 do not directly interact with the antigen but significantly reduce the flexibility of CDR H3 in the bound and unbound state of SM5-1 through intramolecular side chain interactions. Thus, these residues most likely alleviate unfavorable binding entropies associated with extra-long CDR H3s, and this might represent a common strategy during antibody maturation. Models of entire HCMV gB in different conformational states hint that SM5-1 neutralizes HCMV either by blocking the pre- to postfusion transition of gB or by precluding the interaction with additional effectors such as the gH/gL complex.

  10. The DNA Damage Response Induced by Infection with Human Cytomegalovirus and Other Viruses

    Science.gov (United States)

    E, Xiaofei; Kowalik, Timothy F.

    2014-01-01

    Viruses use different strategies to overcome the host defense system. Recent studies have shown that viruses can induce DNA damage response (DDR). Many of these viruses use DDR signaling to benefit their replication, while other viruses block or inactivate DDR signaling. This review focuses on the effects of DDR and DNA repair on human cytomegalovirus (HCMV) replication. Here, we review the DDR induced by HCMV infection and its similarities and differences to DDR induced by other viruses. As DDR signaling pathways are critical for the replication of many viruses, blocking these pathways may represent novel therapeutic opportunities for the treatment of certain infectious diseases. Lastly, future perspectives in the field are discussed. PMID:24859341

  11. Subgingival Human Cytomegalovirus and Epstein Barr Virus in Patients with Aggressive Periodontitis in Ahwaz, Iran

    Directory of Open Access Journals (Sweden)

    M. Jahangirnezhad

    2009-06-01

    Full Text Available Objective: Assessment of subgingival human cytomegalovirus (HCMV and Epstein Barr virus (EBV-1 in subjects with aggressive periodontitis.Materials and Methods: Samples were obtained from plaques formed in subgingival regions of 26 aggressive periodontitis patients. All specimens were submitted to polymerase chain reaction in order to detect HCMV and EBV-1.Results: HCMV and EBV-1 were observed in 27% and 25% of the participants respectively.Coinfection with both viruses was found in 52% of the patients.Conclusion: Within the limitations of the present study, it can be suggested that HCMV and EBV-1 in subgingival plaques may be associated with aggressive periodontitis. Additionally,concomitant occurrence of these viruses may negatively affect the wellbeing ofperiodontal tissues.

  12. Seroprevalence for Human Cytomegalovirus in samples from dialysis and kidney transplanted patients

    Directory of Open Access Journals (Sweden)

    Patricia Okubo

    2014-12-01

    Full Text Available he aim of this study was at investigating the seroprevalence of Human Cytomegalovirus in dialysis and kidney transplanted patients and the association with hemodialysis and sex. It is a cross sectional study made in 2011 and 2012 in the county of Maringá, PA, Brazil. The studied population consisted of 203 dialysis patients and 53 kidney transplanted patients and the viral identification was made through serological technique. The study found 96% (195 seropositivity for anti-HCMV IgG in dialysis patients and 100% in the transplanted ones; 5% (10 seropositivity for anti-HCMV-IgM in dialysis patients and 37.7% (20 in transplanted ones. Performing Fisher’s exact test, there was no significant association between the seroprevalence of anti-HCMV with hemodialysis and gender. A large number of infected patients suggest the importance of nursing care to avoid cross-infection during routine procedures.

  13. Interplay between Human Cytomegalovirus and Intrinsic/Innate Host Responses: A Complex Bidirectional Relationship

    Science.gov (United States)

    Rossini, Giada; Cerboni, Cristina; Santoni, Angela; Landini, Maria Paola; Landolfo, Santo; Gatti, Deborah; Gribaudo, Giorgio; Varani, Stefania

    2012-01-01

    The interaction between human cytomegalovirus (HCMV) and its host is a complex process that begins with viral attachment and entry into host cells, culminating in the development of a specific adaptive response that clears the acute infection but fails to eradicate HCMV. We review the viral and cellular partners that mediate early host responses to HCMV with regard to the interaction between structural components of virions (viral glycoproteins) and cellular receptors (attachment/entry receptors, toll-like receptors, and other nucleic acid sensors) or intrinsic factors (PML, hDaxx, Sp100, viperin, interferon inducible protein 16), the reactions of innate immune cells (antigen presenting cells and natural killer cells), the numerous mechanisms of viral immunoevasion, and the potential exploitation of events that are associated with early phases of virus-host interplay as a therapeutic strategy. PMID:22701276

  14. Pharmacokinetics and safety of the anti-human cytomegalovirus drug letermovir in subjects with hepatic impairment.

    Science.gov (United States)

    Kropeit, Dirk; McCormick, David; Erb-Zohar, Katharina; Moiseev, Valentin S; Kobalava, Zhanna D; Stobernack, Hans-Peter; Zimmermann, Holger; Rübsamen-Schaeff, Helga

    2017-07-18

    Human cytomegalovirus constitutes a prevalent and serious threat to immunocompromised individuals and requires new treatments. Letermovir is a novel viral-terminase inhibitor that has demonstrated prophylactic/pre-emptive activity against human cytomegalovirus in Phase 2 and 3 transplant trials. As unchanged letermovir is primarily excreted via the liver by bile, this trial aimed to assess the effect of hepatic impairment on letermovir pharmacokinetics. Phase 1, open-label, parallel-group pharmacokinetic and safety comparison of multiple once-daily oral letermovir in female subjects with hepatic impairment and healthy matched controls. For 8 days, subjects with moderate hepatic impairment (n = 8) and their matched healthy controls (n = 9) received 60 mg letermovir/day and those with severe hepatic impairment (n = 8) and their matched healthy controls (n = 8) received 30 mg letermovir/day. Pharmacokinetic parameters were determined from blood samples. For subjects with moderate hepatic impairment, maximal observed concentration at steady state (Css,max ) and the area under the concentration vs. time curve over a dosing interval at steady state (AUCτ,ss ) for total letermovir were 1.37-fold (90% confidence interval: 0.87, 2.17) and 1.59-fold (0.98, 2.57) higher, respectively, than in healthy subjects. For subjects with severe hepatic impairment, Css,max and AUCτ,ss values of total letermovir were 2.34-fold (1.91, 2.88) and 3.82-fold (2.94, 4.97) higher, respectively, compared with healthy subjects. Moderate hepatic impairment increased exposure to letermovir letermovir exposure approximately 4-fold as compared with healthy subjects. Letermovir 60/30 mg/day was generally well-tolerated in subjects with hepatic impairment. © 2017 The British Pharmacological Society.

  15. Cytomegalovirus survival and transferability and the effectiveness of common hand-washing agents against cytomegalovirus on live human hands.

    Science.gov (United States)

    Stowell, Jennifer D; Forlin-Passoni, Daniela; Radford, Kay; Bate, Sheri L; Dollard, Sheila C; Bialek, Stephanie R; Cannon, Michael J; Schmid, D Scott

    2014-01-01

    Congenital cytomegalovirus (CMV) transmission can occur when women acquire CMV while pregnant. Infection control guidelines may reduce risk for transmission. We studied the duration of CMV survival after application of bacteria to the hands and after transfer from the hands to surfaces and the effectiveness of cleansing with water, regular and antibacterial soaps, sanitizer, and diaper wipes. Experiments used CMV AD169 in saliva at initial titers of 1 × 10(5) infectious particles/ml. Samples from hands or surfaces (points between 0 and 15 min) were placed in culture and observed for at least 2 weeks. Samples were also tested using CMV real-time PCR. After application of bacteria to the hands, viable CMV was recovered from 17/20 swabs at 0 min, 18/20 swabs at 1 min, 5/20 swabs at 5 min, and 4/20 swabs at 15 min. After transfer, duration of survival was at least 15 min on plastic (1/2 swabs), 5 min on crackers and glass (3/4 swabs), and 1 min or less on metal and cloth (3/4 swabs); no viable virus was collected from wood, rubber, or hands. After cleansing, no viable virus was recovered using water (0/22), plain soap (0/20), antibacterial soap (0/20), or sanitizer (0/22). Viable CMV was recovered from 4/20 hands 10 min after diaper wipe cleansing. CMV remains viable on hands for sufficient times to allow transmission. CMV may be transferred to surfaces with reduced viability. Hand-cleansing methods were effective at eliminating viable CMV from hands.

  16. Convallatoxin-Induced Reduction of Methionine Import Effectively Inhibits Human Cytomegalovirus Infection and Replication.

    Science.gov (United States)

    Cohen, Tobias; Williams, John D; Opperman, Timothy J; Sanchez, Roberto; Lurain, Nell S; Tortorella, Domenico

    2016-12-01

    Cytomegalovirus (CMV) is a ubiquitous human pathogen that increases the morbidity and mortality of immunocompromised individuals. The current FDA-approved treatments for CMV infection are intended to be virus specific, yet they have significant adverse side effects, including nephrotoxicity and hematological toxicity. Thus, there is a medical need for safer and more effective CMV therapeutics. Using a high-content screen, we identified the cardiac glycoside convallatoxin as an effective compound that inhibits CMV infection. Using a panel of cardiac glycoside variants, we assessed the structural elements critical for anti-CMV activity by both experimental and in silico methods. Analysis of the antiviral effects, toxicities, and pharmacodynamics of different variants of cardiac glycosides identified the mechanism of inhibition as reduction of methionine import, leading to decreased immediate-early gene translation without significant toxicity. Also, convallatoxin was found to dramatically reduce the proliferation of clinical CMV strains, implying that its mechanism of action is an effective strategy to block CMV dissemination. Our study has uncovered the mechanism and structural elements of convallatoxin, which are important for effectively inhibiting CMV infection by targeting the expression of immediate-early genes. Cytomegalovirus is a highly prevalent virus capable of causing severe disease in certain populations. The current FDA-approved therapeutics all target the same stage of the viral life cycle and induce toxicity and viral resistance. We identified convallatoxin, a novel cell-targeting antiviral that inhibits CMV infection by decreasing the synthesis of viral proteins. At doses low enough for cells to tolerate, convallatoxin was able to inhibit primary isolates of CMV, including those resistant to the anti-CMV drug ganciclovir. In addition to identifying convallatoxin as a novel antiviral, limiting mRNA translation has a dramatic impact on CMV infection

  17. Activation of nucleotide oligomerization domain 2 (NOD2 by human cytomegalovirus initiates innate immune responses and restricts virus replication.

    Directory of Open Access Journals (Sweden)

    Arun Kapoor

    Full Text Available Nucleotide-binding oligomerization domain 2 (NOD2 is an important innate immune sensor of bacterial pathogens. Its induction results in activation of the classic NF-κB pathway and alternative pathways including type I IFN and autophagy. Although the importance of NOD2 in recognizing RNA viruses has recently been identified, its role in sensing DNA viruses has not been studied. We report that infection with human cytomegalovirus (HCMV results in significant induction of NOD2 expression, beginning as early as 2 hours post infection and increasing steadily 24 hours post infection and afterwards. Infection with human herpesvirus 1 and 2 does not induce NOD2 expression. While the HCMV-encoded glycoprotein B is not required for NOD2 induction, a replication competent virion is necessary. Lentivirus-based NOD2 knockdown in human foreskin fibroblasts (HFFs and U373 glioma cells leads to enhanced HCMV replication along with decreased levels of interferon beta (IFN-β and the pro-inflammatory cytokine, IL8. NOD2 induction in HCMV-infected cells activates downstream NF-κB and interferon pathways supported by reduced nuclear localization of NF-κB and pIRF3 in NOD2 knockdown HFFs. Stable overexpression of NOD2 in HFFs restricts HCMV replication in association with increased levels of IFN-β and IL8. Similarly, transient overexpression of NOD2 in U373 cells or its downstream kinase, RIPK2, results in decreased HCMV replication and enhanced cytokine responses. However, overexpression of a mutant NOD2, 3020insC, associated with severe Crohn's disease, results in enhanced HCMV replication and decreased levels of IFN-β in U373 cells. These results show for the first time that NOD2 plays a significant role in HCMV replication and may provide a model for studies of HCMV recognition by the host cell and HCMV colitis in Crohn's disease.

  18. Permissive human cytomegalovirus infection of a first trimester extravillous cytotrophoblast cell line

    Directory of Open Access Journals (Sweden)

    LaMarca Heather L

    2004-11-01

    Full Text Available Abstract Human cytomegalovirus (HCMV is the leading cause of congenital viral infection in the United States and Europe. Despite the significant morbidity associated with prenatal HCMV infection, little is known about how the virus infects the fetus during pregnancy. To date, primary human cytotrophoblasts (CTBs have been utilized to study placental HCMV infection and replication; however, the minimal mitotic potential of these cells restricts experimentation to a few days, which may be problematic for mechanistic studies of the slow-replicating virus. The aim of this study was to determine whether the human first trimester CTB cell line SGHPL-4 was permissive for HCMV infection and therefore could overcome such limitations. HCMV immediate early (IE protein expression was detected as early as 3 hours post-infection in SGHPL-4 cells and progressively increased as a function of time. HCMV growth assays revealed the presence of infectious virus in both cell lysates and culture supernatants, indicating that viral replication and the release of progeny virus occurred. Compared to human fibroblasts, viral replication was delayed in CTBs, consistent with previous studies reporting delayed viral kinetics in HCMV-infected primary CTBs. These results indicate that SGHPL-4 cells are fully permissive for the complete HCMV replicative cycle. Our findings suggest that these cells may serve as useful tools for future mechanistic studies of HCMV pathogenesis during early pregnancy.

  19. Human Cytomegalovirus Particles Treated with Specific Antibodies Induce Intrinsic and Adaptive but Not Innate Immune Responses.

    Science.gov (United States)

    Wu, Zeguang; Qin, Ruifang; Wang, Li; Bosso, Matteo; Scherer, Myriam; Stamminger, Thomas; Hotter, Dominik; Mertens, Thomas; Frascaroli, Giada

    2017-11-15

    Human cytomegalovirus (HCMV) persistently infects 40% to 100% of the human population worldwide. Experimental and clinical evidence indicates that humoral immunity to HCMV plays an important role in restricting virus dissemination and protecting the infected host from disease. Specific immunoglobulin preparations from pooled plasma of adults selected for high titers of HCMV antibodies have been used for the prevention of CMV disease in transplant recipients and pregnant women. Even though incubation of HCMV particles with these preparations leads to the neutralization of viral infectivity, it is still unclear whether the antibody-treated HCMV particles (referred to here as HCMV-Ab) enter the cells and modulate antiviral immune responses. Here we demonstrate that HCMV-Ab did enter macrophages. HCMV-Ab did not initiate the expression of immediate early antigens (IEAs) in macrophages, but they induced an antiviral state and rendered the cells less susceptible to HCMV infection upon challenge. Resistance to HCMV infection seemed to be due to the activation of intrinsic restriction factors and was independent of interferons. In contrast to actively infected cells, autologous NK cells did not degranulate against HCMV-Ab-treated macrophages, suggesting that these cells may not be eliminated by innate effector cells. Interestingly, HCMV-Ab-treated macrophages stimulated the proliferation of autologous adaptive CD4+ and CD8+ T cells. Our findings not only expand the current knowledge on virus-antibody immunity but may also be relevant for future vaccination strategies.IMPORTANCE Human cytomegalovirus (HCMV), a common herpesvirus, establishes benign but persistent infections in immunocompetent hosts. However, in subjects with an immature or dysfunctional immune system, HCMV is a major cause of morbidity and mortality. Passive immunization has been used in different clinical settings with variable clinical results. Intravenous hyperimmune globulin preparations (IVIg) are

  20. Susceptibilities of Human Cytomegalovirus Clinical Isolates to BAY38-4766, BAY43-9695, and Ganciclovir

    Science.gov (United States)

    McSharry, James J.; McDonough, Ann; Olson, Betty; Hallenberger, Sabine; Reefschlaeger, Juergen; Bender, Wolfgang; Drusano, George L.

    2001-01-01

    BAY38-4766 and BAY43-9695 are nonnucleosidic compounds with activities against human cytomegalovirus (HCMV). Two phenotypic assays were used to determine the drug susceptibilities of 36 HCMV clinical isolates to the BAY compounds and ganciclovir. Using either assay, both BAY compounds at a concentration of approximately 1 μM inhibited the replication of all 36 HCMV clinical isolates, including 11 ganciclovir-resistant clinical isolates, by 50%. PMID:11557492

  1. Seroprevalence of Hepatitis C, Hepatitis B, Cytomegalovirus, and Human Immunodeficiency Viruses in Multitransfused Thalassemic Children in Upper Egypt

    OpenAIRE

    Mahmoud, Ramadan A.; El-Mazary, Abdel-Azeem M.; Khodeary, Ashraf

    2016-01-01

    Background. Frequent blood transfusions in thalassemia major children expose them to the risk of transfusion-transmitted infections (TTIs). The aim of this study was to estimate the prevalence of hepatitis C virus (HCV), hepatitis B virus (HBV), human immunodeficiency virus (HIV), and cytomegalovirus (CMV) in thalassemic children attending the Pediatrics Departments of both Sohag and Minia Universities of Upper Egypt, during the period from May 2014 to May 2015. Methods. Serum samples were sc...

  2. Adenovirus E1A/E1B Transformed Amniotic Fluid Cells Support Human Cytomegalovirus Replication

    Directory of Open Access Journals (Sweden)

    Natascha Krömmelbein

    2016-02-01

    Full Text Available The human cytomegalovirus (HCMV replicates to high titers in primary human fibroblast cell cultures. A variety of primary human cells and some tumor-derived cell lines do also support permissive HCMV replication, yet at low levels. Cell lines established by transfection of the transforming functions of adenoviruses have been notoriously resistant to HCMV replication and progeny production. Here, we provide first-time evidence that a permanent cell line immortalized by adenovirus type 5 E1A and E1B (CAP is supporting the full HCMV replication cycle and is releasing infectious progeny. The CAP cell line had previously been established from amniotic fluid cells which were likely derived from membranes of the developing fetus. These cells can be grown under serum-free conditions. HCMV efficiently penetrated CAP cells, expressed its immediate-early proteins and dispersed restrictive PML-bodies. Viral DNA replication was initiated and viral progeny became detectable by electron microscopy in CAP cells. Furthermore, infectious virus was released from CAP cells, yet to lower levels compared to fibroblasts. Subviral dense bodies were also secreted from CAP cells. The results show that E1A/E1B expression in transformed cells is not generally repressive to HCMV replication and that CAP cells may be a good substrate for dense body based vaccine production.

  3. Genomic analysis of chimeric human cytomegalovirus vaccine candidates derived from strains Towne and Toledo.

    Science.gov (United States)

    Suárez, Nicolás M; Lau, Betty; Kemble, George M; Lee, Ronzo; Mocarski, Edward S; Wilkinson, Gavin W G; Adler, Stuart P; McVoy, Michael A; Davison, Andrew J

    2017-08-01

    Human cytomegalovirus (HCMV) is an important opportunistic pathogen in immunocompromised patients and a major cause of congenital birth defects when acquired in utero. In the 1990s, four chimeric viruses were constructed by replacing genome segments of the high passage Towne strain with segments of the low passage Toledo strain, with the goal of obtaining live attenuated vaccine candidates that remained safe but were more immunogenic than the overly attenuated Towne vaccine. The chimeras were found to be safe when administered to HCMV-seronegative human volunteers, but to differ significantly in their ability to induce seroconversion. This suggests that chimera-specific genetic differences impacted the ability to replicate or persist in vivo and the consequent ability to induce an antibody response. To identify specific genomic breakpoints between Towne and Toledo sequences and establish whether spontaneous mutations or rearrangements had occurred during construction of the chimeras, complete genome sequences were determined. No major deletions or rearrangements were observed, although a number of unanticipated mutations were identified. However, no clear association emerged between the genetic content of the chimeras and the reported levels of vaccine-induced HCMV-specific humoral or cellular immune responses, suggesting that multiple genetic determinants are likely to impact immunogenicity. In addition to revealing the genome organization of the four vaccine candidates, this study provided an opportunity to probe the genetics of HCMV attenuation in humans. The results may be valuable in the future design of safe live or replication-defective vaccines that optimize immunogenicity and efficacy.

  4. Human Cytomegalovirus US28 Is Important for Latent Infection of Hematopoietic Progenitor Cells

    Science.gov (United States)

    Humby, Monica S.

    2015-01-01

    ABSTRACT Human cytomegalovirus (HCMV) resides latently in hematopoietic progenitor cells (HPCs). During latency, only a subset of HCMV genes is transcribed, including one of the four virus-encoded G protein-coupled receptors (GPCRs), US28. Although US28 is a multifunctional lytic protein, its function during latency has remained undefined. We generated a panel of US28 recombinant viruses in the bacterial artificial chromosome (BAC)-derived clinical HCMV strain TB40/E-mCherry. We deleted the entire US28 open reading frame (ORF), deleted all four of the viral GPCR ORFs, or deleted three of the HCMV GPCRs but not the US28 wild-type protein. Using these recombinant viruses, we assessed the requirement for US28 during latency in the Kasumi-3 in vitro latency model system and in primary ex vivo-cultured CD34+ HPCs. Our data suggest that US28 is required for latency as infection with viruses lacking the US28 ORF alone or in combination with the remaining HCMV-encoded GPCR results in transcription from the major immediate early promoter, the production of extracellular virions, and the production of infectious virus capable of infecting naive fibroblasts. The other HCMV GPCRs are not required for this phenotype as a virus expressing only US28 but not the remaining virus-encoded GPCRs is phenotypically similar to that of wild-type latent infection. Finally, we found that US28 copurifies with mature virions and is expressed in HPCs upon virus entry although its expression at the time of infection does not complement the US28 deletion latency phenotype. This work suggests that US28 protein functions to promote a latent state within hematopoietic progenitor cells. IMPORTANCE Human cytomegalovirus (HCMV) is a widespread pathogen that, once acquired, remains with its host for life. HCMV remains latent, or quiescent, in cells of the hematopoietic compartment and upon immune challenge can reactivate to cause disease. HCMV-encoded US28 is one of several genes expressed during

  5. Cell Surface THY-1 Contributes to Human Cytomegalovirus Entry via a Macropinocytosis-Like Process.

    Science.gov (United States)

    Li, Qingxue; Fischer, Elizabeth; Cohen, Jeffrey I

    2016-11-01

    Previously we showed that THY-1 has a critical role in the initial stage of infection of certain cell types with human cytomegalovirus (HCMV) and that THY-1 is important for HCMV-mediated activation of phosphatidylinositol 3-kinase (PI3K)/Akt during virus entry. THY-1 is known to interact with integrins and is a major cargo protein of clathrin-independent endocytic vesicles. Since macropinocytosis involves integrin signaling, is PI3K/Akt dependent, and is a clathrin-independent endocytic process, we determined whether THY-1 has a role in HCMV entry by macropinocytosis. Using electron microscopy in two cell lines that support HCMV infection in a THY-1-dependent manner, we found that HCMV enters these cells by a macropinocytosis-like process. THY-1 associated with HCMV virions on the cell surface and colocalized with virus inside macropinosomes. 5-(N-Ethyl-N-isopropyl)amiloride (EIPA) and soluble THY-1 blocked HCMV infection in the cell lines by ≥80% and 60%, respectively. HCMV entry into the cells triggered increased influx of extracellular fluid, a marker of macropinocytosis, and this increased fluid uptake was inhibited by EIPA and by soluble THY-1. Blocking actin depolymerization, Na + /H + exchange, PI3K, and Pak1 kinase, which are critical for macropinocytosis, impaired HCMV infection. Neither internalized HCMV virions nor THY-1 in virus-infected cells colocalized with transferrin as determined by confocal microscopy, indicating that clathrin-mediated endocytosis was not involved in THY-1-associated virus entry. These results suggest that HCMV has adapted to utilize THY-1, a cargo protein of clathrin-independent endocytotic vesicles, to facilitate efficient entry into certain cell types by a macropinocytosis-like process. Human cytomegalovirus (HCMV) infects over half of the population and is the most common infectious cause of birth defects. The virus is the most important infection occurring in transplant recipients. The mechanism of how HCMV enters cells

  6. Extensive genome-wide variability of human cytomegalovirus in congenitally infected infants.

    Directory of Open Access Journals (Sweden)

    Nicholas Renzette

    2011-05-01

    Full Text Available Research has shown that RNA virus populations are highly variable, most likely due to low fidelity replication of RNA genomes. It is generally assumed that populations of DNA viruses will be less complex and show reduced variability when compared to RNA viruses. Here, we describe the use of high throughput sequencing for a genome wide study of viral populations from urine samples of neonates with congenital human cytomegalovirus (HCMV infections. We show that HCMV intrahost genomic variability, both at the nucleotide and amino acid level, is comparable to many RNA viruses, including HIV. Within intrahost populations, we find evidence of selective sweeps that may have resulted from immune-mediated mechanisms. Similarly, genome wide, population genetic analyses suggest that positive selection has contributed to the divergence of the HCMV species from its most recent ancestor. These data provide evidence that HCMV, a virus with a large dsDNA genome, exists as a complex mixture of genome types in humans and offer insights into the evolution of the virus.

  7. Transcriptome-wide characterization of human cytomegalovirus in natural infection and experimental latency.

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    Cheng, Shu; Caviness, Katie; Buehler, Jason; Smithey, Megan; Nikolich-Žugich, Janko; Goodrum, Felicia

    2017-11-20

    The transcriptional program associated with herpesvirus latency and the viral genes regulating entry into and exit from latency are poorly understood and controversial. Here, we developed and validated a targeted enrichment platform and conducted large-scale transcriptome analyses of human cytomegalovirus (HCMV) infection. We used both an experimental hematopoietic cell model of latency and cells from naturally infected, healthy human subjects (clinical) to define the breadth of viral genes expressed. The viral transcriptome derived from experimental infection was highly correlated with that from clinical infection, validating our experimental latency model. These transcriptomes revealed a broader profile of gene expression during infection in hematopoietic cells than previously appreciated. Further, using recombinant viruses that establish a nonreactivating, latent-like or a replicative infection in CD34+ hematopoietic progenitor cells, we defined classes of low to moderately expressed genes that are differentially regulated in latent vs. replicative states of infection. Most of these genes have yet to be studied in depth. By contrast, genes that were highly expressed, were expressed similarly in both latent and replicative infection. From these findings, a model emerges whereby low or moderately expressed genes may have the greatest impact on regulating the switch between viral latency and replication. The core set of viral genes expressed in natural infection and differentially regulated depending on the pattern of infection provides insight into the HCMV transcriptome associated with latency in the host and a resource for investigating virus-host interactions underlying persistence.

  8. Molecular detection of cytomegalovirus, herpes simplex virus 2, human papillomavirus 16-18 in Turkish pregnants.

    Science.gov (United States)

    Dinc, Bedia; Bozdayi, Gulendam; Biri, Aydan; Kalkanci, Ayse; Dogan, Bora; Bozkurt, Nuray; Rota, Seyyal

    2010-01-01

    Human cytomegalovirus (CMV) is the most common cause of viral intrauterine infections in the world. Herpes simplex virus type 2 (HSV-2) and human papillomavirus (HPV) are the main agents of viral sexually transmitted diseases, which cause genital ulcers and genital warts, respectively. HPV infection has been linked to the majority of the anogenital malignancies. The aim of this study was to detect the existence of CMV, HSV-2 and HPV type 16-18 in Turkish pregnants by using sensitive molecular assays. One hundred thirty-four women (18-41 years old; mean age ± SD: 27 ± 8) applied to outpatient clinic of Obstetrics and Gynecology, in between 18th - 22nd weeks of their pregnancy and a control group of 99 healthy women (15-39 years old; mean age ± SD: 24 ± 8) were included in the study. Cervical smear samples were used for DNA extraction. CMV, HSV-2 and HPV 16-18 detections were carried out by real time PCR and in house PCR method, respectively. Three patients (3/134; 2.2%) were found to be positive for each HPV and HSV-2. Dual infection with HPV and HSV was found in just one patient. HPV 18 was detected in all positive samples. CMV was found to be positive in two patients (2/134; 1.4 %). HPV, HSV and CMV must be screened due to high prevalence of these viruses in pregnants by using sensitive molecular methods.

  9. Molecular detection of cytomegalovirus, herpes simplex virus 2, human papillomavirus 16-18 in Turkish pregnants

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    Bedia Dinc

    Full Text Available OBJECTIVE: Human cytomegalovirus (CMV is the most common cause of viral intrauterine infections in the world. Herpes simplex virus type 2 (HSV-2 and human papillomavirus (HPV are the main agents of viral sexually transmitted diseases, which cause genital ulcers and genital warts, respectively. HPV infection has been linked to the majority of the anogenital malignancies. The aim of this study was to detect the existence of CMV, HSV-2 and HPV type 16-18 in Turkish pregnants by using sensitive molecular assays. METHODS: One hundred thirty-four women (18-41 years old; mean age ± SD: 27 ± 8 applied to outpatient clinic of Obstetrics and Gynecology, in between 18th - 22nd weeks of their pregnancy and a control group of 99 healthy women (15-39 years old; mean age ± SD: 24 ± 8 were included in the study. Cervical smear samples were used for DNA extraction. CMV, HSV-2 and HPV 16-18 detections were carried out by real time PCR and in house PCR method, respectively. RESULTS: Three patients (3/134; 2.2% were found to be positive for each HPV and HSV-2. Dual infection with HPV and HSV was found in just one patient. HPV 18 was detected in all positive samples. CMV was found to be positive in two patients (2/134; 1.4 %. CONCLUSION: HPV, HSV and CMV must be screened due to high prevalence of these viruses in pregnants by using sensitive molecular methods.

  10. Release of human cytomegalovirus from latency by a KAP1/TRIM28 phosphorylation switch.

    Science.gov (United States)

    Rauwel, Benjamin; Jang, Suk Min; Cassano, Marco; Kapopoulou, Adamandia; Barde, Isabelle; Trono, Didier

    2015-04-07

    Human cytomegalovirus (HCMV) is a highly prevalent pathogen that induces life-long infections notably through the establishment of latency in hematopoietic stem cells (HSC). Bouts of reactivation are normally controlled by the immune system, but can be fatal in immuno-compromised individuals such as organ transplant recipients. Here, we reveal that HCMV latency in human CD34(+) HSC reflects the recruitment on the viral genome of KAP1, a master co-repressor, together with HP1 and the SETDB1 histone methyltransferase, which results in transcriptional silencing. During lytic infection, KAP1 is still associated with the viral genome, but its heterochromatin-inducing activity is suppressed by mTOR-mediated phosphorylation. Correspondingly, HCMV can be forced out of latency by KAP1 knockdown or pharmacological induction of KAP1 phosphorylation, and this process can be potentiated by activating NFkB with TNF-α. These results suggest new approaches both to curtail CMV infection and to purge the virus from organ transplants.

  11. Germline V-genes sculpt the binding site of a family of antibodies neutralizing human cytomegalovirus

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    Thomson, Christy A.; Bryson, Steve; McLean, Gary R.; Creagh, A. Louise; Pai, Emil F.; Schrader, John W. (Toronto); (UBC)

    2008-10-17

    Immunoglobulin genes are generated somatically through specialized mechanisms resulting in a vast repertoire of antigen-binding sites. Despite the stochastic nature of these processes, the V-genes that encode most of the antigen-combining site are under positive evolutionary selection, raising the possibility that V-genes have been selected to encode key structural features of binding sites of protective antibodies against certain pathogens. Human, neutralizing antibodies to human cytomegalovirus that bind the AD-2S1 epitope on its gB envelope protein repeatedly use a pair of well-conserved, germline V-genes IGHV3-30 and IGKV3-11. Here, we present crystallographic, kinetic and thermodynamic analyses of the binding site of such an antibody and that of its primary immunoglobulin ancestor. These show that these germline V-genes encode key side chain contacts with the viral antigen and thereby dictate key structural features of the hypermutated, high-affinity neutralizing antibody. V-genes may thus encode an innate, protective immunological memory that targets vulnerable, invariant sites on multiple pathogens.

  12. Human Cytomegalovirus Encoded Homologs of Cytokines, Chemokines and their Receptors: Roles in Immunomodulation

    Science.gov (United States)

    McSharry, Brian P.; Avdic, Selmir; Slobedman, Barry

    2012-01-01

    Human cytomegalovirus (HCMV), the largest human herpesvirus, infects a majority of the world’s population. Like all herpesviruses, following primary productive infection, HCMV establishes a life-long latent infection, from which it can reactivate years later to produce new, infectious virus. Despite the presence of a massive and sustained anti-HCMV immune response, productively infected individuals can shed virus for extended periods of time, and once latent infection is established, it is never cleared from the host. It has been proposed that HCMV must therefore encode functions which help to evade immune mediated clearance during productive virus replication and latency. Molecular mimicry is a strategy used by many viruses to subvert and regulate anti-viral immunity and HCMV has hijacked/developed a range of functions that imitate host encoded immunomodulatory proteins. This review will focus on the HCMV encoded homologs of cellular cytokines/chemokines and their receptors, with an emphasis on how these virus encoded homologs may facilitate viral evasion of immune clearance. PMID:23202490

  13. Two novel human cytomegalovirus NK cell evasion functions target MICA for lysosomal degradation.

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    Ceri A Fielding

    2014-05-01

    Full Text Available NKG2D plays a major role in controlling immune responses through the regulation of natural killer (NK cells, αβ and γδ T-cell function. This activating receptor recognizes eight distinct ligands (the MHC Class I polypeptide-related sequences (MIC A andB, and UL16-binding proteins (ULBP1-6 induced by cellular stress to promote recognition cells perturbed by malignant transformation or microbial infection. Studies into human cytomegalovirus (HCMV have aided both the identification and characterization of NKG2D ligands (NKG2DLs. HCMV immediate early (IE gene up regulates NKGDLs, and we now describe the differential activation of ULBP2 and MICA/B by IE1 and IE2 respectively. Despite activation by IE functions, HCMV effectively suppressed cell surface expression of NKGDLs through both the early and late phases of infection. The immune evasion functions UL16, UL142, and microRNA(miR-UL112 are known to target NKG2DLs. While infection with a UL16 deletion mutant caused the expected increase in MICB and ULBP2 cell surface expression, deletion of UL142 did not have a similar impact on its target, MICA. We therefore performed a systematic screen of the viral genome to search of addition functions that targeted MICA. US18 and US20 were identified as novel NK cell evasion functions capable of acting independently to promote MICA degradation by lysosomal degradation. The most dramatic effect on MICA expression was achieved when US18 and US20 acted in concert. US18 and US20 are the first members of the US12 gene family to have been assigned a function. The US12 family has 10 members encoded sequentially through US12-US21; a genetic arrangement, which is suggestive of an 'accordion' expansion of an ancestral gene in response to a selective pressure. This expansion must have be an ancient event as the whole family is conserved across simian cytomegaloviruses from old world monkeys. The evolutionary benefit bestowed by the combinatorial effect of US18 and US

  14. Salivary glands and human congenital cytomegalovirus infection: What happens in early fetal life?

    Science.gov (United States)

    Gabrielli, Liliana; Bonasoni, Maria Paola; Chiereghin, Angela; Piccirilli, Giulia; Santini, Donatella; Pavia, Claudia; Turello, Gabriele; Squarzoni, Diego; Lazzarotto, Tiziana

    2017-02-01

    Salivary glands are a site of human cytomegalovirus (CMV) replication, latency, and persistence. Prolonged secretion of virus in saliva for months following a primary infection contribute to horizontal transmission. In order to better understand the early effects of CMV on salivary glands and the mechanisms of viral persistent replication, submandibular glands of six CMV congenitally infected fetuses at 21 weeks gestation were studied. Three fetuses at the same gestational age from CMV-seronegative women were compared as negative controls. Tissue viral load and the type of inflammatory infiltrate were evaluated. Moreover, development and branching of salivary glands, the number of myoepithelial cells, cellular proliferation, and expression of secretory proteins of the saliva (Gross Cystic Disease Fluid Protein-15 and lysozyme) were studied. A low viral load and rare CMV-positive cells associated with T CD8 cytotoxic lymphocytes were observed. Branching was impaired with a decrease in terminal acinar structures, the number of myoepithelial cells, and cellular proliferation were reduced. In addition, a compromised secretion of defense proteins involved in the oral humoral immunity was observed. These findings suggest that CMV may affect salivary glands, impairing structure development and secretion of defense proteins, probably responsible for the prolonged viral shedding in saliva. J. Med. Virol. 89:318-323, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  15. The Microtubule Inhibitor Podofilox Inhibits an Early Entry Step of Human Cytomegalovirus

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    Tobias Cohen

    2016-10-01

    Full Text Available Human cytomegalovirus is a ubiquitous β-herpesvirus that infects many different cell types through an initial binding to cell surface receptors followed by a fusion event at the cell membrane or endocytic vesicle. A recent high-throughput screen to identify compounds that block a step prior to viral gene expression identified podofilox as a potent and nontoxic inhibitor. Time-of-addition studies in combination with quantitative-PCR analysis demonstrated that podofilox limits an early step of virus entry at the cell surface. Podofilox was also able to drastically reduce infection by herpes simplex 1, an α-herpesvirus with a very similar entry process to CMV. Podofilox caused a reduced maximal plateau inhibition of infection by viruses with single step binding processes prior to fusion-like Newcastle disease virus, Sendai virus, and influenza A virus or viruses that enter via endocytosis like vesicular stomatitis virus and a clinical-like strain of CMV. These results indicate that microtubules appear to be participating in the post-binding step of virus entry including the pre- and post-penetration events. Modulation of the plasma membrane is required to promote virus entry for herpesviruses, and that podofilox, unlike colchicine or nocodazole, is able to preferentially target microtubule networks at the plasma membrane.

  16. Superresolution Imaging of Human Cytomegalovirus vMIA Localization in Sub-Mitochondrial Compartments

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    Bhuvanendran, Shivaprasad; Salka, Kyle; Rainey, Kristin; Sreetama, Sen Chandra; Williams, Elizabeth; Leeker, Margretha; Prasad, Vidhya; Boyd, Jonathan; Patterson, George H.; Jaiswal, Jyoti K.; Colberg-Poley, Anamaris M.

    2014-01-01

    The human cytomegalovirus (HCMV) viral mitochondria-localized inhibitor of apoptosis (vMIA) protein, traffics to mitochondria-associated membranes (MAM), where the endoplasmic reticulum (ER) contacts the outer mitochondrial membrane (OMM). vMIA association with the MAM has not been visualized by imaging. Here, we have visualized this by using a combination of confocal and superresolution imaging. Deconvolution of confocal microscopy images shows vMIA localizes away from mitochondrial matrix at the Mitochondria-ER interface. By gated stimulated emission depletion (GSTED) imaging, we show that along this interface vMIA is distributed in clusters. Through multicolor, multifocal structured illumination microscopy (MSIM), we find vMIA clusters localize away from MitoTracker Red, indicating its OMM localization. GSTED and MSIM imaging show vMIA exists in clusters of ~100–150 nm, which is consistent with the cluster size determined by Photoactivated Localization Microscopy (PALM). With these diverse superresolution approaches, we have imaged the clustered distribution of vMIA at the OMM adjacent to the ER. Our findings directly compare the relative advantages of each of these superresolution imaging modalities for imaging components of the MAM and sub-mitochondrial compartments. These studies establish the ability of superresolution imaging to provide valuable insight into viral protein location, particularly in the sub-mitochondrial compartments, and into their clustered organization. PMID:24721787

  17. Superresolution Imaging of Human Cytomegalovirus vMIA Localization in Sub-Mitochondrial Compartments

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    Shivaprasad Bhuvanendran

    2014-04-01

    Full Text Available The human cytomegalovirus (HCMV viral mitochondria-localized inhibitor of apoptosis (vMIA protein, traffics to mitochondria-associated membranes (MAM, where the endoplasmic reticulum (ER contacts the outer mitochondrial membrane (OMM. vMIA association with the MAM has not been visualized by imaging. Here, we have visualized this by using a combination of confocal and superresolution imaging. Deconvolution of confocal microscopy images shows vMIA localizes away from mitochondrial matrix at the Mitochondria-ER interface. By gated stimulated emission depletion (GSTED imaging, we show that along this interface vMIA is distributed in clusters. Through multicolor, multifocal structured illumination microscopy (MSIM, we find vMIA clusters localize away from MitoTracker Red, indicating its OMM localization. GSTED and MSIM imaging show vMIA exists in clusters of ~100–150 nm, which is consistent with the cluster size determined by Photoactivated Localization Microscopy (PALM. With these diverse superresolution approaches, we have imaged the clustered distribution of vMIA at the OMM adjacent to the ER. Our findings directly compare the relative advantages of each of these superresolution imaging modalities for imaging components of the MAM and sub-mitochondrial compartments. These studies establish the ability of superresolution imaging to provide valuable insight into viral protein location, particularly in the sub-mitochondrial compartments, and into their clustered organization.

  18. Double-stranded RNA binding by the human cytomegalovirus PKR antagonist TRS1.

    Science.gov (United States)

    Bierle, Craig J; Semmens, Kathryn M; Geballe, Adam P

    2013-07-20

    Protein Kinase R (PKR) inhibits translation initiation following double-stranded RNA (dsRNA) binding and thereby represses viral replication. Human cytomegalovirus (HCMV) encodes two noncanonical dsRNA binding proteins, IRS1 and TRS1, and the expression of at least one of these PKR antagonists is essential for HCMV replication. In this study, we investigated the role of dsRNA binding by TRS1 in PKR inhibition. We found that purified TRS1 binds specifically to dsRNA with an affinity lower than that of PKR. Point mutants in the TRS1 dsRNA binding domain that were deficient in rescuing the replication of vaccinia virus lacking its PKR antagonist E3L were unable to bind to dsRNA but retained the ability bind to PKR. Thus TRS1 binding to dsRNA and to PKR are separable. Overall, our results are most consistent with a model in which TRS1 binds simultaneously to both dsRNA and PKR to inhibit PKR activation. Copyright © 2013 Elsevier Inc. All rights reserved.

  19. RO0504985 is an inhibitor of CMGC kinase proteins and has anti-human cytomegalovirus activity.

    Science.gov (United States)

    Strang, Blair L

    2017-08-01

    Public-private partnerships allow many previously unavailable compounds to be screened for antiviral activity. Here a screening method was used to identify an oxindole compound, RO0504985, from a Roche kinase inhibitor library that inhibited human cytomegalovirus (HCMV) protein production. RO0504985 was previously described as an inhibitor of cyclin-dependent kinase 2 (CDK2). However, using kinase selectivity assays it was found that RO0504985 was an inhibitor of several CMGC group kinase proteins, including CDK2. Using virus yield reduction assays it was observed that RO0504985 inhibited replication of different HCMV strains at low micromolar concentrations. Western blotting was used to investigate how RO0504985 inhibited HCMV replication. Treatment of HCMV infected cells with RO0504985 inhibited production of the immediate early viral IE2 proteins and the late viral protein pp28. Thus, RO0504985 inhibited HCMV replication by preventing production of specific HCMV proteins necessary for virus replication. Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.

  20. Genomic and Functional Characteristics of Human Cytomegalovirus Revealed by Next-Generation Sequencing

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    Sijmons, Steven; Van Ranst, Marc; Maes, Piet

    2014-01-01

    The complete genome of human cytomegalovirus (HCMV) was elucidated almost 25 years ago using a traditional cloning and Sanger sequencing approach. Analysis of the genetic content of additional laboratory and clinical isolates has lead to a better, albeit still incomplete, definition of the coding potential and diversity of wild-type HCMV strains. The introduction of a new generation of massively parallel sequencing technologies, collectively called next-generation sequencing, has profoundly increased the throughput and resolution of the genomics field. These increased possibilities are already leading to a better understanding of the circulating diversity of HCMV clinical isolates. The higher resolution of next-generation sequencing provides new opportunities in the study of intrahost viral population structures. Furthermore, deep sequencing enables novel diagnostic applications for sensitive drug resistance mutation detection. RNA-seq applications have changed the picture of the HCMV transcriptome, which resulted in proof of a vast amount of splicing events and alternative transcripts. This review discusses the application of next-generation sequencing technologies, which has provided a clearer picture of the intricate nature of the HCMV genome. The continuing development and application of novel sequencing technologies will further augment our understanding of this ubiquitous, but elusive, herpesvirus. PMID:24603756

  1. Regulation of Human Cytomegalovirus Transcription in Latency: Beyond the Major Immediate-Early Promoter

    Science.gov (United States)

    Reeves, Matthew; Sinclair, John

    2013-01-01

    Lytic infection of differentiated cell types with human cytomegalovirus (HCMV) results in the temporal expression of between 170–200 open reading frames (ORFs). A number of studies have demonstrated the temporal regulation of these ORFs and that this is orchestrated by both viral and cellular mechanisms associated with the co-ordinated recruitment of transcription complexes and, more recently, higher order chromatin structure. Importantly, HCMV, like all herpes viruses, establishes a lifelong latent infection of the host—one major site of latency being the undifferentiated haematopoietic progenitor cells resident in the bone marrow. Crucially, the establishment of latency is concomitant with the recruitment of cellular enzymes that promote extensive methylation of histones bound to the major immediate early promoter. As such, the repressive chromatin structure formed at the major immediate early promoter (MIEP) elicits inhibition of IE gene expression and is a major factor involved in maintenance of HCMV latency. However, it is becoming increasingly clear that a distinct subset of viral genes is also expressed during latency. In this review, we will discuss the mechanisms that control the expression of these latency-associated transcripts and illustrate that regulation of these latency-associated promoters is also subject to chromatin mediated regulation and that the instructive observations previously reported regarding the negative regulation of the MIEP during latency are paralleled in the regulation of latent gene expression. PMID:23736881

  2. Human cytomegalovirus-encoded US28 may act as a tumor promoter in colorectal cancer.

    Science.gov (United States)

    Cai, Zhen-Zhai; Xu, Jian-Gang; Zhou, Yu-Hui; Zheng, Ji-Hang; Lin, Ke-Zhi; Zheng, Shu-Zhi; Ye, Meng-Si; He, Yun; Liu, Chang-Bao; Xue, Zhan-Xiong

    2016-03-07

    To assess human cytomegalovirus-encoded US28 gene function in colorectal cancer (CRC) pathogenesis. Immunohistochemical analysis was performed to determine US28 expression in 103 CRC patient samples and 98 corresponding adjacent noncancerous samples. Patient data were compared by age, sex, tumor location, histological grade, Dukes' stage, and overall mean survival time. In addition, the US28 gene was transiently transfected into the CRC LOVO cell line, and cell proliferation was assessed using a cell counting kit-8 assay. Cell cycle analysis by flow cytometry and a cell invasion transwell assay were also carried out. US28 levels were clearly higher in CRC tissues (38.8%) than in adjacent noncancerous samples (7.1%) (P = 0.000). Interestingly, elevated US28 amounts in CRC tissues were significantly associated with histological grade, metastasis, Dukes' stage, and overall survival (all P < 0.05); meanwhile, US28 expression was not significantly correlated with age, sex or tumor location. In addition, multivariate Cox regression data revealed US28 level as an independent CRC prognostic marker (P = 0.000). LOVO cells successfully transfected with the US28 gene exhibited higher viability, greater chemotherapy resistance, accelerated cell cycle progression, and increased invasion ability. US28 expression is predictive of poor prognosis and may promote CRC.

  3. Human Cytomegalovirus Secretome Contains Factors That Induce Angiogenesis and Wound Healing

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    Dumortier, Jerome; Streblow, Daniel N.; Moses, Ashlee V.; Jacobs, Jon M.; Kreklywich, Craig N.; Camp, David G.; Smith, Richard D.; Orloff, Susan L.; Nelson, Jay

    2008-07-01

    Human cytomegalovirus (HCMV) is implicated in the acceleration of a number of vascular diseases including transplant vascular sclerosis (TVS), the lesion associated with chronic rejection (CR) of solid organ transplants. Although the virus persists in the allograft throughout the course of disease, few cells are directly infected by CMV. This observation is in contrast to the global effects that CMV has on the acceleration of TVS/CR, suggesting that CMV infection indirectly promotes the vascular disease process. Recent transcriptome analysis of CMV-infected heart allografts indicates that the virus induces cytokines and growth factors associated with angiogenesis (AG) and wound healing (WH), suggesting that CMV may accelerate TVS/CR through the induction and secretion of AG/WH factors from infected cells. We analyzed virus-free supernatants from HCMV-infected cells (HCMV secretomes) for growth factors, by mass spectrometry and immunoassays, and found that the HCMV secretome contains over 1,000 cellular proteins, many of which are involved in AG/WH. Importantly, functional assays demonstrated that CMV but not herpes simplex virus secretomes not only induce AG/WH but also promote neovessel stabilization and endothelial cell survival for 2 weeks. These findings suggest that CMV acceleration of TVS occurs through virus-induced growth factors and cytokines in the CMV secretome.

  4. Impact of Persistent Cytomegalovirus Infection on Dynamic Changes in Human Immune System Profile.

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    Rosanna Vescovini

    Full Text Available Human cytomegalovirus (HCMV imprints the immune system after primary infection, however its effect during chronic infection still needs to be deciphered. In this study we report the variation of blood cell count along with anti-HCMV IgG and T cell responses to pp-65 and IE-1 antigens, that occurred after an interval of five years in a cohort of 25 seropositive healthy adults. We found increased anti-viral IgG antibody responses and intracellular interferon-gamma secreting CD8+ T cell responses to pp-65: a result consistent with memory inflation. With the only exception of shortage in naive CD8+ T cells most memory T cell subsets as well as total CD8+ T cells, T cells, lymphocytes, monocytes and leukocytes had increased. By contrast, none of the cell types tested were found to have increased in 14 subjects stably seronegative. Rather, in addition to a shortage in naive CD8+ T cells, also memory T cell subsets and most other cell types decreased, either in a statistically significant or non-significant manner. The trend of T cell pool representation with regard to CD4/CD8 ratio was in the opposing directions depending on HCMV serology. Globally, this study demonstrates different dynamic changes of most blood cell types depending on presence or absence of HCMV infection. Therefore, HCMV plays a continual role in modulating homeostasis of blood T cells and a broader expanding effect on other cell populations of lymphoid and myeloid origin.

  5. Impact of Persistent Cytomegalovirus Infection on Dynamic Changes in Human Immune System Profile.

    Science.gov (United States)

    Vescovini, Rosanna; Telera, Anna Rita; Pedrazzoni, Mario; Abbate, Barbara; Rossetti, Pietro; Verzicco, Ignazio; Arcangeletti, Maria Cristina; Medici, Maria Cristina; Calderaro, Adriana; Volpi, Riccardo; Sansoni, Paolo; Fagnoni, Francesco Fausto

    2016-01-01

    Human cytomegalovirus (HCMV) imprints the immune system after primary infection, however its effect during chronic infection still needs to be deciphered. In this study we report the variation of blood cell count along with anti-HCMV IgG and T cell responses to pp-65 and IE-1 antigens, that occurred after an interval of five years in a cohort of 25 seropositive healthy adults. We found increased anti-viral IgG antibody responses and intracellular interferon-gamma secreting CD8+ T cell responses to pp-65: a result consistent with memory inflation. With the only exception of shortage in naive CD8+ T cells most memory T cell subsets as well as total CD8+ T cells, T cells, lymphocytes, monocytes and leukocytes had increased. By contrast, none of the cell types tested were found to have increased in 14 subjects stably seronegative. Rather, in addition to a shortage in naive CD8+ T cells, also memory T cell subsets and most other cell types decreased, either in a statistically significant or non-significant manner. The trend of T cell pool representation with regard to CD4/CD8 ratio was in the opposing directions depending on HCMV serology. Globally, this study demonstrates different dynamic changes of most blood cell types depending on presence or absence of HCMV infection. Therefore, HCMV plays a continual role in modulating homeostasis of blood T cells and a broader expanding effect on other cell populations of lymphoid and myeloid origin.

  6. Impact of Persistent Cytomegalovirus Infection on Dynamic Changes in Human Immune System Profile

    Science.gov (United States)

    Vescovini, Rosanna; Telera, Anna Rita; Pedrazzoni, Mario; Abbate, Barbara; Rossetti, Pietro; Verzicco, Ignazio; Arcangeletti, Maria Cristina; Medici, Maria Cristina; Calderaro, Adriana; Volpi, Riccardo; Sansoni, Paolo; Fagnoni, Francesco Fausto

    2016-01-01

    Human cytomegalovirus (HCMV) imprints the immune system after primary infection, however its effect during chronic infection still needs to be deciphered. In this study we report the variation of blood cell count along with anti-HCMV IgG and T cell responses to pp-65 and IE-1 antigens, that occurred after an interval of five years in a cohort of 25 seropositive healthy adults. We found increased anti-viral IgG antibody responses and intracellular interferon-gamma secreting CD8+ T cell responses to pp-65: a result consistent with memory inflation. With the only exception of shortage in naive CD8+ T cells most memory T cell subsets as well as total CD8+ T cells, T cells, lymphocytes, monocytes and leukocytes had increased. By contrast, none of the cell types tested were found to have increased in 14 subjects stably seronegative. Rather, in addition to a shortage in naive CD8+ T cells, also memory T cell subsets and most other cell types decreased, either in a statistically significant or non-significant manner. The trend of T cell pool representation with regard to CD4/CD8 ratio was in the opposing directions depending on HCMV serology. Globally, this study demonstrates different dynamic changes of most blood cell types depending on presence or absence of HCMV infection. Therefore, HCMV plays a continual role in modulating homeostasis of blood T cells and a broader expanding effect on other cell populations of lymphoid and myeloid origin. PMID:26990192

  7. Outcome of cytomegalovirus retinitis in immunocompromised patients without Human Immunodeficiency Virus treated with intravitreal ganciclovir injection.

    Science.gov (United States)

    Agarwal, Aniruddha; Kumari, Neha; Trehan, Amita; Khadwal, Alka; Dogra, Mangat R; Gupta, Vishali; Sharma, Aman; Gupta, Amod; Singh, Ramandeep

    2014-09-01

    To study the outcomes of treatment with intravitreal ganciclovir injection for cytomegalovirus (CMV) retinitis in patients without Human Immunodeficiency Virus (HIV) infection. In this retrospective cohort study, demographic and clinical characteristics of patients with CMV retinitis without HIV were noted. Patients received intravitreal ganciclovir injection (2 mg/0.1 ml) alone until quiescence. The outcome measures were time taken for the lesions to heal, number of injections, change in best-corrected visual acuity (BCVA), recurrence of retinitis, occurrence of immune recovery uveitis (IRU) or injection-related complications and retinal detachment (RD). 18 eyes of ten patients (six males) with mean age of 33.7 years from June 2004 to March 2013 were included. Thirteen eyes with active lesions (mean BCVA of 0.51 ± 0.41) received 5.54 ± 3.36 intravitreal ganciclovir injections with complete healing within 1.81 ± 1.25 months. The final BCVA was 0.43 ± 0.52. IRU was observed in six eyes (33.33%) and RD developed in one eye. One eye had recurrence 1 month after stopping ganciclovir injections. The rest of the patients had recurrence-free follow-up at 9.46 ± 12.42 months. Non-HIV patients with CMV retinitis can be successfully treated with intravitreal ganciclovir injection alone, avoiding the systemic side effects of systemic anti-CMV therapy.

  8. Detection of Human Cytomegalovirus in Different Histopathological Types of Glioma in Iraqi Patients

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    Haidar A. Shamran

    2015-01-01

    Full Text Available Human Cytomegalovirus (HCMV is an endemic herpes virus that reemerges in cancer patients enhancing oncogenic potential. HCMV infection is associated with certain types of cancer morbidity such as glioblastomas. HCMV, like all other herpes viruses, has the ability to remain latent within the body of the host and can contribute in chronic inflammation. To determine the role of HCMV in glioma pathogenesis, paraffin-embedded blocks from glioma patients (n=50 and from benign meningioma patients (n=30 were obtained and evaluated by immunohistochemistry and polymerase chain reaction for the evidence of HCMV antigen expression and the presence of viral DNA. We detected HCMV antigen and DNA for IEI-72, pp65, and late antigen in 33/36, 28/36, and 26/36 in glioblastoma multiforme patients whereas 12/14, 10/14, and 9/14 in anaplastic astrocytoma patients, respectively. Furthermore, 84% of glioma patients were positive for immunoglobulin G (IgG compared to 72.5% among control samples (P=0.04. These data indicate the presence of the HCMV virus in a high percentage of glioma samples demonstrating distinct histopathological grades and support previous reports showing the presence of HCMV infection in glioma tissue. These studies demonstrate that detection of low-levels of latent viral infections may play an active role in glioma development and pathogenesis.

  9. Presentation of an immunodominant immediate-early CD8+ T cell epitope resists human cytomegalovirus immunoevasion.

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    Stefanie Ameres

    Full Text Available Control of human cytomegalovirus (HCMV depends on CD8+ T cell responses that are shaped by an individual's repertoire of MHC molecules. MHC class I presentation is modulated by a set of HCMV-encoded proteins. Here we show that HCMV immunoevasins differentially impair T cell recognition of epitopes from the same viral antigen, immediate-early 1 (IE-1, that are presented by different MHC class I allotypes. In the presence of immunoevasins, HLA-A- and HLA-B-restricted T cell clones were ineffective, but HLA-C*0702-restricted T cell clones recognized and killed infected cells. Resistance of HLA-C*0702 to viral immunoevasins US2 and US11 was mediated by the alpha3 domain and C-terminal region of the HLA heavy chain. In healthy donors, HLA-C*0702-restricted T cells dominated the T cell response to IE-1. The same HLA-C allotype specifically protected infected cells from attack by NK cells that expressed a corresponding HLA-C-specific KIR. Thus, allotype-specific viral immunoevasion allows HCMV to escape control by NK cells and HLA-A- and HLA-B-restricted T cells, while the virus becomes selectively vulnerable to an immunodominant population of HLA-C-restricted T cells. Our work identifies a T cell population that may be of particular efficiency in HCMV-specific immunotherapy.

  10. The Cyclin-Dependent Kinase Ortholog pUL97 of Human Cytomegalovirus Interacts with Cyclins

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    Laura Graf

    2013-12-01

    Full Text Available The human cytomegalovirus (HCMV-encoded protein kinase, pUL97, is considered a cyclin-dependent kinase (CDK ortholog, due to shared structural and functional characteristics. The primary mechanism of CDK activation is binding to corresponding cyclins, including cyclin T1, which is the usual regulatory cofactor of CDK9. This study provides evidence of direct interaction between pUL97 and cyclin T1 using yeast two-hybrid and co-immunoprecipitation analyses. Confocal immunofluorescence revealed partial colocalization of pUL97 with cyclin T1 in subnuclear compartments, most pronounced in viral replication centres. The distribution patterns of pUL97 and cyclin T1 were independent of HCMV strain and host cell type. The sequence domain of pUL97 responsible for the interaction with cyclin T1 was between amino acids 231–280. Additional co-immunoprecipitation analyses showed cyclin B1 and cyclin A as further pUL97 interaction partners. Investigation of the pUL97-cyclin T1 interaction in an ATP consumption assay strongly suggested phosphorylation of pUL97 by the CDK9/cyclin T1 complex in a substrate concentration-dependent manner. This is the first demonstration of interaction between a herpesviral CDK ortholog and cellular cyclins.

  11. High frequency of Human Cytomegalovirus DNA in the Liver of Infants with Extrahepatic Neonatal Cholestasis

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    Escanhoela Cecília AF

    2005-12-01

    Full Text Available Abstract Background Biliary atresia (BA is the most severe hepatic disorder in newborns and its etiopathogenesis remains unknown. Viral involvement has been proposed, including the human cytomegalovirus (HCMV. The aims of the study were to use the polymerase chain reaction (PCR to screen the liver tissue of infants with extrahepatic cholestasis for HCMV and to correlate the results with serological antibodies against HCMV and histological findings. Methods A retrospective study in a tertiary care setting included 35 patients (31 BA, 1 BA associated with a choledochal cyst, 2 congenital stenosis of the distal common bile duct and 1 hepatic cyst. HCMV serology was determined by ELISA. Liver and porta hepatis were examined histologically. Liver samples from infants and a control group were screened for HCMV DNA. Results Twelve patients had HCMV negative serology, 9 were positive for IgG antibodies and 14 were positive for IgG and IgM. Nine liver and seven porta hepatis samples were positive for HCMV DNA but none of the control group were positive (general frequency of positivity was 34.3% – 12/35. There was no correlation between HCMV positivity by PCR and the histological findings. The accuracy of serology for detecting HCMV antibodies was low. Conclusion These results indicate an elevated frequency of HCMV in pediatric patients with extrahepatic neonatal cholestasis. They also show the low accuracy of serological tests for detecting active HCMV infection and the lack of correlation between HCMV positivity by PCR and the histopathological changes.

  12. Human cytomegalovirus infection contributes to glioma disease progression via upregulating endocan expression.

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    Xing, Yan; Wang, Yisong; Wang, Shijie; Wang, Xin; Fan, Dongying; Zhou, Dabiao; An, Jing

    2016-11-01

    The etiology of malignant glioma remains unclear. To examine the association between glioma and human cytomegalovirus (HCMV) infection and the possible mechanism through which HCMV contributes to malignant glioma, we investigated the expression of HCMV components and an angiogenesis marker, endocan, in 79 glioma specimens and 8 control brain samples. HCMV pp65 protein and DNA were detected in 65.8% (52 of 79) and 54.4% (43 of 79) of glioma specimens, respectively. The positive rate and expression levels of pp65 were significantly correlated with the glioma grades. The endocan expression was detected in 78.5% (62 of 79) of glioma specimens, and elevated endocan immunoreactivity was also significantly associated with high-grade glioma. The pp65 was predominantly detected and colocalized with endocan in the cytoplasm of tumor cells. Importantly, there was a significant positive correlation in detection rates between those 2 proteins. In control samples, neither HCMV pp65 nor endocan expression was detected. Moreover, the serum endocan levels in glioma patients were markedly higher than that in healthy subjects. In in vitro study, HCMV infection induced the expression of interleukin 6 and tumor necrosis factor-α in human glioblastoma U87 MG (U87) cells and human umbilical vein endothelial cells (HUVECs). Furthermore, elevated endocan levels were also observed in HCMV-infected U87 cells and HUVECs and antiviral treatment with ganciclovir reduced the endocan expression. These results suggest HCMV infection leads to glioma progression through an upregulation of endocan and the secretion of inflammatory cytokines. Thus, anti-HCMV treatment may represent a potentially novel therapeutic strategy for glioma. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  13. Synergistic inhibition of human cytomegalovirus replication by interferon-alpha/beta and interferon-gamma

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    Morris Cindy A

    2005-02-01

    Full Text Available Abstract Background Recent studies have shown that gamma interferon (IFN-γ synergizes with the innate IFNs (IFN-α and IFN-β to inhibit herpes simplex virus type 1 (HSV-1 replication in vitro. To determine whether this phenomenon is shared by other herpesviruses, we investigated the effects of IFNs on human cytomegalovirus (HCMV replication. Results We have found that as with HSV-1, IFN-γ synergizes with the innate IFNs (IFN-α/β to potently inhibit HCMV replication in vitro. While pre-treatment of human foreskin fibroblasts (HFFs with IFN-α, IFN-β or IFN-γ alone inhibited HCMV plaque formation by ~30 to 40-fold, treatment with IFN-α and IFN-γ or IFN-β and IFN-γ inhibited HCMV plaque formation by 163- and 662-fold, respectively. The generation of isobole plots verified that the observed inhibition of HCMV plaque formation and replication in HFFs by IFN-α/β and IFN-γ was a synergistic interaction. Additionally, real-time PCR analyses of the HCMV immediate early (IE genes (IE1 and IE2 revealed that IE mRNA expression was profoundly decreased in cells stimulated with IFN-α/β and IFN-γ (~5-11-fold as compared to vehicle-treated cells. Furthermore, decreased IE mRNA expression was accompanied by a decrease in IE protein expression, as demonstrated by western blotting and immunofluorescence. Conclusion These findings suggest that IFN-α/β and IFN-γ synergistically inhibit HCMV replication through a mechanism that may involve the regulation of IE gene expression. We hypothesize that IFN-γ produced by activated cells of the adaptive immune response may potentially synergize with endogenous type I IFNs to inhibit HCMV dissemination in vivo.

  14. Human cytomegalovirus exploits interferon-induced transmembrane proteins to facilitate morphogenesis of the virion assembly compartment.

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    Xie, Maorong; Xuan, Baoqin; Shan, Jiaoyu; Pan, Deng; Sun, Yamei; Shan, Zhao; Zhang, Jinping; Yu, Dong; Li, Bin; Qian, Zhikang

    2015-03-01

    Recently, interferon-induced transmembrane proteins (IFITMs) have been identified to be key effector molecules in the host type I interferon defense system. The invasion of host cells by a large range of RNA viruses is inhibited by IFITMs during the entry step. However, the roles of IFITMs in DNA virus infections have not been studied in detail. In this study, we report that human cytomegalovirus (HCMV), a large human DNA virus, exploits IFITMs to facilitate the formation of the virion assembly compartment (vAC) during infection of human fibroblasts. We found that IFITMs were expressed constitutively in human embryonic lung fibroblasts (MRC5 cells). HCMV infection inhibited IFITM protein accumulation in the later stages of infection. Overexpression of an IFITM protein in MRC5 cells slightly enhanced HCMV production and knockdown of IFITMs by RNA interference reduced the virus titer by about 100-fold on day 8 postinfection, according to the findings of a virus yield assay at a low multiplicity of infection. Virus gene expression and DNA synthesis were not affected, but the typical round structure of the vAC was not formed after the suppression of IFITMs, thereby resulting in defective virion assembly and the production of less infectious virion particles. Interestingly, the replication of herpes simplex virus, a human herpesvirus that is closely related to HCMV, was not affected by the suppression of IFITMs in MRC5 cells. These results indicate that IFITMs are involved in a specific pathway required for HCMV replication. HCMV is known to repurpose the interferon-stimulated genes (ISGs) viperin and tetherin to facilitate its replication. Our results expand the range of ISGs that can be exploited by HCMV for its replication. This is also the first report of a proviral function of IFITMs in DNA virus replication. In addition, whereas previous studies showed that IFITMs modulate virus entry, which is a very early stage in the virus life cycle, we identified a new

  15. Human cytomegalovirus infection interferes with the maintenance and differentiation of trophoblast progenitor cells of the human placenta.

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    Tabata, Takako; Petitt, Matthew; Zydek, Martin; Fang-Hoover, June; Larocque, Nicholas; Tsuge, Mitsuru; Gormley, Matthew; Kauvar, Lawrence M; Pereira, Lenore

    2015-05-01

    Human cytomegalovirus (HCMV) is a major cause of birth defects that include severe neurological deficits, hearing and vision loss, and intrauterine growth restriction. Viral infection of the placenta leads to development of avascular villi, edema, and hypoxia associated with symptomatic congenital infection. Studies of primary cytotrophoblasts (CTBs) revealed that HCMV infection impedes terminal stages of differentiation and invasion by various molecular mechanisms. We recently discovered that HCMV arrests earlier stages involving development of human trophoblast progenitor cells (TBPCs), which give rise to the mature cell types of chorionic villi-syncytiotrophoblasts on the surfaces of floating villi and invasive CTBs that remodel the uterine vasculature. Here, we show that viral proteins are present in TBPCs of the chorion in cases of symptomatic congenital infection. In vitro studies revealed that HCMV replicates in continuously self-renewing TBPC lines derived from the chorion and alters expression and subcellular localization of proteins required for cell cycle progression, pluripotency, and early differentiation. In addition, treatment with a human monoclonal antibody to HCMV glycoprotein B rescues differentiation capacity, and thus, TBPCs have potential utility for evaluation of the efficacies of novel antiviral antibodies in protecting and restoring placental development. Our results suggest that HCMV replicates in TBPCs in the chorion in vivo, interfering with the earliest steps in the growth of new villi, contributing to virus transmission and impairing compensatory development. In cases of congenital infection, reduced responsiveness of the placenta to hypoxia limits the transport of substances from maternal blood and contributes to fetal growth restriction. Human cytomegalovirus (HCMV) is a leading cause of birth defects in the United States. Congenital infection can result in permanent neurological defects, mental retardation, hearing loss, visual

  16. Generation of a novel human cytomegalovirus bacterial artificial chromosome tailored for transduction of exogenous sequences.

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    Close, William L; Bhandari, Amit; Hojeij, Marwa; Pellett, Philip E

    2017-10-15

    The study of herpesviruses, including human cytomegalovirus (HCMV), is complicated by viral genome complexity and inefficient methods for genetic manipulation in tissue culture. Reverse genetics of herpesviruses has been facilitated by propagating their genomes in E. coli as bacterial artificial chromosomes (BACs), which enables complex and precise genetic manipulation using bacterial recombinational systems. Internal capsid volume imposes a strict limit on the length of genome that can be packaged efficiently. This necessitates deletion of presumably nonessential segments of the viral genome to allow for incorporation of the E. coli mini-F plasmid propagation sequence. To avoid deleting viral genes, several BACs utilize a Cre/LoxP system to self-excise the mini-F sequence upon reconstitution of virus in tissue culture. Here, we describe the adaptation of Cre/LoxP to modify the mini-F sequence of the HCMV TB40/E BAC, thus generating a new self-excisable BAC, TB40/E/Cre. After excision of the E. coli propagation sequence, a 2.7 kbp genome length deficit is created due to a preexisting deletion within the US2-US6 coding region. We exploited this deficit and an FKBP12 protein destabilization domain (ddFKBP) to create a novel gene transduction system for studying exogenous proteins during HCMV infection. Using TB40/E/Cre, we: i) found genome length-associated differences in growth and ii) demonstrated its utility as a system capable of efficient transduction of exogenous proteins and regulation of their accumulation over periods as short as 2h. TB40/E/Cre is a powerful tool of broad applicability that can be adapted to study HCMV replication and cell biology in a variety of contexts. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Diagnosis and Management of Human Cytomegalovirus Infection in the Mother, Fetus, and Newborn Infant

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    Revello, Maria Grazia; Gerna, Giuseppe

    2002-01-01

    Human cytomegalovirus (HCMV) is the leading cause of congenital viral infection and mental retardation. HCMV infection, while causing asymptomatic infections in most immunocompetent subjects, can be transmitted during pregnancy from the mother with primary (and also recurrent) infection to the fetus. Hence, careful diagnosis of primary infection is required in the pregnant woman based on the most sensitive serologic assays (immunoglobulin M [IgM] and IgG avidity assays) and conventional virologic and molecular procedures for virus detection in blood. Maternal prognostic markers of fetal infection are still under investigation. If primary infection is diagnosed in a timely manner, prenatal diagnosis can be offered, including the search for virus and virus components in fetal blood and amniotic fluid, with fetal prognostic markers of HCMV disease still to be defined. However, the final step for definite diagnosis of congenital HCMV infection is detection of virus in the blood or urine in the first 1 to 2 weeks of life. To date, treatment of congenital infection with antiviral drugs is only palliative both prior to and after birth, whereas the only efficacious preventive measure seems to be the development of a safe and immunogenic vaccine, including recombinant, subunit, DNA, and peptide-based vaccines now under investigation. The following controversial issues are discussed in the light of the most recent advances in the field: the actual perception of the problem; universal serologic screening before pregnancy; the impact of correct counseling on decision making by the couple involved; the role of prenatal diagnosis in ascertaining transmission of virus to the fetus; the impact of preconceptional and periconceptional infections on the prevalence of congenital infection; and the prevalence of congenitally infected babies born to mothers who were immune prior to pregnancy compared to the number born to mothers undergoing primary infection during pregnancy. PMID

  18. A third component of the human cytomegalovirus terminase complex is involved in letermovir resistance.

    Science.gov (United States)

    Chou, Sunwen

    2017-12-01

    Letermovir is a human cytomegalovirus (CMV) terminase inhibitor that was clinically effective in a Phase III prevention trial. In vitro studies have shown that viral mutations conferring letermovir resistance map primarily to the UL56 component of the terminase complex and uncommonly to UL89. After serial culture of a baseline CMV laboratory strain under letermovir, mutation was observed in a third terminase component in 2 experiments, both resulting in amino acid substitution P91S in gene UL51 and adding to a pre-existing UL56 mutation. Recombinant phenotyping indicated that P91S alone conferred 2.1-fold increased letermovir resistance (EC50) over baseline, and when combined with UL56 mutation S229F or R369M, multiplied the level of resistance conferred by those mutations by 3.5-7.7-fold. Similarly a combination of UL56 mutations S229F, L254F and L257I selected in the same experiment conferred 54-fold increased letermovir EC50 over baseline, but 290-fold when combined with UL51 P91S. The P91S mutant was not perceptibly growth impaired. Although pUL51 is essential for normal function of the terminase complex, its biological significance is not well understood. Letermovir resistance mutations mapping to 3 separate genes, and their multiplier effect on the level of resistance, suggest that the terminase components interactively contribute to the structure of a letermovir antiviral target. The diagnostic importance of the UL51 P91S mutation arises from its potential to augment the letermovir resistance of some UL56 mutations at low fitness cost. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Effectiveness of PCR and Immunofluorescence Techniques for Detecting Human Cytomegalovirus in Blood and Bronchoalveolar Lavage Fluid.

    Science.gov (United States)

    Roży, A; Duk, K; Szumna, B; Skrońska, P; Gawryluk, D; Chorostowska-Wynimko, J

    Current diagnostic methods allow a rapid and reliable detection of active human cytomegalovirus (hCMV) infection by identifying the presence of pp65 CMV antigen or CMV DNA in peripheral blood and affected organs. The goal of this study was to evaluate the effectiveness of CMV detection in blood and organ-specific biological material, such as bronchoalveolar lavage fluid (BALF), by comparing two standard diagnostic methods, immunofluorescence (IF) and the real-time polymerase chain reaction (PCR). We evaluated 25 patients with concomitant respiratory disease who were referred to our hospital for diagnosis due to suspected acute CMV infection. The presence of hCMV was concomitantly evaluated by IF and PCR in 16 peripheral blood samples. In two patients, we observed positive results for both IF and PCR, and in two other patients the results were discordant. Of 11 patients, CMV DNA was detected in six BALF samples, and in one blood plasma sample. Real-time PCR detected CMV DNA in 54.6 % of BALF samples and 12.0 % of blood samples, while indirect IF testing confirmed antigenemia in 12.5 % of blood samples. The results from our study suggest that the IF method is as effective as PCR for detecting an ongoing CMV infection in blood samples. However, real-time PCR was much more effective at detecting CMV DNA in BALF compared to blood samples. Our results suggest that the biological material being tested during CMV diagnosis should be derived directly from the virally infected organ(s).

  20. Cytomegalovirus and cancer after kidney transplantation: Role of the human leukocyte antigen system?

    Science.gov (United States)

    Wong, Germaine; Chakera, Aron; Chapman, Jeremy R; Chadban, Steve C; Pilmore, Helen; Craig, Jonathan C; Lim, Wai H

    2017-02-01

    The role of cytomegalovirus (CMV) in cancer development after transplantation remains uncertain. We aimed to determine the association between donor and recipient CMV serological status and the risk of cancer development after kidney transplantation. Using data from the Australian and New Zealand Dialysis and Transplant (ANZDATA) Registry, we assessed the association between CMV donor/recipient (D/R) serological status and the risk of solid organ cancers in primary adult deceased-donor kidney transplant patients between 1990 and 2012. Of 8140 recipients, a total of 895 (11%) recipients developed incident cancers during a follow-up time of 51 555 person-years. Human leukocyte antigen (HLA) mismatches was an effect modifier between CMV serological status and cancer (P=.03 for interaction). In recipients who have received 0-2 HLA-ABDR mismatched kidneys, the adjusted hazard ratios for cancer incidence among those with CMV D-/R-, CMV D-/R+, and CMV D+/R- were 0.47 (95% confidence interval [CI]: 0.24-0.91), 1.42 (95% CI: 0.97-2.07), and 1.02 (95% CI: 0.67-1.57), respectively compared with the reference of CMV D+/R+. A similar association was not observed in those with >2 HLA-ABDR mismatches. CMV D-/R- status was associated with a reduced risk of cancer in kidney transplant recipients who have received well-matched renal allografts, suggesting a potential role of HLA matching in cancer development. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  1. The immunology of human cytomegalovirus latency: could latent infection be cleared by novel immunotherapeutic strategies?

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    Wills, Mark R; Poole, Emma; Lau, Betty; Krishna, Ben; Sinclair, John H

    2015-01-01

    While the host immune response following primary human cytomegalovirus (HCMV) infection is generally effective at stopping virus replication and dissemination, virus is never cleared by the host and like all herpesviruses, persists for life. At least in part, this persistence is known to be facilitated by the ability of HCMV to establish latency in myeloid cells in which infection is essentially silent with, importantly, a total lack of new virus production. However, although the viral transcription programme during latency is much suppressed, a number of viral genes are expressed during latent infection at the protein level and many of these have been shown to have profound effects on the latent cell and its environment. Intriguingly, many of these latency-associated genes are also expressed during lytic infection. Therefore, why the same potent host immune responses generated during lytic infection to these viral gene products are not recognized during latency, thereby allowing clearance of latently infected cells, is far from clear. Reactivation from latency is also a major cause of HCMV-mediated disease, particularly in the immune compromised and immune naive, and is also likely to be a major source of virus in chronic subclinical HCMV infection which has been suggested to be associated with long-term diseases such as atherosclerosis and some neoplasias. Consequently, understanding latency and why latently infected cells appear to be immunoprivileged is crucial for an understanding of the pathogenesis of HCMV and may help to design strategies to eliminate latent virus reservoirs, at least in certain clinical settings. PMID:25132454

  2. Identification of Proteins in Human Cytomegalovirus (HCMV) Particles: the HCMV Proteome

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    Varnum, Susan M.; Streblow, Daniel N.; Monroe, Matthew E.; Smith, Patricia; Auberry, Kenneth J.; Pasa-Tolic, Liljiana; Wang, Dai; Camp, David G.; Rodland, Karin D.; Wiley, H S.; Britt, William; Shenk, Thomas; Smith, Richard D.; Nelson, Jay

    2004-10-15

    Human cytomegalovirus (HCMV), a member of the herpes virus family, is a large complex enveloped virus composed of both viral and cellular gene products. While the sequence of the HCMV genome has been known for over a decade, the full set of viral and cellular proteins that compose the HCMV virion are unknown. To approach this problem we have utilized gel-free two-dimensional capillary liquid chromatography-tandem mass spectrometry (MS/MS) and Fourier transform ion cyclotron resonance MS to identify and determine the relative abundances of viral and cellular proteins in purified HCMV AD169 virions and dense bodies. Analysis of the proteins from purified HCMV virion preparations has indicated that the particle contains significantly more viral proteins than previously known. In this study, we identified 71 HCMV-encoded proteins that included 12 proteins encoded by known viral open reading frames (ORFs) previously not associated with virions and 12 proteins from novel viral ORFs. Analysis of the relative abundance of HCMV proteins indicated that the predominant virion protein was the pp65 tegument protein and that gM rather than gB was the most abundant glycoprotein. We have also identified over 70 host cellular proteins in HCMV virions, which include cellular structural proteins, enzymes, and chaperones. In addition, analysis of HCMV dense bodies indicated that these viral particles are composed of 29 viral proteins with a reduced quantity of cellular proteins in comparison to HCMV virions. This study provides the first comprehensive quantitative analysis of the viral and cellular proteins that compose infectious particles of a large complex virus.

  3. Artesunate demonstrates in vitro synergism with several antiviral agents against human cytomegalovirus.

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    Drouot, Emilien; Piret, Jocelyne; Boivin, Guy

    2016-01-01

    Human cytomegalovirus (HCMV) infections remain a major problem in immunocompromised patients. Three antiviral agents, ganciclovir (GCV), foscarnet (FOS) and cidofovir (CDV), are currently approved for the treatment of HCMV infections. They all target the viral DNA polymerase and are associated with significant side effects. Combinations of novel antiviral compounds acting on different targets such as artesunate (ART) with currently approved drugs or eventually letermovir or maribavir (MBV) may result in synergistic effects. Here, we evaluated the in vitro activity of a series of two-drug combinations against a wild-type recombinant HCMV strain by the Gaussia luciferase (GLuc) reporter assay. The in vitro activity of each drug was first tested individually against HCMV by using the GLuc reporter assay. The activity of two-drug combinations consisting of ART and currently approved drugs, as well as letermovir or MBV, was then analysed by the Chou-Talalay method. The concentrations of GCV, FOS, CDV and ART that reduced the GLuc activity by 50% (EC50 values) were 3.92 ±1.64 µM, 62.45 ±8.39 µM, 0.68 ±0.19 µM and 3.86 ±1.25 µM, respectively, whereas those of MBV and letermovir were 64 ±22 nM and 2.50 ±0.83 nM, respectively. The combination of ART with GCV, CDV or MBV was associated with synergism, whereas combination of ART with FOS or letermovir resulted in moderate synergism. As expected, the combination of MBV with GCV was antagonistic. These results suggest that the combination of ART with the antiviral agents tested in this study could be an interesting strategy for the treatment of HCMV infections to reduce toxicity and drug-resistance development.

  4. Rapid Ganciclovir Susceptibility Assay Using Flow Cytometry for Human Cytomegalovirus Clinical Isolates

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    McSharry, James J.; Lurain, Nell S.; Drusano, George L.; Landay, Alan L.; Notka, Mostafa; O’Gorman, Maurice R. G.; Weinberg, Adriana; Shapiro, Howard M.; Reichelderfer, Patricia S.; Crumpacker, Clyde S.

    1998-01-01

    Rapid, quantitative, and objective determination of the susceptibilities of human cytomegalovirus (HCMV) clinical isolates to ganciclovir has been assessed by an assay that uses a fluorochrome-labeled monoclonal antibody to an HCMV immediate-early antigen and flow cytometry. Analysis of the ganciclovir susceptibilities of 25 phenotypically characterized clinical isolates by flow cytometry demonstrated that the 50% inhibitory concentrations (IC50s) of ganciclovir for 19 of the isolates were between 1.14 and 6.66 μM, with a mean of 4.32 μM (±1.93) (sensitive; IC50 less than 7 μM), the IC50s for 2 isolates were 8.48 and 9.79 μM (partially resistant), and the IC50s for 4 isolates were greater than 96 μM (resistant). Comparative analysis of the drug susceptibilities of these clinical isolates by the plaque reduction assay gave IC50s of less than 6 μM, with a mean of 2.88 μM (±1.40) for the 19 drug-sensitive isolates, IC50s of 6 to 8 μM for the partially resistant isolates, and IC50s of greater than 12 μM for the four resistant clinical isolates. Comparison of the IC50s for the drug-susceptible and partially resistant clinical isolates obtained by the flow cytometry assay with the IC50s obtained by the plaque reduction assay showed an acceptable correlation (r2 = 0.473; P = 0.001), suggesting that the flow cytometry assay could substitute for the more labor-intensive, subjective, and time-consuming plaque reduction assay. PMID:9736557

  5. Flow Cytometric Determination of Ganciclovir Susceptibilities of Human Cytomegalovirus Clinical Isolates

    Science.gov (United States)

    McSharry, James M.; Lurain, Nell S.; Drusano, George L.; Landay, Alan; Manischewitz, Jody; Nokta, Mostafa; O’Gorman, Maurice; Shapiro, Howard M.; Weinberg, Adriana; Reichelderfer, Patricia; Crumpacker, Clyde

    1998-01-01

    A flow cytometric assay has been developed for the measurement of susceptibilities to ganciclovir of laboratory strains and clinical isolates of human cytomegalovirus (HCMV). The assay uses fluorochrome-labeled monoclonal antibodies to HCMV immediate-early and late antigens to identify HCMV-infected cells and flow cytometry to detect and quantitate the number of antigen-positive cells. By this assay, the 50 and 90% inhibitory concentrations (IC50 and IC90, respectively) of ganciclovir for the AD169 strain of HCMV were 1.7 and 9.2 μM, respectively, and the IC50 for the ganciclovir-resistant D6/3/1 derivative of the AD169 strain was greater than 12 μM. The ganciclovir susceptibilities of 17 HCMV clinical isolates were also determined by flow cytometric analysis of the effect of ganciclovir on late-antigen synthesis in HCMV-infected cells. The average IC50 of ganciclovir for drug-sensitive HCMV clinical isolates was 3.79 μM (±2.60). The plaque-reduction assay for these clinical isolates yielded an average IC50 of 2.80 μM (±1.46). Comparison of the results of the flow cytometry assays with those obtained from the plaque-reduction assays demonstrated acceptable bias and precision. Flow cytometric and plaque-reduction analysis of cells infected with ganciclovir-resistant clinical isolates failed to show a reduction in the percentage of late-antigen-positive cells or PFU, even at 96 μM ganciclovir. The flow cytometric assay for determining ganciclovir susceptibility of HCMV is quantitative, and objective, and potentially automatable, and its results are reproducible among laboratories. PMID:9542916

  6. Stress-inducible alternative translation initiation of human cytomegalovirus latency protein pUL138.

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    Grainger, Lora; Cicchini, Louis; Rak, Michael; Petrucelli, Alex; Fitzgerald, Kerry D; Semler, Bert L; Goodrum, Felicia

    2010-09-01

    We have previously characterized a 21-kDa protein encoded by UL138 (pUL138) as a viral factor inherent to low-passage strains of human cytomegalovirus (HCMV) that is required for latent infection in vitro. pUL138 is encoded on 3.6-, 2.7-, and 1.4-kb 3' coterminal transcripts that are produced during productive and latent infections. pUL138 is encoded at the 3' end of each transcript and is preceded by an extensive 5' sequence (approximately 0.5 to 2.5 kb) containing several putative open reading frames (ORFs). We determined that three putative ORFs upstream of UL138 (UL133, UL135, and UL136) encode proteins. The UL138 transcripts are polycistronic, such that each transcript expresses pUL138 in addition to the most-5' ORF. The upstream coding sequences (CDS) present a significant challenge for the translation of pUL138 in mammalian cells. We hypothesized that sequences 5' of UL138 mediate translation initiation of pUL138 by alternative strategies. Accordingly, a 663-nucloetide (nt) sequence overlapping the UL136 CDS supported expression of a downstream cistron in a bicistronic reporter system. We did not detect cryptic promoter activity or RNA splicing events that could account for downstream cistron expression. These data are consistent with the sequence element functioning as an internal ribosome entry site (IRES). Interestingly, pUL138 expression from the 3.6- and 2.7-kb transcripts was induced by serum stress, which concomitantly inhibited normal cap-dependent translation. Our work suggests that an alternative and stress-inducible strategy of translation initiation ensures expression of pUL138 under a variety of cellular contexts. The UL138 polycistronic transcripts serve to coordinate the expression of multiple proteins, including a viral determinant of HCMV latency.

  7. Human cytomegalovirus interleukin-10 enhances matrigel invasion of MDA-MB-231 breast cancer cells.

    Science.gov (United States)

    Valle Oseguera, Cendy A; Spencer, Juliet V

    2017-01-01

    While some risk factors for breast cancer are well-known, the influence of other factors, particularly virus infection, remains unclear. Human cytomegalovirus (HCMV) is widespread in the general population, and both molecular and epidemiological evidence has indicated links between HCMV and breast cancer. The HCMV protein cmvIL-10 is a potent suppressor of immune function that has also been shown to promote proliferation and migration of breast cancer cells. In this study, the impact of cmvIL-10 on tumor cell invasion through a simulated basement membrane was investigated. MDA-MB-231 breast cancer cells exhibited invasion through a matrigel layer that was significantly enhanced in the presence of either purified cmvIL-10 or supernatants from HCMV-infected cells containing secreted cmvIL-10. Transcriptional profiling revealed that cmvIL-10 altered expression of several genes implicated in metastasis. Exposure to cmvIL-10 resulted in higher MMP-3 mRNA levels, greater protein expression, and increased enzymatic activity. Treatment with cmvIL-10 also increased expression of both urokinase plasminogen receptor (uPAR) and plasminogen activator inhibitor-1 (PAI-1), which can stimulate MMP-3 activity and have previously been identified as poor prognostic markers in breast cancer patients. Finally, MDA-MB-231 cells treated with cmvIL-10 showed significant downregulation of metastasis suppressor 1 (MTSS1), a scaffolding protein that regulates cytoskeletal rearrangements and is frequently lost in metastatic tumors. HCMV, and in particular the secreted viral cytokine, cmvIL-10, can induce cellular changes that facilitate cell migration and invasion. These findings indicate that HCMV may be associated with promoting the malignant spread of breast cancer cells and suggest that antiviral treatment may be a useful complement to chemotherapy in some patients.

  8. Frequency of herpes simplex virus, cytomegalovirus and human papillomavirus DNA in semen.

    Science.gov (United States)

    Aynaud, Olivier; Poveda, Jean-Dominique; Huynh, Bernard; Guillemotonia, Aline; Barrasso, Renzo

    2002-08-01

    Herpes simplex virus (HSV-2) and cytomegalovirus (CMV) infections produce brain damage in the newborn, and human papillomavirus (HPV) plays a role in cervical carcinogenesis. To assess the frequency of herpes virus and HPV in semen and its role in transmission, semen from 111 male partners of women with histologically-detected genital HPV infection was analysed for HSV, CMV and HPV infection. We used cell culture to detect HSV and CMV, and polymerase chain reaction (PCR) for HPV. Virological findings in the sperm were correlated to the presence or absence of HPV-associated genital lesions and to the viral type. Viral cultures yielded HSV-2 DNA in 9% and CMV DNA in 6.3% of cases. No correlation was established with a history of clinically apparent infection for HSV. HPV-DNA was detected in 23.4% of semen by PCR techniques: in 48% of subjects with urethral lesions, in 22% of patients with penile lesions, in 2% of patients without HPV-associated lesions. HPV-DNA type 16 was detected in 3.6% of cases. Patients with a positive HPV semen sample and penile or urethral lesions had the same HPV type detected in the two specimens. The study shows a high detection of clinically inapparent HSV and CMV, but does not confirm high HPV prevalence in semen from men without detectable lesions. Our study also suggests that the mechanism for semen contamination by HPV is the exfoliation of infected cells from urethral lesions during semen ejaculation, and probably, by abrasion from penile lesions. This could result in the contamination of semen used in assisted reproductive technology.

  9. Quantitative membrane proteomics reveals a role for tetraspanin enriched microdomains during entry of human cytomegalovirus.

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    Kasinath Viswanathan

    Full Text Available Human cytomegalovirus (HCMV depends on and modulates multiple host cell membrane proteins during each stage of the viral life cycle. To gain a global view of the impact of HCMV-infection on membrane proteins, we analyzed HCMV-induced changes in the abundance of membrane proteins in fibroblasts using stable isotope labeling with amino acids (SILAC, membrane fractionation and protein identification by two-dimensional liquid chromatography and tandem mass spectrometry. This systematic approach revealed that CD81, CD44, CD98, caveolin-1 and catenin delta-1 were down-regulated during infection whereas GRP-78 was up-regulated. Since CD81 downregulation was also observed during infection with UV-inactivated virus we hypothesized that this tetraspanin is part of the viral entry process. Interestingly, additional members of the tetraspanin family, CD9 and CD151, were also downregulated during HCMV-entry. Since tetraspanin-enriched microdomains (TEM cluster host cell membrane proteins including known CMV receptors such as integrins, we studied whether TEMs are required for viral entry. When TEMs were disrupted with the cholesterol chelator methyl-β-cylcodextrin, viral entry was inhibited and this inhibition correlated with reduced surface levels of CD81, CD9 and CD151, whereas integrin levels remained unchanged. Furthermore, simultaneous siRNA-mediated knockdown of multiple tetraspanins inhibited viral entry whereas individual knockdown had little effect suggesting essential, but redundant roles for individual tetraspanins during entry. Taken together, our data suggest that TEM act as platforms for receptors utilized by HCMV for entry into cells.

  10. Inhibition of Ganciclovir-Susceptible and -Resistant Human Cytomegalovirus Clinical Isolates by the Benzimidazole l-Riboside 1263W94

    Science.gov (United States)

    McSharry, James J.; McDonough, Ann; Olson, Betty; Talarico, Christine; Davis, Michele; Biron, Karen K.

    2001-01-01

    The average 50% inhibitory concentration (IC50) values for AD169 were 0.22 ± 0.09 μM 1263W94 and 5.36 ± 0.12 μM ganciclovir. For 35 human cytomegalovirus (HCMV) clinical isolates the average IC50 was 0.42 ± 0.09 μM 1263W94, and for 26 ganciclovir-susceptible HCMV clinical isolates the average IC50 was 3.78 ± 1.62 μM ganciclovir. Nine HCMV clinical isolates that were resistant to ganciclovir were completely susceptible to 1263W94. PMID:11687477

  11. Mutations in the carboxyl-terminal hydrophobic sequence of human cytomegalovirus glycoprotein B alter transport and protein chaperone binding.

    OpenAIRE

    Zheng, Z.; Maidji, E; Tugizov, S; Pereira, L

    1996-01-01

    Human cytomegalovirus glycoprotein B (gB) plays a role in the fusion of the virion envelope with the host cell membrane and in syncytium formation in infected cells. Hydrophobic sequences at the carboxyl terminus, amino acids (aa) 714 to 771, anchor gB in the lipid bilayer, but the unusual length of this domain suggests that it may serve another role in gB structure. To explore the function(s) of this region, we deleted aa 717 to 747 (gB deltaI mutation), aa 751 to 771 (gB deltaII mutation), ...

  12. FIBROBLASTS, EPITHELIAL-CELLS, ENDOTHELIAL-CELLS AND SMOOTH-MUSCLE CELLS ARE MAJOR TARGETS OF HUMAN CYTOMEGALOVIRUS-INFECTION IN LUNG AND GASTROINTESTINAL TISSUES

    NARCIS (Netherlands)

    SINZGER, C; GREFTE, A; PLACHTER, B; GOUW, ASH; THE, TH; JAHN, G

    High titre replication of human cytomegalovirus (HCMV) in cell culture is restricted to primary human fibroblasts. During acute infection in vivo, HCMV nucleic acids and antigens have been found in various organs. Using only morphological criteria, inconsistent data have been reported about the cell

  13. Transient Oral Human Cytomegalovirus Infections Indicate Inefficient Viral Spread from Very Few Initially Infected Cells.

    Science.gov (United States)

    Mayer, Bryan T; Krantz, Elizabeth M; Swan, David; Ferrenberg, James; Simmons, Karen; Selke, Stacy; Huang, Meei-Li; Casper, Corey; Corey, Lawrence; Wald, Anna; Schiffer, Joshua T; Gantt, Soren

    2017-06-15

    Cytomegalovirus (CMV) is acquired by the oral route in children, and primary infection is associated with abundant mucosal replication, as well as the establishment of latency in myeloid cells that results in lifelong infection. The efficiency of primary CMV infection in humans following oral exposure, however, is unknown. We consistently detected self-limited, low-level oral CMV shedding events, which we termed transient CMV infections, in a prospective birth cohort of 30 highly exposed CMV-uninfected infants. We estimated the likelihood of transient oral CMV infections by comparing their observed frequency to that of established primary infections, characterized by persistent high-level shedding, viremia, and seroconversion. We developed mathematical models of viral dynamics upon initial oral CMV infection and validated them using clinical shedding data. Transient infections comprised 76 to 88% of oral CMV shedding events. For this high percentage of transient infections to occur, we identified two mathematical prerequisites: a very small number of initially infected oral cells (1 to 4) and low viral infectivity (<1.5 new cells infected/cell). These observations indicate that oral CMV infection in infants typically begins with a single virus that spreads inefficiently to neighboring cells. Thus, although the incidence of CMV infection is high during infancy, our data provide a mechanistic framework to explain why multiple CMV exposures are typically required before infection is successfully established. These findings imply that a sufficiently primed immune response could prevent CMV from establishing latent infection in humans and support the achievability of a prophylactic CMV vaccine. IMPORTANCE CMV infects the majority of the world's population and is a major cause of birth defects. Developing a vaccine to prevent CMV infection would be extremely valuable but would be facilitated by a better understanding of how natural human CMV infection is acquired. We

  14. Human cytomegalovirus reactivation from latency: validation of a "switch" model in vitro.

    Science.gov (United States)

    Arcangeletti, Maria-Cristina; Vasile Simone, Rosita; Rodighiero, Isabella; De Conto, Flora; Medici, Maria-Cristina; Maccari, Clara; Chezzi, Carlo; Calderaro, Adriana

    2016-10-22

    Human cytomegalovirus (HCMV) is an opportunistic pathogen leading to severe and even fatal diseases in 'at-risk' categories of individuals upon primary infection or the symptomatic reactivation of the endogenous virus. The mechanisms which make the virus able to reactivate from latency are still matter of intense study. However, the very low number of peripheral blood monocytes (an important latent virus reservoir) harbouring HCMV DNA makes it very difficult to obtain adequate viral quantities to use in such studies. Thus, the aim of the present study was to demonstrate the usefulness of human THP-1 monocytes, mostly employed as HCMV latent or lytic infection system, as a reactivation model. THP-1 monocytes were infected with HCMV TB40E strain (latency model) at multiplicities of infection (MOI) of 0.5, 0.25 or 0.125. After infection, THP-1 aliquots were differentiated into macrophages (reactivation model). Infections were carried out for 30 h, 4, 6 and 7 days. Viral DNA evaluation was performed with viable and UV-inactivated virus by q-Real-Time PCR. RNA extracted from latency and reactivation models at 7 days post-infection (p.i.) was subjected to RT-PCR to analyse viral latency and lytic transcripts. To perform viral progeny analysis and titration, the culture medium from infected THP-1 latency and reactivation models (7 days p.i.) was used to infect human fibroblasts; it was also checked for the presence of exosomes. For viral progeny analysis experiments, the Towne strain was also used. Our results showed that, while comparable TB40E DNA amounts were present in both latent and reactivation models at 30 h p.i., gradually increased quantities of viral DNA were only evident in the latter model at 4, 6, 7 days p.i.. The completion of the lytic cycle upon reactivation was also proved by the presence of HCMV lytic transcripts and an infectious viral yield at 7 days p.i. Our data demonstrate the effectiveness of THP-1 cells as a "switch" model for studying the

  15. Immediate-early gene region of human cytomegalovirus trans-activates the promoter of human immunodeficiency virus

    Energy Technology Data Exchange (ETDEWEB)

    Davis, M.G.; Kenney, S.C.; Kamine, J.; Pagano, J.S.; Huang, E.S.

    1987-12-01

    Almost all homosexual patients with acquired immunodeficiency syndrome are also actively infected with human cytomegalovirus (HCMV). The authors have hypothesized that an interaction between HCMV and human immunodeficiency virus (HIV), the agent that causes acquired immunodeficiency syndrome, may exist at a molecular level and contribute to the manifestations of HIV infection. In this report, they demonstrate that the immediate-early gene region of HCMV, in particular immediate-early region 2, trans-activates the expression of the bacterial gene chloramphenicol acetyltransferase that is fused to the HIV long terminal repeat and carried by plasmid pHIV-CAT. The HCMV immediate-early trans-activator increases the level of mRNA from the plamid pHIV-CAT. The sequences of HIV that are responsive to trans-activation by the HDMV immediate-early region are distinct from HIV sequences that are required for response to the HIV tat. The stimulation of HIV gene expression by HDMV gene functions could enhance the consequences of HIV infection in persons with previous or concurrent HCMV infection.

  16. Quantitative analysis of human herpesvirus-6 and human cytomegalovirus in blood and saliva from patients with acute leukemia.

    Science.gov (United States)

    Nefzi, Faten; Ben Salem, Nabil Abid; Khelif, Abderrahim; Feki, Salma; Aouni, Mahjoub; Gautheret-Dejean, Agnès

    2015-03-01

    Human herpesvirus-6 (HHV-6) and human cytomegalovirus (HCMV) DNAs were quantified by real-time PCR assays in blood and saliva obtained from 50 patients with acute leukemia at the time of diagnosis (50 of each matrix), aplasia (65 of each matrix), remission (55 of each matrix), and relapse (20 of each matrix) to evaluate which biological matrix was more suitable to identify a viral reactivation, search for a possible link between HHV-6 and HCMV reactivations, and evaluate the relations between viral loads and count of different leukocyte types in blood. The median HHV-6 loads were 136; 219; 226, and 75 copies/million cells in blood at diagnosis, aplasia, remission and relapse, respectively. The HCMV loads were 193 and 317 copies/million cells in blood at diagnosis and remission. In the saliva samples, the HHV-6 loads were 22,165; 15,238; 30,214, and 17,454 copies/million cells at diagnosis, aplasia, remission, and relapse, respectively. The HCMV loads were 8,991; 1,461; 2,980, and 4,283 copies/million cells at diagnosis, aplasia, remission, and relapse, respectively. The HHV-6 load in the blood was correlated to the counts of polymorphonuclear leukocytes (R(2)  = 0.5; P blood in the detection of HHV-6 or HCMV reactivations. The HHV-6 and HCMV reactivations were linked only in saliva. © 2014 Wiley Periodicals, Inc.

  17. Human cytomegalovirus infection leads to elevated levels of transplant arteriosclerosis in a humanized mouse aortic xenograft model.

    Science.gov (United States)

    Abele-Ohl, S; Leis, M; Wollin, M; Mahmoudian, S; Hoffmann, J; Müller, R; Heim, C; Spriewald, B M; Weyand, M; Stamminger, T; Ensminger, S M

    2012-07-01

    Recent findings emphasized an important role of human cytomegalovirus (HCMV) infection in the development of transplant arteriosclerosis. Therefore, the aim of this study was to develop a human peripheral blood lymphocyte (hu-PBL)/Rag-2(-/-) γc(-/-) mouse-xenograft-model to investigate both immunological as well as viral effector mechanisms in the progression of transplant arteriosclerosis. For this, sidebranches from the internal mammary artery were recovered during coronary artery bypass graft surgery, tissue-typed and infected with HCMV. Then, size-matched sidebranches were implanted into the infrarenal aorta of Rag-2(-/-) γc(-/-) mice. The animals were reconstituted with human peripheral blood mononuclear cells (PBMCs) 7 days after transplantation. HCMV-infection was confirmed by Taqman-PCR and immunofluorescence analyses. Arterial grafts were analyzed by histology on day 40 after transplantation. PBMC-reconstituted Rag-2(-/-) γc(-/-) animals showed splenic chimerism levels ranging from 1-16% human cells. After reconstitution, Rag-2(-/-) γc(-/-) mice developed human leukocyte infiltrates in their grafts and vascular lesions that were significantly elevated after infection. Cellular infiltration revealed significantly increased ICAM-1 and PDGF-R-β expression after HCMV-infection of the graft. Arterial grafts from unreconstituted Rag-2(-/-) γc(-/-) recipients showed no vascular lesions. These data demonstrate a causative relationship between HCMV-infection as an isolated risk factor and the development of transplant-arteriosclerosis in a humanized mouse arterial-transplant-model possibly by elevated ICAM-1 and PDGF-R-β expression. © Copyright 2012 The American Society of Transplantation and the American Society of Transplant Surgeons.

  18. Efficacy and Mechanism of Action of Low Dose Emetine against Human Cytomegalovirus.

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    Rupkatha Mukhopadhyay

    2016-06-01

    Full Text Available Infection with human cytomegalovirus (HCMV is a threat for pregnant women and immunocompromised hosts. Although limited drugs are available, development of new agents against HCMV is desired. Through screening of the LOPAC library, we identified emetine as HCMV inhibitor. Additional studies confirmed its anti-HCMV activities in human foreskin fibroblasts: EC50-40±1.72 nM, CC50-8±0.56 μM, and selectivity index of 200. HCMV inhibition occurred after virus entry, but before DNA replication, and resulted in decreased expression of viral proteins. Synergistic virus inhibition was achieved when emetine was combined with ganciclovir. In a mouse CMV (MCMV model, emetine was well-tolerated, displayed long half-life, preferential distribution to tissues over plasma, and effectively suppressed MCMV. Since the in vitro anti-HCMV activity of emetine decreased significantly in low-density cells, a mechanism involving cell cycle regulation was suspected. HCMV inhibition by emetine depended on ribosomal processing S14 (RPS14 binding to MDM2, leading to disruption of HCMV-induced MDM2-p53 and MDM2-IE2 interactions. Irrespective of cell density, emetine induced RPS14 translocation into the nucleus during infection. In infected high-density cells, MDM2 was available for interaction with RPS14, resulting in disruption of MDM2-p53 interaction. However, in low-density cells the pre-existing interaction of MDM2-p53 could not be disrupted, and RPS14 could not interact with MDM2. In high-density cells the interaction of MDM2-RPS14 resulted in ubiquitination and degradation of RPS14, which was not observed in low-density cells. In infected-only or in non-infected emetine-treated cells, RPS14 failed to translocate into the nucleus, hence could not interact with MDM2, and was not ubiquitinated. HCMV replicated similarly in RPS14 knockdown or control cells, but emetine did not inhibit virus replication in the former cell line. The interaction of MDM2-p53 was maintained

  19. Human Cytomegalovirus Vaccine Based on the Envelope gH/gL Pentamer Complex

    Science.gov (United States)

    Martinez, Joy; Campo, John; Johnson, Erica; Flechsig, Christin; Newell, Maegan; Tran, Elaine; Ortiz, Jose; La Rosa, Corinna; Herrmann, Andreas; Longmate, Jeff; Chakraborty, Rana; Barry, Peter A.; Diamond, Don J.

    2014-01-01

    Human Cytomegalovirus (HCMV) utilizes two different pathways for host cell entry. HCMV entry into fibroblasts requires glycoproteins gB and gH/gL, whereas HCMV entry into epithelial and endothelial cells (EC) requires an additional complex composed of gH, gL, UL128, UL130, and UL131A, referred to as the gH/gL-pentamer complex (gH/gL-PC). While there are no established correlates of protection against HCMV, antibodies are thought to be important in controlling infection. Neutralizing antibodies (NAb) that prevent gH/gL-PC mediated entry into EC are candidates to be assessed for in vivo protective function. However, these potent NAb are predominantly directed against conformational epitopes derived from the assembled gH/gL-PC. To address these concerns, we constructed Modified Vaccinia Ankara (MVA) viruses co-expressing all five gH/gL-PC subunits (MVA-gH/gL-PC), subsets of gH/gL-PC subunits (gH/gL or UL128/UL130/UL131A), or the gB subunit from HCMV strain TB40/E. We provide evidence for cell surface expression and assembly of complexes expressing full-length gH or gB, or their secretion when the corresponding transmembrane domains are deleted. Mice or rhesus macaques (RM) were vaccinated three times with MVA recombinants and serum NAb titers that prevented 50% infection of human EC or fibroblasts by HCMV TB40/E were determined. NAb responses induced by MVA-gH/gL-PC blocked HCMV infection of EC with potencies that were two orders of magnitude greater than those induced by MVA expressing gH/gL, UL128-UL131A, or gB. In addition, MVA-gH/gL-PC induced NAb responses that were durable and efficacious to prevent HCMV infection of Hofbauer macrophages, a fetal-derived cell localized within the placenta. NAb were also detectable in saliva of vaccinated RM and reached serum peak levels comparable to NAb titers found in HCMV hyperimmune globulins. This vaccine based on a translational poxvirus platform co-delivers all five HCMV gH/gL-PC subunits to achieve robust humoral

  20. Rapid In Vitro Evolution of Human Cytomegalovirus UL56 Mutations That Confer Letermovir Resistance

    OpenAIRE

    Chou, Sunwen

    2015-01-01

    Letermovir (LMV) is an experimental cytomegalovirus terminase inhibitor undergoing phase 3 clinical trials. Viral mutations have been described at UL56 codons 231 to 369 that confer widely variable levels of LMV resistance. In this study, 15 independent experiments propagating an exonuclease mutant viral strain in escalating LMV concentrations replicated 6 of the 7 published UL56 mutations and commonly elicited additional resistance-conferring mutations at UL56 codons 231, 236, 237, 244, 257,...

  1. Human cytomegalovirus-induces cytokine changes in the placenta with implications for adverse pregnancy outcomes.

    Directory of Open Access Journals (Sweden)

    Stuart T Hamilton

    Full Text Available Human cytomegalovirus (CMV infection of the developing fetus can result in adverse pregnancy outcomes including death in utero. Fetal injury results from direct viral cytopathic damage to the CMV-infected fetus, although evidence suggests CMV placental infection may indirectly cause injury to the fetus, possibly via immune dysregulation with placental dysfunction. This study investigated the effects of CMV infection on expression of the chemokine MCP-1 (CCL2 and cytokine TNF-α in placentae from naturally infected stillborn babies, and compared these changes with those found in placental villous explant histocultures acutely infected with CMV ex vivo. Tissue cytokine protein levels were assessed using quantitative immunohistochemistry. CMV-infected placentae from stillborn babies had significantly elevated MCP-1 and TNF-α levels compared with uninfected placentae (p = 0.001 and p = 0.007, which was not observed in placentae infected with other microorganisms (p = 0.62 and p = 0.71 (n = 7 per group. Modelling acute clinical infection using ex vivo placental explant histocultures showed infection with CMV laboratory strain AD169 (0.2 pfu/ml caused significantly elevated expression of MCP-1 and TNF-α compared with uninfected explants (p = 0.0003 and p<0.0001 (n = 25 per group. Explant infection with wild-type Merlin at a tenfold lower multiplicity of infection (0.02 pfu/ml, caused a significant positive correlation between increased explant infection and upregulation of MCP-1 and TNF-α expression (p = 0.0001 and p = 0.017. Cytokine dysregulation has been associated with adverse outcomes of pregnancy, and can negatively affect placental development and function. These novel findings demonstrate CMV infection modulates the placental immune environment in vivo and in a multicellular ex vivo model, suggesting CMV-induced cytokine modulation as a potential initiator and/or exacerbator of placental and fetal injury.

  2. Combined genetic variants of human cytomegalovirus envelope glycoproteins as congenital infection markers.

    Science.gov (United States)

    Arcangeletti, Maria-Cristina; Vasile Simone, Rosita; Rodighiero, Isabella; De Conto, Flora; Medici, Maria-Cristina; Martorana, Davide; Chezzi, Carlo; Calderaro, Adriana

    2015-11-26

    Human cytomegalovirus (HCMV) is still considered to be the main viral cause of birth defects and long-term neurological and sensory sequelae following congenital infection. Several Authors sustain a key role of HCMV envelope glycoproteins, such as gB, gN and gO - mainly involved in cell targeting, viral penetration and spread - as putative virulence factors. The genes coding for these glycoproteins possess hypervariable regions, resulting in a number of genetic variants in circulating clinical strains. Considering that the genetic polymorphisms underlying the specific differences between gB, gN and gO genotypes can influence the ability of HCMV to preferentially target specific host cells, it is very likely that they play an important role in defining HCMV infection outcome. In the present study, we analysed HCMV gB, gN and gO gene polymorphisms in viral strains isolated from paediatric patients with congenital or post-natal infection, to investigate whether specific genetic variants may be associated with congenital infection. The restriction fragment polymorphisms of genes coding for HCMV gB (UL55), gN (UL73) and gO (UL74) were investigated by analysing viral DNA extracted from 40 urine samples of as many paediatric patients with congenital or post-natal HCMV infection. Randomly selected samples were subjected to DNA sequencing and phylogenetic analysis. Statistical analysis was performed using Fisher's exact test to assess the significance of single and combined glycoprotein genotypes frequency distribution. Statistical significance was considered at a P <0.05. While gB genomic variants were quite homogeneously represented in both paediatric groups, the gN4 genotype significantly prevailed in congenitally infected children (89.5 %) vs post-natally infected children (47.6 %), with a predominance of the gN4c variant (47.4 %). A similar trend was observed for gO3 (52.6 % vs 19 %). Concerning genotypes association, a statistically significant (P = 0.037) gN4-gO3

  3. Differences in Growth Properties among Two Human Cytomegalovirus Glycoprotein O Genotypes

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    Julia Kalser

    2017-08-01

    Full Text Available Glycoprotein O (gO of the human cytomegalovirus (HCMV is the critical subunit of the envelope trimer gH/gL/gO as it interacts with platelet-derived growth factor alpha receptor upon fibroblast entry, and triggers gB-mediated fusion for fibroblast and epithelial cell infection. Eight genotypes (GT of the highly polymorphic gO gene are described, yet it is unclear whether the distinct GTs differ in their function. Thus, we aimed to elucidate potential functional differences between two highly diverse gO GTs in an otherwise genomically identical HCMV strain. Therefore, resident gO GT1c sequence of strain TB40-BAC4-luc was entirely replaced by gO GT4 of strain Towne and both, GT1c and GT4 viruses, were investigated for their growth properties in fibroblasts and epithelial cells. In addition, two conserved gO cysteines involved in gH/gL/gO stabilization were mutated to serine either in GT1c (C218S and C343S or GT4 (C216S and C336S and their effects on cell-free infectivity were assessed. GT4 viruses displayed a significantly enhanced epithelial cell tropism and this resulted in higher virus release upon replication in epithelial cells when compared to GT1c viruses. Further, when the two cysteines were individually mutated in gO GT1c no impairment in cell-free infectivity was observed. This, however, was in sharp contrast to gO GT4, in which both of the corresponding cysteine mutations led to a substantial reduction in cell-free infectivity which was even more pronounced upon mutation of GT4-C336 than of GT4-C216. In conclusion, these findings provide evidence that the two highly diverse gO genotypes, GT1c and GT4, differ in their functional properties as revealed by their different infection capacities for epithelial cells and by their different responsiveness to mutation of strictly conserved cysteine residues. Thus, it is likely that the gO heterogeneity influences cell-free infectivity of HCMV also in vivo which may have important implications for

  4. Resistance of human cytomegalovirus to ganciclovir/valganciclovir: a comprehensive review of putative resistance pathways.

    Science.gov (United States)

    Komatsu, Takashi E; Pikis, Andreas; Naeger, Lisa K; Harrington, Patrick R

    2014-01-01

    Human cytomegalovirus (HCMV) is a pathogen that can be life-threatening in immunocompromised individuals. Valganciclovir and its parent drug ganciclovir are currently the principle drugs used for the treatment or prevention of HCMV disease. The development of HCMV resistance to ganciclovir/valganciclovir has been documented in treated patients and is associated with the emergence of amino acid substitutions in the viral proteins pUL97, pUL54 or both. Generally, single amino acid substitutions associated with clinical resistance that alone do not confer decreased ganciclovir susceptibility in cell culture have been disregarded as causative or clinically significant. This review focuses on the analysis and mechanisms of antiviral drug resistance to HCMV. We also conducted a review of publicly available clinical and nonclinical data to construct a comprehensive list of pUL97 and pUL54 amino acid substitutions that are associated with a poor clinical response to the first line therapies ganciclovir and valganciclovir, or associated with reduced HCMV ganciclovir susceptibility in cell culture. Over 40 putative ganciclovir/valganciclovir resistance-associated substitutions were identified in this analysis. These include the commonly reported substitutions M460I/V and C592G in pUL97. There were additional substitutions that are not widely considered as ganciclovir/valganciclovir resistance-associated substitutions, including V466M in pUL97 and E315D in pUL54. Some of these ganciclovir/valganciclovir resistance-associated substitutions may confer cross-resistance to other HCMV therapies, such as cidofovir and foscarnet. Based on this review, we propose that there are more potential HCMV ganciclovir/valganciclovir resistance pathways than generally appreciated. The resulting comprehensive list of putative ganciclovir/valganciclovir resistance-associated substitutions provides a foundation for future investigations to characterize the role of specific substitutions or

  5. [Viral infection of herpes simplex, Epstein-Barr, varicela zoster, human papilloma, cytomegalovirus, or adenovirus are not related to sinonasal adenocarcinomas].

    Science.gov (United States)

    Pérez Escuredo, Jhudit; Llorente, José Luis; Melón, Santiago; de Oña, María; García Martínez, Jorge; Alvarez Marcos, César; Hermsen, Mario

    2007-01-01

    Several types of virus have been implicated in the development of head and neck tumors. However, until now sinonasal adenocarcinomas (ACN) have not been studied. The aim of this study is to screen a series of ACN for the presence of a number of viruses known to play a role in cancer. Viral DNA sequences of herpes simplex virus, Epstein-Barr, varicela zoster, human papilloma, cytomegalovirus, and adenovirus were analysed by PCR in 37 primary ACN. Three tumors (8.1%) were positive for Epstein-Barr virus and 1 case (2.7%) for cytomegalovirus. Viral infections do not seem to play a role in the etiology of ACN.

  6. Cell-cycle-dependent localization of human cytomegalovirus UL83 phosphoprotein in the nucleolus and modulation of viral gene expression in human embryo fibroblasts in vitro.

    Science.gov (United States)

    Arcangeletti, Maria-Cristina; Rodighiero, Isabella; Mirandola, Prisco; De Conto, Flora; Covan, Silvia; Germini, Diego; Razin, Sergey; Dettori, Giuseppe; Chezzi, Carlo

    2011-01-01

    The nucleolus is a multifunctional nuclear compartment widely known to be involved in several cellular processes, including mRNA maturation and shuttling to cytoplasmic sites, control of the cell cycle, cell proliferation, and apoptosis; thus, it is logical that many viruses, including herpesvirus, target the nucleolus in order to exploit at least one of the above-mentioned functions. Recent studies from our group demonstrated the early accumulation of the incoming ppUL83 (pp65), the major tegument protein of human cytomegalovirus (HCMV), in the nucleolus. The obtained results also suggested that a functional relationship might exist between the nucleolar localization of pp65, rRNA synthesis, and the development of the lytic program of viral gene expression. Here we present new data which support the hypothesis of a potentially relevant role of HCMV pp65 and its nucleolar localization for the control of the cell cycle by HCMV (arrest of cell proliferation in G1-G1/S), and for the promotion of viral infection. We demonstrated that, although the incoming pp65 amount in the infected cells appears to be constant irrespective of the cell-cycle phase, its nucleolar accumulation is prominent in G1 and G1/S, but very poor in S or G2/M. This correlates with the observation that only cells in G1 and G1/S support an efficient development of the HCMV lytic cycle. We propose that HCMV pp65 might be involved in regulatory/signaling pathways related to nucleolar functions, such as the cell-cycle control. Co-immunoprecipitation experiments have permitted to identify nucleolin as one of the nucleolar partners of pp65.

  7. Complete Genome Sequence of a Cytomegalovirus Towne-BAC (Bacterial Artificial Chromosome) Isolate Maintained in Escherichia?coli for 10?Years and Then Serially Passaged in Human Fibroblasts

    OpenAIRE

    Brechtel, Teal; Tyner, Molly; Tandon, Ritesh

    2013-01-01

    Here, we present the complete genome sequence of a cytomegalovirus, the Towne-BAC (bacterial artificial chromosome) isolate, which was maintained in bacterial cells for 10?years and then serially passaged in human fibroblasts for 10 passages. A total of 132 nucleotide differences were discovered in the Towne sequence compared to the reference sequence (GenBank accession no. AC146851).

  8. Complete Genome Sequence of a Cytomegalovirus Towne-BAC (Bacterial Artificial Chromosome) Isolate Maintained in Escherichia coli for 10 Years and Then Serially Passaged in Human Fibroblasts.

    Science.gov (United States)

    Brechtel, Teal; Tyner, Molly; Tandon, Ritesh

    2013-09-26

    Here, we present the complete genome sequence of a cytomegalovirus, the Towne-BAC (bacterial artificial chromosome) isolate, which was maintained in bacterial cells for 10 years and then serially passaged in human fibroblasts for 10 passages. A total of 132 nucleotide differences were discovered in the Towne sequence compared to the reference sequence (GenBank accession no. AC146851).

  9. CC and CX3C chemokines differentially interact with the N terminus of the human cytomegalovirus-encoded US28 receptor

    DEFF Research Database (Denmark)

    Casarosa, Paola; Waldhoer, Maria; LiWang, Patricia J

    2005-01-01

    Human cytomegalovirus (HCMV) is the causative agent of life-threatening systemic diseases in immunocompromised patients as well as a risk factor for vascular pathologies, like atherosclerosis, in immunocompetent individuals. HCMV encodes a G-protein-coupled receptor (GPCR), referred to as US28...

  10. THE DNA-BINDING PROTEIN PUL57 OF HUMAN CYTOMEGALOVIRUS IS A MAJOR TARGET ANTIGEN FOR THE IMMUNOGLOBULIN-M ANTIBODY-RESPONSE DURING ACUTE INFECTION

    NARCIS (Netherlands)

    VORNHAGEN, R; HINDERER, W; SONNEBORN, HH; BEIN, G; MATTER, L; THE, TH; JAHN, G; PLACHTER, B

    A small polypeptide from pUL57 of human cytomegalovirus was identified as a major target for the immunoglobulin M antibody response. This antigen seems to be superior to antigenic fragments from pp150 and p52 in the identification of sera from acutely infected patients. It may therefore represent an

  11. Transcriptional activation of the major immediate early transcription unit of human cytomegalovirus by heat-shock, arsenite and protein synthesis inhibitors

    NARCIS (Netherlands)

    Geelen, J. L.; Boom, R.; Klaver, G. P.; Minnaar, R. P.; Feltkamp, M. C.; van Milligen, F. J.; Sol, C. J.; van der Noordaa, J.

    1987-01-01

    In Rat-9G cells several copies of the major immediate early (IE) transcription unit (regions 1 and 2) of the human cytomegalovirus (HCMV) are stably integrated. The cells show a repressed phenotype for IE expression but can be induced by inhibition of protein synthesis. In this report we present

  12. The Seroprevalence of Cytomegalovirus Antibodies among ...

    African Journals Online (AJOL)

    Background: Human cytomegalovirus, otherwise called human herpes virus type 5, is a significant cause of morbidity and mortality in pregnancy, and among immunocompromised patients like recipients of organ transplants. Cytomegalovirus is transmissible via blood transfusion, among other parenteral routes. This study ...

  13. Letermovir and inhibitors of the terminase complex: a promising new class of investigational antiviral drugs against human cytomegalovirus

    OpenAIRE

    Melendez DP; Razonable RR

    2015-01-01

    Dante P Melendez,1,2 Raymund R Razonable1,2 1Division of Infectious Diseases, 2William J von Liebig Center for Transplantation and Clinical Regeneration, Mayo Clinic, Rochester, MN, USA Abstract: Infection with cytomegalovirus is prevalent in immunosuppressed patients. In solid organ transplant and hematopoietic stem cell transplant recipients, cytomegalovirus infection is associated with high morbidity and preventable mortality. Prevention and treatment of cytomegalovirus with currently app...

  14. Human Cytomegalovirus-Encoded Receptor US28 Is Expressed in Renal Allografts and Facilitates Viral Spreading In Vitro.

    Science.gov (United States)

    Lollinga, Wouter T; de Wit, Raymond H; Rahbar, Afsar; Vasse, Gwenda F; Davoudi, Belghis; Diepstra, Arjan; Riezebos-Brilman, Annelies; Harmsen, Martin C; Hillebrands, Jan-Luuk; Söderberg-Naucler, Cecilia; van Son, Willem J; Smit, Martine J; Sanders, Jan-Stephan; van den Born, Jacob

    2017-03-01

    Renal transplantation is the preferred treatment for patients with end-stage renal disease. Human cytomegalovirus (HCMV) activation is associated with decreased renal graft function and survival. Human cytomegalovirus encodes several immune modulatory proteins, including the G protein-coupled receptor US28, which scavenges human chemokines and modulates intracellular signaling. Our aim was to identify the expression and localization of US28 in renal allograft biopsies by immunohistochemistry and determine its role in viral spreading in vitro. Immunohistochemistry revealed US28 in 31 of 34 renal transplant biopsies from HCMV-seropositive donors. Expression was independent of HCMV viremia or IgG serostatus. US28 was predominantly expressed in the cytoplasm of vascular smooth muscle cells (VSMCs) and tubular epithelial cells, with a median positivity of 20% and 40%, respectively. Also, US28-positive cells were present within arterial neointima. In contrast to US28, HCMV-encoded immediate early antigen was detected in less than 5% of VSMCs, tubular epithelial cells, interstitial endothelium, interstitial inflammatory infiltrates, and glomerular cells.Primary VSMCs were infected with green fluorescent protein-tagged wild type or US28-deficient HCMV. The viral spreading of US28-deficient HCMV, via culture medium or cell-to-cell transmission, was significantly impeded as shown by green fluorescent protein (ie, infected) cell quantification and quantitative real-time polymerase chain reaction. Additionally, the number and size of foci was smaller. In summary, HCMV-encoded US28 was detected in renal allografts from HCMV-positive donors independent of viremia and serostatus. Also, US28 facilitates HCMV spreading in VSMCs in vitro. Because the vasculature is affected in chronic renal transplant dysfunction, US28 may provide a potential target for therapeutic intervention.

  15. Antagonistic determinants controlling replicative and latent states of human cytomegalovirus infection.

    Science.gov (United States)

    Umashankar, Mahadevaiah; Rak, Michael; Bughio, Farah; Zagallo, Patricia; Caviness, Katie; Goodrum, Felicia D

    2014-06-01

    The mechanisms by which viruses persist and particularly those by which viruses actively contribute to their own latency have been elusive. Here we report the existence of opposing functions encoded by genes within a polycistronic locus of the human cytomegalovirus (HCMV) genome that regulate cell type-dependent viral fates: replication and latency. The locus, referred to as the UL133-UL138 (UL133/8) locus, encodes four proteins, pUL133, pUL135, pUL136, and pUL138. As part of the ULb' region of the genome, the UL133/8 locus is lost upon serial passage of clinical strains of HCMV in cultured fibroblasts and is therefore considered dispensable for replication in this context. Strikingly, we could not reconstitute infection in permissive fibroblasts from bacterial artificial chromosome clones of the HCMV genome where UL135 alone was disrupted. The loss of UL135 resulted in complex phenotypes and could ultimately be overcome by infection at high multiplicities. The requirement for UL135 but not the entire locus led us to hypothesize that another gene in this locus suppressed virus replication in the absence of UL135. The defect associated with the loss of UL135 was largely rescued by the additional disruption of the UL138 latency determinant, indicating a requirement for UL135 for virus replication when UL138 is expressed. In the CD34(+) hematopoietic progenitor model of latency, viruses lacking only UL135 were defective for viral genome amplification and reactivation. Taken together, these data indicate that UL135 and UL138 comprise a molecular switch whereby UL135 is required to overcome UL138-mediated suppression of virus replication to balance states of latency and reactivation. Mechanisms by which viruses persist in their host remain one of the most poorly understood phenomena in virology. Herpesviruses, including HCMV, persist in an incurable, latent state that has profound implications for immunocompromised individuals, including transplant patients. Further

  16. The ND10 Component Promyelocytic Leukemia Protein Acts as an E3 Ligase for SUMOylation of the Major Immediate Early Protein IE1 of Human Cytomegalovirus.

    Science.gov (United States)

    Reuter, Nina; Schilling, Eva-Maria; Scherer, Myriam; Müller, Regina; Stamminger, Thomas

    2017-05-15

    Previous studies identified the nuclear domain 10 (ND10) components promyelocytic leukemia protein (PML), hDaxx, and Sp100 as factors of an intrinsic immune response against human cytomegalovirus (HCMV). This antiviral function of ND10, however, is antagonized by viral effector proteins like IE1p72, which induces dispersal of ND10. Furthermore, we have shown that both major immediate early proteins of HCMV, IE1p72 and IE2p86, transiently colocalize with ND10 subnuclear structures and undergo modification by the covalent attachment of SUMO. Since recent reports indicate that PML acts as a SUMO E3 ligase, we asked whether the SUMOylation of IE1p72 and IE2p86 is regulated by PML. To address this, PML-depleted fibroblasts, as well as cells overexpressing individual PML isoforms, were infected with HCMV. Western blot experiments revealed a clear correlation between the degree of IE1p72 SUMO conjugation and the abundance of PML. On the other hand, the SUMOylation of IE2p86 was not affected by PML. By performing in vitro SUMOylation assays, we were able to provide direct evidence that IE1p72 is a substrate for PML-mediated SUMOylation. Interestingly, disruption of the RING finger domain of PML, which is proposed to confer SUMO E3 ligase activity, abolished PML-induced SUMOylation of IE1p72. In contrast, IE1p72 was still efficiently SUMO modified by a SUMOylation-defective PML mutant, indicating that intact ND10 bodies are not necessary for this effect. Thus, this is the first report that the E3 ligase PML is capable of stimulating the SUMOylation of a viral protein which is supposed to serve as a cellular mechanism to compromise specific functions of IE1p72.IMPORTANCE The major immediate early proteins of human cytomegalovirus, termed IE1p72 and IE2p86, have previously been shown to undergo posttranslational modification by covalent coupling to SUMO moieties at specific lysine residues. However, the enzymatic activities that are responsible for this modification have not

  17. Use of Recombination-Mediated Genetic Engineering for Construction of Rescue Human Cytomegalovirus Bacterial Artificial Chromosome Clones

    Directory of Open Access Journals (Sweden)

    Kalpana Dulal

    2012-01-01

    Full Text Available Bacterial artificial chromosome (BAC technology has contributed immensely to manipulation of larger genomes in many organisms including large DNA viruses like human cytomegalovirus (HCMV. The HCMV BAC clone propagated and maintained inside E. coli allows for accurate recombinant virus generation. Using this system, we have generated a panel of HCMV deletion mutants and their rescue clones. In this paper, we describe the construction of HCMV BAC mutants using a homologous recombination system. A gene capture method, or gap repair cloning, to seize large fragments of DNA from the virus BAC in order to generate rescue viruses, is described in detail. Construction of rescue clones using gap repair cloning is highly efficient and provides a novel use of the homologous recombination-based method in E. coli for molecular cloning, known colloquially as recombineering, when rescuing large BAC deletions. This method of excising large fragments of DNA provides important prospects for in vitro homologous recombination for genetic cloning.

  18. The carboxyl terminus of human cytomegalovirus-encoded 7 transmembrane receptor US28 camouflages agonism by mediating constitutive endocytosis

    DEFF Research Database (Denmark)

    Waldhoer, Maria; Casarosa, Paola; Rosenkilde, Mette M

    2003-01-01

    US28 is one of four 7 transmembrane (7TM) chemokine receptors encoded by human cytomegalovirus and has been shown to both signal and endocytose in a ligand-independent, constitutively active manner. Here we show that the constitutive activity and constitutive endocytosis properties of US28...... that the cytoplasmic tail domain of US28 per se regulates receptor endocytosis, independent of the signaling ability of the core domain of US28. The constitutive endocytic property of the US28 c-tail was transposable to other 7TM receptors, the herpes virus 8-encoded ORF74 and the tachykinin NK1 receptor (ORF74-US28......-ctail and NK1-US28-ctail). Deletion of the US28 C terminus resulted in reduced constitutive endocytosis and consequently enhanced signaling capacity of all receptors tested as assessed by inositol phosphate turnover, NF-kappa B, and cAMP-responsive element-binding protein transcription assays. We...

  19. Human cytomegalovirus (CMV) susceptibility to currently approved antiviral drugs does not impact on CMV terminase complex polymorphism.

    Science.gov (United States)

    Pilorgé, Léa; Burrel, Sonia; Aït-Arkoub, Zaïna; Agut, Henri; Boutolleau, David

    2014-11-01

    Currently approved anti-human cytomegalovirus (CMV) drugs, all targeting the viral DNA polymerase, are associated with significant toxicities and emergence of drug resistance. In this context, CMV terminase complex constitutes a promising target for novel antiviral compounds. In this study, we describe the low natural polymorphism (interstrain identity >97.7% at both nucleotide and amino acid levels) of the terminase subunits pUL56 and pUL89, and the portal protein pUL104, among 63 CMV clinical strains, and we show that the CMV resistance profile to current DNA polymerase inhibitors has no impact on the natural polymorphism of CMV terminase complex. These results support the idea that both CMV clinical strains exhibiting either susceptibility or resistance to current CMV DNA polymerase inhibitors are comparably sensitive to novel inhibitors of CMV terminase complex, such as letermovir. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. Good oral absorption prediction on non-nucleoside benzothiadiazine dioxide human cytomegalovirus inhibitors using combined chromatographic and neuronal network techniques.

    Science.gov (United States)

    Gil, Carmen; Dorronsoro, Isabel; Castro, Ana; Martinez, Ana

    2005-04-01

    The current drugs available against human cytomegalovirus (HCMV) suffer from a number of shortcomings such as toxic side effect, poor bioavailability and/or risk for emergence of drug-resistance virus strains. Due to these limitations, the development of new drugs against HCMV is of great interest. Taking into account the therapeutic potential of benzothiadiazines dioxides (BTD) derivatives, it is most important to know their oral bioavailability because all the current clinical drugs are poorly absorbed. In this work, the utility of CODES neural networks and biopartitioning micellar chromatography (BMC) in predicting pharmacokinetic properties has been used to estimate the oral absorption of BTD derivatives and their efficacy has been verified. The results indicate higher values for BTD derivatives than the currently licensed anti-viral agents.

  1. Interleukin-2 from Adaptive T Cells Enhances Natural Killer Cell Activity against Human Cytomegalovirus-Infected Macrophages.

    Science.gov (United States)

    Wu, Zeguang; Frascaroli, Giada; Bayer, Carina; Schmal, Tatjana; Mertens, Thomas

    2015-06-01

    Control of human cytomegalovirus (HCMV) requires a continuous immune surveillance, thus HCMV is the most important viral pathogen in severely immunocompromised individuals. Both innate and adaptive immunity contribute to the control of HCMV. Here, we report that peripheral blood natural killer cells (PBNKs) from HCMV-seropositive donors showed an enhanced activity toward HCMV-infected autologous macrophages. However, this enhanced response was abolished when purified NK cells were applied as effectors. We demonstrate that this enhanced PBNK activity was dependent on the interleukin-2 (IL-2) secretion of CD4(+) T cells when reexposed to the virus. Purified T cells enhanced the activity of purified NK cells in response to HCMV-infected macrophages. This effect could be suppressed by IL-2 blocking. Our findings not only extend the knowledge on the immune surveillance in HCMV-namely, that NK cell-mediated innate immunity can be enhanced by a preexisting T cell antiviral immunity-but also indicate a potential clinical implication for patients at risk for severe HCMV manifestations due to immunosuppressive drugs, which mainly suppress IL-2 production and T cell responsiveness. Human cytomegalovirus (HCMV) is never cleared by the host after primary infection but instead establishes a lifelong latent infection with possible reactivations when the host's immunity becomes suppressed. Both innate immunity and adaptive immunity are important for the control of viral infections. Natural killer (NK) cells are main innate effectors providing a rapid response to virus-infected cells. Virus-specific T cells are the main adaptive effectors that are critical for the control of the latent infection and limitation of reinfection. In this study, we found that IL-2 secreted by adaptive CD4(+) T cells after reexposure to HCMV enhances the activity of NK cells in response to HCMV-infected target cells. This is the first direct evidence that the adaptive T cells can help NK cells to act

  2. Rapid quantitative PCR assays for the simultaneous detection of herpes simplex virus, varicella zoster virus, cytomegalovirus, Epstein-Barr virus, and human herpesvirus 6 DNA in blood and other clinical specimens

    NARCIS (Netherlands)

    Engelmann, I.; Petzold, D. R.; Kosinska, A.; Hepkema, B. G.; Schulz, T. F.; Heim, A.

    Rapid diagnosis of human herpesvirus primary infections or reactivations is facilitated by quantitative PCRs. Quantitative PCR assays with a standard thermal cycling profile permitting simultaneous detection of herpes simplex virus (HSV), varicella zoster virus (VZV), cytomegalovirus (CMV),

  3. Letermovir and inhibitors of the terminase complex: a promising new class of investigational antiviral drugs against human cytomegalovirus

    Directory of Open Access Journals (Sweden)

    Melendez DP

    2015-08-01

    Full Text Available Dante P Melendez,1,2 Raymund R Razonable1,2 1Division of Infectious Diseases, 2William J von Liebig Center for Transplantation and Clinical Regeneration, Mayo Clinic, Rochester, MN, USA Abstract: Infection with cytomegalovirus is prevalent in immunosuppressed patients. In solid organ transplant and hematopoietic stem cell transplant recipients, cytomegalovirus infection is associated with high morbidity and preventable mortality. Prevention and treatment of cytomegalovirus with currently approved antiviral drugs is often associated with side effects that sometimes preclude their use. Moreover, cytomegalovirus has developed mutations that confer resistance to standard antiviral drugs. During the last decade, there have been calls to develop novel antiviral drugs that could provide better options for prevention and treatment of cytomegalovirus. Letermovir (AIC246 is a highly specific antiviral drug that is currently undergoing clinical development for the management of cytomegalovirus infection. It acts by inhibiting the viral terminase complex. Letermovir is highly potent in vitro and in vivo against cytomegalovirus. Because of a distinct mechanism of action, it does not exhibit cross-resistance with other antiviral drugs. It is predicted to be active against strains that are resistant to ganciclovir, foscarnet, and cidofovir. To date, early-phase clinical trials suggest a very low incidence of adverse effects. Herein, we present a comprehensive review on letermovir, from its postulated novel mechanism of action to the results of most recent clinical studies. Keywords: cytomegalovirus, letermovir, AIC246, terminase, antivirals, transplantation 

  4. SPAM-8, a mouse-human heteromyeloma fusion partner in the production of human monoclonal antibodies. Establishment of a human monoclonal antibody against cytomegalovirus.

    Science.gov (United States)

    Gustafsson, B; Jondal, M; Sundqvist, V A

    1991-01-01

    A heteromyeloma (mouse x human) cell line (SPAM-8) was produced by fusing mouse myeloma cells (SP2/0) with human peripheral blood lymphocytes. The cells were sensitive to aminopterin and resistant to ouabain. The cells showed a doubling time of about 19 hours and a cloning efficiency of 0.8 cells/well (to obtain growth in 50% of wells seeded) using mouse thymocytes as feeder cells. The number of chromosomes was about 86 and 1% of the total DNA was of human origin. Fusion of SPAM-8 cells with lymphocytes prepared from human spleens resulted in approximately one hybridoma per 10(5) seeded lymphocytes. A trioma (human x [mouse x human]) cell line was established by fusing cells of an Epstein-Barr virus-transformed B cell line with SPAM-8 cells. The trioma cells produced antibodies (IgG1, K) against cytomegalovirus, in a concentration of 7 micrograms/ml in spent medium, over a period of six months of continuous culture. The results obtained indicate that the heteromyeloma SPAM-8 may be used as a fusion partner in the production of human monoclonal antibodies.

  5. Modulatory effect of rRNA synthesis and ppUL83 nucleolar compartmentalization on human cytomegalovirus gene expression in vitro.

    Science.gov (United States)

    Arcangeletti, Maria-Cristina; Rodighiero, Isabella; De Conto, Flora; Gatti, Rita; Orlandini, Guido; Ferraglia, Francesca; Motta, Federica; Covan, Silvia; Razin, Sergey V; Dettori, Giuseppe; Chezzi, Carlo

    2009-10-01

    The nucleolus is a nuclear domain involved in the biogenesis of ribosomes, as well as in many other important cellular regulatory activities, such as cell cycle control and mRNA processing. Many viruses, including herpesviruses, are known to exploit the nucleolar compartment during their replication cycle. In a previous study, we demonstrated the preferential targeting and accumulation of the human cytomegalovirus (HCMV) UL83 phosphoprotein (pp65) to the nucleolar compartment and, in particular, to the nucleolar matrix of lytically infected fibroblasts; such targeting was already evident at very early times after infection. Here we have investigated the possible effects of rRNA synthesis inhibition upon the development of HCMV lytic infection, by using either actinomycin D or cisplatin at low concentrations, that are known to selectively inhibit RNA polymerase I activity, whilst leaving RNA polymerase II function unaffected. Following the inhibition of rRNA synthesis by either of the agents used, we observed a significant redistribution of nucleolar proteins within the nucleoplasm and a simultaneous depletion of viral pp65 from the nucleolus; this effect was highly evident in both unextracted cells and in nuclear matrices in situ. Of particular interest, even a brief suppression of rRNA synthesis resulted in a very strong inhibition of the progression of HCMV infection, as was concluded from the absence of accumulation of HCMV major immediate-early proteins within the nucleus of infected cells. These data suggest that a functional relationship might exist between rRNA synthesis, pp65 localization to the nucleolar matrix and the normal development of HCMV lytic infection. (c) 2009 Wiley-Liss, Inc.

  6. Human Cytomegalovirus IE1 Protein Elicits a Type II Interferon-Like Host Cell Response That Depends on Activated STAT1 but Not Interferon-γ

    Science.gov (United States)

    Knoblach, Theresa; Grandel, Benedikt; Seiler, Jana

    2011-01-01

    Human cytomegalovirus (hCMV) is a highly prevalent pathogen that, upon primary infection, establishes life-long persistence in all infected individuals. Acute hCMV infections cause a variety of diseases in humans with developmental or acquired immune deficits. In addition, persistent hCMV infection may contribute to various chronic disease conditions even in immunologically normal people. The pathogenesis of hCMV disease has been frequently linked to inflammatory host immune responses triggered by virus-infected cells. Moreover, hCMV infection activates numerous host genes many of which encode pro-inflammatory proteins. However, little is known about the relative contributions of individual viral gene products to these changes in cellular transcription. We systematically analyzed the effects of the hCMV 72-kDa immediate-early 1 (IE1) protein, a major transcriptional activator and antagonist of type I interferon (IFN) signaling, on the human transcriptome. Following expression under conditions closely mimicking the situation during productive infection, IE1 elicits a global type II IFN-like host cell response. This response is dominated by the selective up-regulation of immune stimulatory genes normally controlled by IFN-γ and includes the synthesis and secretion of pro-inflammatory chemokines. IE1-mediated induction of IFN-stimulated genes strictly depends on tyrosine-phosphorylated signal transducer and activator of transcription 1 (STAT1) and correlates with the nuclear accumulation and sequence-specific binding of STAT1 to IFN-γ-responsive promoters. However, neither synthesis nor secretion of IFN-γ or other IFNs seems to be required for the IE1-dependent effects on cellular gene expression. Our results demonstrate that a single hCMV protein can trigger a pro-inflammatory host transcriptional response via an unexpected STAT1-dependent but IFN-independent mechanism and identify IE1 as a candidate determinant of hCMV pathogenicity. PMID:21533215

  7. Human cytomegalovirus gH stability and trafficking are regulated by ER-associated degradation and transmembrane architecture.

    Science.gov (United States)

    Gardner, Thomas J; Hernandez, Rosmel E; Noriega, Vanessa M; Tortorella, Domenico

    2016-03-30

    The prototypic betaherpesvirus human cytomegalovirus (CMV) establishes life-long persistence within its human host. While benign in healthy individuals, CMV poses a significant threat to the immune compromised, including transplant recipients and neonates. The CMV glycoprotein complex gH/gL/gO mediates infection of fibroblasts, and together with the gH/gL/UL128/130/131 a pentameric complex permits infection of epithelial, endothethial, and myeloid cells. Given the central role of the gH/gL complex during infection, we were interested in studying cellular trafficking of the gH/gL complex through generation of human cells that stably express gH and gL. When expressed alone, CMV gH and gL were degraded through the ER-associated degradation (ERAD) pathway. However, co-expression of these proteins stabilized the polypeptides and enhanced their cell-surface expression. To further define regulatory factors involved in gH/gL trafficking, a CMV gH chimera in which the gH transmembrane and cytoplasmic tail were replaced with that of human CD4 protein permitted cell surface gH expression in absence of gL. We thus demonstrate the ability of distinct cellular processes to regulate the trafficking of viral glycoproteins. Collectively, the data provide insight into the processing and trafficking requirements of CMV envelope protein complexes and provide an example of the co-opting of cellular processes by CMV.

  8. Letermovir and inhibitors of the terminase complex: a promising new class of investigational antiviral drugs against human cytomegalovirus.

    Science.gov (United States)

    Melendez, Dante P; Razonable, Raymund R

    2015-01-01

    Infection with cytomegalovirus is prevalent in immunosuppressed patients. In solid organ transplant and hematopoietic stem cell transplant recipients, cytomegalovirus infection is associated with high morbidity and preventable mortality. Prevention and treatment of cytomegalovirus with currently approved antiviral drugs is often associated with side effects that sometimes preclude their use. Moreover, cytomegalovirus has developed mutations that confer resistance to standard antiviral drugs. During the last decade, there have been calls to develop novel antiviral drugs that could provide better options for prevention and treatment of cytomegalovirus. Letermovir (AIC246) is a highly specific antiviral drug that is currently undergoing clinical development for the management of cytomegalovirus infection. It acts by inhibiting the viral terminase complex. Letermovir is highly potent in vitro and in vivo against cytomegalovirus. Because of a distinct mechanism of action, it does not exhibit cross-resistance with other antiviral drugs. It is predicted to be active against strains that are resistant to ganciclovir, foscarnet, and cidofovir. To date, early-phase clinical trials suggest a very low incidence of adverse effects. Herein, we present a comprehensive review on letermovir, from its postulated novel mechanism of action to the results of most recent clinical studies.

  9. Cloning, Assembly, and Modification of the Primary Human Cytomegalovirus Isolate Toledo by Yeast-Based Transformation-Associated Recombination.

    Science.gov (United States)

    Vashee, Sanjay; Stockwell, Timothy B; Alperovich, Nina; Denisova, Evgeniya A; Gibson, Daniel G; Cady, Kyle C; Miller, Kristofer; Kannan, Krishna; Malouli, Daniel; Crawford, Lindsey B; Voorhies, Alexander A; Bruening, Eric; Caposio, Patrizia; Früh, Klaus

    2017-01-01

    Genetic engineering of cytomegalovirus (CMV) currently relies on generating a bacterial artificial chromosome (BAC) by introducing a bacterial origin of replication into the viral genome using in vivo recombination in virally infected tissue culture cells. However, this process is inefficient, results in adaptive mutations, and involves deletion of viral genes to avoid oversized genomes when inserting the BAC cassette. Moreover, BAC technology does not permit the simultaneous manipulation of multiple genome loci and cannot be used to construct synthetic genomes. To overcome these limitations, we adapted synthetic biology tools to clone CMV genomes in Saccharomyces cerevisiae. Using an early passage of the human CMV isolate Toledo, we first applied transformation-associated recombination (TAR) to clone 16 overlapping fragments covering the entire Toledo genome in Saccharomyces cerevisiae. Then, we assembled these fragments by TAR in a stepwise process until the entire genome was reconstituted in yeast. Since next-generation sequence analysis revealed that the low-passage-number isolate represented a mixture of parental and fibroblast-adapted genomes, we selectively modified individual DNA fragments of fibroblast-adapted Toledo (Toledo-F) and again used TAR assembly to recreate parental Toledo (Toledo-P). Linear, full-length HCMV genomes were transfected into human fibroblasts to recover virus. Unlike Toledo-F, Toledo-P displayed characteristics of primary isolates, including broad cellular tropism in vitro and the ability to establish latency and reactivation in humanized mice. Our novel strategy thus enables de novo cloning of CMV genomes, more-efficient genome-wide engineering, and the generation of viral genomes that are partially or completely derived from synthetic DNA. IMPORTANCE The genomes of large DNA viruses, such as human cytomegalovirus (HCMV), are difficult to manipulate using current genetic tools, and at this time, it is not possible to obtain

  10. Human Cytomegalovirus-Encoded Human Interleukin-10 (IL-10) Homolog Amplifies Its Immunomodulatory Potential by Upregulating Human IL-10 in Monocytes.

    Science.gov (United States)

    Avdic, Selmir; McSharry, Brian P; Steain, Megan; Poole, Emma; Sinclair, John; Abendroth, Allison; Slobedman, Barry

    2016-04-01

    The human cytomegalovirus (HCMV) gene UL111A encodes cytomegalovirus-encoded human interleukin-10 (cmvIL-10), a homolog of the potent immunomodulatory cytokine human interleukin 10 (hIL-10). This viral homolog exhibits a range of immunomodulatory functions, including suppression of proinflammatory cytokine production and dendritic cell (DC) maturation, as well as inhibition of major histocompatibility complex (MHC) class I and class II. Here, we present data showing that cmvIL-10 upregulates hIL-10, and we identify CD14(+)monocytes and monocyte-derived macrophages and DCs as major sources of hIL-10 secretion in response to cmvIL-10. Monocyte activation was not a prerequisite for cmvIL-10-mediated upregulation of hIL-10, which was dose dependent and controlled at the transcriptional level. Furthermore, cmvIL-10 upregulated expression of tumor progression locus 2 (TPL2), which is a regulator of the positive hIL-10 feedback loop, whereas expression of a negative regulator of the hIL-10 feedback loop, dual-specificity phosphatase 1 (DUSP1), remained unchanged. Engagement of the hIL-10 receptor (hIL-10R) by cmvIL-10 led to upregulation of heme oxygenase 1 (HO-1), an enzyme linked with suppression of inflammatory responses, and this upregulation was required for cmvIL-10-mediated upregulation of hIL-10. We also demonstrate an important role for both phosphatidylinositol 3-kinase (PI3K) and STAT3 in the upregulation of HO-1 and hIL-10 by cmvIL-10. In addition to upregulating hIL-10, cmvIL-10 could exert a direct immunomodulatory function, as demonstrated by its capacity to upregulate expression of cell surface CD163 when hIL-10 was neutralized. This study identifies a mechanistic basis for cmvIL-10 function, including the capacity of this viral cytokine to potentially amplify its immunosuppressive impact by upregulating hIL-10 expression. Human cytomegalovirus (HCMV) is a large, double-stranded DNA virus that causes significant human disease, particularly in the

  11. Infection of Cells with Human Cytomegalovirus during S Phase Results in a Blockade to Immediate-Early Gene Expression That Can Be Overcome by Inhibition of the Proteasome

    OpenAIRE

    Elizabeth A Fortunato; Sanchez, Veronica; Judy Y Yen; Spector, Deborah H.

    2002-01-01

    Cells infected with human cytomegalovirus (HCMV) after commencing DNA replication do not initiate viral immediate-early (IE) gene expression and divide before arresting. To determine the nature of this blockade, we examined cells that were infected 24 h after release from G0 using immunofluorescence, laser scanning cytometry, and fluorescence-activated cell sorting (FACS) analysis. Approximately 40 to 50% of the cells had 2N DNA content, became IE+ in the first 12 h, and arrested. Most but no...

  12. The Novel Anticytomegalovirus Compound AIC246 (Letermovir) Inhibits Human Cytomegalovirus Replication through a Specific Antiviral Mechanism That Involves the Viral Terminase ▿

    OpenAIRE

    Goldner, Thomas; Hewlett, Guy; Ettischer, Nicole; Ruebsamen-Schaeff, Helga; Zimmermann, Holger; Lischka, Peter

    2011-01-01

    Human cytomegalovirus (HCMV) remains the leading viral cause of birth defects and life-threatening disease in transplant recipients. All approved antiviral drugs target the viral DNA polymerase and are associated with severe toxicity issues and the emergence of drug resistance. Attempts to discover improved anti-HCMV drugs led to the identification of the small-molecular-weight compound AIC246 (Letermovir). AIC246 exhibits outstanding anti-HCMV activity in vitro and in vivo and currently is u...

  13. Emergence of Multiple Human Cytomegalovirus Ganciclovir-Resistant Mutants with Deletions and Substitutions within the UL97 Gene in a Patient with Severe Combined Immunodeficiency

    Science.gov (United States)

    Wolf, Dana G.; Yaniv, Isaac; Ashkenazi, Shai; Honigman, Alik

    2001-01-01

    Infection with multiple ganciclovir-resistant human cytomegalovirus mutants, containing different substitutions and deletions in the UL97 gene, was found in a patient with severe combined immunodeficiency (SCID) within 3 weeks of ganciclovir therapy. A novel 11-codon deletion at positions 590 to 600 was identified. These unique findings may be related to the nature of the immunodeficiency in the SCID patient. PMID:11158760

  14. Ribonuclease P-mediated inhibition of human cytomegalovirus gene expression and replication induced by engineered external guide sequences.

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    Jiang, Xiaohong; Chen, Yuan-Chuan; Gong, Hao; Trang, Phong; Lu, Sangwei; Liu, Fenyong

    2012-09-01

    External guide sequences (EGSs) are RNA molecules that can bind to a target mRNA and direct ribonuclease P (RNase P), a tRNA processing enzyme, for specific cleavage of the target mRNA. Using an in vitro selection procedure, we have previously generated EGS variants that efficiently direct human RNase P to cleave a target mRNA in vitro. In this study, we constructed EGSs from a variant to target the overlapping region of the mRNAs coding for human cytomegalovirus (HCMV) capsid scaffolding protein (CSP) and assemblin, which are essential for viral capsid formation. The EGS variant was about 40-fold more active in directing human RNase P to cleave the mRNA in vitro than the EGS derived from a natural tRNA. Moreover, a reduction of about 98% and 75% in CSP/assemblin gene expression and a reduction of 7000- and 250-fold in viral growth were observed in HCMV-infected cells that expressed the variant and the tRNA-derived EGS, respectively. Our study shows that the EGS variant is more effective in blocking HCMV gene expression and growth than the tRNA-derived EGS. Moreover, these results demonstrate the utility of highly active EGS RNA variants in gene targeting applications including anti-HCMV therapy.

  15. [DNA-EGS1386 in cells induced RNase P inhibits the expression of human cytomegalovirus UL49 gene].

    Science.gov (United States)

    Cui, Yanwei; Zeng, Zhifeng; Li, Hongjian; Li, Yueqin; Zhou, Qi; Yang, Dan; Zou, Yi; Yang, Guang; Zhou, Tianhong

    2009-11-01

    External Guide Sequences (EGSs) represents a novel nucleic acid based gene interference approach to modulate gene expression. They are oligonucleotides that consist of a sequence complementary to a target mRNA and recruit intracellular RNase P for specific degradation of the target RNA. DNA-based EGS1386 with a size of 12 nt was chemically synthesized to target the mRNA coding for the UL49 gene of human cytomegalovirus (HCMV). The DNA-based EGS1386 molecule efficiently directed human RNase P to cleave the target mRNA sequence in vitro. A reduction of more than 50% in the levels of UL49 expression was observed in human cells treated with the DNA-based EGS1386 targeted UL49 assayed by fluorescent quantization PCR and Western blotting. This results showed that the DNA-EGS1386 can effectively guide the RNase P cut the target mRNA. Therefore, DNA-EGS can develop into a new gene silencing technology and potential of the anti-viral reagents.

  16. Properties of virion transactivator proteins encoded by primate cytomegaloviruses

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    Barry Peter A

    2009-05-01

    Full Text Available Abstract Background Human cytomegalovirus (HCMV is a betaherpesvirus that causes severe disease in situations where the immune system is immature or compromised. HCMV immediate early (IE gene expression is stimulated by the virion phosphoprotein pp71, encoded by open reading frame (ORF UL82, and this transactivation activity is important for the efficient initiation of viral replication. It is currently recognized that pp71 acts to overcome cellular intrinsic defences that otherwise block viral IE gene expression, and that interactions of pp71 with the cell proteins Daxx and ATRX are important for this function. A further property of pp71 is the ability to enable prolonged gene expression from quiescent herpes simplex virus type 1 (HSV-1 genomes. Non-human primate cytomegaloviruses encode homologs of pp71, but there is currently no published information that addresses their effects on gene expression and modes of action. Results The UL82 homolog encoded by simian cytomegalovirus (SCMV, strain Colburn, was identified and cloned. This ORF, named S82, was cloned into an HSV-1 vector, as were those from baboon, rhesus monkey and chimpanzee cytomegaloviruses. The use of an HSV-1 vector enabled expression of the UL82 homologs in a range of cell types, and permitted investigation of their abilities to direct prolonged gene expression from quiescent genomes. The results show that all UL82 homologs activate gene expression, and that neither host cell type nor promoter target sequence has major effects on these activities. Surprisingly, the UL82 proteins specified by non-human primate cytomegaloviruses, unlike pp71, did not direct long term expression from quiescent HSV-1 genomes. In addition, significant differences were observed in the intranuclear localization of the UL82 homologs, and in their effects on Daxx. Strikingly, S82 mediated the release of Daxx from nuclear domain 10 substructures much more rapidly than pp71 or the other proteins tested. All

  17. Prevalence of human cytomegalovirus (HCMV antibodies among patients HIV/AIDS in Kurdistan province, 2015

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    Arezo Omati

    2016-12-01

    Full Text Available Background : Cytomegalovirus (CMV infection is one of the most common causes of death in people with immune deficiency diseases such as: organ or tissue recipients, patients with HIV/AIDS and newborn infants. The aim of this study was to determine the prevalence of anti-CMV antibodies in patients with HIV/AIDS in the Kurdistan province. Materials and Methods: This cross-sectional study carried out on 151 patients with HIV/AIDS covered by behavioral health counseling centers in Kurdistan province, 2015. For all patients the values of CMV antibodies, include IgG and IgM, were determined by ELISA technique and Diagnostic Kits (EIA WELL, Rome, Italy. Data analyses were done by use of t-test and multiple linear regression tests in stata software, version 13. Results: 116 (76.8% and 35 (23.2% of the patients were male and female, respectively. Mean (SD age of the patients were 39.3 (9.1 years. All studied patients were found positive for CMV-IgG (prevalence=100%, whereas only one case (0.7% was found positive for CMV-IgM. The relationship between age and CMV-IgG levels was statistically significant. Conclusion: The results showed that high prevalence of CMV in patients with HIV/AIDS, so in addition to paying more attention to the issue of HIV and CMV co-infection, about value of early antiretroviral treatment conducting further studies seems necessary.

  18. Molecular Imprint of Exposure to Naturally Occurring Genetic Variants of Human Cytomegalovirus on the T cell Repertoire

    Science.gov (United States)

    Smith, Corey; Gras, Stephanie; Brennan, Rebekah M.; Bird, Nicola L.; Valkenburg, Sophie A.; Twist, Kelly-Anne; Burrows, Jacqueline M.; Miles, John J.; Chambers, Daniel; Bell, Scott; Campbell, Scott; Kedzierska, Katherine; Burrows, Scott R.; Rossjohn, Jamie; Khanna, Rajiv

    2014-02-01

    Exposure to naturally occurring variants of herpesviruses in clinical settings can have a dramatic impact on anti-viral immunity. Here we have evaluated the molecular imprint of variant peptide-MHC complexes on the T-cell repertoire during human cytomegalovirus (CMV) infection and demonstrate that primary co-infection with genetic variants of CMV was coincident with development of strain-specific T-cell immunity followed by emergence of cross-reactive virus-specific T-cells. Cross-reactive CMV-specific T cells exhibited a highly conserved public T cell repertoire, while T cells directed towards specific genetic variants displayed oligoclonal repertoires, unique to each individual. T cell recognition foot-print and pMHC-I structural analyses revealed that the cross-reactive T cells accommodate alterations in the pMHC complex with a broader foot-print focussing on the core of the peptide epitope. These findings provide novel molecular insight into how infection with naturally occurring genetic variants of persistent human herpesviruses imprints on the evolution of the anti-viral T-cell repertoire.

  19. Breast Milk Human Cytomegalovirus (CMV) Viral Load and the Establishment of Breast Milk CMV-pp65-Specific CD8 T Cells in Human CMV Infected Mothers.

    Science.gov (United States)

    Moylan, David C; Pati, Sunil K; Ross, Shannon A; Fowler, Karen B; Boppana, Suresh B; Sabbaj, Steffanie

    2017-11-27

    The role of human cytomegalovirus (HCMV)-specific T-cell responses in breast milk of HCMV-seropositive mothers is not well defined. In these studies, we demonstrate that the frequency of cytomegalovirus (CMV)-pp65-specific T-cell responses in peripheral blood mononuclear cells (PBMCs) and breast milk cells (BMCs) is increased for CD8+ T cells in both sample sources when compared with CD4+ T cells. The frequency of pp55-specific CD8 T cells producing interferon γ (IFN-γ) alone or dual IFN-γ/granzyme rB producers is increased in breast milk compared with PBMCs. Lastly, we observed a positive correlation between breast milk viral load and the CD8 pp65-specific response, suggesting that local virus replication drives antigen-specific CD8 T cells into the breast. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

  20. Cytomegalovirus (CMV) and Pregnancy

    Science.gov (United States)

    Cytomegalovirus (CMV) In every pregnancy, a woman starts out with a 3-5% chance of having a ... risk. This sheet talks about whether exposure to cytomegalovirus (CMV) can increase the risk for birth defects ...

  1. Cytomegalovirus (For Parents)

    Science.gov (United States)

    ... Important? Top 10 Homework Tips Raising Confident Kids Cytomegalovirus (CMV) KidsHealth > For Parents > Cytomegalovirus (CMV) Print A A A What's in this ... Diagnosis & Treatment Prevention en español Citomegalovirus About CMV Cytomegalovirus (CMV) is a common infection that can affect ...

  2. Rapid In Vitro Evolution of Human Cytomegalovirus UL56 Mutations That Confer Letermovir Resistance.

    Science.gov (United States)

    Chou, Sunwen

    2015-10-01

    Letermovir (LMV) is an experimental cytomegalovirus terminase inhibitor undergoing phase 3 clinical trials. Viral mutations have been described at UL56 codons 231 to 369 that confer widely variable levels of LMV resistance. In this study, 15 independent experiments propagating an exonuclease mutant viral strain in escalating LMV concentrations replicated 6 of the 7 published UL56 mutations and commonly elicited additional resistance-conferring mutations at UL56 codons 231, 236, 237, 244, 257, 261, 325, and 329. Mutations were first detected earlier in LMV (median, 3 passages) than in 8 parallel experiments with foscarnet (median, 15 passages). As LMV concentrations increased, the typical initial UL56 change F261L, which confers low-grade resistance, combined or was replaced with mutations conferring higher-grade resistance, eventually enabling normal viral growth in 30 μM LMV (>5,000-fold the 50% effective concentration [EC50] for the wild type). At high LMV concentrations, the UL56 changes C325F/R were commonly detected, as well as a combination of changes at codons 236, 257, 329, and/or 369. Recombinant viruses containing individual UL56 mutations and combinations were constructed to confirm their resistance phenotypes and normal growth in cell culture. Several double and triple mutants showed much higher LMV resistance than the respective single mutants, particularly those including changes at both codons 236 and 257. The multiplicity of pathways to high-grade LMV resistance with minimal viral growth impact suggests a low viral genetic barrier and the need for close monitoring during treatment of active infection. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  3. The increased sensitivity of human cytomegalovirus (HCMV) PCR quantitation in whole blood affects reproductive rate (Ro) measurement.

    Science.gov (United States)

    Gurtler, Volker; Mayall, Barrie C; Wang, Jenny; Ghaly-Derias, Shahbano

    2014-02-01

    In order to determine the effect of the increase in sensitivity of HCMV detection in whole blood compared to plasma on reproductive rate (Ro) measurement, an optimized human cytomegalovirus (HCMV) quantitative PCR assay was developed. The results presented in this study are summarized by the following three methodological improvements: (i) at values below the limit of quantitation (LOQ) of 60copies/ml, determination of HCMV load was more sensitive with whole blood than plasma, (ii) for the determination of viral load, whole blood was more sensitive than plasma below 1000copies/ml but little difference was observed above 1000copies/ml and (iii) the measurement of "Reproductive Rate" can be affected by imprecise measurement of HCMV viral load in either plasma or whole blood compartments depending on whether samples were taken from patients on antiviral treatment or from patients where HCMV load was rising. Taken together this study provides methodological improvements suggesting that below HCMV viral load levels of 1000copies/ml (1640IU/ml) both plasma and whole blood should be tested. Crown Copyright © 2013. Published by Elsevier B.V. All rights reserved.

  4. Latency-Associated Expression of Human Cytomegalovirus US28 Attenuates Cell Signaling Pathways To Maintain Latent Infection

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    Benjamin A. Krishna

    2017-12-01

    Full Text Available Reactivation of human cytomegalovirus (HCMV latent infection from early myeloid lineage cells constitutes a threat to immunocompromised or immune-suppressed individuals. Consequently, understanding the control of latency and reactivation to allow targeting and killing of latently infected cells could have far-reaching clinical benefits. US28 is one of the few viral genes that is expressed during latency and encodes a cell surface G protein-coupled receptor (GPCR, which, during lytic infection, is a constitutive cell-signaling activator. Here we now show that in monocytes, which are recognized sites of HCMV latency in vivo, US28 attenuates multiple cell signaling pathways, including mitogen-activated protein (MAP kinase and NF-κB, and that this is required to establish a latent infection; viruses deleted for US28 initiate a lytic infection in infected monocytes. We also show that these monocytes then become potent targets for the HCMV-specific host immune response and that latently infected cells treated with an inverse agonist of US28 also reactivate lytic infection and similarly become immune targets. Consequently, we suggest that the use of inhibitors of US28 could be a novel immunotherapeutic strategy to reactivate the latent viral reservoir, allowing it to be targeted by preexisting HCMV-specific T cells.

  5. Primary human cytomegalovirus infection induces the expansion of virus-specific activated and atypical memory B cells.

    Science.gov (United States)

    Dauby, Nicolas; Kummert, Caroline; Lecomte, Sandra; Liesnard, Corinne; Delforge, Marie-Luce; Donner, Catherine; Marchant, Arnaud

    2014-10-15

    Although neutralizing antibodies play a central role in the control of cytomegalovirus (CMV) dissemination, little is known about the response of B lymphocytes to primary human CMV infection. The proportion, phenotype, specificity, and functionality of B-cell subsets were studied in a cohort of pregnant women with primary CMV infection. CMV-seronegative pregnant women, as well as CMV-seronegative and CMV-seropositive healthy adults, were included as controls. Primary CMV infection was associated with a sustained expansion of activated (CD27(+)CD20(+)CD21(low)) and atypical (CD27(-)CD20(+)CD21(low)) memory B cells (MBCs). Both subsets expressed an effector phenotype, and their proportions were correlated with viremia. Activated MBCs expressed high levels of activation markers and included high frequencies of tumor necrosis α (TNF-α)-producing cells, whereas atypical MBCs expressed high levels of inhibitory receptors and had low TNF-α responses. Fluorescent-labeled antigen experiments indicated that activated and atypical MBCs were enriched in CMV-specific cells. Primary CMV infection mobilizes a large pool of memory B cells that includes activated and atypical MBCs. The functional regulation of CMV-specific MBCs may limit the production of antibodies and the control of viral dissemination. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  6. Early abnormal liver enzyme levels may increase the prevalence of human cytomegalovirus antigenaemia after hematopoietic stem cell transplantation.

    Science.gov (United States)

    Ye, Baning; Zhao, Hong

    2017-04-01

    Objective Human cytomegalovirus (HCMV) infection is common after bone marrow transplantation (BMT), and it increases morbidity and mortality for transplant recipients. HCMV infection may cause hepatitis and elevate the liver enzymes aspartate transferase (AST) and alanine transferase (ALT). This study aimed to analyse the associations between liver enzyme levels and infection with HCMV antigenaemia after BMT. Methods Data from 30 patients after BMT were collected at different time points (0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 4.0, 5.0, and 6.0 months post-transplantation). The patients were divided into the HCMV antigenaemia-positive and HCMV antigenaemia-negative groups according to a peripheral blood pp65 antigen assay. Immunohistochemistry was used to identify HCMV pp65 antigen and conventional methods were used to detect liver enzyme levels. Results Twelve patients were pp65 antigenaemia-positive and 10 patients were positive in the first 3 months post-transplant. Liver enzyme levels were increased after positivity for HCMV antigenaemia (p = 0.034 and p = 0.018 for ALT and AST, respectively). One month before antigenaemia, AST levels were higher in the HCMV antigenaemia-positive group compared with the negative group (p = 0.006). Conclusion HCMV antigenaemia mostly occurs in the early stage of post-BMT and early abnormal liver enzyme levels may increase the chance of HCMV antigenaemia after BMT.

  7. Characterization of Human Cytomegalovirus Genome Diversity in Immunocompromised Hosts by Whole-Genome Sequencing Directly From Clinical Specimens.

    Science.gov (United States)

    Hage, Elias; Wilkie, Gavin S; Linnenweber-Held, Silvia; Dhingra, Akshay; Suárez, Nicolás M; Schmidt, Julius J; Kay-Fedorov, Penelope C; Mischak-Weissinger, Eva; Heim, Albert; Schwarz, Anke; Schulz, Thomas F; Davison, Andrew J; Ganzenmueller, Tina

    2017-06-01

    Advances in next-generation sequencing (NGS) technologies allow comprehensive studies of genetic diversity over the entire genome of human cytomegalovirus (HCMV), a significant pathogen for immunocompromised individuals. Next-generation sequencing was performed on target enriched sequence libraries prepared directly from a variety of clinical specimens (blood, urine, breast milk, respiratory samples, biopsies, and vitreous humor) obtained longitudinally or from different anatomical compartments from 20 HCMV-infected patients (renal transplant recipients, stem cell transplant recipients, and congenitally infected children). De novo-assembled HCMV genome sequences were obtained for 57 of 68 sequenced samples. Analysis of longitudinal or compartmental HCMV diversity revealed various patterns: no major differences were detected among longitudinal, intraindividual blood samples from 9 of 15 patients and in most of the patients with compartmental samples, whereas a switch of the major HCMV population was observed in 6 individuals with sequential blood samples and upon compartmental analysis of 1 patient with HCMV retinitis. Variant analysis revealed additional aspects of minor virus population dynamics and antiviral-resistance mutations. In immunosuppressed patients, HCMV can remain relatively stable or undergo drastic genomic changes that are suggestive of the emergence of minor resident strains or de novo infection.

  8. The host ubiquitin-dependent segregase VCP/p97 is required for the onset of human cytomegalovirus replication.

    Science.gov (United States)

    Lin, Yao-Tang; Prendergast, James; Grey, Finn

    2017-05-01

    The human cytomegalovirus major immediate early proteins IE1 and IE2 are critical drivers of virus replication and are considered pivotal in determining the balance between productive and latent infection. IE1 and IE2 are derived from the same primary transcript by alternative splicing and regulation of their expression likely involves a complex interplay between cellular and viral factors. Here we show that knockdown of the host ubiquitin-dependent segregase VCP/p97, results in loss of IE2 expression, subsequent suppression of early and late gene expression and, ultimately, failure in virus replication. RNAseq analysis showed increased levels of IE1 splicing, with a corresponding decrease in IE2 splicing following VCP knockdown. Global analysis of viral transcription showed the expression of a subset of viral genes is not reduced despite the loss of IE2 expression, including UL112/113. Furthermore, Immunofluorescence studies demonstrated that VCP strongly colocalised with the viral replication compartments in the nucleus. Finally, we show that NMS-873, a small molecule inhibitor of VCP, is a potent HCMV antiviral with potential as a novel host targeting therapeutic for HCMV infection.

  9. Human Cytomegalovirus Inhibits the PARsylation Activity of Tankyrase--A Potential Strategy for Suppression of the Wnt Pathway.

    Science.gov (United States)

    Roy, Sujayita; Liu, Fengjie; Arav-Boger, Ravit

    2015-12-29

    Human cytomegalovirus (HCMV) was reported to downregulate the Wnt/β-catenin pathway. Induction of Axin1, the negative regulator of the Wnt pathway, has been reported as an important mechanism for inhibition of β-catenin. Since Tankyrase (TNKS) negatively regulates Axin1, we investigated the effect of HCMV on TNKS expression and poly-ADP ribose polymerase (PARsylation) activity, during virus replication. Starting at 24 h post infection, HCMV stabilized the expression of TNKS and reduced its PARsylation activity, resulting in accumulation of Axin1 and reduction in its PARsylation as well. General PARsylation was not changed in HCMV-infected cells, suggesting specific inhibition of TNKS PARsylation. Similarly, treatment with XAV939, a chemical inhibitor of TNKS' activity, resulted in the accumulation of TNKS in both non-infected and HCMV-infected cell lines. Reduction of TNKS activity or knockdown of TNKS was beneficial for HCMV, evidenced by its improved growth in fibroblasts. Our results suggest that HCMV modulates the activity of TNKS to induce Axin1, resulting in inhibition of the β-catenin pathway. Since HCMV replication is facilitated by TNKS knockdown or inhibition of its activity, TNKS may serve as an important virus target for control of a variety of cellular processes.

  10. Interleukin-10 blocks in vitro replication of human cytomegalovirus by inhibiting the virus-induced autophagy in MRC5 cells.

    Science.gov (United States)

    Wang, Li; Zhang, Huiping; Qian, Jihong; Wang, Kanqing; Zhu, Jianxing

    2014-06-13

    Interleukin-10 is an important cytokine that regulates immune response. Previous studies have shown that human cytomegalovirus can trigger cell autophagy during the early stages of infection. To our knowledge, whether IL-10 inhibits HCMV-induced autophagy and virus replication has not been studied previously. We investigated whether IL-10 affects cell viability and autophagy under the conditions of starvation and HCMV infection by using the MRC5 cell line. We also explored the role of IL-10-mediated autophagy on HCMV replication. Our data showed that IL-10 inhibited the autophagic flux of the MRC5 cells irrespective of starvation or HCMV infection, and suppressed HCMV replication. The promotion of autophagy with either a pharmacological inducer (rapamycin), or a technique to over-express the BECN1 gene reversed the effect of IL-10 on virus replication. Furthermore, the PI3K/Akt signal pathway was activated when the cells were pretreated with IL-10. Our results indicated that IL-10 can suppress HCMV replication by inhibiting autophagy in host cells during the early stages of infection. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Control of human cytomegalovirus gene expression by differential histone modifications during lytic and latent infection of a monocytic cell line.

    Science.gov (United States)

    Ioudinkova, Elena; Arcangeletti, Maria Cristina; Rynditch, Alla; De Conto, Flora; Motta, Federica; Covan, Silvia; Pinardi, Federica; Razin, Sergey V; Chezzi, Carlo

    2006-12-15

    Non-differentiated THP-1 cells can be infected by human cytomegalovirus (HCMV) Towne strain, which persists in these cells in a non-active (latent) form without undergoing a productive cycle. The same cells become permissive for HCMV lytic infection after induction of cell differentiation by treatment with 12-O-tetradecanoylphorbol-13-acetate. We used this cellular model to study the possible role of histone modifications in the control of HCMV latency. Using chromatin immunoprecipitation with antibodies against histone H3 acetylated or dimethylated in position K9, we demonstrated that in lytically infected cells the HCMV enhancer was associated with heavy acetylated but not dimethylated H3. In the case of latent infection, the HCMV enhancer was associated with neither acetylated nor dimethylated H3. HCMV genes encoding DNA polymerase (early), pp65 (early-late) and pp150 (late) proteins were associated preferentially with acetylated H3 in lytically infected cells and with dimethylated H3 in latently infected cells. These data strongly suggest that K9 methylation of H3 is involved in HCMV gene repression, while association of the above genes with acetylated histones is likely to be necessary for active transcription. It can be postulated that the same histone modifications are used to mark active and repressed genes in both cellular and viral chromatin.

  12. Ribonucleotide reductase inhibitors hydroxyurea, didox, and trimidox inhibit human cytomegalovirus replication in vitro and synergize with ganciclovir.

    Science.gov (United States)

    Bhave, Sukhada; Elford, Howard; McVoy, Michael A

    2013-10-01

    Ganciclovir (GCV) is a deoxyguanosine analog that is effective in inhibiting human cytomegalovirus (HCMV) replication. In infected cells GCV is converted to GCV-triphosphate which competes with dGTP for incorporation into the growing DNA strand by the viral DNA polymerase. Incorporated GCV promotes chain termination as it is an inefficient substrate for elongation. Because viral DNA synthesis also relies on cellular ribonucleotide reductase (RR) to synthesize deoxynucleotides, RR inhibitors are predicted to inhibit HCMV replication. Moreover, as dGTP competes with GCV-triphosphate for incorporation, RR inhibitors may also synergize with GCV by reducing intracellular dGTP levels and there by promoting increased GCV-triphosphate utilization by DNA polymerase. To investigate potential of RR inhibitors as anti-HCMV agents both alone and in combination with GCV, HCMV-inhibitory activities of three RR inhibitors, hydroxyurea, didox, and trimidox, were determined. In both spread inhibition and yield reduction assays RR inhibitors had modest anti-HCMV activity with 50% inhibitory concentrations ranging from 36±1.7 to 221±52μM. However, all three showed significant synergy with GCV at concentrations below their 50% inhibitory and 50% toxic concentrations. These results suggest that combining GCV with relatively low doses of RR inhibitors could significantly potentiate the anti-HCMV activity of GCV in vivo and could improve clinical response to therapy. Copyright © 2013 Elsevier B.V. All rights reserved.

  13. [Different regulation of mitochondrial apoptosis and Bcl-2 gene expression in quescent and proliferative human fibroblasts infected with cytomegalovirus].

    Science.gov (United States)

    Fedorova, N E; Sokolova, T M; Medzhidova, M G; Kushch, A A

    2010-01-01

    The aim of the study was to compare the dynamics ofmitochondrial apoptosis (MA) in cells at different stages of proliferation and with different susceptibility to cytomegalovirus (CMV). It has been found that in quiescent human fibroblasts (HF) CMV regulates MA at the level of bcl-2 gene transcription, exerting both pro- and anti-apoptotic effects. Suppression of bcl-2 transcription is greater in HF-977 line, which is highly susceptible to CMV in comparison with HF-1068 line. The effect of proliferative activity on MA was studied using CMV-infected HF-110044 line at the G0- or S-phase. A direct correlation was established between accumulation of cytochrome c and caspase 3 (MA markers) and production of IE72, pp65 and gB (CMV proteins). In G0-fibrob-lasts, viral replication was highly productive and bcl-2 expression was 10-fold as high as in S-phase cells, in which viral protein production and cell death were much lower. The increased gene transcription and accumulation of Bcl-2 protein enhanced cell viability and provided synthesis of viral proteins. Impaired structure of actin microfilaments, a caspase 3 target, coincided with pronounced suppression of gamma-actin gene in S-phase HF-110044. Our findings provide an insight into CMV-induced mechanisms of MA which lead to rapid death of infected quiescent fibroblasts and to slow death of cells infected at the stage of DNA synthesis.

  14. Active human Cytomegalovirus infection and Glycoprotein B genotypes in Brazilian pediatric renal or hematopoietic stem cell transplantation patients

    Directory of Open Access Journals (Sweden)

    Débora de Campos Dieamant

    2010-03-01

    Full Text Available A prospective analysis of active Human Cytomegalovirus infection (HCMV was conducted on 33 pediatric renal or hematopoietic stem cell post-transplant patients. The HCMV-DNA positive samples were evaluated for the prevalence of different gB subtypes and their subsequent correlation with clinical signs. The surveillance of HCMV active infection was based on the monitoring of antigenemia (AGM and on a nested polymerase chain reaction (N-PCR for the detection of HCMV in the patients studied. Using restriction analysis of the gB gene sequence by PCR-RFLP (Restriction Fragment Length Polymorphism, different HCMV strains could be detected and classified in at least four HCMV genotypes. Thirty-three pediatric recipients of renal or bone marrow transplantation were monitored. Twenty out of thirty-three (60.6% patients demonstrated active HCMV infection. gB1 and gB2 genotypes were more frequent in this population. In this study, we observed that gB2 had correlation with reactivation of HCMV infection and that patients with mixture of genotypes did not show any symptoms of HCMV disease. Future studies has been made to confirm this.

  15. Seroprevalence of Hepatitis C, Hepatitis B, Cytomegalovirus, and Human Immunodeficiency Viruses in Multitransfused Thalassemic Children in Upper Egypt

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    Ramadan A. Mahmoud

    2016-01-01

    Full Text Available Background. Frequent blood transfusions in thalassemia major children expose them to the risk of transfusion-transmitted infections (TTIs. The aim of this study was to estimate the prevalence of hepatitis C virus (HCV, hepatitis B virus (HBV, human immunodeficiency virus (HIV, and cytomegalovirus (CMV in thalassemic children attending the Pediatrics Departments of both Sohag and Minia Universities of Upper Egypt, during the period from May 2014 to May 2015. Methods. Serum samples were screened for hepatitis B surface antigen (HBsAg, anti-HCV, anti-CMV, and anti-HIV type 1 and type 2 using the Vitek Immunodiagnostic Assay System. Results. The frequencies of anti-HCV, HBsAg, anti-CMV, and anti-HIV type 1 and type 2 were found to be 37.11%, 4.12%, 4.12%, 0.00%, and 0.00%, respectively. Seropositivity for anti-HCV, HBsAg, and anti-CMV increased with increasing age of the patients, duration of the disease, serum ferritin level (ng/mL, and liver enzymes (U/L, while it was not significantly associated with gender, frequency of blood transfusion, or the status of splenectomy operation (P>0.05. Conclusion. The frequency of TTIs, especially HCV, is considerably high among Egyptian children with thalassemia major. It is therefore important to implement measures to improve blood transfusion screening, such as polymerase chain reaction, in order to reduce TTIs from blood donor units.

  16. Regulation and gene expression profiling of NKG2D positive human cytomegalovirus-primed CD4+ T-cells

    DEFF Research Database (Denmark)

    Jensen, Helle; Folkersen, Lasse; Skov, Søren

    2012-01-01

    NKG2D is a stimulatory receptor expressed by natural killer (NK) cells, CD8(+) T-cells, and ¿d T-cells. NKG2D expression is normally absent from CD4(+) T-cells, however recently a subset of NKG2D(+) CD4(+) T-cells has been found, which is specific for human cytomegalovirus (HCMV). This particular...... CD4(+) T-cells. These findings provide novel information about the gene expression profile of HCMV-primed NKG2D(+) CD4(+) T-cells, as well as the mechanisms regulating NKG2D cell surface expression....... we used genome-wide analysis of individual genes and gene set enrichment analysis (GSEA) to investigate the gene expression profile of NKG2D(+) CD4(+) T-cells, generated from HCMV-primed CD4(+) T-cells. We show that the HCMV-primed NKG2D(+) CD4(+) T-cells possess a higher differentiated phenotype...... than the NKG2D(-) CD4(+) T-cells, both at the gene expression profile and cytokine profile. The ability to express NKG2D at the cell surface was primarily determined by the activation or differentiation status of the CD4(+) T-cells and not by the antigen presenting cells. We observed a correlation...

  17. Single-dose protection against Plasmodium berghei by a simian adenovirus vector using a human cytomegalovirus promoter containing intron A.

    Science.gov (United States)

    Sridhar, S; Reyes-Sandoval, A; Draper, S J; Moore, A C; Gilbert, S C; Gao, G P; Wilson, J M; Hill, A V S

    2008-04-01

    Human adenovirus serotype 5 (AdH5) vector vaccines elicit strong immune responses to the encoded antigen and have been used in various disease models. We designed AdH5 vectors expressing antigen under the control of a human cytomegalovirus (HCMV) immediate-early promoter containing its intron A sequence. The transcriptional levels of antigen and immune responses to antigen for vectors with the HCMV promoter with the intron A sequence (LP) were greater than those for AdH5 vectors using the HCMV promoter sequence without intron A (SP). We compared an E1E3-deleted AdH5 adenoviral vector, which affords more space for insertion of foreign sequences, and showed it to be as immunogenic as an E1-deleted AdH5 vector. Neutralizing antibodies to AdH5 limit the efficacy of vaccines based on the AdH5 serotype, and simian adenoviral vectors offer an attractive option to overcome this problem. We constructed E1E3-deleted human and simian adenoviral vectors encoding the pre-erythrocytic-stage malarial antigen Plasmodium berghei circumsporozoite protein. We compared the immunogenicity and efficacy of AdC6, a recombinant simian adenovirus serotype 6 vector, in a murine malaria model to those of AdH5 and the poxviral vectors MVA and FP9. AdC6 induced sterile protection from a single dose in 90% of mice, in contrast to AdH5 (25%) and poxviral vectors MVA and FP9 (0%). Adenoviral vectors maintained potent CD8(+) T-cell responses for a longer period after immunization than did poxviral vectors and mainly induced an effector memory phenotype of cells. Significantly, AdC6 was able to maintain protection in the presence of preexisting immunity to AdH5.

  18. Nuclear Matrix Proteins in Human Colon Cancer

    Science.gov (United States)

    Keesee, Susan K.; Meneghini, Marc D.; Szaro, Robert P.; Wu, Ying-Jye

    1994-03-01

    The nuclear matrix is the nonchromatin scaffolding of the nucleus. This structure confers nuclear shape, organizes chromatin, and appears to contain important regulatory proteins. Tissue specific nuclear matrix proteins have been found in the rat, mouse, and human. In this study we compared high-resolution two-dimensional gel electropherograms of nuclear matrix protein patterns found in human colon tumors with those from normal colon epithelia. Tumors were obtained from 18 patients undergoing partial colectomy for adenocarcinoma of the colon and compared with tissue from 10 normal colons. We have identified at least six proteins which were present in 18 of 18 colon tumors and 0 of 10 normal tissues, as well as four proteins present in 0 of 18 tumors and in 10 of 10 normal tissues. These data, which corroborate similar findings of cancer-specific nuclear matrix proteins in prostate and breast, suggest that nuclear matrix proteins may serve as important markers for at least some types of cancer.

  19. Attenuation of chemokine receptor function and surface expression as an immunomodulatory strategy employed by human cytomegalovirus is linked to vGPCR US28

    DEFF Research Database (Denmark)

    Frank, Theresa; Reichel, Anna; Larsen, Olav

    2016-01-01

    Background Some herpesviruses like human cytomegalovirus (HCMV) encode viral G protein-coupled receptors that cause reprogramming of cell signaling to facilitate dissemination of the virus, prevent immune surveillance and establish life-long latency. Human GPCRs are known to function in complex......-delUS28 infected cells, CXCR4 surface expression is not altered in particular at late time points of infection. Conclusions We show that the vGPCR US28 is leading to severely disturbed signaling and surface expression of the chemokine receptor CXCR4 thereby representing an effective mechanism used by vGPCRs...

  20. Resistance of human cytomegalovirus to cyclopropavir maps to a base pair deletion in the open reading frame of UL97.

    Science.gov (United States)

    Gentry, Brian G; Vollmer, Laura E; Hall, Ellie D; Borysko, Katherine Z; Zemlicka, Jiri; Kamil, Jeremy P; Drach, John C

    2013-09-01

    Human cytomegalovirus (HCMV) is a widespread pathogen in the human population, affecting many immunologically immature and immunocompromised patients, and can result in severe complications, such as interstitial pneumonia and mental retardation. Current chemotherapies for the treatment of HCMV infections include ganciclovir (GCV), foscarnet, and cidofovir. However, the high incidences of adverse effects (neutropenia and nephrotoxicity) limit the use of these drugs. Cyclopropavir (CPV), a guanosine nucleoside analog, is 10-fold more active against HCMV than GCV (50% effective concentrations [EC50s] = 0.46 and 4.1 μM, respectively). We hypothesize that the mechanism of action of CPV is similar to that of GCV: phosphorylation to a monophosphate by viral pUL97 protein kinase with further phosphorylation to a triphosphate by endogenous kinases, resulting in inhibition of viral DNA synthesis. To test this hypothesis, we isolated a CPV-resistant virus, sequenced its genome, and discovered that bp 498 of UL97 was deleted. This mutation caused a frameshift in UL97 resulting in a truncated protein that lacks a kinase domain. To determine if this base pair deletion was responsible for drug resistance, the mutation was engineered into the wild-type viral genome, which was then exposed to increasing concentrations of CPV. The results demonstrate that the engineered virus was approximately 72-fold more resistant to CPV (EC50 = 25.8 ± 3.1 μM) than the wild-type virus (EC50 = 0.36 ± 0.11 μM). We conclude, therefore, that this mutation is sufficient for drug resistance and that pUL97 is involved in the mechanism of action of CPV.

  1. Human cytomegalovirus infant infection adversely affects growth and development in maternally HIV-exposed and unexposed infants in Zambia.

    Science.gov (United States)

    Gompels, U A; Larke, N; Sanz-Ramos, M; Bates, M; Musonda, K; Manno, D; Siame, J; Monze, M; Filteau, S

    2012-02-01

    Human immunodeficiency virus (HIV) and human cytomegalovirus (HCMV) coinfections have been shown to increase infant morbidity, mortality, and AIDS progression. In HIV-endemic regions, maternal HIV-exposed but HIV-uninfected infants, which is the majority of children affected by HIV, also show poor growth and increased morbidity. Although nutrition has been examined, the effects of HCMV infection have not been evaluated. We studied the effects of HCMV infection on the growth, development, and health of maternally HIV-exposed and unexposed infants in Zambia. Infants were examined in a cohort recruited to a trial of micronutrient-fortified complementary foods. HIV-infected mothers and infants had received perinatal antiretroviral therapy to prevent mother-to-child HIV transmission. Growth, development, and morbidity were analyzed by linear regression analyses in relation to maternal HIV exposure and HCMV infection, as screened by sera DNA for viremia at 6 months of age and by antibody for infection at 18 months. All HCMV-seropositive infants had decreased length-for-age by 18 months compared with seronegative infants (standard deviation [z]-score difference: -0.44 [95% confidence interval {CI}, -.72 to -.17]; P = .002). In HIV-exposed infants, those who were HCMV positive compared with those who were negative, also had reduced head size (mean z-score difference: -0.72 [95% CI, -1.23 to -.22]; P = .01) and lower psychomotor development (Bayley test score difference: -4.1 [95% CI, -7.8 to -.5]; P = .03). HIV-exposed, HCMV-viremic infants were more commonly referred for hospital treatment than HCMV-negative infants. The effects of HCMV were unaffected by micronutrient fortification. HCMV affects child growth, development, and morbidity of African infants, particularly in those maternally exposed to HIV. HCMV is therefore a risk factor for child health in this region.

  2. Human cytomegalovirus TRS1 protein associates with the 7-methylguanosine mRNA cap and facilitates translation.

    Science.gov (United States)

    Ziehr, Benjamin; Lenarcic, Erik; Vincent, Heather A; Cecil, Chad; Garcia, Benjamin; Shenk, Thomas; Moorman, Nathaniel J

    2015-06-01

    Viruses rely on the host translation machinery for the synthesis of viral proteins. Human cells have evolved sensors that recognize viral RNAs and inhibit mRNA translation in order to limit virus replication. Understanding how viruses manipulate the host translation machinery to gain access to ribosomes and disable the antiviral response is therefore a critical aspect of the host/pathogen interface. In this study, we used a proteomics approach to identify human cytomegalovirus (HCMV) proteins that might contribute to viral mRNA translation. The HCMV TRS1 protein (pTRS1) associated with the 7-methylguanosine mRNA cap, increased the total level of protein synthesis, and colocalized with mRNAs undergoing translation initiation during infection. pTRS1 stimulated translation of a nonviral reporter gene and increased the translation of a reporter containing an HCMV 5' untranslated region (5'UTR) to a greater extent. The preferential effect of pTRS1 on translation of an mRNA containing a viral 5'UTR required the pTRS1 RNA and double-stranded RNA-dependent protein kinase (PKR)-binding domains, and was likely the result of PKR inhibition. However, pTRS1 also stimulated the total level of protein synthesis and translation directed by an HCMV 5'UTR in cells lacking PKR. Thus our results demonstrate that pTRS1 stimulates translation through both PKR-dependent and PKR-independent mechanisms. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. The human cytomegalovirus DNA polymerase processivity factor UL44 is modified by SUMO in a DNA-dependent manner.

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    Elisa Sinigalia

    Full Text Available During the replication of human cytomegalovirus (HCMV genome, the viral DNA polymerase subunit UL44 plays a key role, as by binding both DNA and the polymerase catalytic subunit it confers processivity to the holoenzyme. However, several lines of evidence suggest that UL44 might have additional roles during virus life cycle. To shed light on this, we searched for cellular partners of UL44 by yeast two-hybrid screenings. Intriguingly, we discovered the interaction of UL44 with Ubc9, an enzyme involved in the covalent conjugation of SUMO (Small Ubiquitin-related MOdifier to cellular and viral proteins. We found that UL44 can be extensively sumoylated not only in a cell-free system and in transfected cells, but also in HCMV-infected cells, in which about 50% of the protein resulted to be modified at late times post-infection, when viral genome replication is accomplished. Mass spectrometry studies revealed that UL44 possesses multiple SUMO target sites, located throughout the protein. Remarkably, we observed that binding of UL44 to DNA greatly stimulates its sumoylation both in vitro and in vivo. In addition, we showed that overexpression of SUMO alters the intranuclear distribution of UL44 in HCMV-infected cells, and enhances both virus production and DNA replication, arguing for an important role for sumoylation in HCMV life cycle and UL44 function(s. These data report for the first time the sumoylation of a viral processivity factor and show that there is a functional interplay between the HCMV UL44 protein and the cellular sumoylation system.

  4. Pharmacokinetics and safety of letermovir, a novel anti-human cytomegalovirus drug, in patients with renal impairment.

    Science.gov (United States)

    Kropeit, Dirk; Scheuenpflug, Jürgen; Erb-Zohar, Katharina; Halabi, Atef; Stobernack, Hans-Peter; Hulskotte, Ellen G J; van Schanke, Arne; Zimmermann, Holger; Rübsamen-Schaeff, Helga

    2017-09-01

    Human cytomegalovirus remains a significant issue for immunocompromised patients and existing viral polymerase targeting therapies are associated with significant toxicity. Accordingly, the viral terminase complex inhibitor, letermovir, is in development. We assessed letermovir pharmacokinetics in renal impairment. This was a Phase 1, open-label, nonrandomised trial. Estimated glomerular filtration rate based on the Modification of Diet Renal Disease equation was used to create three groups of eight subjects: healthy function (estimated glomerular filtration rate ≥ 90 ml min(-1)  1.73m(-2) ), moderate (30-59 ml min(-1)  1.73m(-2) ) and severe (letermovir 120 mg was dosed once-daily for 8 days and blood collected for pharmacokinetic analyses. All 24 subjects enrolled completed the trial. Moderate and severe renal impairment increased mean unbound letermovir fractions by 11% and 26%, respectively, vs. healthy subjects. Exposure (AUCτ,ss and Css,max ) was increased with renal impairment [least square mean ratios (90% confidence intervals) total letermovir vs. healthy subjects, AUCτ,ss 192% (143-258%) and 142% (83-243%) for moderate and severe impairment, respectively; Css,max 125% (87-182%) and 106% (75-151%), respectively]. Clearance was decreased vs. healthy subjects. Correlation analyses indicated a correlation between decreasing renal function and increased unbound letermovir concentration (R(2)  = 0.5076, P letermovir 120 mg was well tolerated across groups. Renal impairment increased exposure to letermovir, although age was a confounding factor. © 2017 The British Pharmacological Society.

  5. Is human cytomegalovirus infection associated with hypertension? The United States National Health and Nutrition Examination Survey 1999-2002.

    Science.gov (United States)

    Li, Chao; Samaranayake, Nithushi R; Ong, Kwok Leung; Wong, Hoi Kin; Cheung, Bernard M Y

    2012-01-01

    Recent studies have implicated the human cytomegalovirus (HCMV) as a possible pathogen for causing hypertension. We aimed to study the association between HCMV infection and hypertension in the United States National Health and Nutrition Examination Survey (NHANES). We analyzed data on 2979 men and 3324 women in the NHANES 1999-2002. We included participants aged 16-49 years who had valid data on HCMV infection and hypertension. Of the participants, 54.7% had serologic evidence of HCMV infection and 17.5% had hypertension. There were ethnic differences in the prevalence of HCMV infection (P<0.001) and hypertension (P<0.001). The prevalence of both increased with age (P<0.001). Before adjustment, HCMV seropositivity was significantly associated with hypertension in women (OR=1.63, 95% CI=1.25-2.13, P=0.001) but not in men. After adjustment for race/ethnicity, the association between HCMV seropositivity and hypertension in women remained significant (OR=1.55, 95% CI=1.20-2.02, P=0.002). Further adjustment for body mass index, diabetes status and hypercholesterolemia attenuated the association (OR=1.44, 95% CI=1.10-1.90, P=0.010). However, after adjusting for age, the association was no longer significant (OR=1.24, 95% CI=0.91-1.67, P=0.162). In this nationally representative population-based survey, HCMV seropositivity is associated with hypertension in women in the NHANES population. This association is largely explained by the association of hypertension with age and the increase in past exposure to HCMV with age.

  6. Is Human Cytomegalovirus Infection Associated with Hypertension? The United States National Health and Nutrition Examination Survey 1999–2002

    Science.gov (United States)

    Li, Chao; Samaranayake, Nithushi R.; Ong, Kwok Leung; Wong, Hoi Kin; Cheung, Bernard M. Y.

    2012-01-01

    Purpose Recent studies have implicated the human cytomegalovirus (HCMV) as a possible pathogen for causing hypertension. We aimed to study the association between HCMV infection and hypertension in the United States National Health and Nutrition Examination Survey (NHANES). Methods We analyzed data on 2979 men and 3324 women in the NHANES 1999–2002. We included participants aged 16–49 years who had valid data on HCMV infection and hypertension. Results Of the participants, 54.7% had serologic evidence of HCMV infection and 17.5% had hypertension. There were ethnic differences in the prevalence of HCMV infection (P<0.001) and hypertension (P<0.001). The prevalence of both increased with age (P<0.001). Before adjustment, HCMV seropositivity was significantly associated with hypertension in women (OR = 1.63, 95% CI = 1.25–2.13, P = 0.001) but not in men. After adjustment for race/ethnicity, the association between HCMV seropositivity and hypertension in women remained significant (OR = 1.55, 95% CI = 1.20–2.02, P = 0.002). Further adjustment for body mass index, diabetes status and hypercholesterolemia attenuated the association (OR = 1.44, 95% CI = 1.10–1.90, P = 0.010). However, after adjusting for age, the association was no longer significant (OR = 1.24, 95% CI = 0.91–1.67, P = 0.162). Conclusions In this nationally representative population-based survey, HCMV seropositivity is associated with hypertension in women in the NHANES population. This association is largely explained by the association of hypertension with age and the increase in past exposure to HCMV with age. PMID:22768311

  7. Is human cytomegalovirus infection associated with hypertension? The United States National Health and Nutrition Examination Survey 1999-2002.

    Directory of Open Access Journals (Sweden)

    Chao Li

    Full Text Available PURPOSE: Recent studies have implicated the human cytomegalovirus (HCMV as a possible pathogen for causing hypertension. We aimed to study the association between HCMV infection and hypertension in the United States National Health and Nutrition Examination Survey (NHANES. METHODS: We analyzed data on 2979 men and 3324 women in the NHANES 1999-2002. We included participants aged 16-49 years who had valid data on HCMV infection and hypertension. RESULTS: Of the participants, 54.7% had serologic evidence of HCMV infection and 17.5% had hypertension. There were ethnic differences in the prevalence of HCMV infection (P<0.001 and hypertension (P<0.001. The prevalence of both increased with age (P<0.001. Before adjustment, HCMV seropositivity was significantly associated with hypertension in women (OR=1.63, 95% CI=1.25-2.13, P=0.001 but not in men. After adjustment for race/ethnicity, the association between HCMV seropositivity and hypertension in women remained significant (OR=1.55, 95% CI=1.20-2.02, P=0.002. Further adjustment for body mass index, diabetes status and hypercholesterolemia attenuated the association (OR=1.44, 95% CI=1.10-1.90, P=0.010. However, after adjusting for age, the association was no longer significant (OR=1.24, 95% CI=0.91-1.67, P=0.162. CONCLUSIONS: In this nationally representative population-based survey, HCMV seropositivity is associated with hypertension in women in the NHANES population. This association is largely explained by the association of hypertension with age and the increase in past exposure to HCMV with age.

  8. Role of breast milk in acquisition of cytomegalovirus infection: recent advances.

    Science.gov (United States)

    Schleiss, Mark R

    2006-02-01

    Congenital infection with cytomegalovirus is a major cause of disability in newborns. Recently, there has been increased emphasis on the study of postnatally acquired cytomegalovirus infection. One route by which cytomegalovirus infections are acquired in newborns is via consumption of breast milk from cytomegalovirus-seropositive, lactating mothers. The purpose of this review is to summarize recent studies of breast-milk-acquired cytomegalovirus infections in newborns, particularly in low-birth-weight premature infants. Nearly all cytomegalovirus-seropositive women will reactivate and shed cytomegalovirus during lactation, as demonstrated by sensitive polymerase chain reaction techniques, as well as by viral culture of breast milk. A substantial proportion of infants exposed to cytomegalovirus in breast milk will acquire a primary cytomegalovirus infection. Although acquisition of cytomegalovirus by this route is seldom of consequence in healthy term infants, cytomegalovirus infections in low-birth-weight premature infants have been demonstrated to cause symptomatic illness, including hepatitis, neutropenia, thrombocytopenia, and a 'sepsis-like' state. Cytomegalovirus is commonly shed in human milk, and cytomegalovirus-seropositive women can transmit this infection via breast-feeding. The benefits of breast-feeding greatly outweigh the minimal risk, if any, of infections transmitted to term infants. Caution is warranted, however, in low-birth-weight premature infants, who are at increased risk of cytomegalovirus disease. Interventions to screen breast milk, or to attempt to render breast milk noninfectious through treatments such as freezing, may be warranted in high-risk premature infants.

  9. Drug Susceptibility and Replicative Capacity of Multi-Drug Resistant Recombinant Human Cytomegalovirus Harboring Mutations in UL56 and UL54 Genes.

    Science.gov (United States)

    Piret, Jocelyne; Goyette, Nathalie; Boivin, Guy

    2017-08-14

    Letermovir is an investigational antiviral agent with a novel mechanism of action involving the viral terminase (pUL56). We evaluated the impact of V236M mutation in UL56 gene alone and combined with E756K mutation in UL54 gene on drug susceptibility and viral replicative capacity of recombinant human cytomegalovirus. The double mutant exhibited at least borderline resistance to all antivirals tested (ganciclovir, foscarnet, cidofovir, brincidofovir and letermovir) and replicated less efficiently than the wild-type virus in vitro. Copyright © 2017 American Society for Microbiology.

  10. Oncogenic role of cytomegalovirus in medulloblastoma?

    Science.gov (United States)

    Hortal, Alejandro M; Vermeulen, Jeroen F; Van Hecke, Wim; Bovenschen, Niels

    2017-11-01

    Medulloblastoma is the most common solid tumor among children. Current therapeutic strategies for this malignancy include surgical resection, radiation therapy and chemotherapy. However, these treatments are accompanied with serious side effects such as neurological complications and psychosocial problems, due to the severity of treatment on the developing nervous system. To solve this problem, novel therapeutic approaches are currently being investigated. One of them is targeting human cytomegalovirus in medulloblastoma cancer cells. However, this approach is still under debate, since the presence of cytomegalovirus in medulloblastomas remains controversial. In this review, we discuss the current controversies on the role of cytomegalovirus in medulloblastoma oncogenesis and the potential of cytomegalovirus as a novel (immuno)therapeutic target. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

  11. Inhibition of human cytomegalovirus immediate-early gene expression and replication by the ethyl acetate (EtOAc) fraction of Elaeocarpus sylvestris in vitro.

    Science.gov (United States)

    Bae, Sohee; Kang, Se Chan; Song, Yoon-Jae

    2017-08-29

    In immunocompromised patients, human cytomegalovirus (HCMV) infection can lead to severe, life-threatening diseases, such as pneumonitis, hepatitis, gastrointestinal tract disease, and retinitis. We previously reported that a 70% ethanol extract of Elaeocarpus sylvestris leaves (ESE) inhibits human cytomegalovirus (HCMV) replication in vitro. In the present study, we determined the solvent fraction of ESE that inhibits HCMV replication using activity-guided fractionation. Activity-guided fractionation of ESE was performed to determine the solvent fraction that inhibits HCMV replication. Effects of solvent fractions on HCMV lytic gene expression and major immediate-early (MIE) enhancer/promoter activity were further investigated. Among the solvent fractions tested, the EtOAc fraction of ESE markedly reduced HCMV lytic gene expression and viral replication in vitro without exerting significant cytotoxic effects against human foreskin fibroblasts (HFF). Furthermore, the EtOAc fraction negatively affected HCMV MIE enhancer/promoter activity. Our data collectively indicate that the EtOAc fraction of ESE contains active constituents that inhibit HCMV MIE enhancer/promoter activity and viral replication. The EtOAc fraction of ESE is a good source of novel drug candidates for treatment of HCMV-associated diseases.

  12. Cytomegalovirus Colitis: An Uncommon Mimicker of Common Colitides.

    Science.gov (United States)

    Baniak, Nick; Kanthan, Rani

    2016-08-01

    Cytomegalovirus latency, though ubiquitous in the human population, is known to cause colitis in both immunocompromised and immunocompetent hosts. Furthermore, the clinical, endoscopic, and histologic appearance of cytomegalovirus colitis can mimic that of inflammatory bowel disease, an extremely well-documented disease. In this context, though many reports have looked at inflammatory bowel disease with superimposed cytomegalovirus infection, less attention has been paid to cytomegalovirus as a primary cause of isolated colitis. Owing to the rarity of this phenomenon, it is important to consider this diagnosis and implement proper testing to avoid misdiagnosis and mismanagement.

  13. Human cytomegalovirus latency-associated proteins elicit immune-suppressive IL-10 producing CD4⁺ T cells.

    Directory of Open Access Journals (Sweden)

    Gavin M Mason

    Full Text Available Human cytomegalovirus (HCMV is a widely prevalent human herpesvirus, which, after primary infection, persists in the host for life. In healthy individuals, the virus is well controlled by the HCMV-specific T cell response. A key feature of this persistence, in the face of a normally robust host immune response, is the establishment of viral latency. In contrast to lytic infection, which is characterised by extensive viral gene expression and virus production, long-term latency in cells of the myeloid lineage is characterised by highly restricted expression of viral genes, including UL138 and LUNA. Here we report that both UL138 and LUNA-specific T cells were detectable directly ex vivo in healthy HCMV seropositive subjects and that this response is principally CD4⁺ T cell mediated. These UL138-specific CD4⁺ T cells are able to mediate MHC class II restricted cytotoxicity and, importantly, show IFNγ effector function in the context of both lytic and latent infection. Furthermore, in contrast to CDCD4⁺ T cells specific to antigens expressed solely during lytic infection, both the UL138 and LUNA-specific CD4⁺ T cell responses included CD4⁺ T cells that secreted the immunosuppressive cytokine cIL-10. We also show that cIL-10 expressing CD4⁺ T-cells are directed against latently expressed US28 and UL111A. Taken together, our data show that latency-associated gene products of HCMV generate CD4⁺ T cell responses in vivo, which are able to elicit effector function in response to both lytic and latently infected cells. Importantly and in contrast to CD4⁺ T cell populations, which recognise antigens solely expressed during lytic infection, include a subset of cells that secrete the immunosuppressive cytokine cIL-10. This suggests that HCMV skews the T cell responses to latency-associated antigens to one that is overall suppressive in order to sustain latent carriage in vivo.

  14. Mutual Interplay between the Human Cytomegalovirus Terminase Subunits pUL51, pUL56, and pUL89 Promotes Terminase Complex Formation.

    Science.gov (United States)

    Neuber, Sebastian; Wagner, Karen; Goldner, Thomas; Lischka, Peter; Steinbrueck, Lars; Messerle, Martin; Borst, Eva Maria

    2017-06-15

    Human cytomegalovirus (HCMV) genome encapsidation requires several essential viral proteins, among them pUL56, pUL89, and the recently described pUL51, which constitute the viral terminase. To gain insight into terminase complex assembly, we investigated interactions between the individual subunits. For analysis in the viral context, HCMV bacterial artificial chromosomes carrying deletions in the open reading frames encoding the terminase proteins were used. These experiments were complemented by transient-transfection assays with plasmids expressing the terminase components. We found that if one terminase protein was missing, the levels of the other terminase proteins were markedly diminished, which could be overcome by proteasome inhibition or providing the missing subunit in trans These data imply that sequestration of the individual subunits within the terminase complex protects them from proteasomal turnover. The finding that efficient interactions among the terminase proteins occurred only when all three were present together is reminiscent of a folding-upon-binding principle leading to cooperative stability. Furthermore, whereas pUL56 was translocated into the nucleus on its own, correct nuclear localization of pUL51 and pUL89 again required all three terminase constituents. Altogether, these features point to a model of the HCMV terminase as a multiprotein complex in which the three players regulate each other concerning stability, subcellular localization, and assembly into the functional tripartite holoenzyme.IMPORTANCE HCMV is a major risk factor in immunocompromised individuals, and congenital CMV infection is the leading viral cause for long-term sequelae, including deafness and mental retardation. The current treatment of CMV disease is based on drugs sharing the same mechanism, namely, inhibiting viral DNA replication, and often results in adverse side effects and the appearance of resistant virus strains. Recently, the HCMV terminase has emerged as

  15. Cytomegalovirus Retinitis: A Review.

    Science.gov (United States)

    Port, Alexander D; Orlin, Anton; Kiss, Szilard; Patel, Sarju; D'Amico, Donald J; Gupta, Mrinali P

    2017-05-01

    Cytomegalovirus (CMV) is a ubiquitous DNA herpes virus that causes significant morbidity and mortality in immunocompromised individuals. CMV retinitis is a potentially blinding manifestation of CMV infection that was commonly seen in advanced acquired immunodeficiency syndrome (AIDS) in the era before modern combination antiretroviral therapy era, but is also recognized in patients with immune deficiency from multiple causes. The advent of and advances in antiretroviral therapies for human immunodeficiency virus have decreased the incidence of CMV retinitis by over 90% among AIDS patients, and improved visual outcomes in those affected. The diagnosis is generally a clinical one, and treatment modalities include systemic and intravitreal antiviral medications. Retinal detachment and immune recovery uveitis are sight-threatening complications of CMV retinitis that require specific treatments.

  16. Incidence of human herpes virus-6 and human cytomegalovirus infections in donated bone marrow and umbilical cord blood hematopoietic stem cells

    Directory of Open Access Journals (Sweden)

    Behzad-Behbahani A

    2008-01-01

    Full Text Available This study examined the incidence of human herpes virus-6 (HHV-6 and human cytomegalovirus (HCMV infections that are potentially transmitted to haematopoietic stem cells (HSC transplant recipients via bone marrow (BM or umbilical cord blood (UCB. Bone marrow progenitor cells were collected from 30 allogenic BM donors. UCB HSC were collected from 34 subjects. The extracted DNA was then processed using nested polymerase chain reaction (nPCR technique. HCMV and HHV-6 serological status were determined by enzyme immunoassay (EIA. Nested PCR identified HCMV in 22 (73% of 30 samples of BM progenitor cells but in only eight (23.5% of 34 samples of UBC HSC ( P = 0.001. HHV-6 DNA was detected in 11 (36.6% of 30 BM progenitor cells and in only one (2.9% of 34 UBC cells ( P = 0.002. Both HHV-6 and HCMV infections were determined in nine (26.5% of 34 bone marrow samples. The results indicate that, the risk of HCMV and HHV-6 via BM progenitor cells is higher than transmission by UCB cells ( P= 0.04.

  17. Long and Short Isoforms of the Human Cytomegalovirus UL138 Protein Silence IE Transcription and Promote Latency.

    Science.gov (United States)

    Lee, Song Hee; Caviness, Katie; Albright, Emily R; Lee, Jeong-Hee; Gelbmann, Christopher B; Rak, Mike; Goodrum, Felicia; Kalejta, Robert F

    2016-10-15

    The UL133-138 locus present in clinical strains of human cytomegalovirus (HCMV) encodes proteins required for latency and reactivation in CD34(+) hematopoietic progenitor cells and virion maturation in endothelial cells. The encoded proteins form multiple homo- and hetero-interactions and localize within secretory membranes. One of these genes, UL136 gene, is expressed as at least five different protein isoforms with overlapping and unique functions. Here we show that another gene from this locus, the UL138 gene, also generates more than one protein isoform. A long form of UL138 (pUL138-L) initiates translation from codon 1, possesses an amino-terminal signal sequence, and is a type one integral membrane protein. Here we identify a short protein isoform (pUL138-S) initiating from codon 16 that displays a subcellular localization similar to that of pUL138-L. Reporter, short-term transcription, and long-term virus production assays revealed that both pUL138-L and pUL138-S are able to suppress major immediate early (IE) gene transcription and the generation of infectious virions in cells in which HCMV latency is studied. The long form appears to be more potent at silencing IE transcription shortly after infection, while the short form seems more potent at restricting progeny virion production at later times, indicating that both isoforms of UL138 likely cooperate to promote HCMV latency. Latency allows herpesviruses to persist for the lives of their hosts in the face of effective immune control measures for productively infected cells. Controlling latent reservoirs is an attractive antiviral approach complicated by knowledge deficits for how latently infected cells are established, maintained, and reactivated. This is especially true for betaherpesviruses. The functional consequences of HCMV UL138 protein expression during latency include repression of viral IE1 transcription and suppression of virus replication. Here we show that short and long isoforms of UL138

  18. Complex expression of the UL136 gene of human cytomegalovirus results in multiple protein isoforms with unique roles in replication.

    Science.gov (United States)

    Caviness, Katie; Cicchini, Louis; Rak, Michael; Umashankar, Mahadevaiah; Goodrum, Felicia

    2014-12-01

    Human cytomegalovirus (HCMV) is a complex DNA virus with a 230-kb genome encoding 170 and up to 750 proteins. The upper limit of this coding capacity suggests the evolution of complex mechanisms to substantially increase the coding potential from the 230-kb genome. Our work examines the complexity of one gene, UL136, encoded within the ULb' region of the genome that is lost during serial passage of HCMV in cultured fibroblasts. UL136 is expressed as five protein isoforms. We mapped these isoforms and demonstrate that they originate from both a complex transcriptional profile and, possibly, the usage of multiple translation initiation sites. Intriguingly, the pUL136 isoforms exhibited distinct subcellular distributions with varying association with the Golgi apparatus. The subcellular localization of membrane-bound isoforms of UL136 differed between when they were expressed exogenously and when they were expressed in the context of viral infection, suggesting that the trafficking of these isoforms is mediated by infection-specific factors. While UL136, like most ULb' genes, was dispensable for replication in fibroblasts, the soluble 23- and 19-kDa isoforms suppressed virus replication. In CD34(+) hematopoietic progenitor cells (HPCs) infected in vitro, disruption of the 23- and 19-kDa isoforms resulted in increased replication and a loss of the latency phenotype, similar to the effects of the UL138 latency determinant encoded within the same genetic locus. Our work suggests a complex interplay between the UL136 isoforms which balances viral replication in multiple cell types and likely contributes to the cell type-dependent phenotypes of the UL133/8 locus and the outcome of HCMV infection. HCMV is a significant cause of morbidity in immunocompromised individuals, including transplant patients. The lifelong persistence of the virus results in a high seroprevalence worldwide and may contribute to age-related pathologies, such as atherosclerosis. The mechanisms of

  19. Structural Basis for Clonal Diversity of the Public T Cell Response to a Dominant Human Cytomegalovirus Epitope.

    Science.gov (United States)

    Yang, Xinbo; Gao, Mingming; Chen, Guobing; Pierce, Brian G; Lu, Jinghua; Weng, Nan-Ping; Mariuzza, Roy A

    2015-11-27

    Cytomegalovirus (CMV) is a ubiquitous and persistent human pathogen that is kept in check by CD8(+) cytotoxic T lymphocytes. Individuals expressing the major histocompatibility complex (MHC) class I molecule HLA-A2 produce cytotoxic T lymphocytes bearing T cell receptors (TCRs) that recognize the immunodominant CMV epitope NLVPMVATV (NLV). The NLV-specific T cell repertoire is characterized by a high prevalence of TCRs that are frequently observed in multiple unrelated individuals. These public TCRs feature identical, or nearly identical, complementarity-determining region 3α (CDR3α) and/or CDR3β sequences. The TCRs may express public CDR3α motifs alone, public CDR3β motifs alone, or dual public CDR3αβ motifs. In addition, the same public CDR3α motif may pair with different CDR3β motifs (and the reverse), giving rise to highly diverse NLV-specific TCR repertoires. To investigate the structural underpinnings of this clonal diversity, we determined crystal structures of two public TCRs (C7 and C25) in complex with NLV·HLA-A2. These TCRs utilize completely different CDR3α and CDR3β motifs that, in addition, can associate with multiple variable α and variable β regions in NLV-specific T cell repertoires. The C7·NLV·HLA-A2 and C25·NLV·HLA-A2 complexes exhibit divergent TCR footprints on peptide-MHC such that C25 is more focused on the central portion of the NLV peptide than is C7. These structures combined with molecular modeling show how the public CDR3α motif of C25 may associate with different variable α regions and how the public CDR3α motif of C7 may pair with different CDR3β motifs. This interchangeability of TCR V regions and CDR3 motifs permits multiple structural solutions to binding an identical peptide-MHC ligand and thereby the generation of a clonally diverse public T cell response to CMV. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Structural Basis for Clonal Diversity of the Public T Cell Response to a Dominant Human Cytomegalovirus Epitope*

    Science.gov (United States)

    Yang, Xinbo; Gao, Mingming; Chen, Guobing; Pierce, Brian G.; Lu, Jinghua; Weng, Nan-ping; Mariuzza, Roy A.

    2015-01-01

    Cytomegalovirus (CMV) is a ubiquitous and persistent human pathogen that is kept in check by CD8+ cytotoxic T lymphocytes. Individuals expressing the major histocompatibility complex (MHC) class I molecule HLA-A2 produce cytotoxic T lymphocytes bearing T cell receptors (TCRs) that recognize the immunodominant CMV epitope NLVPMVATV (NLV). The NLV-specific T cell repertoire is characterized by a high prevalence of TCRs that are frequently observed in multiple unrelated individuals. These public TCRs feature identical, or nearly identical, complementarity-determining region 3α (CDR3α) and/or CDR3β sequences. The TCRs may express public CDR3α motifs alone, public CDR3β motifs alone, or dual public CDR3αβ motifs. In addition, the same public CDR3α motif may pair with different CDR3β motifs (and the reverse), giving rise to highly diverse NLV-specific TCR repertoires. To investigate the structural underpinnings of this clonal diversity, we determined crystal structures of two public TCRs (C7 and C25) in complex with NLV·HLA-A2. These TCRs utilize completely different CDR3α and CDR3β motifs that, in addition, can associate with multiple variable α and variable β regions in NLV-specific T cell repertoires. The C7·NLV·HLA-A2 and C25·NLV·HLA-A2 complexes exhibit divergent TCR footprints on peptide-MHC such that C25 is more focused on the central portion of the NLV peptide than is C7. These structures combined with molecular modeling show how the public CDR3α motif of C25 may associate with different variable α regions and how the public CDR3α motif of C7 may pair with different CDR3β motifs. This interchangeability of TCR V regions and CDR3 motifs permits multiple structural solutions to binding an identical peptide-MHC ligand and thereby the generation of a clonally diverse public T cell response to CMV. PMID:26429912

  1. Regulation and gene expression profiling of NKG2D positive human cytomegalovirus-primed CD4+ T-cells.

    Directory of Open Access Journals (Sweden)

    Helle Jensen

    Full Text Available NKG2D is a stimulatory receptor expressed by natural killer (NK cells, CD8(+ T-cells, and γδ T-cells. NKG2D expression is normally absent from CD4(+ T-cells, however recently a subset of NKG2D(+ CD4(+ T-cells has been found, which is specific for human cytomegalovirus (HCMV. This particular subset of HCMV-specific NKG2D(+ CD4(+ T-cells possesses effector-like functions, thus resembling the subsets of NKG2D(+ CD4(+ T-cells found in other chronic inflammations. However, the precise mechanism leading to NKG2D expression on HCMV-specific CD4(+ T-cells is currently not known. In this study we used genome-wide analysis of individual genes and gene set enrichment analysis (GSEA to investigate the gene expression profile of NKG2D(+ CD4(+ T-cells, generated from HCMV-primed CD4(+ T-cells. We show that the HCMV-primed NKG2D(+ CD4(+ T-cells possess a higher differentiated phenotype than the NKG2D(- CD4(+ T-cells, both at the gene expression profile and cytokine profile. The ability to express NKG2D at the cell surface was primarily determined by the activation or differentiation status of the CD4(+ T-cells and not by the antigen presenting cells. We observed a correlation between CD94 and NKG2D expression in the CD4(+ T-cells following HCMV stimulation. However, knock-down of CD94 did not affect NKG2D cell surface expression or signaling. In addition, we show that NKG2D is recycled at the cell surface of activated CD4(+ T-cells, whereas it is produced de novo in resting CD4(+ T-cells. These findings provide novel information about the gene expression profile of HCMV-primed NKG2D(+ CD4(+ T-cells, as well as the mechanisms regulating NKG2D cell surface expression.

  2. Replication stress, DNA damage signalling, and cytomegalovirus infection in human medulloblastomas

    DEFF Research Database (Denmark)

    Bartek, Jiri; Fornara, Olesja; Merchut-Maya, Joanna Maria

    2017-01-01

    of the clinical specimens also showed expression of HCMV immediate early and late proteins, in comparative analyses using three immunohistochemical protocols. Cell culture experiments validated the chronic endogenous replication stress in medulloblastoma cell lines and showed sharply differential, intriguing...... eight established immunohistochemical markers to assess the status of the DDR machinery, we found pronounced endogenous DNA damage signalling (γH2AX marker) and robust constitutive activation of both the ATM-Chk2 and ATR-Chk1 DNA damage checkpoint kinase cascades, yet unexpectedly modest p53 tumour...... responses of normal cells and medulloblastoma cells to HCMV infection, including differential subcellular mislocalization and enhancement of replication stress-related 53BP1 body formation, the latter in cell-non-autonomous manner. Overall, our results strongly indicate that in human medulloblastomas...

  3. World population, human disaster, and nuclear holocaust.

    Science.gov (United States)

    Giddings, J C

    1973-09-01

    Too little attention is being paid to the theoretical mechanics of dealing with the optimum level of human population. Yet without such efforts, population goals are essentially meaningless and there is little incentive to curtail population growth. In an attempt to take a step in that direction, the author discusses a broad class of interrelated phenomena--consisting of various threats to human existence--that hinge strongly on population density. Then he develops simple conceptual models that provide a quantitative population dependence for various threats. These are always expressed as a simple power of population. Only threats of disasters that increase with an increase in population density are considered, eliminating most natural disasters. Threats of the 1st order, such as those from nuclear wastes and chemical pollutants, increase with an increase in population and a resultant increase in production. However, with increasing crowding and dependency upon high energy systems to provide basic resources, many 1st order threats can become threats of the 2nd of higher orders. The burial of nuclear wastes, for example, presents a threat of the 2nd order by affecting 1) the integrity of the geological formation at the burial site; and 2) the quantity of the radioactive material. As the relationship between population and interdependency strengthens, the crucial question will concern the stability of the independent networks; 1 error, due to human frailty or malice, can destroy a chain supplying food, energy, communications, or water. Into this system are thrown nuclear weapons, which, through human error or malice, could demolish the world. To illuminate some of the major elements of the nuclear threat, a model based on the threshold concept of nuclear armaments was created. This concept recognizes that any alliance of people possessing a minimum of technological and economic resources could build and deploy nuclear weapons. Most alliances will constitute no danger

  4. Rapid detection of cytomegalovirus in bronchoalveolar lavage fluid and serum samples by polymerase chain reaction: correlation of virus isolation and clinical outcome for patients with human immunodeficiency virus infection

    DEFF Research Database (Denmark)

    Hansen, K K; Vestbo, Jørgen; Benfield, T

    1997-01-01

    Bronchoalveolar lavage (BAL) fluids and serum samples from 153 patients with pulmonary symptoms who were infected with human immunodeficiency virus (HIV) and underwent BAL were examined for the presence of cytomegalovirus (CMV) by conventional culture and by polymerase chain reaction (PCR...

  5. Dissecting human cytomegalovirus gene function and capsid maturation by ribozyme targeting and electron cryomicroscopy.

    Science.gov (United States)

    Yu, Xuekui; Trang, Phong; Shah, Sanket; Atanasov, Ivo; Kim, Yong-Hwan; Bai, Yong; Zhou, Z Hong; Liu, Fenyong

    2005-05-17

    Human CMV (HCMV) is the leading viral cause of birth defects and causes one of the most common opportunistic infections among transplant recipients and AIDS patients. Cleavage of internal scaffolding proteins by the viral protease (Pr) occurs during HCMV capsid assembly. To gain insight into the mechanism of HCMV capsid maturation and the roles of the Pr in viral replication, an RNase P ribozyme was engineered to target the Pr mRNA and down-regulate its expression by >99%, generating premature Pr-minus capsids. Furthermore, scaffolding protein processing and DNA encapsidation were inhibited by 99%, and viral growth was reduced by 10,000-fold. 3D structural comparison of the Pr-minus and wild-type B capsids by electron cryomicroscopy, at an unprecedented 12.5-angstroms resolution, unexpectedly revealed that the structures are identical in their overall shape and organization. However, the Pr-minus capsid contains tenuous connections between the scaffold and the capsid shell, whereas the wild-type B capsid has extra densities in its core that may represent the viral Pr. Our findings indicate that cleavage of the scaffolding protein is not associated with the morphological changes that occur during capsid maturation. Instead, the protease appears to be required for DNA encapsidation and the subsequent maturation steps leading to infectious progeny. These results therefore provide key insights into an essential step of HCMV infection using an RNase P ribozyme-based inhibition strategy.

  6. New efficient artemisinin derived agents against human leukemia cells, human cytomegalovirus and Plasmodium falciparum: 2nd generation 1,2,4-trioxane-ferrocene hybrids.

    Science.gov (United States)

    Reiter, Christoph; Fröhlich, Tony; Zeino, Maen; Marschall, Manfred; Bahsi, Hanife; Leidenberger, Maria; Friedrich, Oliver; Kappes, Barbara; Hampel, Frank; Efferth, Thomas; Tsogoeva, Svetlana B

    2015-06-05

    In our ongoing search for highly active hybrid molecules exceeding their parent compounds in anticancer, antimalaria as well as antiviral activity and being an alternative to the standard drugs, we present the synthesis and biological investigations of 2nd generation 1,2,4-trioxane-ferrocene hybrids. In vitro tests against the CCRF-CEM leukemia cell line revealed di-1,2,4-trioxane-ferrocene hybrid 7 as the most active compound (IC50 of 0.01 μM). Regarding the activity against the multidrug resistant subline CEM/ADR5000, 1,2,4-trioxane-ferrocene hybrid 5 showed a remarkable activity (IC50 of 0.53 μM). Contrary to the antimalaria activity of hybrids 4-8 against Plasmodium falciparum 3D7 strain with slightly higher IC50 values (between 7.2 and 30.2 nM) than that of their parent compound DHA, hybrids 5-7 possessed very promising activity (IC50 values lower than 0.5 μM) against human cytomegalovirus (HCMV). The application of 1,2,4-trioxane-ferrocene hybrids against HCMV is unprecedented and demonstrated here for the first time. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  7. Low Expression of Estrogen Receptor-α and Progesterone Receptor in Human Breast Cancer Tissues Is Associated With High-Grade Human Cytomegalovirus Protein Expression.

    Science.gov (United States)

    Rahbar, Afsar; Touma, Joel; Costa, Helena; Davoudi, Belghis; Bukholm, Ida Rashid; Sauer, Torill; Vetvik, Katja; Geisler, Jürgen; Söderberg-Naucler, Cecilia

    2017-11-01

    The underlying mechanisms for breast cancer (BC) are largely unknown. We investigated possible correlations between the expression levels of human cytomegalovirus (HCMV) proteins and established histopathological markers of BC, including expression of estrogen receptor (ER)-α, the progesterone receptor (PgR), and HER2. We retrospectively examined paraffin-embedded biopsy specimens of BC (n = 62), ductal carcinoma in situ (n = 19), and adjacent normal breast tissue (n = 42) for HCMV immediate-early protein (IE), HCMV late antigen, HCMV DNA and RNA, and investigated possible correlations between them and expression of ER-α, PgR, and HER2. HCMV DNA and RNA were detected in all examined infiltrating BCs. High-grade positivity for HCMV-IE was detected in 77% of infiltrating BCs, 39% of ductal carcinomas in situ, and 7% of tumor-free breast tissue samples. HCMV expression correlated inversely with ER-α (P = .02) and PgR (P = .003) expression. HER2 expression was also reduced in HCMV-positive samples without reaching a level of statistical significance (P = .09). The negative correlation between high-grade expression HCMV-IE and hormone receptor expression suggests a role for HCMV in hormone receptor-negative BC tumors, possibly by forcing BC cells into a more aggressive phenotype. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Reference gene selection for quantitative real-time PCR analysis in virus infected cells: SARS corona virus, Yellow fever virus, Human Herpesvirus-6, Camelpox virus and Cytomegalovirus infections

    Directory of Open Access Journals (Sweden)

    Müller Marcel A

    2005-02-01

    Full Text Available Abstract Ten potential reference genes were compared for their use in experiments investigating cellular mRNA expression of virus infected cells. Human cell lines were infected with Cytomegalovirus, Human Herpesvirus-6, Camelpox virus, SARS coronavirus or Yellow fever virus. The expression levels of these genes and the viral replication were determined by real-time PCR. Genes were ranked by the BestKeeper tool, the GeNorm tool and by criteria we reported previously. Ranking lists of the genes tested were tool dependent. However, over all, β-actin is an unsuitable as reference gene, whereas TATA-Box binding protein and peptidyl-prolyl-isomerase A are stable reference genes for expression studies in virus infected cells.

  9. Human Factors in Nuclear Power Engineering in Polish Conditions

    OpenAIRE

    Agnieszka Kaczmarek-Kacprzak; Martin Catlow

    2014-01-01

    The paper “Human factors in nuclear power engineering in Polish conditions” focuses on analysis of dynamics of preparing Polish society to build fi rst nuclear power plant in XXI century in Poland. Authors compare experience from constructing nuclear power plant Sizewell B (Great Britain) and Sizewell C, which is in preparation phase with polish nuclear power program. Paper includes aspects e.g. of creating nuclear safety culture and social opinion about investment. Human factors in nuclear p...

  10. Human somatic cell nuclear transfer and cloning.

    Science.gov (United States)

    2012-10-01

    This document presents arguments that conclude that it is unethical to use somatic cell nuclear transfer (SCNT) for infertility treatment due to concerns about safety; the unknown impact of SCNT on children, families, and society; and the availability of other ethically acceptable means of assisted reproduction. This document replaces the ASRM Ethics Committee report titled, "Human somatic cell nuclear transfer (cloning)," last published in Fertil Steril 2000;74:873-6. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  11. Review of EPRI Nuclear Human Factors Program

    Energy Technology Data Exchange (ETDEWEB)

    Hanes, L.F.; O`Brien, J.F. [Electric Power Research Institute, Palo Alto, CA (United States)

    1996-03-01

    The Electric Power Research Institute (EPRI) Human Factors Program, which is part of the EPRI Nuclear Power Group, was established in 1975. Over the years, the Program has changed emphasis based on the shifting priorities and needs of the commercial nuclear power industry. The Program has produced many important products that provide significant safety and economic benefits for EPRI member utilities. This presentation will provide a brief history of the Program and products. Current projects and products that have been released recently will be mentioned.

  12. Activation of PPAR{gamma} by Human Cytomegalovirus for de novo Replication Impairs Migration and Invasiveness of Cytotrophoblast from Early Placenta

    DEFF Research Database (Denmark)

    Rauwel, Benjamin; Mariamé, Bernard; Martin, Hélène

    2010-01-01

    Human cytomegalovirus (HCMV) contributes to pathogenic processes in immuno-suppressed individuals, in fetuses and in neonates. In the present report by using reporter gene activation assays and confocal microscopy in the presence of specific antagonist we show for the first time that HCMV infection...... induced PPARgamma transcriptional activity in infected cells. We demonstrated that PPARgamma antagonist dramatically impaired virus production and that the major immediate-early promoter (MIEP) contained PPAR response elements (PPRE) able to bind PPARgamma, as assessed by electrophoretic mobility shift......, as assessed by using well-established in vitro models of invasive trophoblast i.e. primary cultures of EVCT isolated from first trimester placentas and the EVCT-derived cell line HIPEC. Our data provide new clues to explain how early infection during pregnancy could impair implantation, placentation...

  13. Human Cytomegalovirus IE2 86 kDa Protein Induces STING Degradation and Inhibits cGAMP-Mediated IFN-β Induction

    Directory of Open Access Journals (Sweden)

    Jung-Eun Kim

    2017-09-01

    Full Text Available Stimulator of interferon genes (STING is a critical signaling molecule in the innate immune response against DNA viruses by either directly sensing intracellular DNA or functioning as an adaptor molecule to activate the type I interferon (IFN signaling pathway. We determined the functional interaction between STING and human cytomegalovirus (HCMV. A cDNA library containing 133 HCMV ORFs was screened to identify viral genes that inhibit STING-induced IFN-β promoter activation. Among the screened ORFs, UL122, which encodes the immediate-early 2 86 kDa (IE86 protein, strongly abolished STING-induced IFN-β promoter activation. Interestingly, IE86 protein facilitated the proteasome-dependent degradation of STING and inhibited 2′3′-cGAMP-mediated induction of IFNB1 and CXCL10. Taken together, this study demonstrates the existence of a post-translational regulation of STING by HCMV IE86 protein.

  14. [Mononucleosis caused by cytomegalovirus].

    Science.gov (United States)

    Lajo Plaza, A; del Castillo Martín, F; Martínez Zapico, R

    1990-01-01

    Sixteen cases of mononucleosis due to cytomegalovirus, are presented. The selection of patients was based on clinical criteria. Symptoms are compared with another series of patients affected with mononucleosis by Epstein-Barr virus. We have not found differences comparing the fever, cervical adenopathies and faringoamigdalitis. Differences were significant in hepatomegaly. We conclude that the clinical picture of cytomegalovirus mononucleosis is very similar to those of the Epstein-Barr mononucleosis.

  15. Congenital cytomegalovirus infection

    OpenAIRE

    D'Oronzio, U; Arlettaz, R.; Hagmann, C.

    2015-01-01

    Clinical details of 50 infants with congenital cytomegalovirus infection identified in a prospective study are reported. The mean birthweight, gestational age, and head circumference of children with congenital cytomegalovirus infection were not significantly different from those of controls. Three (6%) had symptoms at birth--two neurological and one pneumonitis. In the first four months of life transient hepatosplenomegaly occurred in two infected children and six suffered interstitial pneum...

  16. The smallest capsid protein mediates binding of the essential tegument protein pp150 to stabilize DNA-containing capsids in human cytomegalovirus.

    Directory of Open Access Journals (Sweden)

    Xinghong Dai

    2013-08-01

    Full Text Available Human cytomegalovirus (HCMV is a ubiquitous herpesvirus that causes birth defects in newborns and life-threatening complications in immunocompromised individuals. Among all human herpesviruses, HCMV contains a much larger dsDNA genome within a similarly-sized capsid compared to the others, and it was proposed to require pp150, a tegument protein only found in cytomegaloviruses, to stabilize its genome-containing capsid. However, little is known about how pp150 interacts with the underlying capsid. Moreover, the smallest capsid protein (SCP, while dispensable in herpes simplex virus type 1, was shown to play essential, yet undefined, role in HCMV infection. Here, by cryo electron microscopy (cryoEM, we determine three-dimensional structures of HCMV capsid (no pp150 and virion (with pp150 at sub-nanometer resolution. Comparison of these two structures reveals that each pp150 tegument density is composed of two helix bundles connected by a long central helix. Correlation between the resolved helices and sequence-based secondary structure prediction maps the tegument density to the N-terminal half of pp150. The structures also show that SCP mediates interactions between the capsid and pp150 at the upper helix bundle of pp150. Consistent with this structural observation, ribozyme inhibition of SCP expression in HCMV-infected cells impairs the formation of DNA-containing viral particles and reduces viral yield by 10,000 fold. By cryoEM reconstruction of the resulting "SCP-deficient" viral particles, we further demonstrate that SCP is required for pp150 functionally binding to the capsid. Together, our structural and biochemical results point to a mechanism whereby SCP recruits pp150 to stabilize genome-containing capsid for the production of infectious HCMV virion.

  17. Refining human T-cell immunotherapy of cytomegalovirus disease: a mouse model with 'humanized' antigen presentation as a new preclinical study tool.

    Science.gov (United States)

    Lemmermann, Niels A W; Reddehase, Matthias J

    2016-12-01

    With the cover headline 'T cells on the attack,' the journal Science celebrated individualized cancer immunotherapy by adoptive transfer of T cells as the 'Breakthrough of the Year' 2013 (J. Couzin-Frankel in Science 342:1432-1433, 2013). It is less well recognized and appreciated that individualized T cell immunotherapy of cytomegalovirus (CMV) infection is approaching clinical application for preventing CMV organ manifestations, interstitial CMV pneumonia in particular. This coincident medical development is particularly interesting as reactivated CMV infection is a major viral complication in the state of transient immunodeficiency after the therapy of hematopoietic malignancies by hematopoietic cell transplantation (HCT). It may thus be attractive to combine T cell immunotherapy of 'minimal residual disease/leukemia (MRD)' and CMV-specific T cell immunotherapy to combat both risks in HCT recipients simultaneously, and ideally with T cells derived from the respective HLA-matched HCT donor. Although clinical trials of human CMV-specific T cell immunotherapy were promising in that the incidence of virus reactivation and disease was found to be reduced with statistical significance, animal models are still instrumental for providing 'proof of concept' by directly documenting the prevention of viral multiple-organ histopathology and organ failure under controlled conditions of the absence versus presence of the therapy, which obviously is not feasible in an individual human patient. Further, animal models can make predictions regarding parameters that determine the efficacy of T cell immunotherapy for improved study design in clinical investigations, and they allow for manipulating host and virus genetics. The latter is of particular value as it opens the possibility for epitope specificity controls that are inherently missing in clinical trials. Here, we review a recently developed new mouse model that is more approximated to human CMV-specific T cell immunotherapy

  18. Cytomegalovirus Colitis in Immunocompetent Patients

    OpenAIRE

    Inayat, Faisal; Hussain, Qulsoom; Shafique, Khurram; Tasleem, Syed H; Hurairah, Abu

    2016-01-01

    Cytomegalovirus colitis is common in immunocompromised patients, but rare in immunocompetent patients. The present study not only represents the colonoscopy and pathological findings, but also applies the method of diagnosing and treating cytomegalovirus colitis in immunocompetent patients.

  19. Cytomegalovirus Colitis in Immunocompetent Patients

    Science.gov (United States)

    Hussain, Qulsoom; Shafique, Khurram; Tasleem, Syed H; Hurairah, Abu

    2016-01-01

    Cytomegalovirus colitis is common in immunocompromised patients, but rare in immunocompetent patients. The present study not only represents the colonoscopy and pathological findings, but also applies the method of diagnosing and treating cytomegalovirus colitis in immunocompetent patients. PMID:27980888

  20. Restriction enzyme analysis of the human cytomegalovirus genome in specimens collected from immunodeficient patients in Belém, State of Pará, Brazil

    Directory of Open Access Journals (Sweden)

    Dorotéa Lobato da Silva

    2011-10-01

    Full Text Available INTRODUCTION: Human cytomegalovirus is an opportunistic betaherpesvirus that causes persistent and serious infections in immunodeficient patients. Recurrent infections occur due to the presence of the virus in a latent state in some cell types. It is possible to examine the virus using molecular methods to aid in the immunological diagnosis and to generate a molecular viral profile in immunodeficient patients. The objective of this study was to characterize cytomegalovirus genotypes and to generate the epidemiological and molecular viral profile in immunodeficient patients. METHODS: A total of 105 samples were collected from immunodeficient patients from the City of Belém, including newborns, hemodialysis patients, transplant recipients and HIV+ patients. An IgG and IgM antibody study was completed using ELISA, and enzymatic analysis by restriction fragment length polymorphism (RFLP was performed to characterize viral genotypes. RESULTS: It was observed that 100% of the patients had IgG antibodies, 87% of which were IgG+/IgM-, consistent with a prior infection profile, 13% were IgG+/IgM+, suggestive of recent infection. The newborn group had the highest frequency (27% of the IgG+/IgM+ profile. By RFLP analysis, only one genotype was observed, gB2, which corresponded to the standard AD169 strain. CONCLUSIONS: The presence of IgM antibodies in new borns indicates that HCMV continues to be an important cause of congenital infection. The low observed genotypic diversity could be attributed to the small sample size because newborns were excluded from the RFLP analysis. This study will be continued including samples from newborns to extend the knowledge of the general and molecular epidemiology of HCMV in immunodeficient patients.

  1. Diagnosis of Cytomegalovirus Infections

    Science.gov (United States)

    Ross, S.A.; Novak, Z.; Pati, S.; Boppana, S.B.

    2013-01-01

    Cytomegalovirus (CMV) is recognized as the most common congenital viral infection in humans and an important cause of morbidity and mortality in immunocompromised hosts. This recognition of the clinical importance of invasive CMV disease in the setting of immunodeficiency and in children with congenital CMV infection has led to the development of new diagnostic procedures for the rapid identification of immunocompromised individuals with CMV disease, as well as fetuses and infants with congenital infection. Diagnosis of acute maternal CMV infection by the presence of IgM and low IgG avidity requires confirmation of fetal infection which is typically performed by CMV PCR of the amniotic fluid. Viral culture of the urine and saliva obtained within the first two weeks of life continue to be the gold standard for diagnosis of congenitally infected infants. PCR assays of dried blood spots from infants have not been shown to have sufficient sensitivity for the identification of most infants with congenital CMV infection. However, saliva PCR assays are currently being assessed as a useful screening method for congenital CMV infection. In the immunocompromised host, newer rapid diagnostic assays such as pp65 antigenemia and real-time CMV PCR of blood or plasma have allowed for preemptive treatment reducing morbidity and mortality. However, lack of standardized real-time PCR protocols hinders the comparison of the data across different centers and the development of uniform guidelines for the management of invasive CMV infections in immunocompromised individuals. PMID:21827433

  2. Evaluation of T Cell Immunity against Human Cytomegalovirus: Impact on Patient Management and Risk Assessment of Vertical Transmission

    Directory of Open Access Journals (Sweden)

    Giulia Freer

    2016-01-01

    Full Text Available Cytomegalovirus (CMV is one of the most common infectious agents, infecting the general population at an early age without causing morbidity most of the time. However, on particular occasions, it may represent a serious risk, as active infection is associated with rejection and disease after solid organ transplantation or fetal transmission during pregnancy. Several methods for CMV diagnosis are available on the market, but because infection is so common, careful selection is needed to discriminate primary infection from reactivation. This review focuses on methods based on CMV-specific T cell reactivity to help monitor the consequences of CMV infection/reactivation in specific categories of patients. This review makes an attempt at discussing the pros and cons of the methods available.

  3. Seroprevalence of cytomegalovirus among pregnant women ...

    African Journals Online (AJOL)

    2014-03-03

    Mar 3, 2014 ... Seroprevalence of CMV-IgG antibodies amongst normal pregnant women in Nigeria. IntJ Womens Health 2011;3:423-8. doi: 10.2147/IJWH.S24850. 18. Zhang LJ, Hanpf P, Rutherford C, Churchill WH,. Crumpacker CS. Detection of human cytomegalovirus. DNA, RNA, and antibody in normal donor blood J.

  4. prevalence of cytomegalovirus antibodies in blood donors

    African Journals Online (AJOL)

    2009-12-02

    Dec 2, 2009 ... 86 (Supplement) December 2009. PREVALENCE OF CYTOMEGALOVIRUS ANTIBODIES IN BLOOD DONORS AT THE NATIONAL BLOOD. TRANSFUSION CENTRE, NAIROBI. D. G. Njeru, MBChB, MMed (Path), Dip. Forensic Med (SA), Registrar, Department of Human Pathology, W. O. Mwanda,. MBChB ...

  5. Seroprevalence of Cytomegalovirus Infection amongst Pregnant ...

    African Journals Online (AJOL)

    This study was conducted to determine the seroprevalence of human cytomegalovirus among pregnant women in Kaduna State, Nigeria. Three hundred and sixty three (363) blood samples were collected from 330 pregnant women attending antenatal clinics and 33 non pregnant women attending the Outpatient ...

  6. In Vitro Drug Combination Studies of Letermovir (AIC246, MK-8228) with Approved Anti-Human Cytomegalovirus (HCMV) and Anti-HIV Compounds in Inhibition of HCMV and HIV Replication

    OpenAIRE

    Wildum, Steffen; Zimmermann, Holger; Lischka, Peter

    2015-01-01

    Despite modern prevention and treatment strategies, human cytomegalovirus (HCMV) remains a common opportunistic pathogen associated with serious morbidity and mortality in immunocompromised individuals, such as transplant recipients and AIDS patients. All drugs currently licensed for the treatment of HCMV infection target the viral DNA polymerase and are associated with severe toxicity issues and the emergence of drug resistance. Letermovir (AIC246, MK-8228) is a new anti-HCMV agent in clinic...

  7. Engineered external guide sequences are highly effective in inducing RNase P for inhibition of gene expression and replication of human cytomegalovirus.

    Science.gov (United States)

    Yang, Yong-Hua; Li, Hongjian; Zhou, Tianhong; Kim, Kihoon; Liu, Fenyong

    2006-01-01

    External guide sequences (EGSs), which are RNA molecules derived from natural tRNAs, bind to a target mRNA and render the mRNA susceptible to hydrolysis by RNase P, a tRNA processing enzyme. Using an in vitro selection procedure, we have previously generated EGS variants that efficiently direct human RNase P to cleave a target mRNA in vitro. In this study, a variant was used to target the overlapping region of the mRNAs encoding human cytomegalovirus (HCMV) essential transcription regulatory factors IE1 and IE2. The EGS variant was approximately 25-fold more active in inducing human RNase P to cleave the mRNA in vitro than the EGS derived from a natural tRNA. Moreover, a reduction of 93% in IE1/IE2 gene expression and a reduction of 3000-fold in viral growth were observed in HCMV-infected cells that expressed the variant, while cells expressing the tRNA-derived EGS exhibited a reduction of 80% in IE1/IE2 expression and an inhibition of 150-fold in viral growth. Our results provide the first direct evidence that EGS variant is highly effective in blocking HCMV gene expression and growth and furthermore, demonstrate the feasibility of developing effective EGS RNA variants for anti-HCMV applications by using in vitro selection procedures.

  8. A prominent role of the human cytomegalovirus UL8 glycoprotein restraining pro-inflammatory cytokine production by myeloid cells at late times during infection.

    Science.gov (United States)

    Pérez-Carmona, Natàlia; Martínez-Vicente, Pablo; Farré, Domènec; Gabaev, Ildar; Messerle, Martin; Engel, Pablo; Angulo, Ana

    2018-02-21

    Human cytomegalovirus (HCMV) persistence in infected individuals relies on a plethora of mechanisms to efficiently reduce host immune responses. To that end, HCMV commits a variety of gene products, some of which have not been identified yet. Here we characterized the UL8 gene, which consists of two exons, sharing the first with the HCMV RL11 family member UL7 UL8 is a transmembrane protein with an N-terminal immunoglobulin (Ig)-like domain in common with UL7 but with an extended stalk and a distinctive cytoplasmic tail. The UL8 open reading frame gives rise to a heavily glycosylated protein, predominantly expressed on the cell surface, from where it can be partially endocytosed and subsequently degraded. Infections with UL8-tagged viruses indicated that UL8 was synthesized with late phase kinetics. By virtue of its highly conserved Ig-like domain, this viral protein interacted with a surface molecule present on activated neutrophils. Notably, when ectopically expressed in THP-1 myeloid cells, UL8 was able to significantly reduce the production of a variety of pro-inflammatory cytokines. Mutations in UL8 indicated that this functional effect was mediated by the cell surface expression of its Ig-like domain. To investigate the impact of the viral protein in the infection context, we engineered HCMVs lacking the UL8 gene, and demonstrated that UL8 decreases the release of a large number of pro-inflammatory factors at late times after infection of THP-1 cells. Our data indicate that UL8 may exert an immunosuppressive role key for HCMV survival in the host. IMPORTANCE Human cytomegalovirus (HCMV) is a major pathogen that causes life-threatening diseases and disabilities in infected newborns and immunocompromised individuals. Containing one of the largest genomes among all reported human viruses, HCMV encodes an impressive repertoire of gene products. However, the functions of a large proportion of them remain still unknown, a fact that complicates the design of new

  9. Neutralization of Diverse Human Cytomegalovirus Strains Conferred by Antibodies Targeting Viral gH/gL/pUL128-131 Pentameric Complex.

    Science.gov (United States)

    Ha, Sha; Li, Fengsheng; Troutman, Matthew C; Freed, Daniel C; Tang, Aimin; Loughney, John W; Wang, Dai; Wang, I-Ming; Vlasak, Josef; Nickle, David C; Rustandi, Richard R; Hamm, Melissa; DePhillips, Pete A; Zhang, Ningyan; McLellan, Jason S; Zhu, Hua; Adler, Stuart P; McVoy, Michael A; An, Zhiqiang; Fu, Tong-Ming

    2017-04-01

    Human cytomegalovirus (HCMV) is the leading cause of congenital viral infection, and developing a prophylactic vaccine is of high priority to public health. We recently reported a replication-defective human cytomegalovirus with restored pentameric complex glycoprotein H (gH)/gL/pUL128-131 for prevention of congenital HCMV infection. While the quantity of vaccine-induced antibody responses can be measured in a viral neutralization assay, assessing the quality of such responses, including the ability of vaccine-induced antibodies to cross-neutralize the field strains of HCMV, remains a challenge. In this study, with a panel of neutralizing antibodies from three healthy human donors with natural HCMV infection or a vaccinated animal, we mapped eight sites on the dominant virus-neutralizing antigen-the pentameric complex of glycoprotein H (gH), gL, and pUL128, pUL130, and pUL131. By evaluating the site-specific antibodies in vaccine immune sera, we demonstrated that vaccination elicited functional antiviral antibodies to multiple neutralizing sites in rhesus macaques, with quality attributes comparable to those of CMV hyperimmune globulin. Furthermore, these immune sera showed antiviral activities against a panel of genetically distinct HCMV clinical isolates. These results highlighted the importance of understanding the quality of vaccine-induced antibody responses, which includes not only the neutralizing potency in key cell types but also the ability to protect against the genetically diverse field strains.IMPORTANCE HCMV is the leading cause of congenital viral infection, and development of a preventive vaccine is a high public health priority. To understand the strain coverage of vaccine-induced immune responses in comparison with natural immunity, we used a panel of broadly neutralizing antibodies to identify the immunogenic sites of a dominant viral antigen-the pentameric complex. We further demonstrated that following vaccination of a replication

  10. Human cytomegalovirus UL55, UL144, and US28 genotype distribution in infants infected congenitally or postnatally.

    Science.gov (United States)

    Paradowska, Edyta; Studzińska, Mirosława; Suski, Patrycja; Kasztelewicz, Beata; Wiśniewska-Ligier, Małgorzata; Zawilińska, Barbara; Gaj, Zuzanna; Nowakowska, Dorota

    2015-10-01

    Cytomegalovirus (CMV) is the most common cause of congenital infection. This pathogen exhibits extensive genetic variability in the genes that encode structural envelope glycoproteins, regulatory proteins, and proteins that contribute to immune evasion. However, the role of specific viral strains in the outcome of congenital CMV infection is unclear. Variation in the UL55 gene encoding glycoprotein B (gB), the UL144 gene encoding TNF α-like receptor, and the US28 gene encoding β-chemokine receptor was determined in 60 newborn infants with congenital CMV infection and 90 infants with postnatal or undefined CMV infection. CMV polymorphisms were studied in relation to disease outcome and viral load. Genotyping was performed by a sequencing analysis of PCR-amplified fragments, and the viral load was measured by quantitative real-time PCR. The results demonstrated that (1) the UL55 and US28 genotype distributions were similar among the group of congenital and postnatal CMV infection; (2) the UL144 B1 genotype was more prevalent in congenital than in postnatal infection and was detected in 70% of newborns with asymptomatic congenital infection; and (3) none of the examined genotype was significantly linked with symptomatic CMV infection. No relationship was observed between genotype and viral load. The results revealed that UL55, UL144, and US28 polymorphisms are not associated with the outcome of CMV infection in infants, but the presence of UL144 B1 genotype might be virological marker of asymptomatic infection at birth. © 2015 Wiley Periodicals, Inc.

  11. Importance of Highly Conserved Peptide Sites of Human Cytomegalovirus gO for Formation of the gH/gL/gO Complex.

    Science.gov (United States)

    Stegmann, Cora; Abdellatif, Mohamed E A; Laib Sampaio, Kerstin; Walther, Paul; Sinzger, Christian

    2017-01-01

    The glycoprotein O (gO) is betaherpesvirus specific. Together with the viral glycoproteins H and L, gO forms a covalent trimeric complex that is part of the viral envelope. This trimer is crucial for cell-free infectivity of human cytomegalovirus (HCMV) but dispensable for cell-associated spread. We hypothesized that the amino acids that are conserved among gOs of different cytomegaloviruses are important for the formation of the trimeric complex and hence for efficient virus spread. In a mutational approach, nine peptide sites, containing all 13 highly conserved amino acids, were analyzed in the context of HCMV strain TB40-BAC4 with regard to infection efficiency and formation of the gH/gL/gO complex. Mutation of amino acids (aa) 181 to 186 or aa 193 to 198 resulted in the loss of the trimer and a complete small-plaque phenotype, whereas mutation of aa 108 or aa 249 to 254 caused an intermediate phenotype. While individual mutations of the five conserved cysteines had little impact, their relevance was revealed in a combined mutation, which abrogated both complex formation and cell-free infectivity. C343 was unique, as it was sufficient and necessary for covalent binding of gO to gH/gL. Remarkably, however, C218 together with C167 rescued infectivity in the absence of detectable covalent complex formation. We conclude that all highly conserved amino acids contribute to the function of gO to some extent but that aa 181 to 198 and cysteines 343, 218, and 167 are particularly relevant. Surprisingly, covalent binding of gO to gH/gL is required neither for its incorporation into virions nor for proper function in cell-free infection. Like all herpesviruses, the widespread human pathogen HCMV depends on glycoproteins gB, gH, and gL for entry into target cells. Additionally, gH and gL have to bind gO in a trimeric complex for efficient cell-free infection. Homologs of gO are shared by all cytomegaloviruses, with 13 amino acids being highly conserved. In a mutational

  12. Hearing Loss and Cytomegalovirus.

    Science.gov (United States)

    Strauss, Melvin

    1997-01-01

    Cytomegalovirus is the most common cause of congenital virally induced hearing loss. Maternal infection is most often asymptomatic as is the infection in the newborn. Hearing loss occurs in both clinically apparent infection and in the asymptomatic infection. Current methods of detection, treatment, and prevention and research efforts are…

  13. Cytomegalovirus Hepatitis During Pregnancy

    Directory of Open Access Journals (Sweden)

    Ying Chan

    1995-01-01

    Full Text Available Background: Although cytomegalovirus (CMV is an uncommon cause of viral hepatitis during pregnancy, a definitive diagnosis is important because of the potential for congenital CMV. In the case reported here, a diagnosis of hepatitis caused by CMV was made after the more common viral pathogens had been ruled out.

  14. Cytomegalovirus en pericarditis

    NARCIS (Netherlands)

    Beekhuis, Johan Willem

    1976-01-01

    Dit proefschrift beschrijft een onderzoek naar de aetiologische betekenis van enkele virussen, in het bijzonder van cytomegalovirus (CMV) voor verschillende vormen van pericarditis. CMV-infecties worden merendeels in de adolescentie en op volwassen leeftijd doorgemaakt en kunnen aanleiding geven tot

  15. Human Mars Surface Mission Nuclear Power Considerations

    Science.gov (United States)

    Rucker, Michelle A.

    2018-01-01

    A key decision facing Mars mission designers is how to power a crewed surface field station. Unlike the solar-powered Mars Exploration Rovers (MER) that could retreat to a very low power state during a Martian dust storm, human Mars surface missions are estimated to need at least 15 kilowatts of electrical (kWe) power simply to maintain critical life support and spacecraft functions. 'Hotel' loads alone for a pressurized crew rover approach two kWe; driving requires another five kWe-well beyond what the Curiosity rover’s Radioisotope Power System (RPS) was designed to deliver. Full operation of a four-crew Mars field station is estimated at about 40 kWe. Clearly, a crewed Mars field station will require a substantial and reliable power source, beyond the scale of robotic mission experience. This paper explores the applications for both fission and RPS nuclear options for Mars.

  16. Factors Associated with Cytomegalovirus Reactivation Following Allogeneic Hematopoietic Stem Cell Transplantation: Human Leukocyte Antigens Might Be Among the Risk Factors

    Directory of Open Access Journals (Sweden)

    Kadir Acar

    2014-09-01

    Full Text Available OBJECTIVE: Cytomegalovirus (CMV is a significant cause of morbidity and mortality in allogeneic hematopoietic stem cell transplantation (AHSCT recipients. Current practice includes prophylactic and preemptive treatment modalities, which have risks, side effects, and costs of their own. There is no established risk scoring system that applies to all patients. We aimed to investigate the risk factors for CMV reactivation in AHSCT recipients. METHODS: We retrospectively analyzed the risk factors for CMV reactivation in 185 consequent AHSCT recipients transplanted between September 2003 and December 2009 at the Stem Cell Transplantation Unit of Gazi University. Besides the standard transplant-related parameters, HLA antigens were also included among the variables analyzed. RESULTS: Despite the very high rate of donor (94.6% and recipient (100% seropositivity, which are the so-called major risk factors in previous reports, our reactivation rate was much lower, with a frequency of 24.9%. The underlying disease, sex, conditioning regimen, and presence of antithymocyte globulin or fludarabine in the conditioning regimen had no impact on reactivation rate. CMV reactivation was significantly more frequent in recipients with graft-versus-host disease (GVHD compared to those without GVHD (p<0.0001. CMV reactivation was significantly more frequent (p<0.05 in patients with HLA-B14, HLADRB1* 01, and HLA-DRB1*13 antigens and less frequent in recipients with HLA-A11 and HLA-DRB1*04 antigens (p<0.05. CONCLUSION: Universal risk factors/scores that apply to all transplant recipients are required for tailored prophylaxis and/ or treatment strategies for CMV reactivation. Uncovering the role of genetic factors, including HLA antigens, as possible risk factors might lead the way to risk-adaptive strategies for adoptive cellular therapy and/or vaccination.

  17. Human cytomegalovirus induces TLR4 signaling components in monocytes altering TIRAP, TRAM and downstream interferon-beta and TNF-alpha expression.

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    Kok-Hooi Yew

    Full Text Available Using TLR pathways, primary human cytomegalovirus (HCMV induces innate responses including the production of inflammatory cytokines. Mounting evidence suggests that LPS recognition by TLR4/MD2/CD14 results in differential utilization of TIRAP-TRAF6 and TRAM-TRIF signaling, thereby leading to transcriptional activation of various cytokine genes. However, relative roles of the TLR4/MD2/CD14 complex and its adaptor proteins TIRAP and TRAM involved in regulating monocyte responses to HCMV are incomplete. Here, we provided evidence supporting the notion that the TLR4/MD2/CD14 complex contributes notably to HCMV-induced signaling and subsequent cytokine production in monocytes. In particular, induction of both IL-6 and IL-8 is associated with elevated TIRAP and reduced TRAM mRNA expression. The latter may serve in a compensatory pathway that yields a robust IFN response when TIRAP signaling is blocked in monocytes incubated with Toledo strain HCMV. Inhibitory studies using antisense oligonucleotides or neutralizing antibodies indicate that IL-6 induction by TLR4/MD2 complex is important for the activation of endogenous CD14 which later acts in concert or synergy with TLR4/MD2 as a factor resulting in IL-8 gene expression. We further show that exogenous recombinant CD14 can potentiate innate immune response via TLR4-dependent and possibly via TLR9-dependent pathways to promote enhanced expression/production of IL-8 and IFN-β, respectively.

  18. Toll-like receptor 4 is involved in the cell cycle modulation and required for effective human cytomegalovirus infection in THP-1 macrophages

    Energy Technology Data Exchange (ETDEWEB)

    Arcangeletti, Maria-Cristina, E-mail: mariacristina.arcangeletti@unipr.it [Department of Clinical and Experimental Medicine, University of Parma, Parma (Italy); Germini, Diego; Rodighiero, Isabella [Department of Clinical and Experimental Medicine, University of Parma, Parma (Italy); Mirandola, Prisco [Department of Biomedical, Biotechnological and Translational Sciences, University of Parma, Parma (Italy); De Conto, Flora; Medici, Maria-Cristina [Department of Clinical and Experimental Medicine, University of Parma, Parma (Italy); Gatti, Rita [Department of Biomedical, Biotechnological and Translational Sciences, University of Parma, Parma (Italy); Chezzi, Carlo; Calderaro, Adriana [Department of Clinical and Experimental Medicine, University of Parma, Parma (Italy)

    2013-05-25

    Suitable host cell metabolic conditions are fundamental for the effective development of the human cytomegalovirus (HCMV) lytic cycle. Indeed, several studies have demonstrated the ability of this virus to interfere with cell cycle regulation, mainly by blocking proliferating cells in G1 or G1/S. In the present study, we demonstrate that HCMV deregulates the cell cycle of THP-1 macrophages (a cell line irreversibly arrested in G0) by pushing them into S and G2 phases. Moreover, we show that HCMV infection of THP-1 macrophages leads to Toll-like receptor 4 (TLR4) activation. Since various studies have indicated TLR4 to be involved in promoting cell proliferation, here we investigate the possible role of TLR4 in the observed HCMV-induced cell cycle perturbation. Our data strongly support TLR4 as a mediator of HCMV-triggered cell cycle activation in THP-1 macrophages favouring, in turn, the development of an efficient viral lytic cycle. - Highlights: ► We studied HCMV infection impact on THP-1 macrophage cell cycle. ► We analysed the role played by Toll-like receptor (TLR) 4 upon HCMV infection. ► HCMV pushes THP-1 macrophages (i.e. resting cells) to re-enter the cell cycle. ► TLR4 pathway inhibition strongly affects the effectiveness of HCMV replication. ► TLR4 pathway inhibition significantly decreases HCMV-induced cell cycle re-entry.

  19. Human cytomegalovirus UL97 kinase is involved in the mechanism of action of methylenecyclopropane analogs with 6-ether and -thioether substitutions.

    Science.gov (United States)

    Komazin-Meredith, Gloria; Chou, Sunwen; Prichard, Mark N; Hartline, Caroll B; Cardinale, Steven C; Comeau, Katelyn; Williams, John D; Khan, Atiyya R; Peet, Norton P; Bowlin, Terry L

    2014-01-01

    Methylenecyclopropane nucleoside (MCPN) analogs are being investigated for treatment of human cytomegalovirus (HCMV) infection because of favorable preclinical data and limited ganciclovir cross-resistance. Monohydroxymethyl MCPNs bearing ether and thioether functionalities at the purine 6 position have antiviral activity against herpes simplex virus (HSV) and varicella-zoster virus (VZV) in addition to HCMV. The role of the HCMV UL97 kinase in the mechanism of action of these derivatives was examined. When tested against a kinase-inactive UL97 K355M virus, a moderate 5- to 7-fold increase in 50% effective concentration (EC50) was observed, in comparison to a 13- to 25-fold increase for either cyclopropavir or ganciclovir. Serial propagation of HCMV under two of these compounds selected for three novel UL97 mutations encoding amino acid substitutions D456N, C480R,and Y617del. When transferred to baseline laboratory HCMV strains, these mutations individually conferred resistance to all of the tested MCPNs, ganciclovir, and maribavir. However, the engineered strains also demonstrated severe growth defects and abnormal cytopathic effects similar to the kinase-inactive mutant. Expressed and purified UL97 kinase showed in vitro phosphorylation of the newly tested MCPNs. Thus, HCMV UL97 kinase is involved in the antiviral action of these MCPNs, but the in vitro selection of UL97-defective viruses suggests that their activity against more typical ganciclovir-resistant growth-competent UL97 mutants may be relatively preserved.

  20. Evaluation of a Probe-Based PCR-ELISA System for Simultaneous Semi Quantitative Detection and Genotyping of Human Cytomegalovirus (HCMV) Infection in Clinical Specimens.

    Science.gov (United States)

    Talkhabifard, Majid; Javid, Naeme; Moradi, Abdolvahab; Ghaemi, Amir; Tabarraei, Alijan

    2017-01-01

    Human cytomegalovirus (HCMV) is a common opportunistic pathogen that causes serious complications in immunosuppressed patients and infected newborns. In this study, PCR-ELISA was optimized for semi-quantitative detection of infection in clinical specimens and simultaneous genotyping of glycoprotein B for 4 major genotypes, due to its significance. During DIG-labeling PCR, a pair of primers amplifies a fragment of variable region of the glycoprotein B encoding sequence. Under optimized conditions, labeled Target amplicons hybridize to biotinated specific probes and are detected in an ELISA system. PCR-ELISA system showed specific performance with detection limit of approximately 100 copies of CMV DNA. The linear correlation was observed between the PCR-ELISA results (OD) and logarithmic scale of CMV (r=0.979). Repeatability of PCR-ELISA detection system for intra-assay and inter-assay was evaluated for negative and positive samples. In optimized conditions of hybridization, differentiation between genotypes of glycoprotein B was feasible using genotype-specific probes in PCR-ELISA genotyping system. In comparison with sequencing method, genotyping system was confirmed with kappa index of 1. PCR-ELISA is proposed as an applicable and reliable technique for semi-quantitative diagnosis and typing of the infection. This technique is flexible to apply in a variety of molecular fields.

  1. The cellular protein MCM3AP is required for inhibition of cellular DNA synthesis by the IE86 protein of human cytomegalovirus.

    Directory of Open Access Journals (Sweden)

    Emma Poole

    Full Text Available Like all DNA viruses, human cytomegalovirus (HCMV infection is known to result in profound effects on host cell cycle. Infection of fibroblasts with HCMV is known to induce an advance in cell cycle through the G(0-G(1 phase and then a subsequent arrest of cell cycle in early S-phase, presumably resulting in a cellular environment optimum for high levels of viral DNA replication whilst precluding replication of cellular DNA. Although the exact mechanisms used to arrest cell cycle by HCMV are unclear, they likely involve a number of viral gene products and evidence points to the ability of the virus to prevent licensing of cellular DNA synthesis. One viral protein known to profoundly alter cell cycle is the viral immediate early 86 (IE86 protein--an established function of which is to initially drive cells into early S phase but then inhibit cellular DNA synthesis. Here we show that, although IE86 interacts with the cellular licensing factor Cdt1, it does not inhibit licensing of cellular origins. Instead, IE86-mediated inhibition of cellular DNA synthesis requires mini-chromosome-maintenance 3 (MCM3 associated protein (MCM3AP, which can cause subsequent inhibition of initiation of cellular DNA synthesis in a licensing-independent manner.

  2. Dendritic cell-induced apoptosis of human cytomegalovirus-infected fibroblasts promotes cross-presentation of pp65 to CD8+ T cells.

    Science.gov (United States)

    Mandron, Marie; Martin, Hélène; Bonjean, Béatrice; Lulé, Jacqueline; Tartour, Eric; Davrinche, Christian

    2008-01-01

    An efficient host response to human cytomegalovirus (HCMV) infection may depend on rapid sensing of the infection by the innate immune response prior to deployment of viral immunosubversive functions. Control of HCMV dissemination could be ensured by apoptosis of cells immediately following infection. In the present report, it is demonstrated that changes in the ratio of c-FLIP to FLICE contributed to early sensitivity of HCMV-infected MRC5 fibroblasts to tumour necrosis factor alpha (TNF-alpha), providing an innate response to infection. Dendritic cells (DCs) co-cultured with HCMV-infected MRC5 cells acquired the ability to secrete TNF-alpha in an amount sufficient to kill infected fibroblasts. Blockage of TNF-alpha binding to its receptor on MRC5 cells with soluble TNF-R reduced the number of dead, HCMV-infected fibroblasts ingested by DCs, thus highlighting the impact of the apoptotic state of infected cells for efficient loading of DCs. Those DCs loaded with antigens available early in infection, such as input virion-associated pp65, could then engage antigen processing for cross-presentation to specific CD8(+) T cells. Cross-presentation was impaired when MRC5 cells were treated with the pan-caspase inhibitor ZVAD before co-culture with DCs. Altogether, our data suggest that the innate killing capacity of DCs at the early stage of infection plays a role in the activation of anti-HCMV CD8(+) T cells.

  3. Toll-like receptor 4 is involved in the cell cycle modulation and required for effective human cytomegalovirus infection in THP-1 macrophages.

    Science.gov (United States)

    Arcangeletti, Maria-Cristina; Germini, Diego; Rodighiero, Isabella; Mirandola, Prisco; De Conto, Flora; Medici, Maria-Cristina; Gatti, Rita; Chezzi, Carlo; Calderaro, Adriana

    2013-05-25

    Suitable host cell metabolic conditions are fundamental for the effective development of the human cytomegalovirus (HCMV) lytic cycle. Indeed, several studies have demonstrated the ability of this virus to interfere with cell cycle regulation, mainly by blocking proliferating cells in G1 or G1/S. In the present study, we demonstrate that HCMV deregulates the cell cycle of THP-1 macrophages (a cell line irreversibly arrested in G0) by pushing them into S and G2 phases. Moreover, we show that HCMV infection of THP-1 macrophages leads to Toll-like receptor 4 (TLR4) activation. Since various studies have indicated TLR4 to be involved in promoting cell proliferation, here we investigate the possible role of TLR4 in the observed HCMV-induced cell cycle perturbation. Our data strongly support TLR4 as a mediator of HCMV-triggered cell cycle activation in THP-1 macrophages favouring, in turn, the development of an efficient viral lytic cycle. Copyright © 2013 Elsevier Inc. All rights reserved.

  4. Epigenetically repressing human cytomegalovirus lytic infection and reactivation from latency in THP-1 model by targeting H3K9 and H3K27 histone demethylases.

    Directory of Open Access Journals (Sweden)

    Xin Gan

    Full Text Available Human Cytomegalovirus (hCMV infects a broad range of the population and establishes life-long latency in the infected individuals. Periodically the latently infected virus can reactivate and becomes a significant cause of morbidity and mortality in immunocompromised individuals. In latent infection, the viral genome is suppressed in a heterochromatic state and viral gene transcription is silenced. Upon reactivation, the repressive chromatin is remodeled to an active form, allowing viral lytic gene transcription, initiated by the expression of viral Immediate Early (IE genes. During this process, a number of histone modification enzymes, including histone demethylases (HDMs, play important roles in driving IE expression, but the mechanisms involved are not fully understood. To get a better understanding of these mechanisms, we focused on two HDMs, KDM4 and KDM6, which reverse the repressive histone H3-lysine 9 and lysine 27 methylation, respectively. Our studies show that in lytic infection, both demethylases are important in the activation of viral IE gene expression. Simultaneous disruption of both via genetic or chemical methods leads to severely impaired viral IE gene expression and viral replication. Additionally, in an experimental latency-reactivation model in THP-1 cells, the KDM6 family member JMJD3 is induced upon viral reactivation and its knockdown resulted in reduced IE gene transcription. These findings suggest pharmacological inhibition of these HDMs may potentially block hCMV lytic infection and reactivation, and control the viral infection associated diseases, which are of significant unmet medical needs.

  5. Herpes simplex virus and cytomegalovirus co-infection presenting as exuberant genital ulcer in a woman infected with human immunodeficiency virus.

    Science.gov (United States)

    Gouveia, A I; Borges-Costa, J; Soares-Almeida, L; Sacramento-Marques, M; Kutzner, H

    2014-12-01

    In patients infected with human immunodeficiency virus (HIV), genital herpes can result in severe and atypical clinical presentations, and can become resistant to aciclovir treatment. Rarely, these manifestations may represent concurrent herpes simplex virus (HSV) with other agents. We report a 41-year-old black woman with HIV who presented with extensive and painful ulceration of the genitalia. Histological examination of a biopsy sample was suggestive of herpetic infection, and intravenous aciclovir was started, but produced only partial improvement. PCR was performed on the biopsy sample, and both HSV and cytomegalovirus (CMV) DNA was detected. Oral valganciclovir was started with therapeutic success. CMV infection is common in patients infected with HIV, but its presence in mucocutaneous lesions is rarely reported. This case exemplifies the difficulties of diagnosis of genital ulcers in patients infected with HIV. The presence of exuberant and persistent HSV genital ulcers in patients with HIV should also raise suspicions of the presence of co-infection with other organisms such as CMV. © 2014 British Association of Dermatologists.

  6. Comparison of the performance of polymerase chain reaction and pp65 antigenemia for the detection of human cytomegalovirus in immunosuppressed patients

    Directory of Open Access Journals (Sweden)

    Patrícia Borba Martiny

    2011-06-01

    Full Text Available INTRODUCTION: Human cytomegalovirus (HCMV is often reactive in latently infected immunosuppressed patients. Accordingly, HCMV remains one of the most common infections following solid organ and hemopoietic stem cell transplantations, resulting in significant morbidity, graft loss and occasional mortality. The early diagnosis of HCMV disease is important in immunosuppressed patients, since in these individuals, preemptive treatment is useful. The objective of this study was to compare the performance of the in-house qualitative polymerase chain reaction (PCR and pp65 antigenemia to HCMV infection in immunosuppressed patients in the Hospital de Clínicas of Porto Alegre (HCPA. METHODS: A total of 216 blood samples collected between August 2006 and January 2007 were investigated. RESULTS: Among the samples analyzed, 81 (37.5% were HCMV-positive by PCR, while 48 (22.2% were positive for antigenemia. Considering antigenemia as the gold standard, sensitivity, specificity, positive predictive values and negative predictive values for PCR were 87.5%, 76.8%, 51.8% and 95.5% respectively. CONCLUSIONS: These results demonstrated that qualitative PCR has high sensitivity and negative predictive value (NPV. Consequently PCR is especially indicated for the initial diagnosis of HCMV infection. In the case of preemptive treatment strategy, identification of patients at high-risk for HCMV disease is fundamental and PCR can be useful tool.

  7. Human cytomegalovirus-induced NKG2C(hi) CD57(hi) natural killer cells are effectors dependent on humoral antiviral immunity.

    Science.gov (United States)

    Wu, Zeguang; Sinzger, Christian; Frascaroli, Giada; Reichel, Johanna; Bayer, Carina; Wang, Li; Schirmbeck, Reinhold; Mertens, Thomas

    2013-07-01

    Recent studies indicate that expansion of NKG2C-positive natural killer (NK) cells is associated with human cytomegalovirus (HCMV); however, their activity in response to HCMV-infected cells remains unclear. We show that NKG2C(hi) CD57(hi) NK cells gated on CD3(neg) CD56(dim) cells can be phenotypically identified as HCMV-induced NK cells that can be activated by HCMV-infected cells. Using HCMV-infected autologous macrophages as targets, we were able to show that these NKG2C(hi) CD57(hi) NK cells are highly responsive to HCMV-infected macrophages only in the presence of HCMV-specific antibodies, whereas they are functionally poor effectors of natural cytotoxicity. We further demonstrate that NKG2C(hi) CD57(hi) NK cells are intrinsically responsive to signaling through CD16 cross-linking. Our findings show that the activity of pathogen-induced innate immune cells can be enhanced by adaptive humoral immunity. Understanding the activity of NKG2C(hi) CD57(hi) NK cells against HCMV-infected cells will be of relevance for the further development of adoptive immunotherapy.

  8. Distinct functional domains within the acidic cluster of tegument protein pp28 required for trafficking and cytoplasmic envelopment of human cytomegalovirus.

    Science.gov (United States)

    Seo, Jun-Young; Jeon, Hyejin; Hong, Sookyung; Britt, William J

    2016-10-01

    Human cytomegalovirus UL99-encoded tegument protein pp28 contains a 16 aa acidic cluster that is required for pp28 trafficking to the assembly compartment (AC) and the virus assembly. However, functional signals within the acidic cluster of pp28 remain undefined. Here, we demonstrated that an acidic cluster rather than specific sorting signals was required for trafficking to the AC. Recombinant viruses with chimeric pp28 proteins expressing non-native acidic clusters exhibited delayed viral growth kinetics and decreased production of infectious virus, indicating that the native acidic cluster of pp28 was essential for wild-type virus assembly. These results suggested that the acidic cluster of pp28 has distinct functional domains required for trafficking and for efficient virus assembly. The first half (aa 44-50) of the acidic cluster was sufficient for pp28 trafficking, whereas the native acidic cluster consisting of aa 51-59 was required for the assembly of wild-type levels of infectious virus.

  9. Detection of cytomegalovirus DNA in serum correlates with clinical cytomegalovirus retinitis in AIDS

    DEFF Research Database (Denmark)

    Hansen, K K; Ricksten, A; Hofmann, B

    1994-01-01

    The high sensitivity of nested polymerase chain reaction (PCR) offers the possibility of rapid detection of cytomegalovirus (CMV) DNA in serum. Five consecutive serum samples were examined from 52 human immunodeficiency virus (HIV)-seropositive patients (19 of whom had clinically presumed diagnosis...

  10. Development of human resources for Indian nuclear power ...

    Indian Academy of Sciences (India)

    This paper discusses the requirements of the human resource development programme for nuclear energy, the challenges in the way of its realization, its national and international status and traces the history of nuclear education in the country. It brings out the linkage of human resource development programme with the ...

  11. Nuclear power in human medicine; Kernenergie in der Humanmedizin

    Energy Technology Data Exchange (ETDEWEB)

    Kuczera, Bernhard

    2012-01-15

    The public widely associate nuclear power with the megawatt dimensions of nuclear power plants in which nuclear power is released and used for electricity production. While this use of nuclear power for electricity generation is rejected by part of the population adopting the polemic attitude of ''opting out of nuclear,'' the application of nuclear power in medicine is generally accepted. The appreciative, positive term used in this case is nuclear medicine. Both areas, nuclear medicine and environmentally friendly nuclear electricity production, can be traced back to one common origin, i.e. the ''Atoms for Peace'' speech by U.S. President Eisenhower to the U.N. Plenary Assembly on December 8, 1953. The methods of examination and treatment in nuclear medicine are illustrated in a few examples from the perspective of a nuclear engineer. Nuclear medicine is a medical discipline dealing with the use of radionuclides in humans for medical purposes. This is based on 2 principles, namely that the human organism is unable to distinguish among different isotopes in metabolic processes, and the radioactive substances are employed in amounts so small that metabolic processes will not be influenced. As in classical medicine, the application of these principles serves two complementary purposes: diagnosis and therapy. (orig.)

  12. The Role of RhoA, RhoB and RhoC GTPases in Cell Morphology, Proliferation and Migration in Human Cytomegalovirus (HCMV Infected Glioblastoma Cells

    Directory of Open Access Journals (Sweden)

    Melpomeni Tseliou

    2016-01-01

    Full Text Available Background/Aims: Rho GTPases are crucial regulators of the actin cytoskeleton, membrane trafficking and cell signaling and their importance in cell migration and invasion is well- established. The human cytomegalovirus (HCMV is a widespread pathogen responsible for generally asymptomatic and persistent infections in healthy people. Recent evidence indicates that HCMV gene products are expressed in over 90% of malignant type glioblastomas (GBM. In addition, the HCMV Immediate Early-1 protein (IE1 is expressed in >90% of tumors analyzed. Methods: RhoA, RhoB and RhoC were individually depleted in U373MG glioblastoma cells as well as U373MG cells stably expressing the HCMV IE1 protein (named U373MG-IE1 cells shRNA lentivirus vectors. Cell proliferation assays, migration as well as wound-healing assays were performed in uninfected and HCMV-infected cells. Results: The depletion of RhoA, RhoB and RhoC protein resulted in significant alterations in the morphology of the uninfected cells, which were further enhanced by the cytopathic effect caused by HCMV. Furthermore, in the absence or presence of HCMV, the knockdown of RhoB and RhoC proteins decreased the proliferation rate of the parental and the IE1-expressing glioblastoma cells, whereas the knockdown of RhoA protein in the HCMV infected cell lines restored their proliferation rate. In addition, wound healing assays in U373MG cells revealed that depletion of RhoA, RhoB and RhoC differentially reduced their migration rate, even in the presence or the absence of HCMV. Conclusion: Collectively, these data show for the first time a differential implication of Rho GTPases in morphology, proliferation rate and motility of human glioblastoma cells during HCMV infection, further supporting an oncomodulatory role of HCMV depending on the Rho isoforms' state.

  13. Human cytomegalovirus entry into dendritic cells occurs via a macropinocytosis-like pathway in a pH-independent and cholesterol-dependent manner.

    Science.gov (United States)

    Haspot, Fabienne; Lavault, Amélie; Sinzger, Christian; Laib Sampaio, Kerstin; Stierhof, York-Dieter; Pilet, Paul; Bressolette-Bodin, Céline; Halary, Franck

    2012-01-01

    Human cytomegalovirus (HCMV) is a ubiquitous herpesvirus that is able to infect fibroblastic, epithelial, endothelial and hematopoietic cells. Over the past ten years, several groups have provided direct evidence that dendritic cells (DCs) fully support the HCMV lytic cycle. We previously demonstrated that the C-type lectin dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) has a prominent role in the docking of HCMV on monocyte-derived DCs (MDDCs). The DC-SIGN/HCMV interaction was demonstrated to be a crucial and early event that substantially enhanced infection in trans, i.e., from one CMV-bearing cell to another non-infected cell (or trans-infection), and rendered susceptible cells fully permissive to HCMV infection. Nevertheless, nothing is yet known about how HCMV enters MDDCs. In this study, we demonstrated that VHL/E HCMV virions (an endothelio/dendrotropic strain) are first internalized into MDDCs by a macropinocytosis-like process in an actin- and cholesterol-dependent, but pH-independent, manner. We observed the accumulation of virions in large uncoated vesicles with endosomal features, and the virions remained as intact particles that retained infectious potential for several hours. This trans-infection property was specific to MDDCs because monocyte-derived macrophages or monocytes from the same donor were unable to allow the accumulation of and the subsequent transmission of the virus. Together, these data allowed us to delineate the early mechanisms of the internalization and entry of an endothelio/dendrotropic HCMV strain into human MDDCs and to propose that DCs can serve as a "Trojan horse" to convey CMV from entry sites to other locations that may favor the occurrence of either latency or acute infection.

  14. Human cytomegalovirus entry into dendritic cells occurs via a macropinocytosis-like pathway in a pH-independent and cholesterol-dependent manner.

    Directory of Open Access Journals (Sweden)

    Fabienne Haspot

    Full Text Available Human cytomegalovirus (HCMV is a ubiquitous herpesvirus that is able to infect fibroblastic, epithelial, endothelial and hematopoietic cells. Over the past ten years, several groups have provided direct evidence that dendritic cells (DCs fully support the HCMV lytic cycle. We previously demonstrated that the C-type lectin dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN has a prominent role in the docking of HCMV on monocyte-derived DCs (MDDCs. The DC-SIGN/HCMV interaction was demonstrated to be a crucial and early event that substantially enhanced infection in trans, i.e., from one CMV-bearing cell to another non-infected cell (or trans-infection, and rendered susceptible cells fully permissive to HCMV infection. Nevertheless, nothing is yet known about how HCMV enters MDDCs. In this study, we demonstrated that VHL/E HCMV virions (an endothelio/dendrotropic strain are first internalized into MDDCs by a macropinocytosis-like process in an actin- and cholesterol-dependent, but pH-independent, manner. We observed the accumulation of virions in large uncoated vesicles with endosomal features, and the virions remained as intact particles that retained infectious potential for several hours. This trans-infection property was specific to MDDCs because monocyte-derived macrophages or monocytes from the same donor were unable to allow the accumulation of and the subsequent transmission of the virus. Together, these data allowed us to delineate the early mechanisms of the internalization and entry of an endothelio/dendrotropic HCMV strain into human MDDCs and to propose that DCs can serve as a "Trojan horse" to convey CMV from entry sites to other locations that may favor the occurrence of either latency or acute infection.

  15. Human Factors in Nuclear Power Engineering in Polish Conditions

    Directory of Open Access Journals (Sweden)

    Agnieszka Kaczmarek-Kacprzak

    2014-09-01

    Full Text Available The paper “Human factors in nuclear power engineering in Polish conditions” focuses on analysis of dynamics of preparing Polish society to build fi rst nuclear power plant in XXI century in Poland. Authors compare experience from constructing nuclear power plant Sizewell B (Great Britain and Sizewell C, which is in preparation phase with polish nuclear power program. Paper includes aspects e.g. of creating nuclear safety culture and social opinion about investment. Human factors in nuclear power engineering are as well important as relevant economical and technical factors, but very often negligible. In Poland where history about Czarnobyl is still alive, and social opinion is created on emotions after accident in Fukushima, human factors are crucial and should be under comprehensive consideration.

  16. Human factors aspects of advanced instrumentation in the nuclear industry

    Energy Technology Data Exchange (ETDEWEB)

    Carter, R.J.

    1989-01-01

    An important consideration in regards to the use of advanced instrumentation in the nuclear industry is the interface between the instrumentation system and the human. A survey, oriented towards identifying the human factors aspects of digital instrumentation, was conducted at a number of United States (US) and Canadian nuclear vendors and utilities. Human factors issues, subsumed under the categories of computer-generated displays, controls, organizational support, training, and related topics were identified. 20 refs., 2 tabs.

  17. Loss of the Human Cytomegalovirus US16 Protein Abrogates Virus Entry into Endothelial and Epithelial Cells by Reducing the Virion Content of the Pentamer.

    Science.gov (United States)

    Luganini, Anna; Cavaletto, Noemi; Raimondo, Stefania; Geuna, Stefano; Gribaudo, Giorgio

    2017-06-01

    The human cytomegalovirus (HCMV) US12 gene family encodes a group of predicted seven-transmembrane proteins whose functions have yet to be established. While inactivation of individual US12 members in laboratory strains of HCMV does not affect viral replication in fibroblasts, disruption of the US16 gene in the low-passage-number TR strain prevents viral growth in endothelial and epithelial cells. In these cells, the US16-null viruses fail to express immediate early (IE), early (E), and late (L) viral proteins due to a defect which occurs prior to IE gene expression. Here, we show that this defective phenotype is a direct consequence of deficiencies in the entry of US16-null viruses in these cell types due to an impact on the gH/gL/UL128/UL130/UL131A (pentamer) complex. Indeed, viral particles released from fibroblasts infected with US16-null viruses were defective for the pentamer, thus preventing entry during infections of endothelial and epithelial cells. A link between pUS16 and the pentamer was further supported by the colocalization of pUS16 and pentamer proteins within the cytoplasmic viral assembly compartment (cVAC) of infected fibroblasts. Deletion of the C-terminal tail of pUS16 reproduced the defective growth phenotype and alteration of virion composition as US16-null viruses. However, the pentamer assembly and trafficking to the cVAC were not affected by the lack of the C terminus of pUS16. Coimmunoprecipitation results then indicated that US16 interacts with pUL130 but not with the mature pentamer or gH/gL/gO. Together, these results suggest that pUS16 contributes to the tropism of HCMV by influencing the content of the pentamer into virions.IMPORTANCE Human cytomegalovirus (HCMV) is major pathogen in newborns and immunocompromised individuals. A hallmark of HCMV pathogenesis is its ability to productively replicate in an exceptionally broad range of target cells. The virus infects a variety of cell types by exploiting different forms of the envelope

  18. A Tyrosine-Based Trafficking Motif of the Tegument Protein pUL71 Is Crucial for Human Cytomegalovirus Secondary Envelopment.

    Science.gov (United States)

    Dietz, Andrea N; Villinger, Clarissa; Becker, Stefan; Frick, Manfred; von Einem, Jens

    2018-01-01

    The human cytomegalovirus (HCMV) tegument protein pUL71 is required for efficient secondary envelopment and accumulates at the Golgi compartment-derived viral assembly complex (vAC) during infection. Analysis of various C-terminally truncated pUL71 proteins fused to enhanced green fluorescent protein (eGFP) identified amino acids 23 to 34 as important determinants for its Golgi complex localization. Sequence analysis and mutational verification revealed the presence of an N-terminal tyrosine-based trafficking motif (YXXΦ) in pUL71. This led us to hypothesize a requirement of the YXXΦ motif for the function of pUL71 in infection. Mutation of both the tyrosine residue and the entire YXXΦ motif resulted in an altered distribution of mutant pUL71 at the plasma membrane and in the cytoplasm during infection. Both YXXΦ mutant viruses exhibited similarly decreased focal growth and reduced virus yields in supernatants. Ultrastructurally, mutant-virus-infected cells exhibited impaired secondary envelopment manifested by accumulations of capsids undergoing an envelopment process. Additionally, clusters of capsid accumulations surrounding the vAC were observed, similar to the ultrastructural phenotype of a UL71-deficient mutant. The importance of endocytosis and thus the YXXΦ motif for targeting pUL71 to the Golgi complex was further demonstrated when clathrin-mediated endocytosis was inhibited either by coexpression of the C-terminal part of cellular AP180 (AP180-C) or by treatment with methyl-β-cyclodextrin. Both conditions resulted in a plasma membrane accumulation of pUL71. Altogether, these data reveal the presence of a functional N-terminal endocytosis motif that is an important determinant for intracellular localization of pUL71 and that is furthermore required for the function of pUL71 during secondary envelopment of HCMV capsids at the vAC. IMPORTANCE Human cytomegalovirus (HCMV) is the leading cause of birth defects among congenital virus infections and can

  19. Monitoring of Human Cytomegalovirus and Virus-Specific T-Cell Response in Young Patients Receiving Allogeneic Hematopoietic Stem Cell Transplantation

    Science.gov (United States)

    Lilleri, Daniele; Gerna, Giuseppe; Zelini, Paola; Chiesa, Antonella; Rognoni, Vanina; Mastronuzzi, Angela; Giorgiani, Giovanna; Zecca, Marco; Locatelli, Franco

    2012-01-01

    In allogeneic hematopoietic stem-cell transplantation (HSCT) recipients, outcome of human cytomegalovirus (HCMV) infection results from balance between viral load/replication and pathogen-specific T-cell response. Using a cut-off of 30,000 HCMV DNA copies/ml blood for pre-emptive therapy and cut-offs of 1 and 3 virus-specific CD4+ and CD8+ T cells/µl blood for T-cell protection, we conducted in 131 young patients a prospective 3-year study aimed at verifying whether achievement of such immunological cut-offs protects from HCMV disease. In the first three months after transplantation, 55/89 (62%) HCMV-seropositive patients had infection and 36/55 (65%) were treated pre-emptively, whereas only 7/42 (17%) HCMV-seronegative patients developed infection and 3/7 (43%) were treated. After 12 months, 76 HCMV-seropositive and 9 HCMV-seronegative patients (cumulative incidence: 90% and 21%, respectively) displayed protective HCMV-specific immunity. Eighty of these 85 (95%) patients showed spontaneous control of HCMV infection without additional treatment. Five patients after reaching protective T-cell levels needed pre-emptive therapy, because they developed graft-versus-host disease (GvHD). HSCT recipients reconstituting protective levels of HCMV-specific T-cells in the absence of GvHD are no longer at risk for HCMV disease, at least within 3 years after transplantation. The decision to treat HCMV infection in young HSCT recipients may be taken by combining virological and immunological findings. PMID:22848556

  20. Human cytomegalovirus prevalence and distribution of glycoprotein B, O genotypes among hospitalized children with respiratory infections in West China, 2009-2014.

    Science.gov (United States)

    Chen, Jia-Yi; Zheng, Tian-Li; Zhou, Tao; Hu, Peng-Wei; Huang, Meng-Jiao; Xu, Xin; Pei, Xiao-Fang

    2016-11-01

    Human cytomegalovirus (HCMV) is an important pathogen causing morbidity and mortality in children. HCMV prevalence in children with respiratory infections has not been investigated in West China. Previous studies have suggested that glycoproteins genotypes may be associated with different clinical presentations, but the associations were controversial. The aim of this study was to determine the prevalence of HCMV infection in children with respiratory infections, the distributions of gB, gO genotypes among these isolates and their potential predictive roles for the development of symptoms in children. A total of 1709 respiratory specimens were obtained from hospitalised children with respiratory symptoms from 2009 to 2014 for the confirmation of HCMV infection. Glycoprotein B,O genotyping was carried out by multiplex nested PCR and sequencing. The overall infection rate was 10.8%, and dominant genotypes were gB1 (74.2%) and gO1 (37.1%). Clinical characteristics differed between infants and children >1 year of age. Infants infected with HCMV had a higher frequency of fever (P < 0.001), cough (P < 0.001), rhinorrhea (P < 0.001), expectoration (P = 0.001) and diarrhoea (P = 0.005). Children <1 year age infected with gB1 had a higher rate of cough (P = 0.0192). Infants infected with HCMV had a severe clinical outcome. gB1 may negatively associate with clinical presentations and quality of life in these children. The prevalence of HCMV infection and genotype distribution emphasises the importance of HCMV screening, vaccination and control for transmission. © 2016 John Wiley & Sons Ltd.

  1. Intrapulmonary Human Cytomegalovirus Replication in Lung Transplant Recipients Is Associated With a Rise of CCL-18 and CCL-20 Chemokine Levels.

    Science.gov (United States)

    Weseslindtner, Lukas; Görzer, Irene; Roedl, Kevin; Küng, Erik; Jaksch, Peter; Klepetko, Walter; Puchhammer-Stöckl, Elisabeth

    2017-01-01

    In lung transplant recipients (LTRs), human cytomegalovirus (HCMV) DNA detection in the bronchoalveolar lavage fluid (BALF) indicates HCMV replication in the pulmonary compartment. Such local HCMV replication episodes may remain asymptomatic or may lead to symptomatic HCMV disease. Here, we investigated LTRs with intrapulmonary HCMV replication for the chemokines CCL-18 and CCL-20. In particular, we analyzed whether these chemokines rise in the allograft and/or the blood and are associated with HCMV disease. CCL-18 and CCL-20 levels were quantitated by ELISA in BALF and serum samples from 60 LTRs. During the posttransplant follow-up, these LTRs displayed HCMV DNA detection in the BALF by PCR, whereas other infectious agents were undetectable. Furthermore, we investigated samples from 10 controls who did not display any HCMV replication episode during the follow-up. HCMV replication in the allograft was associated with a significant increase of CCL-18 and CCL-20 BALF levels (P < 0.001, Wilcoxon signed-rank test) and a significant rise of CCL-20 (P < 0.0001, Wilcoxon signed-rank test) but not of CCL-18 in the blood. In controls, no such chemokine increase was observed. Furthermore, CCL-18 BALF levels were significantly higher in 8 LTRs who additionally developed HCMV disease, as compared with the other 52 patients in whom HCMV replication remained asymptomatic (P < 0.001, Mann-Whitney U test). HCMV replication in the allograft causes an intrapulmonary increase of CCL-18 and CCL-20 and a systemic rise of CCL-20 serum levels. Strong intrapulmonary CCL-18 responses are associated with symptomatic HCMV disease, proposing that CCL-18 BALF levels could serve as a marker.

  2. The human cytomegalovirus UL11 protein interacts with the receptor tyrosine phosphatase CD45, resulting in functional paralysis of T cells.

    Directory of Open Access Journals (Sweden)

    Ildar Gabaev

    2011-12-01

    Full Text Available Human cytomegalovirus (CMV exerts diverse and complex effects on the immune system, not all of which have been attributed to viral genes. Acute CMV infection results in transient restrictions in T cell proliferative ability, which can impair the control of the virus and increase the risk of secondary infections in patients with weakened or immature immune systems. In a search for new immunomodulatory proteins, we investigated the UL11 protein, a member of the CMV RL11 family. This protein family is defined by the RL11 domain, which has homology to immunoglobulin domains and adenoviral immunomodulatory proteins. We show that pUL11 is expressed on the cell surface and induces intercellular interactions with leukocytes. This was demonstrated to be due to the interaction of pUL11 with the receptor tyrosine phosphatase CD45, identified by mass spectrometry analysis of pUL11-associated proteins. CD45 expression is sufficient to mediate the interaction with pUL11 and is required for pUL11 binding to T cells, indicating that pUL11 is a specific CD45 ligand. CD45 has a pivotal function regulating T cell signaling thresholds; in its absence, the Src family kinase Lck is inactive and signaling through the T cell receptor (TCR is therefore shut off. In the presence of pUL11, several CD45-mediated functions were inhibited. The induction of tyrosine phosphorylation of multiple signaling proteins upon TCR stimulation was reduced and T cell proliferation was impaired. We therefore conclude that pUL11 has immunosuppressive properties, and that disruption of T cell function via inhibition of CD45 is a previously unknown immunomodulatory strategy of CMV.

  3. The Novel Anticytomegalovirus Compound AIC246 (Letermovir) Inhibits Human Cytomegalovirus Replication through a Specific Antiviral Mechanism That Involves the Viral Terminase ▿

    Science.gov (United States)

    Goldner, Thomas; Hewlett, Guy; Ettischer, Nicole; Ruebsamen-Schaeff, Helga; Zimmermann, Holger; Lischka, Peter

    2011-01-01

    Human cytomegalovirus (HCMV) remains the leading viral cause of birth defects and life-threatening disease in transplant recipients. All approved antiviral drugs target the viral DNA polymerase and are associated with severe toxicity issues and the emergence of drug resistance. Attempts to discover improved anti-HCMV drugs led to the identification of the small-molecular-weight compound AIC246 (Letermovir). AIC246 exhibits outstanding anti-HCMV activity in vitro and in vivo and currently is undergoing a clinical phase IIb trial. The initial mode-of-action studies suggested that the drug acts late in the HCMV replication cycle via a mechanism distinct from that of polymerase inhibitors. Here, we extend our mode-of-action analyses and report that AIC246 blocks viral replication without inhibiting the synthesis of progeny HCMV DNA or viral proteins. The genotyping of mutant viruses that escaped AIC246 inhibition uncovered distinct point mutations in the UL56 subunit of the viral terminase complex. Marker transfer analyses confirmed that these mutations were sufficient to mediate AIC246 resistance. The mapping of drug resistance to open reading frame UL56 suggests that viral DNA processing and/or packaging is targeted by AIC246. In line with this, we demonstrate that AIC246 affects the formation of proper unit-length genomes from viral DNA concatemers and interferes with virion maturation. However, since AIC246-resistant viruses do not exhibit cross-resistance to previously published terminase inhibitors, our data suggest that AIC246 interferes with HCMV DNA cleavage/packaging via a molecular mechanism that is distinct from that of other compound classes known to target the viral terminase. PMID:21752907

  4. The novel anticytomegalovirus compound AIC246 (Letermovir) inhibits human cytomegalovirus replication through a specific antiviral mechanism that involves the viral terminase.

    Science.gov (United States)

    Goldner, Thomas; Hewlett, Guy; Ettischer, Nicole; Ruebsamen-Schaeff, Helga; Zimmermann, Holger; Lischka, Peter

    2011-10-01

    Human cytomegalovirus (HCMV) remains the leading viral cause of birth defects and life-threatening disease in transplant recipients. All approved antiviral drugs target the viral DNA polymerase and are associated with severe toxicity issues and the emergence of drug resistance. Attempts to discover improved anti-HCMV drugs led to the identification of the small-molecular-weight compound AIC246 (Letermovir). AIC246 exhibits outstanding anti-HCMV activity in vitro and in vivo and currently is undergoing a clinical phase IIb trial. The initial mode-of-action studies suggested that the drug acts late in the HCMV replication cycle via a mechanism distinct from that of polymerase inhibitors. Here, we extend our mode-of-action analyses and report that AIC246 blocks viral replication without inhibiting the synthesis of progeny HCMV DNA or viral proteins. The genotyping of mutant viruses that escaped AIC246 inhibition uncovered distinct point mutations in the UL56 subunit of the viral terminase complex. Marker transfer analyses confirmed that these mutations were sufficient to mediate AIC246 resistance. The mapping of drug resistance to open reading frame UL56 suggests that viral DNA processing and/or packaging is targeted by AIC246. In line with this, we demonstrate that AIC246 affects the formation of proper unit-length genomes from viral DNA concatemers and interferes with virion maturation. However, since AIC246-resistant viruses do not exhibit cross-resistance to previously published terminase inhibitors, our data suggest that AIC246 interferes with HCMV DNA cleavage/packaging via a molecular mechanism that is distinct from that of other compound classes known to target the viral terminase.

  5. Distribution of human Cytomegalovirus gB genotypes in samples from pediatric patients in Parma during the period 1995-2003

    Directory of Open Access Journals (Sweden)

    Maria Cristina Arcangeletti

    2011-03-01

    Full Text Available Background. Human Cytomegalovirus (HCMV infection is a leading cause of developmental disability and late neurological sequelae in children. Several literature data indicate that HCMV pericapsidic glycoprotein B (gB is highly immunogenic and is involved in virus-cell interaction.The gB gene has hypervariable regions producing four genotypes (gB1, gB2, gB3, gB4; however, the correlation between gB genotypes and HCMV infection outcome remains unclear. Objectives. The main goal of this study was that of evaluating the distribution of HCMV gB genotypes in samples from pediatric population in Parma with congenital, perinatal or post-natal infections, in order to find a correlation between viral gB genotypes and the clinical outcome of the infection. Study design. Forty eight urine samples, selected between 1999 and 2003 and stored at -80°C, underwent DNA extraction, nested PCR amplification of a gB gene region and restriction polymorphism analysis (RFLP. Results. The gB genotypes distribution in the considered pediatric population was as follows: gB1 was the most diffused (45.83% followed by gB2 (22.92%, gB3 (16.67% and gB4 (14.58%. Conclusions. In the considered population, gB1 was the most represented genotype and was often found in congenital and perinatal symptomatic infections, as well as in post-natal, asymptomatic infections.

  6. Human cytomegalovirus tegument protein pp65 is detected in all intra- and extra-axial brain tumours independent of the tumour type or grade.

    Directory of Open Access Journals (Sweden)

    Sylwia Libard

    Full Text Available Human cytomegalovirus (HCMV has been indicated being a significant oncomodulator. Recent reports have suggested that an antiviral treatment alters the outcome of a glioblastoma. We analysed the performance of commercial HCMV-antibodies applying the immunohistochemical (IHC methods on brain sample obtained from a subject with a verified HCMV infection, on samples obtained from 14 control subjects, and on a tissue microarray block containing cores of various brain tumours. Based on these trials, we selected the best performing antibody and analysed a cohort of 417 extra- and intra-axial brain tumours such as gliomas, medulloblastomas, primary diffuse large B-cell lymphomas, and meningiomas. HCMV protein pp65 immunoreactivity was observed in all types of tumours analysed, and the IHC expression did not depend on the patient's age, gender, tumour type, or grade. The labelling pattern observed in the tumours differed from the labelling pattern observed in the tissue with an active HCMV infection. The HCMV protein was expressed in up to 90% of all the tumours investigated. Our results are in accordance with previous reports regarding the HCMV protein expression in glioblastomas and medulloblastomas. In addition, the HCMV protein expression was seen in primary brain lymphomas, low-grade gliomas, and in meningiomas. Our results indicate that the HCMV protein pp65 expression is common in intra- and extra-axial brain tumours. Thus, the assessment of the HCMV expression in tumours of various origins and pathologically altered tissue in conditions such as inflammation, infection, and even degeneration should certainly be facilitated.

  7. Cis and trans acting factors involved in human cytomegalovirus experimental and natural latent infection of CD14 (+) monocytes and CD34 (+) cells.

    Science.gov (United States)

    Rossetto, Cyprian C; Tarrant-Elorza, Margaret; Pari, Gregory S

    2013-01-01

    The parameters involved in human cytomegalovirus (HCMV) latent infection in CD14 (+) and CD34 (+) cells remain poorly identified. Using next generation sequencing we deduced the transcriptome of HCMV latently infected CD14 (+) and CD34 (+) cells in experimental as well as natural latency settings. The gene expression profile from natural infection in HCMV seropositive donors closely matched experimental latency models, and included two long non-coding RNAs (lncRNAs), RNA4.9 and RNA2.7 as well as the mRNAs encoding replication factors UL84 and UL44. Chromatin immunoprecipitation assays on experimentally infected CD14 (+) monocytes followed by next generation sequencing (ChIP-Seq) were employed to demonstrate both UL84 and UL44 proteins interacted with the latent viral genome and overlapped at 5 of the 8 loci identified. RNA4.9 interacts with components of the polycomb repression complex (PRC) as well as with the MIE promoter region where the enrichment of the repressive H3K27me3 mark suggests that this lncRNA represses transcription. Formaldehyde Assisted Isolation of Regulatory Elements (FAIRE), which identifies nucleosome-depleted viral DNA, was used to confirm that latent mRNAs were associated with actively transcribed, FAIRE analysis also showed that the terminal repeat (TR) region of the latent viral genome is depleted of nucleosomes suggesting that this region may contain an element mediating viral genome maintenance. ChIP assays show that the viral TR region interacts with factors associated with the pre replication complex and a plasmid subclone containing the HCMV TR element persisted in latently infected CD14 (+) monocytes, strongly suggesting that the TR region mediates viral chromosome maintenance.

  8. Cis and trans acting factors involved in human cytomegalovirus experimental and natural latent infection of CD14 (+ monocytes and CD34 (+ cells.

    Directory of Open Access Journals (Sweden)

    Cyprian C Rossetto

    Full Text Available The parameters involved in human cytomegalovirus (HCMV latent infection in CD14 (+ and CD34 (+ cells remain poorly identified. Using next generation sequencing we deduced the transcriptome of HCMV latently infected CD14 (+ and CD34 (+ cells in experimental as well as natural latency settings. The gene expression profile from natural infection in HCMV seropositive donors closely matched experimental latency models, and included two long non-coding RNAs (lncRNAs, RNA4.9 and RNA2.7 as well as the mRNAs encoding replication factors UL84 and UL44. Chromatin immunoprecipitation assays on experimentally infected CD14 (+ monocytes followed by next generation sequencing (ChIP-Seq were employed to demonstrate both UL84 and UL44 proteins interacted with the latent viral genome and overlapped at 5 of the 8 loci identified. RNA4.9 interacts with components of the polycomb repression complex (PRC as well as with the MIE promoter region where the enrichment of the repressive H3K27me3 mark suggests that this lncRNA represses transcription. Formaldehyde Assisted Isolation of Regulatory Elements (FAIRE, which identifies nucleosome-depleted viral DNA, was used to confirm that latent mRNAs were associated with actively transcribed, FAIRE analysis also showed that the terminal repeat (TR region of the latent viral genome is depleted of nucleosomes suggesting that this region may contain an element mediating viral genome maintenance. ChIP assays show that the viral TR region interacts with factors associated with the pre replication complex and a plasmid subclone containing the HCMV TR element persisted in latently infected CD14 (+ monocytes, strongly suggesting that the TR region mediates viral chromosome maintenance.

  9. Ganciclovir penetrates into the cerebrospinal fluid of an infant with congenital cytomegalovirus infection.

    Science.gov (United States)

    Natale, Fabio; Bizzarri, Bianca; Cardi, Veronica; Gaeta, Aurelia; Villani, Paola; Liuzzi, Giuseppina; De Curtis, Mario

    2015-03-31

    Currently, there is no evidence whether ganciclovir, or its oral prodrug valganciclovir, penetrates into the cerebrospinal fluid of human infants treated for congenital cytomegalovirus infection. Here, we report a case study providing evidence that ganciclovir, administered as valganciclovir, reaches the infant's cerebrospinal fluid when used at the currently recommended dose for congenital cytomegalovirus infection.

  10. Modelling human resource requirements for the nuclear industry in Europe

    Energy Technology Data Exchange (ETDEWEB)

    Roelofs, Ferry [Nuclear Research and Consultancy Group (NRG) (Netherlands); Flore, Massimo; Estorff, Ulrik von [Joint Research Center (JRC) (Netherlands)

    2017-11-15

    The European Human Resource Observatory for Nuclear (EHRO-N) provides the European Commission with essential data related to supply and demand for nuclear experts in the EU-28 and the enlargement and integration countries based on bottom-up information from the nuclear industry. The objective is to assess how the supply of experts for the nuclear industry responds to the needs for the same experts for present and future nuclear projects in the region. Complementary to the bottom-up approach taken by the EHRO-N team at JRC, a top-down modelling approach has been taken in a collaboration with NRG in the Netherlands. This top-down modelling approach focuses on the human resource requirements for operation, construction, decommissioning, and efforts for long term operation of nuclear power plants. This paper describes the top-down methodology, the model input, the main assumptions, and the results of the analyses.

  11. Initial Human Response to Nuclear Radiation

    Science.gov (United States)

    1982-04-01

    Nuclear Excursion," New England Journal of Medicine, Vol. 272, No. 15, 15 April 1965, pp. 755-761. On 24 July 1964, at a United Nuclear Corporation plant... nematological depression, days 24 to 34, the patient remained well. On day 36 a mouth infection was treated with orally administeied penicillin. By day...l Journal , II April 1953, pp. 802-805. Fifty patients were given a single therapeutic X-ray dose of about 150 rads. They were primarily afflicted

  12. Promotion of Bilateral Cooperative Programs in Nuclear Human Resources Development

    Energy Technology Data Exchange (ETDEWEB)

    Lee, E. J.; Han, K. W.; Nam, Y. M. (and others)

    2009-08-15

    The purpose of this project is strengthening of bilateral cooperation with those countries for sharing Korea's technology, and providing of education and training on Korean experience regarding national nuclear policy, technology self reliance, and technology itself, in the field of nuclear power generation and the application of radioisotopes and radiation. This project covers an analysis on the need of nuclear human resource development in countries having interest in the introduction of nuclear power and/or promotion of the use of nuclear energy, and provision of courses on 'nuclear power policy, planning and management' and 'design and operation of nuclear research reactor, and application of radiation technology' along with the country specific needs. Education and training of key members in nuclear energy development from Egypt: It was implemented through bilateral cooperation and support by KOICA program. The first part, which targeted staff members from Egypt Nuclear Commission, was held for 2 months providing a KOICA course on policy, planning and management for nuclear power project, and second part was on the job training in Korea Hydro and Nuclear Power and Korea Institute of Nuclear Safety, KAERI respectively. On the job training of 1 scientist from Vietnam was implemented on the basis of bilateral cooperation in a research laboratory on radioactive waste treatment technology, at KAERI. Education and training for scientists from South East RCA countries were carried out for 11 participants from Vietnam, Thailand, Indonesia, China, Pakistan, Malaysia, Philippines, and Bangladesh. The course dealt with nuclear research reactor and radiation application technology. Development of nuclear education and training programs for key persons involved in nuclear power projects from countries of Middle East: The developed program consists of 15 courses addressing 3 technical levels, i.e. high level policy makers, middle level project

  13. Human Cytomegalovirus miR-UL112-3p Targets TLR2 and Modulates the TLR2/IRAK1/NFκB Signaling Pathway.

    Directory of Open Access Journals (Sweden)

    Igor Landais

    2015-05-01

    Full Text Available Human Cytomegalovirus (HCMV encodes multiple microRNAs (miRNAs whose functions are just beginning to be uncovered. Using in silico approaches, we identified the Toll-Like Receptor (TLR innate immunity pathway as a possible target of HCMV miRNAs. Luciferase reporter assay screens further identified TLR2 as a target of HCMV miR-UL112-3p. TLR2 plays a major role in innate immune response by detecting both bacterial and viral ligands, including HCMV envelope proteins gB and gH. TLR2 activates a variety of signal transduction routes including the NFκB pathway. Furthermore, TLR2 plays an important role in controlling CMV infection both in humans and in mice. Immunoblot analysis of cells transfected with a miR-UL112-3p mimic revealed that endogenous TLR2 is down-regulated by miR-UL112-3p with similar efficiency as a TLR2-targeting siRNA (siTLR2. We next found that TLR2 protein level decreases at late times during HCMV infection and correlates with miR-UL112-3p accumulation in fibroblasts and monocytic THP1 cells. Confirming direct miR-UL112-3p targeting, down-regulation of endogenous TLR2 was not observed in cells infected with HCMV mutants deficient in miR-UL112-3p expression, but transfection of miR-UL112-3p in these cells restored TLR2 down-regulation. Using a NFκB reporter cell line, we found that miR-UL112-3p transfection significantly inhibited NFκB-dependent luciferase activity with similar efficiency as siTLR2. Consistent with this observation, miR-UL112-3p transfection significantly reduced the expression of multiple cytokines (IL-1β, IL-6 and IL-8 upon stimulation with a TLR2 agonist. Finally, miR-UL112-3p transfection significantly inhibited the TLR2-induced post-translational activation of IRAK1, a kinase located in the upstream section of the TLR2/NFκB signaling axis. To our knowledge, this is the first identified mechanism of TLR2 modulation by HCMV and is the first report of functional targeting of TLR2 by a viral miRNA. These

  14. Preliminary Evaluation of Next-Generation Sequencing Performance Relative to qPCR and In Vitro Cell Culture Tests for Human Cytomegalovirus.

    Science.gov (United States)

    Ng, Siemon H S; Azizi, Ali; Edamura, Kerrie; Malott, Rebecca J; Charlebois, Robert L; Logvinoff, Carine; Schreiber, Martha; Mallet, Laurent; Gisonni-Lex, Lucy

    2014-01-01

    To compare the performances of conventional in vitro indicator cell culture, quantitative polymerase chain reaction (qPCR), and next-generation sequencing (NGS) as detection methods for adventitious agents, a preliminary assessment was performed using human cytomegalovirus (HCMV) as a model virus. HCMV was spiked into a crude viral harvest at various concentrations and inoculated onto MRC-5 cell monolayers. The cultures were observed for cytopathic effects (CPEs) as per the compendial method requirements, and samples were taken at various time points for analysis by qPCR or NGS. When using NGS, the detection of HCMV is 10 fold more sensitive than observed using the conventional CPE endpoint method. It may be possible for qPCR to achieve the detection level demonstrated by NGS, but further optimization of the technique would be required. In addition, NGS was able to detect HCMV in the initial inoculum when it was spiked in at 10 CCID50/mL, suggesting the potential to eliminate cell culture amplification with an NGS-based assay. This study highlights the advantage of NGS as a surrogate broad-spectrum technology for the detection of adventitious agents in biologics. Human cytomeglovirus (HCMV) is highly prevalent in the general population and can lead to serious health issues in both immumocompromised individuals and/or newborns. The testing of HCMV in biological materials is stipulated by multiple regulatory agencies where HCMV is a potential risk. This test involves inoculating cell lines that are susceptible to HCMV infection, incubating the cultures for 28 days, and observing the cells for signs of viral infection. Next-generation sequencing and quantitative polymerase chain reaction (qPCR) are two technologies that can potentially shorten the extended 28 day cell culture incubation. In this study, we compared the sensitivity of the compendial cell culture assay with NGS and qPCR for the detection of HCMV. Our results show that NGS can potentially achieve a

  15. Human Cytomegalovirus Immediate-Early 1 Protein Rewires Upstream STAT3 to Downstream STAT1 Signaling Switching an IL6-Type to an IFNγ-Like Response.

    Directory of Open Access Journals (Sweden)

    Thomas Harwardt

    2016-07-01

    Full Text Available The human cytomegalovirus (hCMV major immediate-early 1 protein (IE1 is best known for activating transcription to facilitate viral replication. Here we present transcriptome data indicating that IE1 is as significant a repressor as it is an activator of host gene expression. Human cells induced to express IE1 exhibit global repression of IL6- and oncostatin M-responsive STAT3 target genes. This repression is followed by STAT1 phosphorylation and activation of STAT1 target genes normally induced by IFNγ. The observed repression and subsequent activation are both mediated through the same region (amino acids 410 to 445 in the C-terminal domain of IE1, and this region serves as a binding site for STAT3. Depletion of STAT3 phenocopies the STAT1-dependent IFNγ-like response to IE1. In contrast, depletion of the IL6 receptor (IL6ST or the STAT kinase JAK1 prevents this response. Accordingly, treatment with IL6 leads to prolonged STAT1 instead of STAT3 activation in wild-type IE1 expressing cells, but not in cells expressing a mutant protein (IE1dl410-420 deficient for STAT3 binding. A very similar STAT1-directed response to IL6 is also present in cells infected with a wild-type or revertant hCMV, but not an IE1dl410-420 mutant virus, and this response results in restricted viral replication. We conclude that IE1 is sufficient and necessary to rewire upstream IL6-type to downstream IFNγ-like signaling, two pathways linked to opposing actions, resulting in repressed STAT3- and activated STAT1-responsive genes. These findings relate transcriptional repressor and activator functions of IE1 and suggest unexpected outcomes relevant to viral pathogenesis in response to cytokines or growth factors that signal through the IL6ST-JAK1-STAT3 axis in hCMV-infected cells. Our results also reveal that IE1, a protein considered to be a key activator of the hCMV productive cycle, has an unanticipated role in tempering viral replication.

  16. Novel Human Cytomegalovirus Viral Chemokines, vCXCL-1s, Display Functional Selectivity for Neutrophil Signaling and Function

    NARCIS (Netherlands)

    Heo, Jinho; Dogra, Pranay; Masi, Tom J; Pitt, Elisabeth A; de Kruijf, Petra; Smit, Martine J; Sparer, Tim E

    2015-01-01

    Human CMV (HCMV) uses members of the hematopoietic system including neutrophils for dissemination throughout the body. HCMV encodes a viral chemokine, vCXCL-1, that is postulated to attract neutrophils for dissemination within the host. The gene encoding vCXCL-1, UL146, is one of the most variable

  17. Sociodemographic factors associated with IgG and IgM seroprevalence for human cytomegalovirus infection in adult populations of Pakistan: a seroprevalence survey

    Directory of Open Access Journals (Sweden)

    Saira Ibrahim

    2016-10-01

    Full Text Available Abstract Background The seroprevalence of human cytomegalovirus (HCMV infection ranges from 30 to 90 % in developed countries. Reliable estimates of HCMV seroprevalence are not available for Pakistan. This study determined the seroprevalence and sociodemographic factors associated with HCMV infection in adult populations of Karachi, Pakistan. Methods A seroprevalence survey was conducted on 1000 adults, including residents of two semi-urban communities, and visitors to a government and a private hospital. Questionnaire-based interviews were conducted. Sera were analysed for HCMV-specific IgG and IgM. Chi-square or Fisher’s exact test was used for comparing sociodemographic variables against seropositivity of HCMV-IgG or IgM. Multiple logistic regression modeling was performed for IgG seroprevalence and adjusted odds ratios were computed. Results The seroprevalence of HCMV-IgG and IgM was 93.2 and 4.3 % respectively. 95.3 % of individuals who were IgM seropositive were also seropositive for IgG. Around 6 % (15/250 of women of childbearing age remained uninfected and were therefore susceptible to primary infection. HCMV-IgG seroprevalence was associated with being female (p = 0.001, increasing age (p = 0.002 and crowding index (p = 0.003 and also with lower levels of both education (p < 0.001 and income (p = 0.008. Seroprevalence also differed significantly by marital status (p = 0.008 and sampling location (p < 0.001. A logistic regression model for HCMV-IgG seroprevalence showed associations with being female (OR = 1.89; 95 % CI: 1.10–3.25, increasing age (OR = 3.95; 95 % CI: 1.79–8.71 and decreasing income (OR = 0.72; 95 % CI: 0.54–0.96. A strong association was observed between increased seroprevalence of HCMV-IgM and decreasing household size (p = 0.008. Conclusions Seroprevalence of HCMV is very high in Pakistan, although 6 % of women of childbearing age remain at risk of primary

  18. Sociodemographic factors associated with IgG and IgM seroprevalence for human cytomegalovirus infection in adult populations of Pakistan: a seroprevalence survey.

    Science.gov (United States)

    Ibrahim, Saira; Siddiqui, Anwar A; Siddiqui, Amna R; Ahmed, Waquaruddin; Moss, Paul A H; Lalani, El-Nasir M A

    2016-10-22

    The seroprevalence of human cytomegalovirus (HCMV) infection ranges from 30 to 90 % in developed countries. Reliable estimates of HCMV seroprevalence are not available for Pakistan. This study determined the seroprevalence and sociodemographic factors associated with HCMV infection in adult populations of Karachi, Pakistan. A seroprevalence survey was conducted on 1000 adults, including residents of two semi-urban communities, and visitors to a government and a private hospital. Questionnaire-based interviews were conducted. Sera were analysed for HCMV-specific IgG and IgM. Chi-square or Fisher's exact test was used for comparing sociodemographic variables against seropositivity of HCMV-IgG or IgM. Multiple logistic regression modeling was performed for IgG seroprevalence and adjusted odds ratios were computed. The seroprevalence of HCMV-IgG and IgM was 93.2 and 4.3 % respectively. 95.3 % of individuals who were IgM seropositive were also seropositive for IgG. Around 6 % (15/250) of women of childbearing age remained uninfected and were therefore susceptible to primary infection. HCMV-IgG seroprevalence was associated with being female (p = 0.001), increasing age (p = 0.002) and crowding index (p = 0.003) and also with lower levels of both education (p < 0.001) and income (p = 0.008). Seroprevalence also differed significantly by marital status (p = 0.008) and sampling location (p < 0.001). A logistic regression model for HCMV-IgG seroprevalence showed associations with being female (OR = 1.89; 95 % CI: 1.10-3.25), increasing age (OR = 3.95; 95 % CI: 1.79-8.71) and decreasing income (OR = 0.72; 95 % CI: 0.54-0.96). A strong association was observed between increased seroprevalence of HCMV-IgM and decreasing household size (p = 0.008). Seroprevalence of HCMV is very high in Pakistan, although 6 % of women of childbearing age remain at risk of primary infection. The IgM seropositivity observed in some individuals

  19. Association of SNPs from IL1A, IL1B, and IL6 Genes with Human Cytomegalovirus Infection Among Pregnant Women.

    Science.gov (United States)

    Wujcicka, Wioletta Izabela; Wilczyński, Jan Szczęsny; Nowakowska, Dorota Ewa

    2017-05-01

    The study was aimed to estimate the role and prevalence rates of genotypes, haplotypes, and alleles, located within the single-nucleotide polymorphisms (SNPs) of interleukin (IL) 1A, IL1B, and IL6 genes, in the occurrence and development of human cytomegalovirus (HCMV) infection among pregnant women. A research was conducted in 129 pregnant women, out of whom, 65 were HCMV infected and 64 were age-matched control uninfected individuals. HCMV DNA was quantitated for UL55 gene by the real-time Q PCR in the body fluids. The genotypic statuses within the SNPs were determined by nested PCR-RFLP assays and confirmed, by sequencing for randomly selected representative PCR products. A relationship between the genotypes and alleles, as well as haplotypes and multiple variants in the studied polymorphisms, and the occurrence of HCMV infection in pregnant women, was determined using a logistic regression model. TT genotype within IL1A polymorphism significantly decreased the risk of HCMV infection (OR 0.32, 95% CI 0.09-1.05; p ≤ 0.050). Considering IL6 SNP, the prevalence rate of GC genotype was significantly decreased among the HCMV infected, compared to the uninfected control individuals (OR 0.45, 95% CI 0.21-0.99; p ≤ 0.050). Moreover, CC homozygotic status in IL6 SNP, found in pregnant women, significantly decreased the risk of congenital infection with HCMV in their offsprings (OR 0.12; p ≤ 0.050). In multiple SNP analysis, TC haplotype within the IL1 polymorphisms significantly decreased the risk of the infection in pregnant women (OR 0.38 95% CI 0.15-0.96; p ≤ 0.050). In addition, TTG complex variants for all the studied polymorphisms and TG variants for IL1B and IL6 SNPs were significantly more prevalent among the infected offsprings with symptomatic congenital cytomegaly than among the asymptomatic cases (p ≤ 0.050). In conclusion, the analyzed IL1A -889 C>T, IL1B +3954 C>T, and IL6 -174 G>C polymorphisms may be associated with the

  20. Phenotype and specificity of T cells in primary human cytomegalovirus infection during pregnancy: IL-7Rpos long-term memory phenotype is associated with protection from vertical transmission.

    Science.gov (United States)

    Mele, Federico; Fornara, Chiara; Jarrossay, David; Furione, Milena; Arossa, Alessia; Spinillo, Arsenio; Lanzavecchia, Antonio; Gerna, Giuseppe; Sallusto, Federica; Lilleri, Daniele

    2017-01-01

    Congenital human cytomegalovirus (HCMV) infection is the major cause of birth defects and a precise definition of the HCMV-specific T-cell response in primary infection may help define reliable correlates of immune protection during pregnancy. In this study, a high throughput method was used to define the frequency of CD4+ and CD8+ T cells specific for four HCMV proteins in the naïve compartment of seronegative subjects and the effector/memory compartments of subjects with primary/remote HCMV infection. The naïve repertoire displayed comparable frequencies of T cells that were reactive with HCMV structural (pp65, gB and the pentamer gHgLpUL128L) and non-structural (IE-1) proteins. Whereas, following natural infection, the majority of effector/memory CD4+ and CD8+ T cells recognized either gB or IE-1, respectively, and pp65. The pattern of T cell reactivity was comparable at early and late stages of infection and in pregnant women with primary HCMV infection transmitting or not transmitting the virus to the fetus. At an early stage of primary infection, about 50% of HCMV-reactive CD4+ T cells were long-term IL-7Rpos memory cells, while 6-12 months later, the frequency of these cells increased to 70%, approaching 100% in remote infections. In contrast, only 10-20% of HCMV-specific CD8+ T cells were long-term memory cells up to 12 months after infection onset, thereafter increasing to 70% in remote infections. Interestingly, a significantly higher frequency of HCMV-specific CD4+ T cells with a long-term IL-7Rpos memory phenotype was observed in non-transmitting compared to transmitting women. These findings indicate that immunodominance in HCMV infection is not predetermined in the naïve compartment, but is the result of virus-host interactions and suggest that prompt control of HCMV infection in pregnancy is associated with the rapid development of long-term IL-7Rpos memory HCMV-specific CD4+ T cells and a low risk of virus transmission to the fetus.

  1. Roadmap for human resources for expanded Indian nuclear industry

    Energy Technology Data Exchange (ETDEWEB)

    Singh, R.K. [Bhabha Atomic Research Centre, Mumbai (India); Indian Nuclear Society (India); Srinivasan, G.R.; Goyal, O.P. [Bhabha Atomic Research Centre, Mumbai (India)

    2011-07-01

    This paper deals with detailed requirement of human resources for all phases of nuclear power plant, for the manufacturing sector and the probable roadmap for achieving India's target. The accident in Fukushima has brought out that only nuclear power that avoids being a threat to the health and safety of the population and the environmental will be acceptable to the society and for this to be achieved human resources could be a single major contributor. India has ambitious plan of achieving 20,000MW by 2020 and 63,000MW by 2050. It is felt out of the three resources men, material and money; the critical shortage would be human resources both in quality and quantity. As per IAEA report (Publication of 2008 edition of energy, electricity and nuclear power estimates for the period of 2030), nuclear capacity must grow to at least 1.8 times current capacity by 2030 if global temperature rises are to be kept at 2°C. Objective of recruiting and training human resources for Indian Industry can be as follows: a) For catering domestic market. b) For catering international market later on for nuclear industries outside India. As India will be an important future international player. The above would require a multiplication of human resources by nearly seven times. In addition it has to be wholesome covering all levels and all skills and all disciplines and stages covering the whole nuclear cycle including regulators. Human resources are required for design and engineering, construction, commissioning, operation, manufacturing and for support services. The manpower for these has to be trained to achieve high quality of nuclear standards. Presently Indian Department of Atomic Energy(DAE) runs several training schools giving one year Post Graduate, tailor made courses. This needs to be multiplied by Joint efforts. Training should be on 'SAT (Systematic Approach to Training)' methodology to ensure focussed, specific, needed to culminate in safe, reliable and

  2. Biology and pathogenesis of cytomegalovirus in periodontal disease.

    Science.gov (United States)

    Contreras, Adolfo; Botero, Javier Enrique; Slots, Jørgen

    2014-02-01

    Human periodontitis is associated with a wide range of bacteria and viruses and with complex innate and adaptive immune responses. Porphyromonas gingivalis, Tannerella forsythia, Aggregatibacter actinomycetemcomitans, Treponema denticola, cytomegalovirus and other herpesviruses are major suspected pathogens of periodontitis, and a combined herpesvirus-bacterial periodontal infection can potentially explain major clinical features of the disease. Cytomegalovirus infects periodontal macrophages and T-cells and elicits a release of interleukin-1β and tumor necrosis factor-α. These proinflammatory cytokines play an important role in the host defense against the virus, but they also have the potential to induce alveolar bone resorption and loss of periodontal ligament. Gingival fibroblasts infected with cytomegalovirus also exhibit diminished collagen production and release of an increased level of matrix metalloproteinases. This article reviews innate and adaptive immunity to cytomegalovirus and suggests that immune responses towards cytomegalovirus can play roles in controlling, as well as in exacerbating, destructive periodontal disease. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. Human factors engineering plan for reviewing nuclear plant modernization programs

    Energy Technology Data Exchange (ETDEWEB)

    O' Hara, John; Higgins, James [Brookhaven National Laboratory, Upton, NY (United States)

    2004-12-01

    The Swedish Nuclear Power Inspectorate reviews the human factors engineering (HFE) aspects of nuclear power plants (NPPs) involved in the modernization of the plant systems and control rooms. The purpose of a HFE review is to help ensure personnel and public safety by verifying that accepted HFE practices and guidelines are incorporated into the program and nuclear power plant design. Such a review helps to ensure the HFE aspects of an NPP are developed, designed, and evaluated on the basis of a structured top-down system analysis using accepted HFE principles. The review addresses eleven HFE elements: HFE Program Management, Operating Experience Review, Functional Requirements Analysis and Allocation, Task Analysis, Staffing, Human Reliability Analysis, Human-System Interface Design, Procedure Development, Training Program Development, Human Factors Verification and Validation, and Design Implementation.

  4. Cytomegalovirus in Immunosuppressed Patients: A Silent and ...

    African Journals Online (AJOL)

    Cytomegalovirus (CMV) is a recognized cause of morbidity and mortality among immunocompromised individuals. This review will concentrate on understanding the pathogenesis, clinical manifestations and laboratory diagnostic options for CMV infection. Keywords: Review, Cytomegalovirus, Immunosuppressed ...

  5. Human factor engineering applied to nuclear power plant design

    Energy Technology Data Exchange (ETDEWEB)

    Manrique, A. [TECNATOM SA, BWR General Electric Business Manager, Madrid (Spain); Valdivia, J.C. [TECNATOM SA, Operation Engineering Project Manager, Madrid (Spain); Jimenez, A. [TECNATOM SA, Operation Engineering Div. Manager, Madrid (Spain)

    2001-07-01

    For the design and construction of new nuclear power plants as well as for maintenance and operation of the existing ones new man-machine interface designs and modifications are been produced. For these new designs Human Factor Engineering must be applied the same as for any other traditional engineering discipline. Advantages of implementing adequate Human Factor Engineering techniques in the design of nuclear reactors have become not only a fact recognized by the majority of engineers and operators but also an explicit requirement regulated and mandatory for the new designs of the so called advanced reactors. Additionally, the big saving achieved by a nuclear power plant having an operating methodology which significantly decreases the risk of operating errors makes it necessary and almost vital its implementation. The first step for this is preparing a plan to incorporate all the Human Factor Engineering principles and developing an integral design of the Instrumentation and Control and Man-machine interface systems. (author)

  6. Mood stabilizers inhibit cytomegalovirus infection.

    Science.gov (United States)

    Ornaghi, Sara; Davis, John N; Gorres, Kelly L; Miller, George; Paidas, Michael J; van den Pol, Anthony N

    2016-12-01

    Cytomegalovirus (CMV) infection can generate debilitating disease in immunocompromised individuals and neonates. It is also the most common infectious cause of congenital birth defects in infected fetuses. Available anti-CMV drugs are partially effective but are limited by some toxicity, potential viral resistance, and are not recommended for fetal exposure. Valproate, valpromide, and valnoctamide have been used for many years to treat epilepsy and mood disorders. We report for the first time that, in contrast to the virus-enhancing actions of valproate, structurally related valpromide and valnoctamide evoke a substantial and specific inhibition of mouse and human CMV in vitro. In vivo, both drugs safely attenuate mouse CMV, improving survival, body weight, and developmental maturation of infected newborns. The compounds appear to act by a novel mechanism that interferes with CMV attachment to the cell. Our work provides a novel potential direction for CMV therapeutics through repositioning of agents already approved for use in psychiatric disorders. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. The human granulocyte nucleus: Unusual nuclear envelope and heterochromatin composition.

    Science.gov (United States)

    Olins, Ada L; Zwerger, Monika; Herrmann, Harald; Zentgraf, Hanswalter; Simon, Amos J; Monestier, Marc; Olins, Donald E

    2008-05-01

    The human blood granulocyte (neutrophil) is adapted to find and destroy infectious agents. The nucleus of the human neutrophil has a segmented appearance, consisting of a linear or branched array of three or four lobes. Adequate levels of lamin B receptor (LBR) are necessary for differentiation of the lobulated nucleus. The levels of other components of the nuclear envelope may also be important for nuclear shape determination. In the present study, immunostaining and immunoblotting procedures explored the levels of various components of the nuclear envelope and heterochromatin, comparing freshly isolated human neutrophils with granulocytic forms of HL-60 cells, a tissue culture model system. In comparison to granulocytic HL-60 cells, blood neutrophil nuclear envelopes contain low-to-negligible amounts of LBR, lamins A/C, B1 and B2, LAP2beta and emerin. Surprisingly, a "mitotic" chromosome marker, H3(S10)phos, is elevated in neutrophil nuclei, compared to granulocytic HL-60 cells. Furthermore, neutrophil nuclei appear to be more fragile to methanol fixation, than observed with granulocytic HL-60 cells. Thus, the human neutrophil nucleus appears to be highly specialized, possessing a paucity of nuclear envelope-stabilizing proteins. In consequence, the neutrophil nucleus appears to be very malleable, supporting rapid migration through tight tissue spaces.

  8. Summary of the II International Symposium on Cytomegalovirus

    NARCIS (Netherlands)

    de Jong, M. D.; Galasso, G. J.; Gazzard, B.; Griffiths, P. D.; Jabs, D. A.; Kern, E. R.; Spector, S. A.

    1998-01-01

    Human cytomegalovirus (HCMV) is a highly species-specific DNA virus belonging to the Betaherpesvirinae subfamily of the herpesviridae family. Like other herpesviruses, primary infection with HCMV is followed by persistence of the virus in a latent form. The sites of latency are still largely

  9. Cytomegalovirus infections among low birth weight infants in a ...

    African Journals Online (AJOL)

    Human Cytomegalovirus (HCMV) is one of the leading causes of congenital infections, which can lead to severe foetal anomalies or even foetal loss. In order to determine the incidence of congenital HCMV infection in low birth weight neonates and the most prevalent genotype, cord blood samples were collected from a ...

  10. Technological Advances, Human Performance, and the Operation of Nuclear Facilities

    Science.gov (United States)

    Corrado, Jonathan K.

    Many unfortunate and unintended adverse industrial incidents occur across the United States each year, and the nuclear industry is no exception. Depending on their severity, these incidents can be problematic for people, the facilities, and surrounding environments. Human error is a contributing factor in many such incidents. This dissertation first explored the hypothesis that technological changes that affect how operators interact within the systems of the nuclear facilities exacerbate the cost of incidents caused by human error. I conducted a review of nuclear incidents in the United States from 1955 through 2010 that reached Level 3 (serious incident) or higher on the International Nuclear Events Scale (INES). The cost of each incident at facilities that had recently undergone technological changes affecting plant operators' jobs was compared to the cost of events at facilities that had not undergone changes. A t-test determined a statistically significant difference between the two groups, confirming the hypothesis. Next, I conducted a follow-on study to determine the impact of the incorporation of new technologies into nuclear facilities. The data indicated that spending more money on upgrades increased the facility's capacity as well as the number of incidents reported, but the incident severity was minor. Finally, I discuss the impact of human error on plant operations and the impact of evolving technology on the 21st-century operator, proposing a methodology to overcome these challenges by applying the systems engineering process.

  11. Coinfection of Epstein-Barr virus, cytomegalovirus, herpes simplex virus, human papillomavirus and anal intraepithelial neoplasia in HIV patients in Amazon, Brazil

    Directory of Open Access Journals (Sweden)

    Adriana Gonçalves Daumas Pinheiro Guimarães

    2012-03-01

    Full Text Available OBJECTIVE: The prevention of anal cancer is a goal of worldwide Aids support centers. Despite the efforts that have been made and progress in the antiretroviral therapy, effective disease control remains elusive. Difficulty in preventing anal cancer may result from the ineffectiveness of highly active antiretroviral therapy on the human papillomavirus (HPV since the coinfection with HIV and HPV appears to increase the risk of HPV-infected cells, becoming cancerous. METHODS: We evaluated 69 HIV-positive and 30 HIV-negative male patients who underwent cytological evaluation by RT-PCR for the presence of HPV, Epstein-Barr virus, cytomegalovirus and herpes virus types (HSV 1 and 2, and histopathology analysis of the anal canal. RESULTS: The prevalence of anal intraepithelial neoplasia was 35% and it was restricted to HIV-positive patients. Patients infected with high-risk HPV and with fewer than 50 TCD4 cells/µL showed an anal intraepithelial neoplasia rate of 85.7% compared to those with TCD4 cells >200 cells/µL (pOBJETIVO: A prevenção do câncer anal tem sido aplicada pelos centros de apoio a pacientes com Aids em todo o mundo. Apesar dos esforços empregados, o eficaz controle da doença permanece distante. A dificuldade na prevenção do câncer anal pode resultar, em parte, da ineficácia da ação da terapia antirretroviral sobre o papilomavírus humano (HPV, pois a coinfecção com HIV e HPV parece aumentar o risco das células infectadas pelo HPV em tornarem-se cancerosas. MÉTODOS: Foram avaliados 69 HIV-positivos e 30 pacientes HIV-negativos do sexo masculino, que foram submetidos à avaliação citológica anal por real time-PCR para a presença de HPV, vírus Epstein-Barr, citomegalovírus e herpes vírus tipos (HSV 1 e 2 além da análise histopatológica de fragmento de mucosa do canal anal. RESULTADOS: A prevalência de neoplasia intraepitelial anal foi de 35% e foi restrita a pacientes HIV-positivos. Os pacientes infectados com o

  12. Phenotypic characterization of two naturally occurring human Cytomegalovirus sequence polymorphisms located in a distinct region of ORF UL56 known to be involved in in vitro resistance to letermovir.

    Science.gov (United States)

    Goldner, Thomas; Zimmermann, Holger; Lischka, Peter

    2015-04-01

    Letermovir is a new drug in Phase 3 clinical development for the prevention of human Cytomegalovirus (HCMV) infections in hematopoietic-stem-cell transplant recipients (HSCT). In contrast to marketed anti-HCMV drugs which all target the viral DNA polymerase, letermovir's novel mode of action targets the UL56 subunit of the viral terminase complex. Consistently letermovir resistance has mapped in vitro to a distinct region within ORF UL56 (amino acid 230-370). Here we used marker transfer to demonstrate that two naturally occurring UL56 sequence variants within this region, located directly adjacent to sites known to mediate letermovir resistance in vitro (D242G and A327V) represent normal interstrain polymorphisms unrelated to drug-resistance. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. Human and organizational factors in nuclear safety; Factores humanos y organizativos en la seguridad nuclear

    Energy Technology Data Exchange (ETDEWEB)

    Garcia, A.; Barrientos, M.; Gil, B.

    2015-07-01

    Nuclear installations are socio technical systems where human and organizational factors, in both utilities and regulators, have a significant impact on safety. Three Mile Island (TMI) accident, original of several initiatives in the human factors field, nevertheless became a lost opportunity to timely acquire lessons related to the upper tiers of the system. Nowadays, Spanish nuclear installations have integrated in their processes specialists and activities in human and organizational factors, promoted by the licensees After many years of hard work, Spanish installations have achieved a better position to face new challenges, such as those posed by Fukushima. With this experience, only technology-centered action plan would not be acceptable, turning this accident in yet another lost opportunity. (Author)

  14. Quality management in the nuclear industry: the human factor

    Energy Technology Data Exchange (ETDEWEB)

    1990-01-01

    In the nuclear industry it is vital to understand the 'human factor' with regard to plant performance and plant safety. A proper management system ensures that personnel perform their duties correctly. 'Quality Management in the Nuclear Industry: the Human Factor', was a conference organized by the Institution of Mechanical Engineers in October 1990. The conference covered a wide range of topics on an international level including: standards, licensing and regulatory procedures; selection assessment and training of personnel; feedback from experience of good practice and of deviations; management and support of personnel performance; modelling and evaluation of human factors. The papers presented at the conference are contained in this volume. All twenty papers are indexed separately. (author).

  15. Human Factors Considerations in New Nuclear Power Plants: Detailed Analysis.

    Energy Technology Data Exchange (ETDEWEB)

    OHara,J.; Higgins, J.; Brown, W.; Fink, R.

    2008-02-14

    This Nuclear Regulatory Commission (NRC) sponsored study has identified human-performance issues in new and advanced nuclear power plants. To identify the issues, current industry developments and trends were evaluated in the areas of reactor technology, instrumentation and control technology, human-system integration technology, and human factors engineering (HFE) methods and tools. The issues were organized into seven high-level HFE topic areas: Role of Personnel and Automation, Staffing and Training, Normal Operations Management, Disturbance and Emergency Management, Maintenance and Change Management, Plant Design and Construction, and HFE Methods and Tools. The issues where then prioritized into four categories using a 'Phenomena Identification and Ranking Table' methodology based on evaluations provided by 14 independent subject matter experts. The subject matter experts were knowledgeable in a variety of disciplines. Vendors, utilities, research organizations and regulators all participated. Twenty issues were categorized into the top priority category. This Brookhaven National Laboratory (BNL) technical report provides the detailed methodology, issue analysis, and results. A summary of the results of this study can be found in NUREG/CR-6947. The research performed for this project has identified a large number of human-performance issues for new control stations and new nuclear power plant designs. The information gathered in this project can serve as input to the development of a long-term strategy and plan for addressing human performance in these areas through regulatory research. Addressing human-performance issues will provide the technical basis from which regulatory review guidance can be developed to meet these challenges. The availability of this review guidance will help set clear expectations for how the NRC staff will evaluate new designs, reduce regulatory uncertainty, and provide a well-defined path to new nuclear power plant

  16. Human nucleoporins promote HIV-1 docking at the nuclear pore, nuclear import and integration.

    Directory of Open Access Journals (Sweden)

    Francesca Di Nunzio

    Full Text Available The nuclear pore complex (NPC mediates nucleo-cytoplasmic transport of macromolecules and is an obligatory point of passage and functional bottleneck in the replication of some viruses. The Human Immunodeficiency Virus (HIV has evolved the required mechanisms for active nuclear import of its genome through the NPC. However the mechanisms by which the NPC allows or even assists HIV translocation are still unknown. We investigated the involvement of four key nucleoporins in HIV-1 docking, translocation, and integration: Nup358/RanBP2, Nup214/CAN, Nup98 and Nup153. Although all induce defects in infectivity when depleted, only Nup153 actually showed any evidence of participating in HIV-1 translocation through the nuclear pore. We show that Nup358/RanBP2 mediates docking of HIV-1 cores on NPC cytoplasmic filaments by interacting with the cores and that the C-terminus of Nup358/RanBP2 comprising a cyclophilin-homology domain contributes to binding. We also show that Nup214/CAN and Nup98 play no role in HIV-1 nuclear import per se: Nup214/CAN plays an indirect role in infectivity read-outs through its effect on mRNA export, while the reduction of expression of Nup98 shows a slight reduction in proviral integration. Our work shows the involvement of nucleoporins in diverse and functionally separable steps of HIV infection and nuclear import.

  17. Cytomegalovirus glycoprotein B genotyping in ocular fluids and blood of AIDS patients with cytomegalovirus retinitis

    NARCIS (Netherlands)

    Peek, R.; Verbraak, F.; Bruinenberg, M.; van der Lelij, A.; van den Horn, G.; Kijlstra, A.

    1998-01-01

    To determine the frequency of cytomegalovirus glycoprotein B (gB) genotypes in clinical samples of ocular fluids of patients with acquired immune deficiency syndrome (AIDS) who have cytomegalovirus retinitis and to compare these with the cytomegalovirus gB genotype in paired peripheral blood

  18. Nuclear Accidents: Consequences for Human, Society and Energy Sector

    Directory of Open Access Journals (Sweden)

    L. A. Bolshov

    2016-01-01

    Full Text Available The article examines radiation and hygienic regulations with regard to the elimination of consequences of the Chernobyl NPP accident in the context of relationships with other aspects, primarily socio-economic and political factors. This experience is reasonable to take into account when defining criteria in other regulatory fields, for example, in radioactive waste classification and remediation of areas. The article presents an analysis of joint features and peculiarities of nuclear accidents in the industry and energy sectors. It is noted that the scale of global consequences of the Chernobyl NPP accident is defined by the large-scale release of radioactivity into the environment, as well as an affiliation of the nuclear installation with the energy sector. Large-scale radiation accidents affect the most diverse spheres of human activities, what, in its turn, evokes the reverse reaction from the society and its institutions, including involvement of political means of settlement. If the latter is seeing for criteria that are scientifically justified and feasible, then the preconditions for minimizing socio-economic impacts are created. In other cases, political decisions, such as nuclear units’ shutdown and phasing out of nuclear energy, appear to be an economic price which society, as a whole and a single industry sector, pay to compensate the negative public response. The article describes fundamental changes in approaches to ensure nuclear and radiation safety that occurred after the Chernobyl NPP accident. Multiple and negative consequences of the Chernobyl accident for human and society are balanced to some extent by a higher level of operational safety, emergency preparedness, and life-cycle safety. The article indicates that harmonization and ensuring consistency of regulations that involve different aspects of nuclear and radiation safety are important to implement practical solutions to the nuclear legacy problems. The

  19. History of the molecular biology of cytomegaloviruses.

    Science.gov (United States)

    Stinski, Mark F

    2014-01-01

    The history of the molecular biology of cytomegaloviruses from the purification of the virus and the viral DNA to the cloning and expression of the viral genes is reviewed. A key genetic element of cytomegalovirus (the CMV promoter) contributed to our understanding of eukaryotic cell molecular biology and to the development of lifesaving therapeutic proteins. The study of the molecular biology of cytomegaloviruses also contributed to the development of antivirals to control the viral infection.

  20. Pluripotent stem cells are protected from cytomegalovirus infection at multiple points: implications of a new pathogenesis for congenital anomaly caused by cytomegalovirus.

    Science.gov (United States)

    Kawasaki, Hideya

    2012-09-01

    In humans, the cytomegalovirus (CMV) is the most significant cause of intrauterine infections that cause congenital anomalies. Intrauterine infection with human CMV is thought to be responsible for a variety of abnormalities, including mental retardation, microcephaly, developmental delay, seizure disorders, and cerebral palsy, depending on the timing of the fetal infection, the infectious route, and the virulence of the virus. In addition to the adaptive immune system, the embryo has potential resistance to CMV during early embryogenesis. Embryonic stem (ES) cells are more resistant to CMV than most other cell types, although the mechanism responsible for this resistance is not well understood. ES cells allow approximately 20-fold less murine CMV (MCMV) DNA to enter the nucleus than mouse embryonic fibroblasts (MEFs), and this inhibition occurs in a multistep manner. In situ hybridization showed that ES cell nuclei had significantly less MCMV DNA than MEF nuclei. This finding appears to be supported by the fact that ES cells express less heparan sulfate, β1-integrin, and vimentin and have fewer nuclear pores than differentiated cells such as MEF. This may reduce the ability of MCMV to attach to and enter the cellular membrane, translocate to the nucleus, and cross the nuclear membrane in pluripotent stem cells (ES-induced pluripotent stem cells). This finding may indicate a new pathogenesis for the congenital anomaly caused by CMV. © 2012 The Author. Congenital Anomalies © 2012 Japanese Teratology Society.

  1. Microgravity Analogues of Herpes Virus Pathogenicity: Human Cytomegalovirus (hCMV) and Varicella Zoster (VZV) Infectivity in Human Tissue Like Assemblies (TLAs)

    Science.gov (United States)

    Goodwin, T. J.; McCarthy, M.; Albrecht, T.; Cohrs, R.

    2009-01-01

    The old adage we are our own worst enemies may perhaps be the most profound statement ever made when applied to man s desire for extraterrestrial exploration and habitation of Space. Consider the immune system protects the integrity of the entire human physiology and is comprised of two basic elements the adaptive or circulating and the innate immune system. Failure of the components of the adaptive system leads to venerability of the innate system from opportunistic microbes; viral, bacteria, and fungal, which surround us, are transported on our skin, and commonly inhabit the human physiology as normal and imunosuppressed parasites. The fine balance which is maintained for the preponderance of our normal lives, save immune disorders and disease, is deregulated in microgravity. Thus analogue systems to study these potential Risks are essential for our progress in conquering Space exploration and habitation. In this study we employed two known physiological target tissues in which the reactivation of hCMV and VZV occurs, human neural and lung systems created for the study and interaction of these herpes viruses independently and simultaneously on the innate immune system. Normal human neural and lung tissue analogues called tissue like assemblies (TLAs) were infected with low MOIs of approximately 2 x 10(exp -5) pfu hCMV or VZV and established active but prolonged low grade infections which spanned .7-1.5 months in length. These infections were characterized by the ability to continuously produce each of the viruses without expiration of the host cultures. Verification and quantification of viral replication was confirmed via RT_PCR, IHC, and confocal spectral analyses of the respective essential viral genomes. All host TLAs maintained the ability to actively proliferate throughout the entire duration of the experiments as is analogous to normal in vivo physiological conditions. These data represent a significant advance in the ability to study the triggering

  2. Nuclear and cytoplasmic LIMK1 enhances human breast cancer progression

    Directory of Open Access Journals (Sweden)

    Gutierrez-Hartmann Arthur

    2011-06-01

    Full Text Available Abstract Background LIM kinase 1 (LIMK1 is expressed in both cytoplasmic and nuclear compartments, and is a key regulator of cytoskeletal organization involved in cell migration and proliferation. LIMK1 levels are increased in several human cancers, with LIMK1 over-expression in prostate and breast cancer cells leading to tumor progression. While it has been presumed that the mechanism by which LIMK1 promotes cancer progression is via its cytoplasmic effects, the role of nuclear vs cytoplasmic LIMK1 in the tumorigenic process has not been examined. Results To determine if cytoplasmic or nuclear LIMK1 expression correlated with breast cancer, we performed immunohistochemical (IHC analysis of breast tissue microarrays (TMAs, The IHC analysis of breast TMAs revealed that 76% of malignant breast tissue samples strongly expressed LIMK1 in the cytoplasm, with 52% of these specimens also expressing nuclear LIMK1. Only 48% of benign breast samples displayed strong cytoplasmic LIMK1 expression and 27% of these expressed nuclear LIMK1. To investigate the respective roles of cytoplamsic and nuclear LIMK1 in breast cancer progression, we targeted GFP-LIMK1 to cytoplasmic and nuclear subcellular compartments by fusing nuclear export signals (NESs or nuclear localization sequences (NLS, respectively, to the amino-terminus of GFP-LIMK1. Stable pools of MDA-MB-231 cells were generated by retroviral transduction, and fluorescence microscopy revealed that GFP alone (control and GFP-LIMK1 were each expressed in both the cytoplasm and nucleus of MDA-MB-231 cells, whereas NLS-GFP-LIMK1 was expressed in the nucleus and NES-GFP-LIMK1 was expressed in the cytoplasm. Western blot analyses revealed equal expression of GFP-LIMK1 and NES-GFP-LIMK1, with NLS-GFP-LIMK1 expression being less but equal to endogenous LIMK1. Also, Western blotting revealed increased levels of phospho-cofilin, phospho-FAK, phospho-paxillin, phospho-Src, phospho-AKT, and phospho-Erk1/2 in cells

  3. Antiviral treatment of cytomegalovirus infection: an update.

    Science.gov (United States)

    Härter, Georg; Michel, Detlef

    2012-04-01

    This editorial summarizes recent developments in the management of ganciclovir-resistant human cytomegalovirus (HCMV) infections. All current drugs available for systemic treatment, including ganciclovir (GCV), valganciclovir, foscarnet and cidofovir, target the viral polymerase. However, all such compounds are hampered by dose-related toxicities and the emergence of resistance. Different approaches (e.g., PCR-based direct sequencing, pyrosequencing, mass spectrometry-based comparative sequencing) allow the fast detection of resistant HCMV and are well suited to therapy monitoring. However, more studies are required on the dynamic of mixed HCMV populations under drug pressure. Alternate antiviral compounds with new mechanisms of action, such as artesunate, leflunomid, letermovir and maribavir, are now being investigated in clinical studies. An advantage of some of the new substances is lesser toxicity issues, which might lead to new prophylactic and treatment strategies.

  4. Cytomegalovirus Treatment Strategy After a Liver Transplant: Preemptive Therapy or Prophylaxis for Cytomegalovirus Seropositive Donor and Recipient.

    Science.gov (United States)

    Lindner, Kirsten; Anthoni, Christoph; Beckebaum, Susanne; Senninger, Norbert; Hölzen, Jens Peter; Wolters, Heiner

    2016-08-01

    Cytomegalovirus infections cause the most frequent infection after solid-organ transplant. While Cytomegalovirus prophylaxis is established in high-risk patients (donor+/ recipient-), data on Cytomegalovirus prophylaxis in other serostatus constellation are rare. The aim of this study was to evaluate the influence of Cytomegalovirus treatment strategy after a liver transplant (preemptive therapy vs general prophylaxis) in the largest group of patients: Cytomegalovirus seropositive donor and recipient. Forty-seven seropositive recipients of seropositive donor liver transplants (D+/R+, 2005-2012) were included in this retrospective study. Twenty-one patients received oral valganciclovir as Cytomegalovirus prophylaxis 100 days after transplant. Cytomegalovirus infection and Cytomegalovirus disease were monitored during the first 6 months. A Cytomegalovirus infection could be detected in 4 out of 47 patients (8.5%), including Cytomegalovirus disease in 2 patients (Cytomegalovirus pneumonia and Cytomegalovirus-CNS disease). Three of these patients received no Cytomegalovirus prophylaxis (P = .408). Eight patients developed a graft failure; this occurred more frequently among patients without Cytomegalovirus prophylaxis (P = .044). Patients receiving Cytomegalovirus prophylaxis more often developed leukopenia. No difference was seen regarding the number of platelets, hemoglobin, and creatinine. Cytomegalovirus prophylaxis can minimize the risk of Cytomegalovirus reactivation and graft failure. However, disadvantages of the prophylaxis as leukopenia should be considered.

  5. 21 CFR 866.3175 - Cytomegalovirus serological reagents.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Cytomegalovirus serological reagents. 866.3175... Cytomegalovirus serological reagents. (a) Identification. Cytomegalovirus serological reagents are devices that consist of antigens and antisera used in serological tests to identify antibodies to cytomegalovirus in...

  6. Prevalence of Cytomegalovirus in Iraqi Children

    Directory of Open Access Journals (Sweden)

    Sevan Najem Alwan

    2017-11-01

    Full Text Available The majority of children with congenital cytomegalovirus are born to cytomegalovirus seropositive women. However, the likelihood of congenital infection with disability is highest for children whose mothers were cytomegalovirus seronegative acquired infection during pregnancy. Objectives: to provide first nationally estimate of cytomegalovirus seropositivity among Iraqi children under five years of age. Materials and Methods was used total of 1000 hospitalized children under five years of age form different geographical area in Iraq were enrolled in this study. The numbers of children were collected by proportional allocation for each selected governorate according to total number of participant children. Kuppuswamy scale has been used to measure the socioeconomic status for children. Serum samples were obtained from each subject participate in this study, cytomegalovirus infection was defined as IgM antibody positive by electro-chemiluminescence Immunoassay techniques. The results show that the current study revealed a prevalence of cytomegalovirus specific IgM as a serum marker was 5.4% among children under five years of age. Positive cytomegalovirus was associated with low socioeconomic status, maternal bad obstetric history, and lower age of children, while the infection was not associated to geographical distribution and gender. By stratified the children into symptomatic and asymptomatic according to the signs and symptoms of cytomegalovirus congenital and acquired infection at time of sample collection, 15% and 0.9% proved to have positive specific IgM among symptomatic and asymptomatic children, respectively. Jaundice was the most predominant clinical sign 56% among symptomatic infected children, followed by hepatosplenomegaly 41.7%. Conclusion: The data provide in the current study strongly encourage routine testing for cytomegalovirus Antibodies among pregnant women in Iraq. Infants diagnosed to be sub-clinically infected with

  7. Cytomegalovirus reactivation in ICU patients.

    Science.gov (United States)

    Papazian, Laurent; Hraiech, Sami; Lehingue, Samuel; Roch, Antoine; Chiche, Laurent; Wiramus, Sandrine; Forel, Jean-Marie

    2016-01-01

    Approximately 20 years have passed since we reported our results of histologically proven cytomegalovirus (CMV) pneumonia in non-immunocompromised ICU patients. Even if there are more recent reports suggesting that CMV may worsen the outcomes for ICU patients, there is no definite answer to this question: is CMV a potential pathogen for ICU patients or is it simply a bystander? We will describe the pathophysiology of active CMV infection and the most recent insights concerning the epidemiological aspects of these reactivations. Cytomegalovirus can be pathogenic by a direct organ insult (such as for the lung), by decreasing host defences against other microorganisms and/or by enhancing the body's inflammatory response (as in acute respiratory distress syndrome). The incidence of active CMV infection is dependent on the diagnostic method used. Using the most sophisticated available biological tools, the incidence can reach 15-20% of ICU patients (20-40% in ICU patients with positive CMV serology). In adequately powered cohorts of patients, active CMV infection appears to be associated with worse outcomes for mechanically ventilated ICU patients. There is no absolute direct proof of a negative impact of active CMV infection on the health outcomes of mechanically ventilated patients. Prospective randomized trials are lacking. Future trials should examine the potential benefits for health outcomes of using antiviral treatments. Such treatments could be prophylactic, pre-emptive or used only when there is an end-organ disease. Cytomegalovirus infection may affect health outcomes for ICU patients. Additional prospective trials are necessary to confirm this hypothesis.

  8. Human based roots of failures in nuclear events investigations

    Energy Technology Data Exchange (ETDEWEB)

    Ziedelis, Stanislovas; Noel, Marc; Strucic, Miodrag [Commission of the European Communities, Petten (Netherlands). European Clearinghouse on Operational Experience Feedback for Nuclear Power Plants

    2012-10-15

    This paper aims for improvement of quality of the event investigations in the nuclear industry through analysis of the existing practices, identifying and removing the existing Human and Organizational Factors (HOF) and management related barriers. It presents the essential results of several studies performed by the European Clearinghouse on Operational Experience. Outcomes of studies are based on survey of currently existing event investigation practices typical for nuclear industry of 12 European countries, as well as on insights from analysis of numerous event investigation reports. System of operational experience feedback from information based on event investigation results is not enough effective to prevent and even to decrease frequency of recurring events due to existing methodological, HOF-related and/or knowledge management related constraints. Besides that, several latent root causes of unsuccessful event investigation are related to weaknesses in safety culture of personnel and managers. These weaknesses include focus on costs or schedule, political manipulation, arrogance, ignorance, entitlement and/or autocracy. Upgrades in safety culture of organization's personnel and its senior management especially seem to be an effective way to improvement. Increasing of competencies, capabilities and level of independency of event investigation teams, elaboration of comprehensive software, ensuring of positive approach, adequate support and impartiality of management could also facilitate for improvement of quality of the event investigations. (orig.)

  9. Human and environmental nuclear toxicology; Toxicologie nucleaire environnementale et humaine

    Energy Technology Data Exchange (ETDEWEB)

    Menager, M.Th.; Garnier-Laplace, J.; Goyffon, M

    2009-07-01

    When the civil nuclear is in expansion, this book takes stock of the situation on the geochemical processes implied the accumulation of radioactive or stable chemical elements and on the toxicity and all that is ensuing. It is important to answer at the best to increasing demands of toxicology, ecotoxicology for evaluation and management of environmental and sanitary risks. This work is divided in five parts: the behaviour of chemical species in biosphere, particularly for the living, the cell molecular mechanisms of interactions among the living, their consequences on human health and the ecosystems, the state of knowledge, element by element and the new development of technology opened with the knowledge progress. The fields of application, goes from the detection to the treatments of contaminations, from environment to man. (N.C.)

  10. Peripheral Blood Leukocytes and Serum Nested Polymerase Chain Reaction Are Complementary Methods for Monitoring Active Cytomegalovirus Infection in Transplant Patients

    Directory of Open Access Journals (Sweden)

    PD Andrade

    2013-01-01

    Full Text Available BACKGROUND: Human cytomegalovirus is an important cause of morbidity and mortality in immunocompromised patients. Qualitative polymerase chain reaction (PCR has proven to be a sensitive and effective technique in defining active cytomegalovirus infection, in addition to having low cost and being a useful test for situations in which there is no need for quantification. Real-time PCR has the advantage of quantification; however, the high cost of this methodology makes it impractical for routine use.

  11. Analysis and isolation of cytomegalovirus DNA by field inversion gel electrophoresis

    NARCIS (Netherlands)

    van den Berg, F. M.; Jiwa, M.; Rook, R.; Geelen, J. L.

    1988-01-01

    High molecular weight human cytomegalovirus (CMV) DNA was isolated from agarose-embedded infected human diploid cells by employing field inversion gel electrophoresis. A high yield of CMV DNA molecules was obtained within 1 week of infecting the cell culture. Labelling of the viral DNA with biotin

  12. Identification of human appendicitis by in vitro nuclear magnetic resonance.

    Science.gov (United States)

    Jacobs, D O; Settle, R G; Clarke, J R; Trerotola, S O; Sachdeva, A K; Wolf, G L; Rombeau, J L

    1990-02-01

    The spin-lattice relaxation time, T1 as determined by nuclear magnetic resonance techniques, correlates positively with tissue water content. The latter relationship has been observed in rabbits with experimentally induced appendicitis whose inflamed appendiceal tissues had significantly higher T1's and water contents than tissue from normal controls. The present experiment studied these relationships in humans. Tissue water content and T1 were measured on appendiceal tissue from 10 patients with documented appendicitis and from 6 controls without the disease. All T1's were determined within 30-60 min of removal of the appendix at operation. The mean in vitro T1 of appendiceal tissue from patients with appendicitis was significantly higher than that of controls (527 +/- 15 msec versus 430 +/- 17 msec, mean +/- SEM, P less than 0.002). In addition, a strong positive correlation was noted between T1 and tissue water content (r = 0.70, P less than 0.01). Based on these findings, the use of in vivo magnetic resonance imaging techniques to detect human appendicitis noninvasively warrants investigation.

  13. seroprevalence of cytomegalovirus infection amongst pregnant ...

    African Journals Online (AJOL)

    boaz

    were collected from 330 pregnant women attending antenatal clinics and 33 non pregnant women attending the Outpatient. Department in three ... Keywords: Seroprevalence, Cytomegalovirus, IgG, Pregnant women, Kaduna, Nigeria. SEROPREVALENCE ... thrombocytopenia, gastrointestinal ulceration, hepatitis and ...

  14. Latency-Associated Viral Interleukin-10 (IL-10) Encoded by Human Cytomegalovirus Modulates Cellular IL-10 and CCL8 Secretion during Latent Infection through Changes in the Cellular MicroRNA hsa-miR-92a

    Science.gov (United States)

    Poole, Emma; Avdic, Selmir; Hodkinson, Jemima; Jackson, Sarah; Wills, Mark; Slobedman, Barry

    2014-01-01

    ABSTRACT The UL111A gene of human cytomegalovirus encodes a viral homologue of the cellular immunomodulatory cytokine interleukin 10 (cIL-10), which, due to alternative splicing, results in expression of two isoforms designated LAcmvIL-10 (expressed during both lytic and latent infection) and cmvIL-10 (identified only during lytic infection). We have analyzed the functions of LAcmvIL-10 during latent infection of primary myeloid progenitor cells and found that LAcmvIL-10 is responsible, at least in part, for the known increase in secretion of cellular IL-10 and CCL8 in the secretomes of latently infected cells. This latency-associated increase in CCL8 expression results from a concomitant LAcmvIL-10-mediated suppression of the expression of the cellular microRNA (miRNA) hsa-miR-92a, which targets CCL8 directly. Taking the data together, we show that the previously observed downregulation of hsa-miR-92a and upregulation of CCL8 during HCMV latent infection of myeloid cells are intimately linked via the latency-associated expression of LAcmvIL-10. IMPORTANCE HCMV latency causes significant morbidity and mortality in immunocompromised individuals, yet HCMV is carried silently (latently) in 50 to 90% of the population. Understanding how HCMV maintains infection for the lifetime of an infected individual is critical for the treatment of immunocompromised individuals suffering with disease as a result of HCMV. In this study, we analyze one of the proteins that are expressed during the “latent” phase of HCMV, LAcmvIL-10, and find that the expression of the gene modulates the microenvironment of the infected cell, leading to evasion of the immune system. PMID:25253336

  15. The downregulation of GFI1 by the EZH2-NDY1/KDM2B-JARID2 axis and by human cytomegalovirus (HCMV associated factors allows the activation of the HCMV major IE promoter and the transition to productive infection.

    Directory of Open Access Journals (Sweden)

    George Sourvinos

    2014-05-01

    Full Text Available Earlier studies had suggested that epigenetic mechanisms play an important role in the control of human cytomegalovirus (HCMV infection. Here we show that productive HCMV infection is indeed under the control of histone H3K27 trimethylation. The histone H3K27 methyltransferase EZH2, and its regulators JARID2 and NDY1/KDM2B repress GFI1, a transcriptional repressor of the major immediate-early promoter (MIEP of HCMV. Knocking down EZH2, NDY1/KDM2B or JARID2 relieves the repression and results in the upregulation of GFI1. During infection, the incoming HCMV rapidly downregulates the GFI1 mRNA and protein in both wild-type cells and in cells in which EZH2, NDY1/KDM2B or JARID2 were knocked down. However, since the pre-infection levels of GFI1 in the latter cells are significantly higher, the virus fails to downregulate it to levels permissive for MIEP activation and viral infection. Following the EZH2-NDY1/KDM2B-JARID2-independent downregulation of GFI1 in the early stages of infection, the virus also initiates an EZH2-NDY1/ΚDM2Β-JARID2-dependent program that represses GFI1 throughout the infection cycle. The EZH2 knockdown also delays histone H3K27 trimethylation in the immediate early region of HCMV, which is accompanied by a drop in H3K4 trimethylation that may contribute to the shEZH2-mediated repression of the major immediate early HCMV promoter. These data show that HCMV uses multiple mechanisms to allow the activation of the HCMV MIEP and to prevent cellular mechanisms from blocking the HCMV replication program.

  16. In vitro drug combination studies of Letermovir (AIC246, MK-8228) with approved anti-human cytomegalovirus (HCMV) and anti-HIV compounds in inhibition of HCMV and HIV replication.

    Science.gov (United States)

    Wildum, Steffen; Zimmermann, Holger; Lischka, Peter

    2015-01-01

    Despite modern prevention and treatment strategies, human cytomegalovirus (HCMV) remains a common opportunistic pathogen associated with serious morbidity and mortality in immunocompromised individuals, such as transplant recipients and AIDS patients. All drugs currently licensed for the treatment of HCMV infection target the viral DNA polymerase and are associated with severe toxicity issues and the emergence of drug resistance. Letermovir (AIC246, MK-8228) is a new anti-HCMV agent in clinical development that acts via a novel mode of action and has demonstrated anti-HCMV activity in vitro and in vivo. For the future, drug combination therapies, including letermovir, might be indicated under special medical conditions, such as the emergence of multidrug-resistant virus strains in transplant recipients or in HCMV-HIV-coinfected patients. Accordingly, knowledge of the compatibility of letermovir with other HCMV or HIV antivirals is of medical importance. Here, we evaluated the inhibition of HCMV replication by letermovir in combination with all currently approved HCMV antivirals using cell culture checkerboard assays. In addition, the effects of letermovir on the antiviral activities of selected HIV drugs, and vice versa, were analyzed. Using two different mathematical techniques to analyze the experimental data, (i) additive effects were observed for the combination of letermovir with anti-HCMV drugs and (ii) no interaction was found between letermovir and anti-HIV drugs. Since none of the tested drug combinations significantly antagonized letermovir efficacy (or vice versa), our findings suggest that letermovir may offer the potential for combination therapy with the tested HCMV and HIV drugs. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  17. Latency-associated viral interleukin-10 (IL-10) encoded by human cytomegalovirus modulates cellular IL-10 and CCL8 Secretion during latent infection through changes in the cellular microRNA hsa-miR-92a.

    Science.gov (United States)

    Poole, Emma; Avdic, Selmir; Hodkinson, Jemima; Jackson, Sarah; Wills, Mark; Slobedman, Barry; Sinclair, John

    2014-12-01

    The UL111A gene of human cytomegalovirus encodes a viral homologue of the cellular immunomodulatory cytokine interleukin 10 (cIL-10), which, due to alternative splicing, results in expression of two isoforms designated LAcmvIL-10 (expressed during both lytic and latent infection) and cmvIL-10 (identified only during lytic infection). We have analyzed the functions of LAcmvIL-10 during latent infection of primary myeloid progenitor cells and found that LAcmvIL-10 is responsible, at least in part, for the known increase in secretion of cellular IL-10 and CCL8 in the secretomes of latently infected cells. This latency-associated increase in CCL8 expression results from a concomitant LAcmvIL-10-mediated suppression of the expression of the cellular microRNA (miRNA) hsa-miR-92a, which targets CCL8 directly. Taking the data together, we show that the previously observed downregulation of hsa-miR-92a and upregulation of CCL8 during HCMV latent infection of myeloid cells are intimately linked via the latency-associated expression of LAcmvIL-10. HCMV latency causes significant morbidity and mortality in immunocompromised individuals, yet HCMV is carried silently (latently) in 50 to 90% of the population. Understanding how HCMV maintains infection for the lifetime of an infected individual is critical for the treatment of immunocompromised individuals suffering with disease as a result of HCMV. In this study, we analyze one of the proteins that are expressed during the "latent" phase of HCMV, LAcmvIL-10, and find that the expression of the gene modulates the microenvironment of the infected cell, leading to evasion of the immune system. Copyright © 2014 Poole et al.

  18. Off-the-Shelf Virus-Specific T Cells to Treat BK Virus, Human Herpesvirus 6, Cytomegalovirus, Epstein-Barr Virus, and Adenovirus Infections After Allogeneic Hematopoietic Stem-Cell Transplantation.

    Science.gov (United States)

    Tzannou, Ifigeneia; Papadopoulou, Anastasia; Naik, Swati; Leung, Kathryn; Martinez, Caridad A; Ramos, Carlos A; Carrum, George; Sasa, Ghadir; Lulla, Premal; Watanabe, Ayumi; Kuvalekar, Manik; Gee, Adrian P; Wu, Meng-Fen; Liu, Hao; Grilley, Bambi J; Krance, Robert A; Gottschalk, Stephen; Brenner, Malcolm K; Rooney, Cliona M; Heslop, Helen E; Leen, Ann M; Omer, Bilal

    2017-11-01

    Purpose Improvement of cure rates for patients treated with allogeneic hematopoietic stem-cell transplantation (HSCT) will require efforts to decrease treatment-related mortality from severe viral infections. Adoptively transferred virus-specific T cells (VSTs) generated from eligible, third-party donors could provide broad antiviral protection to recipients of HSCT as an immediately available off-the-shelf product. Patient and Methods We generated a bank of VSTs that recognized five common viral pathogens: Epstein-Barr virus (EBV), adenovirus (AdV), cytomegalovirus (CMV), BK virus (BKV), and human herpesvirus 6 (HHV-6). The VSTs were administered to 38 patients with 45 infections in a phase II clinical trial. Results A single infusion produced a cumulative complete or partial response rate of 92% (95% CI, 78.1% to 98.3%) overall and the following rates by virus: 100% for BKV (n = 16), 94% for CMV (n = 17), 71% for AdV (n = 7), 100% for EBV (n = 2), and 67% for HHV-6 (n = 3). Clinical benefit was achieved in 31 patients treated for one infection and in seven patients treated for multiple coincident infections. Thirteen of 14 patients treated for BKV-associated hemorrhagic cystitis experienced complete resolution of gross hematuria by week 6. Infusions were safe, and only two occurrences of de novo graft-versus host disease (grade 1) were observed. VST tracking by epitope profiling revealed persistence of functional VSTs of third-party origin for up to 12 weeks. Conclusion The use of banked VSTs is a feasible, safe, and effective approach to treat severe and drug-refractory infections after HSCT, including infections from two viruses (BKV and HHV-6) that had never been targeted previously with an off-the-shelf product. Furthermore, the multispecificity of the VSTs ensures extensive antiviral coverage, which facilitates the treatment of patients with multiple infections.

  19. Disseminated Cytomegalovirus: Report of a Case

    OpenAIRE

    Mosquera Klinger, Gabriel Alonso; Hospital Universitario San Ignacio; Guevara Pulido, Fredy Orlando; Hospital Universitario San Ignacio; Álvarez Moreno, Carlos

    2013-01-01

    This article presents a case of a man without previous history of disease, which initially consulted for abdominal pain. Acute cholecystitis is documented in findings of cytomegalovirus disease. In addition, it has many manifestations, including respiratory symptoms, gastrointestinal bleeding, pancytopenia. Was documented HIV infection, and serological and histological signs of cytomegalovirus, citomegalovirus diagnosis is made in the context of widespread HIV infection. El artículo presen...

  20. Cytomegalovirus Infection and Pre-Eclampsia

    Directory of Open Access Journals (Sweden)

    Rădulescu Carmen

    2016-06-01

    Full Text Available Introduction: Pre-eclampsia is a pregnancy-specific disease characterized by hypertension after 20 weeks of gestation and proteinuria. It is a major cause of maternal and perinatal morbidity and mortality. The pathogenesis of pre-eclampsia is not completely understood. In our study we investigated if there is a potential link between cytomegalovirus infection and pre-eclampsia and if cytomegalovirus infection is the triggering factor of pre-eclampsia.

  1. Frosted branch angiitis as a result of immune recovery uveitis in a patient with cytomegalovirus retinitis.

    Science.gov (United States)

    Leeamornsiri, Supinda; Choopong, Pitipol; Tesavibul, Nattaporn

    2013-06-22

    Since the introduction of Highly Active Antiretroviral Therapy (HAART), AIDs related morbidity and mortality have declined. However, the advent of HAART brought the new problem of immune recovery inflammatory syndrome. Cytomegalovirus retinitis remains the most common cause of visual loss in AIDs patients. Some patients with cytomegalovirus retinitis who experienced immune recovery as a consequence of HAART develop worsening of visual symptoms from immune recovery uveitis (IRU). We report a case of cytomegalovirus retinitis and AIDs who developed an unusual presentation of IRU after the initiation of HAART. A 40-year-old woman presented with a history of blurry vision in the right eye. She was diagnosed with human immunodeficiency virus infection and cytomegalovirus retinitis, treated with intravitreal injections of ganciclovir. The retinitis improved. One week after HAART initiation, she developed IRU, characterized by increased intraocular inflammation, extensive frosted branch angiitis and cystoid macular edema. The CD4+ T lymphocyte count increased from 53 to 107 cells/mm3. Systemic prednisolone with continuation of HAART and intravitreal injections of ganciclovir were given with significant improvement. Atypical presentation of IRU, characterized by extensive frosted branch angiitis and increased intraocular inflammation may occur in immunocompromised patients with cytomegalovirus retinitis who experienced immune recovery. The time from HAART initiation to develop IRU may vary from days to months. This case demonstrated a very rapidly developed IRU which should be recognized and appropriately managed to avoid permanent damage of the eye.

  2. Preparation of human resources for future nuclear energy using FBNR as the instrument of learning

    Energy Technology Data Exchange (ETDEWEB)

    Sefidvash, Farhang; Espinoza, Patricio; Guerrero, Victor Hugo [Escuela Politecnica Nacional (EPN), Quito (Ecuador); and others

    2015-11-15

    An increasing number of developing countries are showing interest to become the emerging countries to nuclear energy. Most of these countries lack human resources and adequate infrastructures to enter such a venture. The principle objective of activities of FBNR Group is to train human resources for the countries that at the present lack the necessary conditions, but aim at the future clean and safe nuclear energy through the fourth generation and INPRO compatible nuclear reactors. The preparation for the future nuclear energy is done through development of innovative nuclear reactor that meets the INPRO philosophies and criteria. These countries may or may not have decided as yet to utilize nuclear energy, but are interested to gain a strong educational foundation for their future. The research and development of a small innovative nuclear reactor FBNR is used as the instrument for learning. The young scientists will learn how to be innovative with the vision of INPRO philosophy and criteria.

  3. Prevalence of cytomegalovirus antibodies in blood donars at the ...

    African Journals Online (AJOL)

    Background: Cytomegalovirus (CMV) infection in susceptible patients is associated with serious morbidity and a high mortality. Transmission of cytomegalovirus infection through blood transfusion is markedly reduced by transfusion of CMV seronegative blood products, or by transfusion of leucodepleted blood products.

  4. Cytomegalovirus Vaccines: Current Status and Future Prospects

    Science.gov (United States)

    Anderholm, KM; Bierle, CJ; Schleiss, MR

    2017-01-01

    Congenital human cytomegalovirus (HCMV) infection can result in in severe and permanent neurological injury in newborns, and vaccine development is accordingly a major public health priority. HCMV can also cause disease in solid organ (SOT) and hematopoietic stem cell transplant (HSCT) recipients, and a vaccine would be valuable in prevention of viremia and end-organ disease in these populations. Currently there is no licensed HCMV vaccine, but progress toward this goal has been made in recent clinical trials. A recombinant HCMV glycoprotein B (gB) vaccine has been shown to have some efficacy in prevention of infection in young women and adolescents, and provided benefit to HCMV-seronegative SOT recipients. Similarly, DNA vaccines based on gB and the immunodominant T-cell target, pp65 (ppUL83), have been shown to reduce viremia in HSCT patients. This review provides an overview of HCMV vaccine candidates in various stages of development, as well as an update on the current status of ongoing clinical trials. Protective correlates of vaccine-induced immunity may be different for pregnant woman and transplant patients. As more knowledge emerges about correlates of protection, the ultimate licensure of HCMV vaccines may reflect the uniqueness of the target populations being immunized. PMID:27882457

  5. The downmodulation of the foreign body reaction by cytomegalovirus encoded interleukin-10

    NARCIS (Netherlands)

    van Putten, S.M.; Wubben, M.; Hennink, W.E.; van Luyna, M.J.A.; Harmsen, M.C.

    2009-01-01

    The foreign body reaction (FBR) is of great importance for the function and turnover of biomaterial scaffolds. The development of biological tools that modulate the FBR will augment scaffold functionality and benefit regenerative medicine. The human cytomegalovirus encodes a functional homolog of

  6. Investigation of the Role of the Cytomegalovirus as a Respiratory Pathogen in HIV-Infected Patients

    Directory of Open Access Journals (Sweden)

    Rafael E de la Hoz

    1996-01-01

    Full Text Available OBJECTIVE: To investigate the occurrence of cytomegalovirus (CMV pneumonitis in the setting of human immunodeficiency virus (HIV infection and whether the presence of CMV as copathogen is associated with increased clinical severity or short term mortality in patients with Pneumocystis carinii pneumonia.

  7. Acute rejection before cytomegalovirus infection enhances von Willebrand factor and soluble VCAM-1 in blood

    NARCIS (Netherlands)

    Kas-Deelen, AM; Harmsen, MC; de Maar, EF; Oost-Kort, WW; Tervaert, JWC; van der Meer, J; van Son, WJ; The, TH

    2000-01-01

    Background. Human cytomegalovirus (HCMV) infections in transplantation patients are associated with vascular endothelial damage. This is reflected by the appearance of cytomegalic endothelial cells (CECs) and noninfected endothelial cells (ECs) in blood. To get more insight in the extent of vascular

  8. Novel Chemokine-Based Immunotoxins for Potent and Selective Targeting of Cytomegalovirus Infected Cells

    DEFF Research Database (Denmark)

    Spiess, Katja; Jeppesen, Mads G.; Malmgaard-Clausen, Mikkel

    2017-01-01

    of human cytomegalovirus (HCMV) infections. US28 is expressed on virus-infected cells and scavenge chemokines by rapid internalization. The chemokine-based fusion-toxin protein (FTP) consisted of a variant (F49A) of CX3CL1 specifically targeting US28 linked to the catalytic domain of Pseudomonas exotoxin...

  9. Associations between anti-cytomegalovirus IgG responses and health effect biomarkers

    Science.gov (United States)

    Human cytomegalovirus (HCMV) is a member of the herpes simplex virus family that infects approximately 50% of US adults. HCMV is transmitted from person to person through bodily fluids, congenitally or from donors to transplant recipients. It causes a lifelong latent infection th...

  10. Perianal Plaques of Cytomegalovirus in a Patient with Central Nervous System Lymphoma

    Directory of Open Access Journals (Sweden)

    Scott K. Heysell

    2012-01-01

    Full Text Available Cutaneous manifestations of cytomegalovirus (CMV in patients without human immunodeficiency virus remain rare. Perianal CMV may be observed due to periodic fecal shedding but may be confused for other pathogens, and definitive diagnosis requires histopathologic examination. An instructive case is described, and the literature reviewed.

  11. Human cytomegalovirus and Epstein-Barr virus infection impact on {sup 18}F-FDG PET/CT SUVmax, CT volumetric and KRAS-based parameters of patients with locally advanced rectal cancer treated with neoadjuvant therapy

    Energy Technology Data Exchange (ETDEWEB)

    Sole, Claudio V. [Instituto de Radiomedicina, Department of Radiation Oncology, Santiago (Chile); School of Medicine Complutense University, Madrid (Spain); Calvo, Felipe A. [Hospital General Universitario Gregorio Maranon, Department of Oncology, Madrid (Spain); School of Medicine Complutense University, Madrid (Spain); Hospital General Universitario Gregorio Maranon, Institute for Sanitary Research, Madrid (Spain); Ferrer, Carlos [Hospital Provincial de Castellon, Institute of Oncology, Castellon de la Plana (Spain); School of Medicine Cardenal Herrera-CEU University, Castellon de la Plana (Spain); Alvarez, Emilio [School of Medicine Complutense University, Madrid (Spain); Hospital General Universitario Gregorio Maranon, Department of Pathology, Madrid (Spain); Hospital General Universitario Gregorio Maranon, Institute for Sanitary Research, Madrid (Spain); Carreras, Jose L. [School of Medicine Complutense University, Madrid (Spain); Hospital General Universitario Gregorio Maranon, Department of Radiology and Medical Physics, Madrid (Spain); Ochoa, Enrique [Hospital Provincial de Castellon, Institute of Oncology, Castellon de la Plana (Spain)

    2014-10-01

    It has long been debated whether human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV) are associated with rectal cancer. The gene products of HCMV and EBV contribute to cell-cycle progression, mutagenesis, angiogenesis and immune evasion. The aim of this prospective study was to analyse the association between infection of a tumour by HCMV and EBV and clinical, histological, metabolic ({sup 18}F-FDG uptake), volumetric (from CT) and molecular (KRAS status) features and long-term outcomes in a homogeneously treated group of patients with locally advanced rectal cancer. HCMV and EBV were detected in pretreatment biopsies using polymerase chain reaction (PCR). The Cox proportional hazards regression model was used to explore associations between viral infection and disease-free survival (DFS) and overall survival (OS). We analysed 37 patients with a median follow-up of 74 months (range 5-173 months). Locoregional control, OS and DFS at 5 years were 93 %, 74 % and 71 %, respectively. Patients with HCMV/EBV coinfection had a significantly higher maximum standardized uptake value than patients without viral coinfection (p = 0.02). Significant differences were also observed in staging and percentage relative reduction in tumour volume between patients with and without HCMV infection (p < 0.01) and EBV infection (p < 0.01). KRAS wildtype status was significantly more frequently observed in patients with EBV infection (p <0.01) and HCMV/EBV co-infection (p = 0.04). No significant differences were observed in OS or DFS between patients with and without EBV infection (p = 0.88 and 0.73), HCMV infection (p = 0.84 and 0.79), and EBV/CMV coinfection (p = 0.24 and 0.39). This pilot study showed that viral infections were associated with metabolic staging differences, and differences in the evolution of metabolic and volumetric parameters and KRAS mutations. Further findings of specific features will help determine the best candidates for metabolic and volumetric staging and

  12. "pp65 antigenemia and real time polymerase chain reaction (PCR based-study to determine the prevalence of human cytomegalovirus (HCMV in kidney donors and recipients with follow-up studies."

    Directory of Open Access Journals (Sweden)

    Vijayakumar Ramanathan

    2010-11-01

    Full Text Available Abstract Background The present study was undertaken to determine the rate of occurrence of Human cytomegalovirus (HCMV among kidney transplant recipients and donors by application of direct detection methods and to understand HCMV infection/disease development among transplanted patients as a prospective study. Results Peripheral blood samples collected from 76 kidney donors and 76 recipients from September 2007 to August 2009 were subjected to pp65 antigenemia and Quantitative real-time PCR (qRT-PCR assays. Data were analyzed under Group A, B and C. Group A was further divided into sub-groups I, II, III, IV, and V for better understanding. Three, one and two donors in sub-group I, III, IV of Group A tested positive for real time PCR respectively. One recipient from group III tested positive for HCMV by qRT- PCR prior transplantation and remained positive one month post-transplantation. Three other recipients, tested negative prior to transplantation became positive a month after transplantation. Group B consisted of 18 donor-recipient pairs and one of the donor tested positive for HCMV by qRT-PCR. Eight recipients tested positive for HCMV one month after transplantation. The pp65 positivity and HCMV DNA load was high among group C recipients who mostly had symptoms of active disease. Significantly high values of pp65 antigenemia were observed among recipients of sub-group II (non-parametric chi-square test p = 0.007. Positive correlation between pp65 antigenemia and qRT-PCR value was observed. Thirty three of the recipients with disease treated with Valgancyclovir showed improved clinical outcome. Conclusion Our study showed that a significant proportion of kidney recipients develop HCMV infection following renal transplantation in spite of the absence of HCMV among donors. pp65 antigenemia assay and qRT- PCR methods can be applied to detect HCMV among kidney donors and recipients to monitor development of disease and these assays were

  13. Conserved retinoblastoma protein-binding motif in human cytomegalovirus UL97 kinase minimally impacts viral replication but affects susceptibility to maribavir

    Directory of Open Access Journals (Sweden)

    Chou Sunwen

    2009-01-01

    Full Text Available Abstract The UL97 kinase has been shown to phosphorylate and inactivate the retinoblastoma protein (Rb and has three consensus Rb-binding motifs that might contribute to this activity. Recombinant viruses containing mutations in the Rb-binding motifs generally replicated well in human foreskin fibroblasts with only a slight delay in replication kinetics. Their susceptibility to the specific UL97 kinase inhibitor, maribavir, was also examined. Mutation of the amino terminal motif, which is involved in the inactivation of Rb, also renders the virus hypersensitive to the drug and suggests that the motif may play a role in its mechanism of action.

  14. Congenital and perinatal cytomegalovirus infection

    Directory of Open Access Journals (Sweden)

    Chun Soo Kim

    2010-01-01

    Full Text Available Cytomegalovirus (CMV is currently the most common agent of congenital infection and the leading infectious cause of brain damage and hearing loss in children. Symptomatic congenital CMV infections usually result from maternal primary infection during early pregnancy. One half of symptomatic infants have cytomegalic inclusion disease (CID, which is characterized by involvement of multiple organs, in particular, the reticuloendothelial and central nervous system (CNS. Moreover, such involvement may or may not include ocular and auditory damage. Approximately 90% of infants with congenital infection are asymptomatic at birth. Preterm infants with perinatal CMV infection can have symptomatic diseases such as pneumonia, hepatitis, and thrombocytopenia. Microcephaly and abnormal neuroradiologic imaging are associated with a poor prognosis. Hearing loss may occur in both symptomatic and asymptomatic infants with congenital infection and may progress through childhood. Congenital infection is defined by the isolation of CMV from infants within the first 3 weeks of life. Ganciclovir therapy can be considered for infants with symptomatic congenital CMV infection involving the CNS. Pregnant women of seronegative state should be counseled on the importance of good hand washing and other control measures to prevent CMV infection. Heat treatment of infected breast milk at 72?#608;for 5 seconds can eliminate CMV completely.

  15. Cytomegalovirus Infection Drives Adaptive Epigenetic Diversification of NK Cells with Altered Signaling and Effector Function

    OpenAIRE

    Schlums, Heinrich; Cichocki, Frank; Tesi, Bianca; Theorell, Jakob; Beziat, Vivien; Holmes, Tim D.; Han, Hongya; Chiang, Samuel C.C.; Foley, Bree; Mattsson, Kristin; Larsson, Stella; Schaffer, Marie; Malmberg, Karl-Johan; Ljunggren, Hans-Gustaf; Miller, Jeffrey S.

    2015-01-01

    The mechanisms underlying human natural killer (NK) cell phenotypic and functional heterogeneity are unknown. Here, we describe the emergence of diverse subsets of human NK cells selectively lacking expression of signaling proteins after human cytomegalovirus (HCMV) infection. The absence of B and myeloid cell-related signaling protein expression in these NK cell subsets correlated with promoter DNA hyperme-thylation. Genome-wide DNA methylation patterns were strikingly similar between HCMV-a...

  16. Country report on human resource development in nuclear field

    Energy Technology Data Exchange (ETDEWEB)

    Wanitsuksombut, Warapon; Noochpramool, Kovit [Office of Atomic Energy for Peace, Bangkok (Thailand)

    2000-12-01

    The short-term plan is to promote utilization of the new research reactor in Thailand. The long-term plan is to gain public understanding and acceptance of nuclear technology. Since 1991, the office has conducted training and seminars in nuclear related field. The major training is in radiation protection, and training in nuclear reactor was at noticeably smaller portion. For ten years of training, 3,649 persons passed different radiation protection courses. Education programs in universities are outlined with the curriculums in the paper. It is clear that the manpower produced in nuclear field in Thailand is inadequate. Further more, most of them are working in limited areas in specific institutes, research laboratories, modern hospitals, and academic teaching. They seldom contact with the public. Hence communication to the public is lacking. After the training course for schoolteachers in our research reactor site, many of them appreciate new knowledge of nuclear technology. They became to realize that they had been involved with the nuclear technology before in their everyday well being. The urgent need is to arrange various suitable courses on research reactor utilization. In this effort, the exchange of information, equipment as well as teaching materials form developed institutes are necessary. The urgent need is a system of qualification for Radiation Protection Officer. By exchange of information and seminars, it may help the country to decide whether the harmonization and accreditation of training courses or the accredited examination is adopted. For long-term achievement, a regular seminar for schoolteacher should be formulated, and a program for social and economics curriculum in nuclear field should be initiated. (Tanaka, Y.)

  17. Human factors design guidelines for maintainability of Department of Energy nuclear facilities

    Energy Technology Data Exchange (ETDEWEB)

    Bongarra, J.P. Jr.; VanCott, H.P.; Pain, R.F.; Peterson, L.R.; Wallace, R.I.

    1985-06-18

    Intent of these guidelines is to provide design and design review teams of DOE nuclear facilities with human factors principles to enhance the design and aid in the inspection of DOE nuclear facilities, systems, and equipment. These guidelines are concerned with design features of DOE nuclear facilities which can potentially affect preventive and corrective maintenance of systems within DOE nuclear facilities. Maintenance includes inspecting, checking, troubleshooting, adjusting, replacing, repairing, and servicing activities. Other factors which influence maintainability such as repair and maintenance suport facilities, maintenance information, and various aspects of the environment are also addressed.

  18. Human Reliability Analysis for Digitized Nuclear Power Plants: Case Study on the LingAo II Nuclear Power Plant

    Directory of Open Access Journals (Sweden)

    Yanhua Zou

    2017-03-01

    Full Text Available The main control room (MCR in advanced nuclear power plants (NPPs has changed from analog to digital control system (DCS. Operation and control have become more automated, centralized, and accurate due to the digitalization of NPPs, which has improved the efficiency and security of the system. New issues associated with human reliability inevitably arise due to the adoption of new accident procedures and digitalization of main control rooms in NPPs. The LingAo II NPP is the first digital NPP in China to apply the state-oriented procedure. In order to address issues related to human reliability analysis for DCS and DCS + state-oriented procedure, the Hunan Institute of Technology conducted a research project based on a cooperative agreement with the LingDong Nuclear Power Co. Ltd. This paper is a brief introduction to the project.

  19. Human resource development for nuclear generation - from the perspective of a utility company

    Science.gov (United States)

    Kahar, Wan Shakirah Wan Abdul; Mostafa, Nor Azlan; Salim, Mohd Faiz

    2017-01-01

    Malaysia is currently in the planning phase of its nuclear power program, with the first unit targeted to be operational in 2030. Training of nuclear power plant (NPP) staffs are usually long and rigorous due to the complexity and safety aspects of nuclear power. As the sole electricity utility in the country, it is therefore essential that Tenaga Nasional Berhad (TNB) prepares early in developing its human resource and nuclear expertise as a potential NPP owner-operator. A utility also has to be prudent in managing its work force efficiently and effectively, while ensuring that adequate preparations are being made to acquire the necessary nuclear knowledge with sufficient training lead time. There are several approaches to training that can be taken by a utility company with no experience in nuclear power. These include conducting feasibility studies and benchmarking exercises, preparing long term human resource development, increasing the exposure on nuclear power technology to both the top management and general staff, and employing the assistance of relevant agencies locally and abroad. This paper discusses the activities done and steps taken by TNB in its human resource development for Malaysia's nuclear power program.

  20. Cytomegalovirus Infection of the Rat Developing Brain In Utero Prominently Targets Immune Cells and Promotes Early Microglial Activation.

    Science.gov (United States)

    Cloarec, Robin; Bauer, Sylvian; Luche, Hervé; Buhler, Emmanuelle; Pallesi-Pocachard, Emilie; Salmi, Manal; Courtens, Sandra; Massacrier, Annick; Grenot, Pierre; Teissier, Natacha; Watrin, Françoise; Schaller, Fabienne; Adle-Biassette, Homa; Gressens, Pierre; Malissen, Marie; Stamminger, Thomas; Streblow, Daniel N; Bruneau, Nadine; Szepetowski, Pierre

    2016-01-01

    Congenital cytomegalovirus infections are a leading cause of neurodevelopmental disorders in human and represent a major health care and socio-economical burden. In contrast with this medical importance, the pathophysiological events remain poorly known. Murine models of brain cytomegalovirus infection, mostly neonatal, have brought recent insights into the possible pathogenesis, with convergent evidence for the alteration and possible involvement of brain immune cells. In order to confirm and expand those findings, particularly concerning the early developmental stages following infection of the fetal brain, we have created a model of in utero cytomegalovirus infection in the developing rat brain. Rat cytomegalovirus was injected intraventricularly at embryonic day 15 (E15) and the brains analyzed at various stages until the first postnatal day, using a combination of gene expression analysis, immunohistochemistry and multicolor flow cytometry experiments. Rat cytomegalovirus infection was increasingly seen in various brain areas including the choroid plexi and the ventricular and subventricular areas and was prominently detected in CD45low/int, CD11b+ microglial cells, in CD45high, CD11b+ cells of the myeloid lineage including macrophages, and in CD45+, CD11b- lymphocytes and non-B non-T cells. In parallel, rat cytomegalovirus infection of the developing rat brain rapidly triggered a cascade of pathophysiological events comprising: chemokines upregulation, including CCL2-4, 7 and 12; infiltration by peripheral cells including B-cells and monocytes at E17 and P1, and T-cells at P1; and microglia activation at E17 and P1. In line with previous findings in neonatal murine models and in human specimen, our study further suggests that neuroimmune alterations might play critical roles in the early stages following cytomegalovirus infection of the brain in utero. Further studies are now needed to determine which role, whether favorable or detrimental, those putative

  1. Origin of nuclear buds and micronuclei in normal and folate-deprived human lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Lindberg, Hanna K. [New Technologies and Risks, Work Environment Development, Finnish Institute of Occupational Health, Topeliuksenkatu 41aA, FI-00250 Helsinki (Finland); Wang Xu [Genome Health and Nutrigenomics Project, CSIRO Human Nutrition, Adelaide BC, SA 5000 (Australia); School of Life Sciences, Yunnan Normal University, Kunming, Yunnan 650092 (China); Jaerventaus, Hilkka [New Technologies and Risks, Work Environment Development, Finnish Institute of Occupational Health, Topeliuksenkatu 41aA, FI-00250 Helsinki (Finland); Falck, Ghita C.-M. [New Technologies and Risks, Work Environment Development, Finnish Institute of Occupational Health, Topeliuksenkatu 41aA, FI-00250 Helsinki (Finland); Norppa, Hannu [New Technologies and Risks, Work Environment Development, Finnish Institute of Occupational Health, Topeliuksenkatu 41aA, FI-00250 Helsinki (Finland)]. E-mail: hannu.norppa@ttl.fi; Fenech, Michael [Genome Health and Nutrigenomics Project, CSIRO Human Nutrition, Adelaide BC, SA 5000 (Australia)

    2007-04-01

    Micronuclei are formed from chromosomes and chromosomal fragments that lag behind in anaphase and are left outside daughter nuclei in telophase. They may also be derived from broken anaphase bridges. Nuclear buds, micronucleus-like bodies attached to the nucleus by a thin nucleoplasmic connection, have been proposed to be generated similarly to micronuclei during nuclear division or in S-phase as a stage in the extrusion of extra DNA, possibly giving rise to micronuclei. To better understand these phenomena, we have characterized the contents of 894 nuclear buds and 1392 micronuclei in normal and folate-deprived 9-day cultures of human lymphocytes using fluorescence in situ hybridization with pancentromeric and pantelomeric DNA probes. Such information has not earlier been available for human primary cells. Surprisingly, there appears to be no previous data on the occurrence of telomeres in micronuclei (or buds) of normal human cells in general. Our results suggest that nuclear buds and micronuclei have partly different mechanistic origin. Interstitial DNA without centromere or telomere label was clearly more prevalent in nuclear buds (43%) than in micronuclei (13%). DNA with only telomere label or with both centromere and telomere label was more frequent in micronuclei (62% and 22%, respectively) than in nuclear buds (44% and 10%, respectively). Folate deprivation especially increased the frequency of nuclear buds and micronuclei harboring telomeric DNA and nuclear buds harboring interstitial DNA but also buds and micronuclei with both centromeric and telomeric DNA. According to the model we propose, that micronuclei in binucleate lymphocytes primarily derive from lagging chromosomes and terminal acentric fragments during mitosis. Most nuclear buds, however, are suggested to originate from interstitial or terminal acentric fragments, possibly representing nuclear membrane entrapment of DNA that has been left in cytoplasm after nuclear division or excess DNA that

  2. Nuclear lactate dehydrogenase modulates histone modification in human hepatocytes

    Energy Technology Data Exchange (ETDEWEB)

    Castonguay, Zachary; Auger, Christopher; Thomas, Sean C.; Chahma, M’hamed; Appanna, Vasu D., E-mail: vappanna@laurentian.ca

    2014-11-07

    Highlights: • Nuclear LDH is up-regulated under oxidative stress. • SIRT1 is co-immunoprecipitated bound to nuclear LDH. • Nuclear LDH is involved in histone deacetylation and epigenetics. - Abstract: It is becoming increasingly apparent that the nucleus harbors metabolic enzymes that affect genetic transforming events. Here, we describe a nuclear isoform of lactate dehydrogenase (nLDH) and its ability to orchestrate histone deacetylation by controlling the availability of nicotinamide adenine dinucleotide (NAD{sup +}), a key ingredient of the sirtuin-1 (SIRT1) deacetylase system. There was an increase in the expression of nLDH concomitant with the presence of hydrogen peroxide (H{sub 2}O{sub 2}) in the culture medium. Under oxidative stress, the NAD{sup +} generated by nLDH resulted in the enhanced deacetylation of histones compared to the control hepatocytes despite no discernable change in the levels of SIRT1. There appeared to be an intimate association between nLDH and SIRT1 as these two enzymes co-immunoprecipitated. The ability of nLDH to regulate epigenetic modifications by manipulating NAD{sup +} reveals an intricate link between metabolism and the processing of genetic information.

  3. Rat pancreatic beta cells and cytomegalovirus infection.

    Science.gov (United States)

    Smelt, Maaike J; Faas, Marijke M; de Haan, Bart J; Hofstede, Jeroen; Cheung, Chi-Wai; van der Iest, Hanna; de Haan, Aalzen; de Vos, Paul

    2010-01-01

    Cytomegalovirus (CMV) infection has been suggested to accelerate beta-cell destruction and thereby to contribute to new-onset diabetes and failure of islet allografts in both humans and rodents. Surprisingly, direct CMV infection of beta cells has received only minor attention. Therefore, we investigated the susceptibility of rat beta cells for rat CMV (RCMV) infection and the direct effects on the regulation of immune cell-activating ligands. Primary rat beta cells, the rat beta-cell line Rin-m5F, and fibroblasts were RCMV-infected in vitro. The viral gene and protein expression levels were determined as a measure for RCMV susceptibility. Gene expression levels of intracellular adhesion molecule 1, lymphocyte function associated antigen 3, rat major histocompatibility complex region A, rat major histocompatibility complex region E, toll like receptor 2, and clustered domain 14 were determined as a measure for cellular immunogenicity. We demonstrate that beta cells are susceptible for RCMV infection but allow only low levels of viral gene expression. In contrast, infected fibroblasts demonstrated productive viral infection and formation of viral progeny. After RCMV infection, beta-cell immunogenicity was markedly increased, as demonstrated by the increased cellular expression of immune cell-activating ligands. Direct beta-cell infection by RCMV and subsequent low-grade viral gene expression may lead to increased immunogenicity of native or transplanted beta cells in vivo. An infection-induced enhanced beta-cell recognizability may have important consequences for beta-cell survival and the development of diabetes or rejection of islet grafts.

  4. Cloning, expression and nuclear localization of human NPM3, a member of the nucleophosmin/nucleoplasmin family of nuclear chaperones

    Directory of Open Access Journals (Sweden)

    Ganguly Amit

    2001-11-01

    Full Text Available Abstract Background Studies suggest that the related proteins nucleoplasmin and nucleophosmin (also called B23, NO38 or numatrin are nuclear chaperones that mediate the assembly of nucleosomes and ribosomes, respectively, and that these activities are accomplished through the binding of basic proteins via their acidic domains. Recently discovered and less well characterized members of this family of acidic phosphoproteins include mouse nucleophosmin/nucleoplasmin 3 (Npm3 and Xenopus NO29. Here we report the cloning and initial characterization of the human ortholog of Npm3. Results Human genomic and cDNA clones of NPM3 were isolated and sequenced. NPM3 lies 5.5 kb upstream of FGF8 and thus maps to chromosome 10q24-26. In addition to amino acid similarities, NPM3 shares many physical characteristics with the nucleophosmin/nucleoplasmin family, including an acidic domain, multiple potential phosphorylation sites and a putative nuclear localization signal. Comparative analyses of 14 members of this family from various metazoans suggest that Xenopus NO29 is a candidate ortholog of human and mouse NPM3, and they further group both proteins closer with the nucleoplasmins than with the nucleophosmins. Northern blot analysis revealed that NPM3 was strongly expressed in all 16 human tissues examined, with especially robust expression in pancreas and testis; lung displayed the lowest level of expression. An analysis of subcellular fractions of NIH3T3 cells expressing epitope-tagged NPM3 revealed that NPM3 protein was localized solely in the nucleus. Conclusions Human NPM3 is an abundant and widely expressed protein with primarily nuclear localization. These biological activities, together with its physical relationship to the chaparones nucleoplasmin and nucleophosmin, are consistent with the proposed function of NPM3 as a molecular chaperone functioning in the nucleus.

  5. Prevalence and associated characteristics of cytomegalovirus (CMV ...

    African Journals Online (AJOL)

    Background: The screening for cytomegalovirus (CMV) specific antibodies is not routine in our setting, thus the transfusion of blood portends high risk for susceptible individuals. Objective: To determine the prevalence of IgG and IgM specific antibodies and associated characteristics in blood donors seen at a referral ...

  6. Seroepidemiology study of cytomegalovirus and rubella among ...

    African Journals Online (AJOL)

    Background: Maternal cytomegalovirus (CMV) and rubella infections have adverse neonatal outcomes. Both CMV and rubella are more widespread in developing countries and in communities with lower socioeconomic status. The aim of this study was to investigate sero-prevalence of CMV and rubella infection and ...

  7. Postnatally acquired cytomegalovirus infections in preterm infants

    NARCIS (Netherlands)

    Nijman, J.

    2013-01-01

    A postnatal cytomegalovirus (CMV) infection is common in very low birth weight infants with an estimated prevalence of 6–59%. Breast milk from CMV seropositive mothers is the main source of postnatal CMV infection. Ninety-six percent of these mothers shed CMV in their breast milk after delivery due

  8. Seroprevalence of cytomegalovirus among pregnant women ...

    African Journals Online (AJOL)

    Background: Primary Cytomegalovirus (CMV) infection during pregnancy is a frequent and serious threat to the fetus. There is no vaccine as such alternative measures are needed to prevent congenital CMV infection. Objective: This study determined CMV Immunoglobulin G (IgG) antibody among pregnant women in order ...

  9. Seroprevalence of cytomegalovirus among some voluntary blood ...

    African Journals Online (AJOL)

    Background: Cytomegalovirus (CMV) is one of the most significant pathogens infecting immuno-suppressed individuals. CMV is transmissible through transfusion of blood components. Aim: The goal of this study was to determine the seroprevalence of antibodies to CMV among blood donors seen at the 37 Military Hospital ...

  10. Systemic lupus erythematous revealed by cytomegalovirus infection ...

    African Journals Online (AJOL)

    Cytomegalovirus (CMV) infection have been described as exacerbing systemic lupus erythematous (SLE). The role of CMV in starting off SLE remains object of debate. We report a severe presentation of SLE revealed by CMV infection with hemophogocytic syndrome. A 22 old women without a history of systemic disease ...

  11. Cytomegalovirus ileocolitis in a rheumatoid arthritis patient: case report and literature review

    Directory of Open Access Journals (Sweden)

    M. S. Dag

    2015-06-01

    Full Text Available Rheumatoid arthritis (RA is an autoimmune, systemic, chronic, inflammatory disease generally treated with various immunosuppressive drugs. Cytomegalovirus (CMV is an opportunistic, viral infection that is commonly seen in immunosuppressed patients. A sixty-four-year old female diagnosed with RA and treated with immunosuppressive agents was admitted to our rheumatology outpatient service with complaints of diarrhea and abdominal pain, which had lasted longer than four weeks. The patient’s colonoscopy revealed inflamed and ulcerated areas in the colon and in the terminal ileum. A biopsy showed intra-nuclear inclusion particles consistent with CMV. We started an oral valganciclovir therapy in this serum-CMV-polymerase chain reactionpositive patient. The concomitant use of immunosuppressive agents and anti-viral drugs eased the patient’s complaints, and the endoscopic picture improved. Consequently, cytomegalovirus ileocolitis in immunosuppressed patients admitted with severe diarrhea must be considered in the differential diagnosis.

  12. Identification of a putative nuclear export signal motif in human NANOG homeobox domain

    Energy Technology Data Exchange (ETDEWEB)

    Park, Sung-Won; Do, Hyun-Jin; Huh, Sun-Hyung; Sung, Boreum [Department of Biomedical Science, College of Life Science, CHA University, Seongnam-si, Gyeonggi-Do (Korea, Republic of); Uhm, Sang-Jun [Department of Animal Science and Biotechnology, Sangji Youngseo College, Wonju, Gwangwon-Do (Korea, Republic of); Song, Hyuk [Department of Animal and Food Bioscience, College of Natural Science, Konkuk University, ChungJu, Chungbuk (Korea, Republic of); Kim, Nam-Hyung [Department of Animal Sciences, Chungbuk National University, Cheongju, Chungbuk (Korea, Republic of); Kim, Jae-Hwan, E-mail: jaehwan_k@cha.ac.kr [Department of Biomedical Science, College of Life Science, CHA University, Seongnam-si, Gyeonggi-Do (Korea, Republic of)

    2012-05-11

    Highlights: Black-Right-Pointing-Pointer We found the putative nuclear export signal motif within human NANOG homeodomain. Black-Right-Pointing-Pointer Leucine-rich residues are important for human NANOG homeodomain nuclear export. Black-Right-Pointing-Pointer CRM1-specific inhibitor LMB blocked the potent human NANOG NES-mediated nuclear export. -- Abstract: NANOG is a homeobox-containing transcription factor that plays an important role in pluripotent stem cells and tumorigenic cells. To understand how nuclear localization of human NANOG is regulated, the NANOG sequence was examined and a leucine-rich nuclear export signal (NES) motif ({sup 125}MQELSNILNL{sup 134}) was found in the homeodomain (HD). To functionally validate the putative NES motif, deletion and site-directed mutants were fused to an EGFP expression vector and transfected into COS-7 cells, and the localization of the proteins was examined. While hNANOG HD exclusively localized to the nucleus, a mutant with both NLSs deleted and only the putative NES motif contained (hNANOG HD-{Delta}NLSs) was predominantly cytoplasmic, as observed by nucleo/cytoplasmic fractionation and Western blot analysis as well as confocal microscopy. Furthermore, site-directed mutagenesis of the putative NES motif in a partial hNANOG HD only containing either one of the two NLS motifs led to localization in the nucleus, suggesting that the NES motif may play a functional role in nuclear export. Furthermore, CRM1-specific nuclear export inhibitor LMB blocked the hNANOG potent NES-mediated export, suggesting that the leucine-rich motif may function in CRM1-mediated nuclear export of hNANOG. Collectively, a NES motif is present in the hNANOG HD and may be functionally involved in CRM1-mediated nuclear export pathway.

  13. Seroprevalence of Human Cytomegalovirus (HCMV) infection in ...

    African Journals Online (AJOL)

    A total of 148 participants were recruited into the study consisting of 72 patients with haematologic malignancy and 76 healthy blood donors. The seroprevalence of anti-CMV IgG was similar in both groups (98.6% and 94.7% respectively) while there was a higher anti-CMV IgM seropositivity in patients with haematologic ...

  14. Human cytomegalovirus infections in premature infants by ...

    African Journals Online (AJOL)

    Freezing breast milk may be protective for the preterm infant until the titer of CMV antibody increases. However clinical importance of CMV infection in premature infants by breast-feeding is still unclear. This minireview focuses on recent advances in the study of CMV infection in premature infants by breastfeeding.

  15. A Proteomics Analysis of Human Cytomegalovirus Particles

    Energy Technology Data Exchange (ETDEWEB)

    Streblow, Daniel N.; Varnum, Susan M.; Smith, Richard D.; Nelson, Jay

    2006-01-01

    While the sequence of the AD169 HCMV genome has been known for several years, the viral and cellular proteins that compose the infectious HCMV virion and entry-competent, non-replicating viral particles such as Dense Bodies (DBs) and Non-Infectious Enveloped Particles (NIEPs) are unknown. To approach this problem we have utilized a gel-free 2-D capillary liquid chromatography (LC)-MS/MS and Fourier transform ion cyclotron resonance (FTICR) mass spectrometry to identify and determine the relative abundance of viral and cellular proteins in purified HCMV AD169 particles. !is study has identified and quantitated the proteins that compose both HCMV virions and DBs. While a number of previously identified proteins were detected by this method the number of viral proteins that compose the HCMV virion was doubled in this study suggesting that over a third of the viral open reading frames are part of an infectious virion. !is chapter will discuss the implications of our findings in relation to what was previously known about HCMV and MCMV virion composition.

  16. Adapting Human Reliability Analysis from Nuclear Power to Oil and Gas Applications

    Energy Technology Data Exchange (ETDEWEB)

    Boring, Ronald Laurids [Idaho National Laboratory

    2015-09-01

    ABSTRACT: Human reliability analysis (HRA), as currently used in risk assessments, largely derives its methods and guidance from application in the nuclear energy domain. While there are many similarities be-tween nuclear energy and other safety critical domains such as oil and gas, there remain clear differences. This paper provides an overview of HRA state of the practice in nuclear energy and then describes areas where refinements to the methods may be necessary to capture the operational context of oil and gas. Many key distinctions important to nuclear energy HRA such as Level 1 vs. Level 2 analysis may prove insignifi-cant for oil and gas applications. On the other hand, existing HRA methods may not be sensitive enough to factors like the extensive use of digital controls in oil and gas. This paper provides an overview of these con-siderations to assist in the adaptation of existing nuclear-centered HRA methods to the petroleum sector.

  17. The Nuclear DNA Sensor IFI16 Acts as a Restriction Factor for Human Papillomavirus Replication through Epigenetic Modifications of the Viral Promoters.

    Science.gov (United States)

    Lo Cigno, Irene; De Andrea, Marco; Borgogna, Cinzia; Albertini, Silvia; Landini, Manuela M; Peretti, Alberto; Johnson, Karen E; Chandran, Bala; Landolfo, Santo; Gariglio, Marisa

    2015-08-01

    The human interferon-inducible IFI16 protein, an innate immune sensor of intracellular DNA, was recently demonstrated to act as a restriction factor for human cytomegalovirus (HCMV) and herpes simplex virus 1 (HSV-1) infection by inhibiting both viral-DNA replication and transcription. Through the use of two distinct cellular models, this study provides strong evidence in support of the notion that IFI16 can also restrict human papillomavirus 18 (HPV18) replication. In the first model, an immortalized keratinocyte cell line (NIKS) was used, in which the IFI16 protein was knocked down through the use of small interfering RNA (siRNA) technology and overexpressed following transduction with the adenovirus IFI16 (AdVIFI16) vector. The second model consisted of U2OS cells transfected by electroporation with HPV18 minicircles. In differentiated IFI16-silenced NIKS-HPV18 cells, viral-load values were significantly increased compared with differentiated control cells. Consistent with this, IFI16 overexpression severely impaired HPV18 replication in both NIKS and U2OS cells, thus confirming its antiviral restriction activity. In addition to the inhibition of viral replication, IFI16 was also able to reduce viral transcription, as demonstrated by viral-gene expression analysis in U2OS cells carrying episomal HPV18 minicircles and HeLa cells. We also provide evidence that IFI16 promotes the addition of heterochromatin marks and the reduction of euchromatin marks on viral chromatin at both early and late promoters, thus reducing both viral replication and transcription. Altogether, these results argue that IFI16 restricts chromatinized HPV DNA through epigenetic modifications and plays a broad surveillance role against viral DNA in the nucleus that is not restricted to herpesviruses. Intrinsic immunity is mediated by cellular restriction factors that are constitutively expressed and active even before a pathogen enters the cell. The host nuclear factor IFI16 acts as a sensor

  18. Cytomegalovirus-induced embryopathology: mouse submandibular salivary gland epithelial-mesenchymal ontogeny as a model

    Directory of Open Access Journals (Sweden)

    Huang Jing

    2006-09-01

    Full Text Available Abstract Background Human studies suggest, and mouse models clearly demonstrate, that cytomegalovirus (CMV is dysmorphic to early organ and tissue development. CMV has a particular tropism for embryonic salivary gland and other head mesenchyme. CMV has evolved to co-opt cell signaling networks so to optimize replication and survival, to the detriment of infected tissues. It has been postulated that mesenchymal infection is the critical step in disrupting organogenesis. If so, organogenesis dependent on epithelial-mesenchymal interactions would be particularly vulnerable. In this study, we chose to model the vulnerability by investigating the cell and molecular pathogenesis of CMV infected mouse embryonic submandibular salivary glands (SMGs. Results We infected E15 SMG explants with mouse CMV (mCMV. Active infection for up to 12 days in vitro results in a remarkable cell and molecular pathology characterized by atypical ductal epithelial hyperplasia, apparent epitheliomesenchymal transformation, oncocytic-like stromal metaplasia, β-catenin nuclear localization, and upregulation of Nfkb2, Relb, Il6, Stat3, and Cox2. Rescue with an antiviral nucleoside analogue indicates that mCMV replication is necessary to initiate and maintain SMG dysmorphogenesis. Conclusion mCMV infection of embryonic mouse explants results in dysplasia, metaplasia, and, possibly, anaplasia. The molecular pathogenesis appears to center around the activation of canonical and, perhaps more importantly, noncanonical NFκB. Further, COX-2 and IL-6 are important downstream effectors of embryopathology. At the cellular level, there appears to be a consequential interplay between the transformed SMG cells and the surrounding extracellular matrix, resulting in the nuclear translocation of β-catenin. From these studies, a tentative framework has emerged within which additional studies may be planned and performed.

  19. Mouse Embryonic Stem Cells Inhibit Murine Cytomegalovirus Infection through a Multi-Step Process

    Science.gov (United States)

    Kawasaki, Hideya; Kosugi, Isao; Arai, Yoshifumi; Iwashita, Toshihide; Tsutsui, Yoshihiro

    2011-01-01

    In humans, cytomegalovirus (CMV) is the most significant infectious cause of intrauterine infections that cause congenital anomalies of the central nervous system. Currently, it is not known how this process is affected by the timing of infection and the susceptibility of early-gestational-period cells. Embryonic stem (ES) cells are more resistant to CMV than most other cell types, although the mechanism responsible for this resistance is not well understood. Using a plaque assay and evaluation of immediate-early 1 mRNA and protein expression, we found that mouse ES cells were resistant to murine CMV (MCMV) at the point of transcription. In ES cells infected with MCMV, treatment with forskolin and trichostatin A did not confer full permissiveness to MCMV. In ES cultures infected with elongation factor-1α (EF-1α) promoter-green fluorescent protein (GFP) recombinant MCMV at a multiplicity of infection of 10, less than 5% of cells were GFP-positive, despite the fact that ES cells have relatively high EF-1α promoter activity. Quantitative PCR analysis of the MCMV genome showed that ES cells allow approximately 20-fold less MCMV DNA to enter the nucleus than mouse embryonic fibroblasts (MEFs) do, and that this inhibition occurs in a multi-step manner. In situ hybridization revealed that ES cell nuclei have significantly less MCMV DNA than MEF nuclei. This appears to be facilitated by the fact that ES cells express less heparan sulfate, β1 integrin, and vimentin, and have fewer nuclear pores, than MEF. This may reduce the ability of MCMV to attach to and enter through the cellular membrane, translocate to the nucleus, and cross the nuclear membrane in pluripotent stem cells (ES/induced pluripotent stem cells). The results presented here provide perspective on the relationship between CMV susceptibility and cell differentiation. PMID:21407806

  20. Functional impairment of Tax-specific but not cytomegalovirus-specific CD8+ T lymphocytes in a minor population of asymptomatic human T-cell leukemia virus type 1-carriers

    Directory of Open Access Journals (Sweden)

    Takamori Ayako

    2011-12-01

    Full Text Available Abstract Background Human T-cell leukemia virus type 1 (HTLV-1 causes adult T-cell leukemia (ATL and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP in a small percentage of infected individuals. ATL is often associated with general immune suppression and an impaired HTLV-1-specific T-cell response, an important host defense system. We previously found that a small fraction of asymptomatic HTLV-1-carriers (AC already showed impaired T-cell responses against the major target antigen, Tax. However, it is unclear whether the impaired HTLV-1 Tax-specific T-cell response in these individuals is an HTLV-1-specific phenomenon, or merely reflects general immune suppression. In this study, in order to characterize the impaired HTLV-1-specific T-cell response, we investigated the function of Tax-specific CD8+ T-cells in various clinical status of HTLV-1 infection. Results By using tetramers consisting of HLA-A*0201, -A*2402, or -A*1101, and corresponding Tax epitope peptides, we detected Tax-specific CD8+ T-cells in the peripheral blood from 87.0% of ACs (n = 20/23 and 100% of HAM/TSP patients (n = 18/18 tested. We also detected Tax-specific CD8+ T-cells in 38.1% of chronic type ATL (cATL patients (n = 8/21, although its frequencies in peripheral blood CD8+ T cells were significantly lower than those of ACs or HAM/TSP patients. Tax-specific CD8+ T-cells detected in HAM/TSP patients proliferated well in culture and produced IFN-γ when stimulated with Tax peptides. However, such functions were severely impaired in the Tax-specific CD8+ T-cells detected in cATL patients. In ACs, the responses of Tax-specific CD8+ T-cells were retained in most cases. However, we found one AC sample whose Tax-specific CD8+ T-cells hardly produced IFN-γ, and failed to proliferate and express activation (CD69 and degranulation (CD107a markers in response to Tax peptide. Importantly, the same AC sample contained cytomegalovirus (CMV pp65-specific CD8+ T

  1. Nuclear resonance scattering measurement of human iron stores

    Energy Technology Data Exchange (ETDEWEB)

    Wielopolski, L.; Ancona, R.C.; Mossey, R.T.; Vaswani, A.N.; Cohn, S.H.

    1985-07-01

    Hepatic iron stores were measured noninvasively in 31 patients (thalassemia, hemodialysis, hemosiderosis, refractory anemia) with suspected iron overload, employing a nuclear resonance scattering (NRS) technique. The thalassemia patients were undergoing desferrioxamine chelation therapy during the NRS measurements. The hemodialysis patients were measured before chelation therapy. Iron levels measured by NRS were in general agreement with those determined in liver biopsies by atomic absorption spectroscopy. In addition, NRS measurements from the thorax of some of these patients suggest that this method may also prove useful for clinical assessment of cardiac iron.

  2. Interaction profiling identifies the human nuclear exosome targeting complex

    DEFF Research Database (Denmark)

    Lubas, Michal Szymon; Christensen, Marianne Spangsberg; Kristiansen, Maiken Søndergaard

    2011-01-01

    The RNA exosome is a conserved degradation machinery, which obtains full activity only when associated with cofactors. The most prominent activator of the yeast nuclear exosome is the RNA helicase Mtr4p, acting in the context of the Trf4p/Air2p/Mtr4p polyadenylation (TRAMP) complex. The existence...... from nucleoli, and consistently NEXT is specifically required for the exosomal degradation of promoter upstream transcripts (PROMPTs). We also detect putative homolog TRAMP subunits hTRF4-2 (Trf4p) and ZCCHC7 (Air2p) in hRRP6 and hMTR4 precipitates. However, at least ZCCHC7 function is restricted...

  3. Human Factors and Information Operation for a Nuclear Power Space Vehicle

    Science.gov (United States)

    Trujillo, Anna C.; Brown-VanHoozer, S. Alenka

    2002-01-01

    This paper describes human-interactive systems needed for a crewed nuclear-enabled space mission. A synthesis of aircraft engine and nuclear power plant displays, biofeedback of sensory input, virtual control, brain mapping for control process and manipulation, and so forth are becoming viable solutions. These aspects must maintain the crew's situation awareness and performance, which entails a delicate function allocation between crew and automation.

  4. Comparison of solar photovoltaic and nuclear reactor power systems for a human-tended lunar observatory

    Science.gov (United States)

    Hickman, J. M.; Bloomfield, H. S.

    1989-01-01

    Photovoltaic and nuclear surface power systems were examined at the 20 to 100 kW power level range for use at a human-tended lunar astronomical observatory, and estimates of the power system masses were made. One system, consisting of an SP-100 thermoelectric nuclear power supply integrated with a lunar lander, is recommended for further study due to its low system mass, potential for modular growth, and applicability to other surface power missions, particularly in the Martian system.

  5. Human factors in maintenance: Development and research in Swedish nuclear power plants

    Energy Technology Data Exchange (ETDEWEB)

    Salo, I. [Lund Univ. (Sweden). Dept. of Psychology; Svensson, Ola [Stockholm Univ. (Sweden). Dept. of Psychology

    2001-11-01

    The present report investigated previously completed, ongoing, and planned research and development projects focusing human factors and maintenance work carried out at Swedish nuclear power plants and SKI. In addition, needs for future research and development works were also investigated. Participants from all nuclear power plants and SKI were included in the study. Participants responded to a set of questions in an interview. The interviews also generated a list of future research and development projects.

  6. Molecular Image Analysis: Quantitative Description and Classification of the Nuclear Lamina in Human Mesenchymal Stem Cells

    OpenAIRE

    Righolt, Christiaan H.; Vered Raz; Vermolen, Bart J.; Dirks, Roeland W; Hans J. Tanke; Young, Ian T

    2011-01-01

    The nuclear lamina is an intermediate filament network that provides a structural framework for the cell nucleus. Changes in lamina structure are found during changes in cell fate such as cell division or cell death and are associated with human diseases. An unbiased method that quantifies changes in lamina shape can provide information on cells undergoing changes in cellular functions. We have developed an image processing methodology that finds and quantifies the 3D structure of the nuclear...

  7. Comparison of solar photovoltaic and nuclear reactor power systems for a human-tended lunar observatory

    Science.gov (United States)

    Hickman, J. M.; Bloomfield, H. S.

    1989-01-01

    Photovoltaic and nuclear surface power systems were examined at the 20 to 100 kW power level range for use at a human-tended lunar astronomical observatory, andestimates of the power system masses were made. One system, consisting of an SP-100 thermoelectric nuclear power supply integrated with a lunar lander, is recommended for further study due to its low system mass, potential for modular growth, and applicability to other surface power missions, particularly in the Martian system.

  8. Safe Operation of Nuclear Power Plants: Impacts of Human and Organisational Factors and Emerging Technologies

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2001-07-01

    In co-operation with the OECD Nuclear Energy Agency (NEA), the Halden Reactor Project organised a Summer School on ''Safe Operation of Nuclear Power Plants: Impacts of Human and Organisational Factors and Emerging Technologies'' in the period August 27-August 31, 2001. The Summer School was intended for scientists, engineers and technicians working for nuclear installations, engineering companies, industry and members of universities and research institutes, who wanted to broaden their nuclear background by getting acquainted with Man-Technology-Organisation-related subjects and issues. The Summer School should also serve to transfer knowledge to the ''young generation'' in the nuclear field. The following presentations were given: (1) Overview of the Nuclear Community and Current issues, (2) The Elements of Safety Culture; Evaluation of Events, (3) Quality Management (QM), (4) Probabilistic Risk Assessment (PSA), (5) Human Behaviour from the Viewpoint of Industrial Psychology, (6) Technical tour of the Halden Project Experimental Facilities, (7) Human Factors in Control Room Design, (8) Computerised Operator Support Systems (COSSs) and (9) Artificial Intelligence; a new Approach. Most of the contributions are overhead figures from spoken lectures.

  9. The Red Queen in mitochondria: cyto-nuclear co-evolution, hybrid breakdown and human disease

    Directory of Open Access Journals (Sweden)

    Jui-Yu eChou

    2015-05-01

    Full Text Available Cyto-nuclear incompatibility, a specific form of Dobzhansky-Muller incompatibility caused by incompatible alleles between mitochondrial and nuclear genomes, has been suggested to play a critical role during speciation. Several features of the mitochondrial genome (mtDNA, including high mutation rate, dynamic genomic structure, and uniparental inheritance, make mtDNA more likely to accumulate mutations in the population. Once mtDNA has changed, the nuclear genome needs to play catch-up due to the intimate interactions between these two genomes. In two populations, if cyto-nuclear co-evolution is driven in different directions, it may eventually lead to hybrid incompatibility. Although cyto-nuclear incompatibility has been observed in a wide range of organisms, it remains unclear what type of mutations drives the co-evolution. Currently, evidence supporting adaptive mutations in mtDNA remains limited. On the other hand, it has been known that some mutations allow mtDNA to propagate more efficiently but compromise the host fitness (described as selfish mtDNA. Arms races between such selfish mtDNA and host nuclear genomes can accelerate cyto-nuclear co-evolution and lead to a phenomenon called the Red Queen Effect. Here, we discuss how the Red Queen Effect may contribute to the frequent observation of cyto-nuclear incompatibility and be the underlying driving force of some human mitochondrial diseases.

  10. The Red Queen in mitochondria: cyto-nuclear co-evolution, hybrid breakdown and human disease.

    Science.gov (United States)

    Chou, Jui-Yu; Leu, Jun-Yi

    2015-01-01

    Cyto-nuclear incompatibility, a specific form of Dobzhansky-Muller incompatibility caused by incompatible alleles between mitochondrial and nuclear genomes, has been suggested to play a critical role during speciation. Several features of the mitochondrial genome (mtDNA), including high mutation rate, dynamic genomic structure, and uniparental inheritance, make mtDNA more likely to accumulate mutations in the population. Once mtDNA has changed, the nuclear genome needs to play catch-up due to the intimate interactions between these two genomes. In two populations, if cyto-nuclear co-evolution is driven in different directions, it may eventually lead to hybrid incompatibility. Although cyto-nuclear incompatibility has been observed in a wide range of organisms, it remains unclear what type of mutations drives the co-evolution. Currently, evidence supporting adaptive mutations in mtDNA remains limited. On the other hand, it has been known that some mutations allow mtDNA to propagate more efficiently but compromise the host fitness (described as selfish mtDNA). Arms races between such selfish mtDNA and host nuclear genomes can accelerate cyto-nuclear co-evolution and lead to a phenomenon called the Red Queen Effect. Here, we discuss how the Red Queen Effect may contribute to the frequent observation of cyto-nuclear incompatibility and be the underlying driving force of some human mitochondrial diseases.

  11. Nuclear DNA Content Varies with Cell Size across Human Cell Types

    Science.gov (United States)

    Gillooly, James F.; Hein, Andrew; Damiani, Rachel

    2015-01-01

    Variation in the size of cells, and the DNA they contain, is a basic feature of multicellular organisms that affects countless aspects of their structure and function. Within humans, cell size is known to vary by several orders of magnitude, and differences in nuclear DNA content among cells have been frequently observed. Using published data, here we describe how the quantity of nuclear DNA across 19 different human cell types increases with cell volume. This observed increase is similar to intraspecific relationships between DNA content and cell volume in other species, and interspecific relationships between diploid genome size and cell volume. Thus, we speculate that the quantity of nuclear DNA content in somatic cells of humans is perhaps best viewed as a distribution of values that reflects cell size distributions, rather than as a single, immutable quantity. PMID:26134319

  12. Nuclear DNA Content Varies with Cell Size across Human Cell Types.

    Science.gov (United States)

    Gillooly, James F; Hein, Andrew; Damiani, Rachel

    2015-07-01

    Variation in the size of cells, and the DNA they contain, is a basic feature of multicellular organisms that affects countless aspects of their structure and function. Within humans, cell size is known to vary by several orders of magnitude, and differences in nuclear DNA content among cells have been frequently observed. Using published data, here we describe how the quantity of nuclear DNA across 19 different human cell types increases with cell volume. This observed increase is similar to intraspecific relationships between DNA content and cell volume in other species, and interspecific relationships between diploid genome size and cell volume. Thus, we speculate that the quantity of nuclear DNA content in somatic cells of humans is perhaps best viewed as a distribution of values that reflects cell size distributions, rather than as a single, immutable quantity. Copyright © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.

  13. The effects of different maceration techniques on nuclear DNA amplification using human bone.

    Science.gov (United States)

    Lee, Esther J; Luedtke, Jennifer G; Allison, Jamie L; Arber, Carolyn E; Merriwether, D Andrew; Steadman, Dawnie Wolfe

    2010-07-01

    Forensic anthropologists routinely macerate human bone for the purposes of identity and trauma analysis, but the heat and chemical treatments used can destroy genetic evidence. As a follow-up to a previous study on nuclear DNA recovery that used pig ribs, this study utilizes human skeletal remains treated with various bone maceration techniques for nuclear DNA amplification using the standard Combined DNA Index System (CODIS) markers. DNA was extracted from 18 samples of human lower leg bones subjected to nine chemical and heat maceration techniques. Genotyping was carried out using the AmpFlSTR COfiler and AmpFlSTR Profiler Plus ID kits. Results showed that heat treatments via microwave or Biz/Na(2)CO(3) in sub-boiling water efficiently macerate bone and produce amplifiable nuclear DNA for genetic analysis. Long-term use of chemicals such as hydrogen peroxide is discouraged as it results in poor bone quality and has deleterious effects on DNA amplification.

  14. Los Alamos National Laboratory Human and Intellectual Capital for Sustaining Nuclear Deterrence

    Energy Technology Data Exchange (ETDEWEB)

    McAlpine, Bradley [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2015-04-01

    This paper provides an overview of the current human and intellectual capital at Los Alamos National Laboratory, through specific research into the statistics and demographics as well as numerous personal interviews at all levels of personnel. Based on this information, a series of recommendations are provided to assist Los Alamos National Laboratory in ensuring the future of the human and intellectual capital for the nuclear deterrence mission. While the current human and intellectual capital is strong it stands on the precipice and action must be taken to ensure Los Alamos National Laboratory maintains leadership in developing and sustaining national nuclear capabilities. These recommendations may be applicable to other areas of the nuclear enterprise, including the Air Force, after further research and study.

  15. A review of the evolution of human reliability analysis methods at nuclear industry

    Energy Technology Data Exchange (ETDEWEB)

    Oliveira, Lécio N. de; Santos, Isaac José A. Luquetti dos; Carvalho, Paulo V.R., E-mail: lecionoliveira@gmail.com, E-mail: luquetti@ien.gov.br, E-mail: paulov@ien.gov.br [Instituto de Engenharia Nuclear (IEN/CNEN-RJ), Rio de Janeiro, RJ (Brazil)

    2017-11-01

    This paper reviews the status of researches on the application of human reliability analysis methods at nuclear industry and its evolution along the years. Human reliability analysis (HRA) is one of the elements used in Probabilistic Safety Analysis (PSA) and is performed as part of PSAs to quantify the likelihood that people will fail to take action, such as errors of omission and errors of commission. Although HRA may be used at lots of areas, the focus of this paper is to review the applicability of HRA methods along the years at nuclear industry, especially in Nuclear Power Plants (NPP). An electronic search on CAPES Portal of Journals (A bibliographic database) was performed. This literature review covers original papers published since the first generation of HRA methods until the ones published on March 2017. A total of 94 papers were retrieved by the initial search and 13 were selected to be fully reviewed and for data extraction after the application of inclusion and exclusion criteria, quality and suitability evaluation according to applicability at nuclear industry. Results point out that the methods from first generation are more used in practice than methods from second generation. This occurs because it is more concentrated towards quantification, in terms of success or failure of human action what make them useful for quantitative risk assessment to PSA. Although the second generation considers context and error of commission in human error prediction, they are not wider used in practice at nuclear industry to PSA. (author)

  16. The human side of the nuclear safety issue

    Energy Technology Data Exchange (ETDEWEB)

    Wallis, L.R. [U.S. Citizens for Total Energy, Sunol, CA (United States)

    1995-12-31

    Historically, scientists have been primarily concerned with the technical integrity of their own speciality within the rest of the technical community. No one saw a need to establish dialogue with the general public, special interest groups or women`s groups about decisions governed by their own professions. Public support, confidence and respect came automatically. As society became more complex, this mode of operation by scientists became even more intense while institutions became more and more unwieldy. At the same time they failed to note a rather dramatic change slowly taking place - the public began making their decisions for them -decisions based many times on propaganda not fact. Left unchallenged, our sons and daughters will grow up believing that nuclear power is the most visible and potent symbol of a technological society gone amok, and it must be destroyed. Fortunately, the scientific community now recognizes that public support, confidence and respect no longer comes automatically; they must be cultivated and earned. (author).

  17. A human factors design of a nuclear plant analyzer

    Energy Technology Data Exchange (ETDEWEB)

    Byeon, Seung Nam; Lee, Dong Hoon; Park, Chan Woo [Kyounghee Univ., Seoul (Korea, Republic of)

    2000-12-15

    The project consists of four key stages as follows : based on the review of various literature, human factors design principles and processes are identified, the literature reviewed in the project includes NUREG-0700, research papers for MMI, human factors handbooks, and laboratory reports, after the design principles and processes are determined, a design checklist is developed to evaluate the user interface of NPA, the design checklist consists of seven different categories such as display screen, menu interface, form-fillin, alphanumeric characters, symbols, color, and highlighting, NPA was tested with the design checklist for conformance to the human factors design principles, the expert reviews are performed to evaluate a graphic user interface of NPA, the application of the design checklist and the subjective opinion of the expert identify the design included in the user interface of NPA, based on the thorough analysis of design defects, design guidelines are recommended to improve the user interface of NPA.

  18. Cytomegalovirus myelitis in perinatally acquired HIV.

    OpenAIRE

    Güngör, T; Funk, M; Linde, R; Jacobi, G; Horn, M; Kreuz, W

    1993-01-01

    A 7 year old child perinatally infected with HIV who died from progressive muscular paralysis and central nervous respiratory failure is described. Cytomegalovirus (CMV) prophylaxis with a special intravenous CMV hyper-immunoglobulin had been successfully conducted for more than four years. Macroscopic and microscopic immunohistochemical examination of the spinal cord revealed a diffuse CMV infiltration of the entire myelon. CMV infected cells were identified as astrocytes, oligodendrocytes, ...

  19. Women's attitudes toward practicing cytomegalovirus prevention behaviors

    OpenAIRE

    Thackeray, Rosemary; Magnusson, Brianna M.

    2016-01-01

    Congenital cytomegalovirus (CMV) infection causes severe disabilities and developmental delays. Women's awareness of CMV is low. Only about half of healthcare providers report counseling women about behaviors to reduce CMV risk and public health education is limited. Routine CMV counseling is not recommend. Providers may lack time to counsel women; other conditions may take priority for counseling; there may be a perception that women are reluctant to follow advice. This cross-sectional descr...

  20. Close but Distinct Regions of Human Herpesvirus 8 Latency-Associated Nuclear Antigen 1 Are Responsible for Nuclear Targeting and Binding to Human Mitotic Chromosomes

    Science.gov (United States)

    Piolot, Tristan; Tramier, Marc; Coppey, Maité; Nicolas, Jean-Claude; Marechal, Vincent

    2001-01-01

    Human herpesvirus 8 is associated with all forms of Kaposi's sarcoma, AIDS-associated body cavity-based lymphomas, and some forms of multicentric Castleman's disease. Herpesvirus 8, like other gammaherpesviruses, can establish a latent infection in which viral genomes are stably maintained as multiple episomes. The latent nuclear antigen (LANA or LNAI) may play an essential role in the stable maintenance of latent episomes, notably by interacting concomitantly with the viral genomes and the metaphase chromosomes, thus ensuring an efficient transmission of the neoduplicated episomes to the daughter cells. To identify the regions responsible for its nuclear and subnuclear localization in interphase and mitotic cells, LNAI and various truncated forms were fused to a variant of green fluorescent protein. This enabled their localization and chromosome binding activity to be studied by low-light-level fluorescence microscopy in living HeLa cells. The results demonstrate that nuclear localization of LNAI is due to a unique signal, which maps between amino acids 24 and 30. Interestingly, this nuclear localization signal closely resembles those identified in EBNA1 from Epstein-Barr virus and herpesvirus papio. A region encompassing amino acids 5 to 22 was further proved to mediate the specific interaction of LNA1 with chromatin during interphase and the chromosomes during mitosis. The presence of putative phosphorylation sites in the chromosome binding sites of LNA1 and EBNA1 suggests that their activity may be regulated by specific cellular kinases. PMID:11264383

  1. Cytomegalovirus as an oncomodulatory agent in the progression of glioma.

    Science.gov (United States)

    Joseph, Gabriel P; McDermott, Ryan; Baryshnikova, Maria A; Cobbs, Charles S; Ulasov, Ilya V

    2017-01-01

    Glioblastoma multiforme (GBM) is the most aggressive neoplastic brain tumor in humans with a median survival of less than 2 years. It is therefore critical to understand the mechanism of glioma progression and to identify future targets for intervention. We investigate the mechanisms of cytomegalovirus as an oncomodulatory agent implicated in glioma progression, as well as immunosuppression. This review provides a comprehensive evaluation of recent investigative developments concerning the role of CMV in cellular processes during glioma growth. The manners in which CMV and its viral products interact with regulatory cellular signaling pathways in the host are of primary interest. Here, we examine some of the most significant oncomodulatory effects that CMV can confer in brain tumors, including the inhibition of apoptosis and promoting the growth of glioma stem cells, which are tightly linked to tumor survival and recurrence. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  2. Cytomegalovirus-induced immunopathology and its clinical consequences

    Science.gov (United States)

    2011-01-01

    Human cytomegalovirus (CMV) is a ubiquitous DNA virus that causes severe disease in patients with immature or impaired immune systems. During active infection, CMV modulates host immunity, and CMV-infected patients often develop signs of immune dysfunction, such as immunosuppression and autoimmune phenomena. Furthermore, active viral infection has been observed in several autoimmune diseases, and case reports have linked primary CMV infection and the onset of autoimmune disorders. In addition, CMV infection promotes allograft rejection and graft-versus-host disease in solid organ and bone marrow transplant recipients, respectively, further implicating CMV in the genesis and maintenance of immunopathological phenomena. The mechanisms by which CMV could induce inhibition of host defense, inflammation, and autoimmunity are discussed, as is the treatment of virus-induced immunopathology with antivirals. PMID:21473750

  3. Collaborative human-machine nuclear non-proliferation analysis

    Energy Technology Data Exchange (ETDEWEB)

    Greitzer, F.L.; Badalamente, R.V.; Stewart, T.S.

    1993-10-01

    The purpose of this paper is to report on the results of a project investigating support concepts for the information treatment needs of the International Atomic Energy Agency (IAEA, also referred to as the Agency) and its attempts to strengthen international safeguards. The aim of the research was to define user/computer interface concepts and intelligent support features that will enhance the analyst`s access to voluminous and diverse information, the ability to recognize and evaluate uncertain data, and the capability to make decisions and recommendations. The objective was to explore techniques for enhancing safeguards analysis through application of (1) more effective user-computer interface designs and (2) advanced concepts involving human/system collaboration. The approach was to identify opportunities for human/system collaboration that would capitalize on human strengths and still accommodate human limitations. This paper documents the findings and describes a concept prototype, Proliferation Analysis Support System (PASS), developed for demonstration purposes. The research complements current and future efforts to enhance the information systems used by the IAEA, but has application elsewhere, as well.

  4. Transient Antiphospholipid Syndrome Associated with Primary Cytomegalovirus Infection: A Case Report and Literature Review

    Directory of Open Access Journals (Sweden)

    Tsuyoshi Nakayama

    2014-01-01

    Full Text Available Viral infection is known to induce transient autoimmunity in humans. Acute cytomegalovirus (CMV infection is implicated in occasional thrombosis formation. We here, for the first time, report a 19-year-old female who had an acute CMV infection, leading to a deep venous thrombosis and a pulmonary embolism along with transient appearance of lupus anticoagulant. The pathological role of antiphospholipid antibodies in CMV-mediated thrombosis is discussed.

  5. Characterization of specific antibodies against cytomegalovirus (CMV)-encoded interleukin 10 produced by 28 % of CMV-seropositive blood donors

    DEFF Research Database (Denmark)

    de Lemos Rieper, Carina; Galle, Pia Søndergaard; Pedersen, Bente Klarlund

    2011-01-01

    Cytomegalovirus (CMV) has evolved multiple immunological evasion strategies, including the encoding of viral interleukin (IL)-10 homologues (cmvIL-10). In this study, cmvIL-10 bound avidly to the same receptors on blood mononuclear cells and was as bio-potent as native human IL-10. Seventeen...

  6. Evaluation of New Commercial Real-Time PCR Quantification Assay for Prenatal Diagnosis of Cytomegalovirus Congenital Infection▿

    Science.gov (United States)

    Ducroux, Aurélie; Cherid, Samira; Benachi, Alexandra; Ville, Yves; Leruez-Ville, Marianne

    2008-01-01

    A new commercial real-time human cytomegalovirus (HCMV) PCR kit was evaluated after automated DNA extraction of 153 amniotic fluids in parallel with an in-house real-time PCR assay. The commercial kit displayed 100% sensitivity/specificity compared to the “in-house” assay and was suitable for prenatal diagnosis of HCMV congenital infection. PMID:18417654

  7. Supra-Additive Expression of Interleukin-6, Interleukin-8 and Basic Fibroblast Growth Factor in Vascular Smooth Muscle Cells Following Coinfection with Chlamydia pneumoniae and Cytomegalovirus as a Novel Link between Infection and Atherosclerosis

    Directory of Open Access Journals (Sweden)

    Dirk Prochnau

    2012-01-01

    Full Text Available BACKGROUND: Chlamydia pneumoniae and human cytomegalovirus (HCMV may be involved in the pathogenesis of atherosclerosis. Prospective studies indicate an increased risk for cardiovascular events in patients with evidence of multiple infections.

  8. Recommendations to the NRC on human engineering guidelines for nuclear power plant maintainability

    Energy Technology Data Exchange (ETDEWEB)

    Badalamente, R.V.; Fecht, B.A.; Blahnik, D.E.; Eklund, J.D.; Hartley, C.S.

    1986-03-01

    This document contains human engineering guidelines which can enhance the maintainability of nuclear power plants. The guidelines have been derived from general human engineering design principles, criteria, and data. The guidelines may be applied to existing plants as well as to plants under construction. They apply to nuclear power plant systems, equipment and facilities, as well as to maintenance tools and equipment. The guidelines are grouped into seven categories: accessibility and workspace, physical environment, loads and forces, maintenance facilities, maintenance tools and equipment, operating equipment design, and information needs. Each chapter of the document details specific maintainability problems encountered at nuclear power plants, the safety impact of these problems, and the specific maintainability design guidelines whose application can serve to avoid these problems in new or existing plants.

  9. Consequences of nuclear accidents for human beings and the environment; Auswirkungen nuklearer Unfaelle auf den Menschen und die Umwelt

    Energy Technology Data Exchange (ETDEWEB)

    Huebner, Felix; Jung, Jennifer Jana; Schultmann, Frank

    2017-03-15

    In the 1950s and 1960s many countries believed that they could become energy self-efficient with the help of nuclear power plants. For this reason, nuclear energy experienced internationally an enormous upswing during these years. In the following decades, many nuclear power plants were planned and built. Despite a few critics, who drew attention to the dangers of nuclear power use, an end to the upturn was not foreseeable. But in 1979, the nuclear accident in Three Mile Island in Harrisburg (USA) caused international attention and the critics of the nuclear power became stronger. However, many Europeans only were aware of the danger by nuclear power plants when in 1986 the nuclear disaster occurred in the Ukrainian nuclear power plant in Chernobyl. The effects of this nuclear disaster were also felt in Central Europe. Recently, the nuclear disaster at the Fukushima nuclear power plant has prompted a nuclear phase-out in Germany and the rest of the world. This work focuses not on the political implications of these nuclear accidents, but rather on the impacts of nuclear accidents on human beings, animals and the environment. In particular, reports on the incidents in Chernobyl and Fukushima as well as studies on their effects are analysed and summarised.

  10. Procedure for conducting a human-reliability analysis for nuclear power plants. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Bell, B.J.; Swain, A.D.

    1983-05-01

    This document describes in detail a procedure to be followed in conducting a human reliability analysis as part of a probabilistic risk assessment when such an analysis is performed according to the methods described in NUREG/CR-1278, Handbook for Human Reliability Analysis with Emphasis on Nuclear Power Plant Applications. An overview of the procedure describing the major elements of a human reliability analysis is presented along with a detailed description of each element and an example of an actual analysis. An appendix consists of some sample human reliability analysis problems for further study.

  11. Molecular Image Analysis : Quantitative Description and Classification of the Nuclear Lamina in Human Mesenchymal Stem Cells

    NARCIS (Netherlands)

    Righolt, C.H.; Raz, V.; Vermolen, B.J.; Dirks, R.W.; Tanke, H.J.; Young, I.T.

    2011-01-01

    The nuclear lamina is an intermediate filament network that provides a structural framework for the cell nucleus. Changes in lamina structure are found during changes in cell fate such as cell division or cell death and are associated with human diseases. An unbiased method that quantifies changes

  12. Nuclear localization of human DNA mismatch repair protein exonuclease 1 (hEXO1)

    DEFF Research Database (Denmark)

    Knudsen, Nina Østergaard; Nielsen, Finn Cilius; Vinther, Lena

    2007-01-01

    Human exonuclease 1 (hEXO1) is implicated in DNA mismatch repair (MMR) and mutations in hEXO1 may be associated with hereditary nonpolyposis colorectal cancer (HNPCC). Since the subcellular localization of MMR proteins is essential for proper MMR function, we characterized possible nuclear locali...

  13. Human performance tools in nuclear power plants. Introduction, implementation and experiences; Human Performance Tools in Kernkraftwerken. Einfuehrung, Umsetzung und Erfahrungen

    Energy Technology Data Exchange (ETDEWEB)

    Dexheimer, Kai; Bassing, Gerd [Dexcon Consulting GmbH, Neuhausen (Switzerland); Kreuzer, Peter [E.ON Kernkraft GmbH, Essenbach (Germany). Kernkraftwerk Isar

    2015-06-01

    The basis of safe nuclear power plant operation (NPP) and a strong safety culture is the professional application of Human Performance Optimisation Tools (HPO). HPO trainings have been carried out by German NPPs for a number of years and recently also by Swiss NPPs. This article describes the origination, the bases, experiences and thereby the special features of the HPO training programme applied by German NPP operators. Moreover, this article provides an outlook on future developments - in particular when considering the requirements of the ongoing phase out of nuclear energy in Germany.

  14. Present status and needs of human resource development in nuclear field in Korea

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Young-Myung; Lee, Eui-Jin [Nuclear Training Center, Korea Atomic Energy Research Institute, Taejon (Korea)

    2000-12-01

    The Nuclear Training Center (NTC) of KAERI (Korea Atomic Energy Research Institute) began training technical personnel in the field of radioisotope utilization and radiation protection during the 1960's. During the first stage of the nation's nuclear power project in the 1970's, the main effort of the Center focused on training those in nuclear power and nuclear engineering. During a stage of increased technical self-reliance in the 1980's, the Center extended its training role to implement more specific training courses on nuclear power and safety fields. Since 1983, the Center has been empowered at the request of government to provide retraining courses for nuclear-related license holders and qualified engineers. The Center has offered IAEA regional training course annually for Asia and Pacific region member states since 1988. Since 1967, the total number of trainees is up to 27,777 as of the end of 1998. KEPCO (Korea Electric Power Corporation) started Nuclear Power Education Center (NPEC) in 1990. The outlines of KEPCO's in-house training programs are presented in the report. The reactor operators, and the persons engaged in nuclear fuel materials, radioisotope or radiation generating devices need particular licenses in accordance with Korean Atomic Energy Laws and Regulation. NTC/KAERI and NPEC/KEPCO should report annual retraining programs for licensed personnel to Ministry Of Science and Technology (MOST) every year. The outlines of projects, which are directly related to human resources development in nuclear field in Korea, are described in the paper. The International Atomic Energy Agency (IAEA) has made efforts to provide training programs for technical personnel of developing countries for the peaceful uses of nuclear energy. Korea has also received lots of assistance for her manpower development from the Agency. Korea is now on the verge of transforming herself from a technology recipient country in some practical and

  15. Present status and needs of human resource development in the nuclear field in the Philippines

    Energy Technology Data Exchange (ETDEWEB)

    Bernido, Corazon C.; Roceles, Pilar C. [Philippine Nuclear Research Institute, Commonwealth Avenue, Diliman, Quezon (Philippines)

    2000-12-01

    The first nuclear power plant was nearing completion. However, due to change in political climate and support for the nuclear power program, this has been mothballed. There is a possibility for the introduction of nuclear power plant in the country's projected energy sources by the year 2020. The country has one research reactor, but at the present time it is undergoing repair and is not operational. The Philippine Nuclear Research Institute (PNRI), an Institute under the Department of Science and Technology (DOST), is the sole government agency mandated by the law to take charge of all matters pertaining to nuclear science and technology, and the regulation of nuclear energy. There is one another government agency, the Radiation Health Service (RHS) of the Department of Health, which is responsible for regulating the use and application of X-rays and non-ionizing radiation. The PNRI conducts national training courses in nuclear science and technology, and radiation protection to users of radioisotopes. Individual courses are outlined in the paper. Up to the present time, around 7,300 have participated in national training courses conducted by PNRI. Distributions of PNRI trainees are: 53 % for industrial, 12 % medical, 12 % for academe, and 23 % for others. Nuclear science and technology education in schools and universities are presented. The International Atomic Energy Agency (IAEA) training activities availed 77 % of the total foreign training from 1993 to 1998; Japan follows next at 20 %; and others comprise the remaining 3 %. An approach to training and human resources development, which could reach out to more target trainees, is Distance Learning. In 1998, as a part of a Regional Cooperative Agreement (RCA) and IAEA project, the Philippines participated in the trial of distance learning modules in radiation protection. The distance learning modules were developed at the Australian Nuclear Science and Technology Organization (ANSTO). These modules will

  16. Identification of a functional nuclear localization signal within the human USP22 protein

    Energy Technology Data Exchange (ETDEWEB)

    Xiong, Jianjun [Key Laboratory of Jiangxi Province for the Systems Bio-Medicine, Jiujiang, Jiangxi Province 332000 (China); College of Basic Medical Science, Jiujiang University, Jiujiang, Jiangxi Province 332000 (China); Wang, Yaqin [Reproductive Medical Center, Renmin Hospital of Wuhan University, Wuhan, Hubei Province 430060 (China); Gong, Zhen [Key Laboratory of Jiangxi Province for the Systems Bio-Medicine, Jiujiang, Jiangxi Province 332000 (China); Liu, Jianyun [College of Basic Medical Science, Jiujiang University, Jiujiang, Jiangxi Province 332000 (China); Li, Weidong, E-mail: lwd626518@163.com [College of Basic Medical Science, Jiujiang University, Jiujiang, Jiangxi Province 332000 (China)

    2014-06-20

    Highlights: • USP22 was accumulated in nucleus. • We identified of a functional USP22 NLS. • The KRRK amino acid residues are indispensable in NLS. • The KRRK motif is conserved in USP22 homologues. - Abstract: Ubiquitin-specific processing enzyme 22 (USP22), a member of the deubiquitinase family, is over-expressed in most human cancers and has been implicated in tumorigenesis. Because it is an enzymatic subunit of the human SAGA transcriptional cofactor, USP22 deubiquitylates histone H2A and H2B in the nucleus, thus participating in gene regulation and cell-cycle progression. However, the mechanisms regulating its nuclear translocation have not yet been elucidated. It was here demonstrated that USP22 is imported into the nucleus through a mechanism mediated by nuclear localization signal (NLS). The bipartite NLS sequence KRELELLKHNPKRRKIT (aa152–168), was identified as the functional NLS for its nuclear localization. Furthermore, a short cluster of basic amino acid residues KRRK within this bipartite NLS plays the primary role in nuclear localization and is evolutionarily conserved in USP22 homologues. In the present study, a functional NLS and the minimal sequences required for the active targeting of USP22 to the nucleus were identified. These findings may provide a molecular basis for the mechanism underlying USP22 nuclear trafficking and function.

  17. Development of a Pilot Program for Human Factors Management in Operating Nuclear Power plants

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jung-Woon; Lee, Yong-Hee; Jang, Tong-Il; Kim, Dae-Ho [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2007-07-01

    The human factors of operating NPPs have been reviewed as a part of Periodic Safety Reviews (PSRs). This human factors PSR covers a wide range of human factors including control room man-machine interfaces (MMIs), procedures, working conditions, qualification, training, information requirements and workload. Korea Atomic Energy Research Institute (KAERI) has performed human factors PSRs from the first PSR for Kori 1. It was determined in 2005 that for a Continuous Operation of the Korean NPPs an enhanced PSR should be performed and issues raised from the PSRs should be resolved. From the results of the PSR for Kori 1, several safety enhancement issues related to human factors were raised. KAERI is working on a resolution of some of the human factors issues for the Korea Hydro and Nuclear Power Co. (KHNP). As a part of the resolution, we are developing a human factors management program (HFMP) for Kori 1. This paper introduces the status of our development of HFMP.

  18. Technology Roadmap Instrumentation, Control, and Human-Machine Interface to Support DOE Advanced Nuclear Energy Programs

    Energy Technology Data Exchange (ETDEWEB)

    Donald D Dudenhoeffer; Burce P Hallbert

    2007-03-01

    Instrumentation, Controls, and Human-Machine Interface (ICHMI) technologies are essential to ensuring delivery and effective operation of optimized advanced Generation IV (Gen IV) nuclear energy systems. In 1996, the Watts Bar I nuclear power plant in Tennessee was the last U.S. nuclear power plant to go on line. It was, in fact, built based on pre-1990 technology. Since this last U.S. nuclear power plant was designed, there have been major advances in the field of ICHMI systems. Computer technology employed in other industries has advanced dramatically, and computing systems are now replaced every few years as they become functionally obsolete. Functional obsolescence occurs when newer, more functional technology replaces or supersedes an existing technology, even though an existing technology may well be in working order.Although ICHMI architectures are comprised of much of the same technology, they have not been updated nearly as often in the nuclear power industry. For example, some newer Personal Digital Assistants (PDAs) or handheld computers may, in fact, have more functionality than the 1996 computer control system at the Watts Bar I plant. This illustrates the need to transition and upgrade current nuclear power plant ICHMI technologies.

  19. Bacterial Artificial Chromosome Clones of Viruses Comprising the Towne Cytomegalovirus Vaccine

    Directory of Open Access Journals (Sweden)

    Xiaohong Cui

    2012-01-01

    Full Text Available Bacterial artificial chromosome (BAC clones have proven invaluable for genetic manipulation of herpesvirus genomes. BAC cloning can also be useful for capturing representative genomes that comprise a viral stock or mixture. The Towne live attenuated cytomegalovirus vaccine was developed in the 1970s by serial passage in cultured fibroblasts. Although its safety, immunogenicity, and efficacy have been evaluated in nearly a thousand human subjects, the vaccine itself has been little studied. Instead, genetic composition and in vitro growth properties have been inferred from studies of laboratory stocks that may not always accurately represent the viruses that comprise the vaccine. Here we describe the use of BAC cloning to define the genotypic and phenotypic properties of viruses from the Towne vaccine. Given the extensive safety history of the Towne vaccine, these BACs provide a logical starting point for the development of next-generation rationally engineered cytomegalovirus vaccines.

  20. Dataset of aqueous humor cytokine profile in HIV patients with Cytomegalovirus (CMV retinitis

    Directory of Open Access Journals (Sweden)

    Jayant Venkatramani Iyer

    2016-09-01

    Full Text Available The data shows the aqueous humor cytokine profiling results acquired in a small cohort of 17 HIV patients clinically diagnosed with Cytomegalovirus retinitis using the FlexMAP 3D (Luminex® platform using the Milliplex Human Cytokine® kit. Aqueous humor samples were collected from these patients at different time points (pre-treatment and at 4-weekly intervals through the 12-week course of intravitreal ganciclovir treatment and 41 cytokine levels were analyzed at each time point. CMV DNA viral load was assessed in 8 patients at different time points throughout the course of ganciclovir treatment. The data described herein is related to the research article entitled “Aqueous humor immune factors and cytomegalovirus (CMV levels in CMV retinitis through treatment - The CRIGSS study” (Iyer et al., 2016 [1]. Cytokine levels against the different time points which indicate the response to the given treatment and against the CMV viral load were analyzed.

  1. Massive alimentary tract bleeding due to cytomegalovirus infection in an elderly patient

    Directory of Open Access Journals (Sweden)

    Bora Koc

    2014-09-01

    Full Text Available In recent years, cytomegalovirus (CMV has been recognized as an important common pathogen in immunocompromized patients. This is due to the increasing number of immunosuppressive medications, intensive cancer chemotherapy use, recurrent transplantations, progressively aging population, and the higher number of human immunodeficiency virus infections. Cytomegalovirus infection especially interests the gastrointestinal tract, anywhere, from the mouth to the anus. Namely, the most commonly affected area is the colon, followed by duodenum, stomach, esophagus and small intestine. The most frequent manifestations of CMV colitis are: diarrhea, fever, gastrointestinal bleeding and abdominal pain. We report here the case of an 82-year-old woman, who was treated for non-Hodgkin lymphoma; she was admitted to the emergency department for abdominal pain and diffuse arthralgia, following massive upper- and lower- gastrointestinal bleeding, due to duodenal and colonic ulcers related to CMV infection.

  2. Congenital Cytomegalovirus Infection in the Absence of Maternal Cytomegalovirus-IgM Antibodies

    NARCIS (Netherlands)

    Gunkel, Julia; van der Knoop, Bloeme J; Nijman, Joppe; De Vries, Linda S.|info:eu-repo/dai/nl/072995408; Manten, Gwendolyn T.R.|info:eu-repo/dai/nl/261633325; Nikkels, Peter G.J.|info:eu-repo/dai/nl/074234692; Murk, Jean Luc; de Vries, Johanna I P; Wolfs, Tom F.W.|info:eu-repo/dai/nl/096507497

    2017-01-01

    Background: Congenital cytomegalovirus (cCMV) infections are the most prevalent intrauterine infections worldwide and are the result of maternal primary or non-primary infections. Early maternal primary infections are thought to carry the highest risk of fetal developmental abnormalities as seen by

  3. A summary of the 2nd workshop on Human Resources Development (HRD) in the nuclear field in Asia. FY2000

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2001-06-01

    The Human Resources Development (HRD) Project was added in 1999 as a Cooperation Activity of 'the Forum for Nuclear Cooperation in Asia (FNCA)' which is organized by Nuclear Committee. The HRD Project supports to solidify the foundation of nuclear development utilization in Asia by promoting human resources development in Asian countries. The principal activity of the HRD Project is to hold the Workshop on Human Resources Development in the Nuclear Field in Asia once a year. The objective of the Workshop is to clarify problems and needs of the human resources development of each country and to support it mutually by exchanging information etc. The report consists of a summary of the 2nd Workshop on Human Resources Development in the Nuclear Field in Asia held on November 27 and 28, 2000 at Tokai Research Establishment of JAERI. (author)

  4. Modeling for climate change in the aspect of nuclear energy priority: Nuclear power energy-based convergence social-humanity analysis

    Energy Technology Data Exchange (ETDEWEB)

    Woo, Tae Ho [Systemix Global Co. Ltd., Seoul, (Korea, Republic of)

    2015-05-15

    Following the industry expansion, the energy consumptions have increased steeply, which have produced the global warming in our lives by carbon production energies. This climate change has provoked significant natural disasters which have damaged to social as well economic matters. Considering the non-carbon production which is the major factor of global warming, nuclear energy is a newly spotlighted source as the green energy source. The climate change factor is affected by the carbon productions made by humans. Then, the nuclear energy increasing rate with the climate change factor affects to the temperature change which is expressed by annual anomaly. Fig. 6 is the protocol for climate change investigation incorporated with the nuclear industry where the climate factor like the temperature is an important index to find out the priority of nuclear energy. The increased environmental pollutions can give the expanding of nuclear energy due to the carbon gas of fossil fuels. This study showed the effectiveness of the nuclear energy by the simulations. The seasonal climate disaster like the very cold winter and very hot summer can increase the necessity of nuclear energy development which could appeal to the general public persons as well as the politicians. So, it is important for the nuclear energy manager to make people understand the importance of the nuclear energy comparing to the oil or coal fuels. The regeneration energy has been considered as the alternative source.

  5. Human performance and its influence on the safety of nuclear installations

    Energy Technology Data Exchange (ETDEWEB)

    Berg, Karl-Heinz; Fechner, Jochen B. [Bundesministerium des Innern, Bonn (Germany)

    2015-05-15

    Nuclear facilities need to be handled in all safety considerations as ''man-machine-system'' as humans considered as liveware are of equally importance ensuring safety as hardware and software. The Federal Minister of the Interior is attempting to give greater focus to this knowledge for the practice of nuclear safety measures. Therefore hardware and software need to be improved according to ergonomic findings and greater importance needs to be issued to the qualification of the operational staff as well as to the qualification of experts consulted by local authorities.

  6. Safety, reliability, risk management and human factors: an integrated engineering approach applied to nuclear facilities

    Energy Technology Data Exchange (ETDEWEB)

    Vasconcelos, Vanderley de; Silva, Eliane Magalhaes Pereira da; Costa, Antonio Carlos Lopes da; Reis, Sergio Carneiro dos [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil)], e-mail: vasconv@cdtn.br, e-mail: silvaem@cdtn.br, e-mail: aclc@cdtn.br, e-mail: reissc@cdtn.br

    2009-07-01

    Nuclear energy has an important engineering legacy to share with the conventional industry. Much of the development of the tools related to safety, reliability, risk management, and human factors are associated with nuclear plant processes, mainly because the public concern about nuclear power generation. Despite the close association between these subjects, there are some important different approaches. The reliability engineering approach uses several techniques to minimize the component failures that cause the failure of the complex systems. These techniques include, for instance, redundancy, diversity, standby sparing, safety factors, and reliability centered maintenance. On the other hand system safety is primarily concerned with hazard management, that is, the identification, evaluation and control of hazards. Rather than just look at failure rates or engineering strengths, system safety would examine the interactions among system components. The events that cause accidents may be complex combinations of component failures, faulty maintenance, design errors, human actions, or actuation of instrumentation and control. Then, system safety deals with a broader spectrum of risk management, including: ergonomics, legal requirements, quality control, public acceptance, political considerations, and many other non-technical influences. Taking care of these subjects individually can compromise the completeness of the analysis and the measures associated with both risk reduction, and safety and reliability increasing. Analyzing together the engineering systems and controls of a nuclear facility, their management systems and operational procedures, and the human factors engineering, many benefits can be realized. This paper proposes an integration of these issues based on the application of systems theory. (author)

  7. Nuclear expression of claudin-3 in human colorectal adenocarcinoma cell lines and tissues.

    Science.gov (United States)

    Tokuhara, Yasunori; Morinishi, Tatsuya; Matsunaga, Toru; Sakai, Manabu; Sakai, Takayoshi; Ohsaki, Hiroyuki; Kadota, Kyuichi; Kushida, Yoshio; Haba, Reiji; Hirakawa, Eiichiro

    2018-01-01

    Claudins are members of a large family of transmembrane proteins, which are essential for the formation of tight junctions and have a significant effect on the biological behavior of tumor progression. Previous studies have demonstrated that several claudins show aberrant expression patterns in numerous types of cancer. The present study investigated the expression and localization of claudin-3 and claudin-7 in human colorectal adenocarcinoma cell lines and tissues. The protein expression levels of claudin-3 and claudin-7 were determined using immunocytochemical and immunohistochemical staining. Claudin-3, but not claudin-7, exhibited nuclear localization in the human colorectal adenocarcinoma Caco-2 and SW620 cell lines. Surgically resected colorectal adenocarcinoma tissue specimens were obtained, and the associations between the expression of claudin-3 or claudin-7 and various clinicopathological parameters were analyzed. The membranous expression rates of claudin-3 and claudin-7 were 58.0 and 50.0%, while their nuclear expression rates were 22.0 and 2.0%, respectively. The membranous expression of claudin-3 and claudin-7 was not associated with any clinicopathological factors, whereas the nuclear expression of claudin-3 was associated with histological type and was significantly increased in colorectal mucinous adenocarcinomas compared with that in well- to moderately-differentiated colorectal adenocarcinomas (P<0.01). However, no associations were observed between the nuclear expression of claudin-7 and any clinicopathological parameter. In conclusion, the nuclear expression of claudin-3 in colorectal mucinous adenocarcinoma may be involved in the biological transformation of tumors. The results from the present study indicated that claudin-3 is an important protein associated with histological type and has potential as a prognostic marker. Although the mechanisms underlying the nuclear localization of claudin-3 in tumorigenesis have not yet been elucidated in

  8. Strongyloides Hyperinfection Syndrome Combined with Cytomegalovirus Infection

    Directory of Open Access Journals (Sweden)

    Fatehi Elnour Elzein

    2016-01-01

    Full Text Available The mortality in Strongyloides hyperinfection syndrome (SHS is alarmingly high. This is particularly common in bone marrow, renal, and other solid organ transplant (SOT patients, where figures may reach up to 50–85%. Immunosuppressives, principally corticosteroids, are the primary triggering factor. In general, the clinical features of Strongyloides stercoralis hyperinfection are nonspecific; therefore, a high index of suspicion is required for early diagnosis and starting appropriate therapy. Although recurrent Gram-negative sepsis and meningitis have been previously reported, the combination of both cytomegalovirus (CMV and strongyloidiasis had rarely been associated. We here describe a patient who survived SHS with recurrent Escherichia coli (E. coli urosepsis and CMV infection.

  9. A HUMAN FACTORS META MODEL FOR U.S. NUCLEAR POWER PLANT CONTROL ROOM MODERNIZATION

    Energy Technology Data Exchange (ETDEWEB)

    Joe, Jeffrey C.

    2017-03-01

    Over the last several years, the United States (U.S.) Department of Energy (DOE) has sponsored human factors research and development (R&D) and human factors engineering (HFE) activities through its Light Water Reactor Sustainability (LWRS) program to modernize the main control rooms (MCR) of commercial nuclear power plants (NPP). Idaho National Laboratory (INL), in partnership with numerous commercial nuclear utilities, has conducted some of this R&D to enable the life extension of NPPs (i.e., provide the technical basis for the long-term reliability, productivity, safety, and security of U.S. NPPs). From these activities performed to date, a human factors meta model for U.S. NPP control room modernization can now be formulated. This paper discusses this emergent HFE meta model for NPP control room modernization, with the goal of providing an integrated high level roadmap and guidance on how to perform human factors R&D and HFE for those in the U.S. nuclear industry that are engaging in the process of upgrading their MCRs.

  10. Establishing a value chain for human factors in nuclear power plantcontrol room modernization

    Energy Technology Data Exchange (ETDEWEB)

    Joe, Jeffrey Clark [Idaho National Laboratory; Thomas, Kenneth David [Idaho National Laboratory; Boring, Ronald Laurids [Idaho National Laboratory

    2015-07-01

    Commercial nuclear power plants in the United States (U.S.) have operated reliably and efficiently for decades. With the life extensions of plants now being planned for operation beyond their original operating licenses, there are opportunities to achieve even greater efficiencies, while maintaining high operational reliabilities, with strategic, risk- and economically-informed, upgrades to plant systems and infrastructure. The U.S. Department of Energy’s Light Water Reactor Sustainability (LWRS) program supports the commercial nuclear industry’s modernization efforts through research and development (R&D) activities across many areas to help establish the technical and economic bases for modernization activities. The Advanced Instrumentation, Information, and Control Systems Technologies pathway is one R&D focus area for the LWRS program, and has researchers at Idaho National Laboratory working with select utility partners to use human factors and instrumentation and controls R&D to help modernize the plant’s main control room. However, some in the nuclear industry have not been as enthusiastic about using human factors R&D to inform life extension decision making. Part of the reason for this may stem from uncertainty decision-makers have regarding how human factors fits into the value chain for nuclear power plant control room modernization. This paper reviews past work that has attempted to demonstrate the value of human factors, and then describes the value chain concept, how it applies to control room modernization, and then makes a case for how and why human factors is an essential link in the modernization value chain.

  11. Handbook of human-reliability analysis with emphasis on nuclear power plant applications. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Swain, A D; Guttmann, H E

    1983-08-01

    The primary purpose of the Handbook is to present methods, models, and estimated human error probabilities (HEPs) to enable qualified analysts to make quantitative or qualitative assessments of occurrences of human errors in nuclear power plants (NPPs) that affect the availability or operational reliability of engineered safety features and components. The Handbook is intended to provide much of the modeling and information necessary for the performance of human reliability analysis (HRA) as a part of probabilistic risk assessment (PRA) of NPPs. Although not a design guide, a second purpose of the Handbook is to enable the user to recognize error-likely equipment design, plant policies and practices, written procedures, and other human factors problems so that improvements can be considered. The Handbook provides the methodology to identify and quantify the potential for human error in NPP tasks.

  12. A development of the Human Factors Assessment Guide for the Study of Erroneous Human Behaviors in Nuclear Power Plants

    Energy Technology Data Exchange (ETDEWEB)

    Oh, Yeon Ju; Lee, Yong Hee; Jang, Tong Il; Kim, Sa Kil [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2014-08-15

    The aim of this paper is to describe a human factors assessment guide for the study of the erroneous characteristic of operators in nuclear power plants (NPPs). We think there are still remaining the human factors issues such as an uneasy emotion, fatigue and stress, varying mental workload situation by digital environment, and various new type of unsafe response to digital interface for better decisions, although introducing an advanced main control room. These human factors issues may not be resolved through the current human reliability assessment which evaluates the total probability of a human error occurring throughout the completion of a specific task. This paper provides an assessment guide for the human factors issues a set of experimental methodology, and presents an assessment case of measurement and analysis especially from neuro physiology approach. It would be the most objective psycho-physiological research technique on human performance for a qualitative analysis considering the safety aspects. This paper can be trial to experimental assessment of erroneous behaviors and their influencing factors, and it can be used as an index for recognition and a method to apply human factors engineering V and V, which is required as a mandatory element of human factor engineering program plan for a NPP design.

  13. Cytomegalovirus Retinitis and the Acquired Immune Deficiency Syndrome: Bench to Bedside: LXVII Edward Jackson Memorial Lecture

    Science.gov (United States)

    Jabs, Douglas A.

    2010-01-01

    Purpose To update information on cytomegalovirus (CMV) retinitis in patients with the acquired immune deficiency syndrome (AIDS) and to integrate information on its pathogenesis and clinical outcomes. Design Literature review. Methods Selected articles from the medical literature, particularly large epidemiologic studies, including the Johns Hopkins Cytomegalovirus Retinitis Cohort Study, the Longitudinal Study of the Ocular Complications of AIDS, and the Cytomegalovirus Retinitis and Viral Resistance Study, were reviewed. Clinical information is discussed in light of knowledge on CMV, its pathogenesis, and its interactions with human immunodeficiency virus (HIV). Results Cytomegalovirus uses several mechanisms to evade the immune system and establish latent infection in immunologically normal hosts. With immune deficiency, such as late-stage AIDS, CMV reactivates, is disseminated to the eye, and establishes a productive infection, resulting in retinal necrosis. HIV and CMV potentiate each other: CMV accelerates HIV disease, and CMV retinitis is associated with increased mortality. Randomized clinical trials have demonstrated the efficacy of treatments for CMV retinitis. Systemically-administered treatment for CMV retinitis decreases AIDS mortality. Highly active antiretroviral therapy (HAART), effectively suppresses HIV replication, resulting in immune recovery, which, if sufficient, controls retinitis without anti-CMV therapy. Resistant CMV, detected in the blood, correlates with resistant virus in the eye and is associated with worse clinical outcomes, including mortality. Host factors, including host genetics and access to care, play a role in the development of CMV retinitis. Conclusions Clinical outcomes of CMV retinitis in patients with AIDS are dependent on characteristics of the virus and host and on HIV–CMV interactions. PMID:21168815

  14. Human-Machine interface for off normal and emergency situations in nuclear power plants

    Energy Technology Data Exchange (ETDEWEB)

    Kwon, Kee Choon [Korea Atomic Energy Research Institute, Taejeon (Korea)

    2000-01-01

    Many nuclear power plants (NPPs) have reported that a high percentage of all major failures in the plants are caused by human errors. Therefore, there has been much focus on elimination of human errors, enhancement of human performance, and general improvement of human machine interface (HMI). Both the utility management and the regulators are demanding improvement in this area. The International Atomic Energy Agency (IAEA) Specialists' Meeting on 'Human-Machine Interface for Off Normal and Emergency Situations in Nuclear Power Plants' was co-organized by the Korea Atomic Energy Research Institute (KAERI) and the Korea Power Engineering Company, INC (KOPEC), and took place in Taejeon, Republic of Korea, 1999 October 26-28. Fifty eight participants, representing nine member countries reviewed recent developments and discussed directions for future efforts in the Human-Machine Interface for Off Normal and Emergency Situations in NPPs. Twenty papers were presented, covering a wide spectrum of technical and scientific subjects including recent experience and benefits from Operational Experience with HMI, Development of HMI System, Licensing Issues for HMI and Future Development and Trends. (Author)

  15. Molecular Image Analysis: Quantitative Description and Classification of the Nuclear Lamina in Human Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Christiaan H. Righolt

    2011-01-01

    Full Text Available The nuclear lamina is an intermediate filament network that provides a structural framework for the cell nucleus. Changes in lamina structure are found during changes in cell fate such as cell division or cell death and are associated with human diseases. An unbiased method that quantifies changes in lamina shape can provide information on cells undergoing changes in cellular functions. We have developed an image processing methodology that finds and quantifies the 3D structure of the nuclear lamina. We show that measurements on such images can be used for cell classification and provide information concerning protein spatial localization in this structure. To demonstrate the efficacy of this method, we compared the lamina of unmanipulated human mesenchymal stem cells (hMSCs at passage 4 to cells activated for apoptosis. A statistically significant classification was found between the two populations.

  16. Molecular image analysis: quantitative description and classification of the nuclear lamina in human mesenchymal stem cells.

    Science.gov (United States)

    Righolt, Christiaan H; Raz, Vered; Vermolen, Bart J; Dirks, Roeland W; Tanke, Hans J; Young, Ian T

    2011-01-01

    The nuclear lamina is an intermediate filament network that provides a structural framework for the cell nucleus. Changes in lamina structure are found during changes in cell fate such as cell division or cell death and are associated with human diseases. An unbiased method that quantifies changes in lamina shape can provide information on cells undergoing changes in cellular functions. We have developed an image processing methodology that finds and quantifies the 3D structure of the nuclear lamina. We show that measurements on such images can be used for cell classification and provide information concerning protein spatial localization in this structure. To demonstrate the efficacy of this method, we compared the lamina of unmanipulated human mesenchymal stem cells (hMSCs) at passage 4 to cells activated for apoptosis. A statistically significant classification was found between the two populations.

  17. Report from the second cytomegalovirus and immunosenescence workshop

    Directory of Open Access Journals (Sweden)

    Wills Mark

    2011-10-01

    Full Text Available Abstract The Second International Workshop on CMV & Immunosenescence was held in Cambridge, UK, 2-4th December, 2010. The presentations covered four separate sessions: cytomegalovirus and T cell phenotypes; T cell memory frequency, inflation and immunosenescence; cytomegalovirus in aging, mortality and disease states; and the immunobiology of cytomegalovirus-specific T cells and effects of the virus on vaccination. This commentary summarizes the major findings of these presentations and references subsequently published work from the presenter laboratory where appropriate and draws together major themes that were subsequently discussed along with new areas of interest that were highlighted by this discussion.

  18. Extraction of human nuclear DNA from feces samples using the QIAamp DNA Stool Mini Kit.

    Science.gov (United States)

    Vandenberg, Nicholas; van Oorschot, Roland A H

    2002-09-01

    The use of a QIAamp DNA Stool Mini Kit (QIAGEN) for extracting human nuclear DNA from feces samples is reported. This method employs a stool lysis buffer and a unique matrix (InhibitEX tablet) to remove PCR inhibitory substances specific to feces samples. DNA extracted from various amounts of stool and from stool samples exposed to different environmental impacts was successfully amplified and typed using the Profiler Plus Amplification Kit and ABI PRISM 310 Genetic Analyser.

  19. Gene-specific repression of proinflammatory cytokines in stimulated human macrophages by nuclear IκBα.

    Science.gov (United States)

    Ghosh, Chandra C; Ramaswami, Sitharam; Juvekar, Ashish; Vu, Hai-Yen; Galdieri, Luciano; Davidson, Dennis; Vancurova, Ivana

    2010-09-15

    We have previously shown that increased nuclear accumulation of IkappaBalpha inhibits NF-kappaB activity and induces apoptosis in human leukocytes. In this study, we wanted to explore the possibility that the nucleocytoplasmic distribution of IkappaBalpha can be used as a therapeutic target for the regulation of NF-kappaB-dependent cytokine synthesis. Treatment of LPS-stimulated human U937 macrophages with an inhibitor of chromosome region maintenance 1-dependent nuclear export, leptomycin B, resulted in the increased nuclear accumulation of IkappaBalpha and inhibition of NF-kappaB DNA binding activity, caused by the nuclear IkappaBalpha-p65 NF-kappaB interaction. Surprisingly, however, whereas mRNA expression and cellular release of TNF-alpha, the beta form of pro-IL-1 (IL-1beta), and IL-6 were inhibited by the leptomycin B-induced nuclear IkappaBalpha, IL-8 mRNA expression and cellular release were not significantly affected. Analysis of in vivo recruitment of p65 NF-kappaB to NF-kappaB-regulated promoters by chromatin immunoprecipitation in U937 cells and human PBMCs indicated that although the p65 recruitment to TNF-alpha, IL-1beta, and IL-6 promoters was inhibited by the nuclear IkappaBalpha, p65 recruitment to IL-8 promoter was not repressed. Chromatin immunoprecipitation analyses using IkappaBalpha and S536 phosphospecific p65 NF-kappaB Abs demonstrated that although the newly synthesized IkappaBalpha induced by postinduction repression is recruited to TNF-alpha, IL-1beta, and IL-6 promoters but not to the IL-8 promoter, S536-phosphorylated p65 is recruited to IL-8 promoter, but not to TNF-alpha, IL-1beta, or IL-6 promoters. Together, these data indicate that the inhibition of NF-kappaB-dependent transcription by nuclear IkappaBalpha in LPS-stimulated macrophages is gene specific and depends on the S536 phosphorylation status of the recruited p65 NF-kappaB.

  20. Letermovir for the management of cytomegalovirus infection.

    Science.gov (United States)

    Bowman, Lyndsey J; Melaragno, Jennifer Iuppa; Brennan, Daniel C

    2017-02-01

    Cytomegalovirus (CMV) is a major cause of morbidity and mortality in immunocompromised patients. Available antivirals are fraught with adverse effects and risk for the development of CMV resistance. Letermovir is a novel antiviral in the late stages of drug development for the treatment and prevention of CMV. Areas covered: A MEDLINE search of the MeSH terms 'letermovir,' 'cytomegalovirus,' 'hematopoietic stem cell transplant,' and 'solid organ transplant,' was last conducted on 15 August 2016. Articles were selected on the basis of their contribution to current knowledge about letermovir. Expert opinion: Letermovir's mechanism of action, pharmacokinetic and pharmacodynamic profile, and favorable efficacy and safety make it an attractive option for both the prevention and treatment of CMV in immunocompromised patients. The lack of cross-resistance with other antivirals and the absence of myelosuppression are two prominent characteristics of letermovir that could support broad use of this product following FDA-approval. One major limitation is its lack of activity against other herpesviruses, which are commonly seen in immunocompromised hosts. We believe that with additional clinical efficacy data, this medication could emerge as a primary option for the prevention and treatment of CMV in the immunocompromised patient population.

  1. Contribution to the human society from the nuclear science and technology

    Energy Technology Data Exchange (ETDEWEB)

    Matsuura, Shojiro [Japan Atomic Energy Research Inst., Tokyo (Japan)

    1999-09-01

    All of us living on this planet feel a hearty gratitude for our being endowed with natural blessings like sunshine, atmosphere, water, green of the mountains and blue of the ocean, etc. From the same point of view nuclear power and radiation are also precious blessings from the nature. To begin with, sunshine originates from the thermonuclear reactions in the sun, and a considerable portion of geothermal energy is assumed to be from natural radioactivity. The effects of natural radiation onto the evolution of life are considered as immeasurably great. The creation of this universe is, in the first place, thought to owe to certain nuclear reactions. The process of the nuclear reaction or radiation itself cannot be perceived by human senses and feeling such as eyesight or hearing. In order to recognize them we must possess powers of understanding, or intelligence, as well as detectors of the specific purpose. However, this may have caused among people the feelings of alienation and fear. Some can be said for cases of bacteria, virus, electricity, and many others. There seems to be good grounds to say that the greatest characteristic of the modern civilization is that it has evolved, so far, the quality of human life adopting what man can recognize by means of intelligence and detectors' combination, in addition to his senses and feelings. Typical examples of this are radioactivity and radiation both of which were discovered in the end of the 19th century and, provoked by this, the nuclear physics achieved an immense progress in consequence. Based on these, the nuclear science and technology have been developed with a giant step and exerted their powerful influence on all over the world in this century. This characteristic is supposed to permeate into the human society of the 21st century more widely and deeply. The nuclear science and technology have become to play a significant role in science research, as an energy source and in industry and medicine. In the

  2. Production of human CD59-transgenic pigs by embryonic germ cell nuclear transfer

    Energy Technology Data Exchange (ETDEWEB)

    Ahn, Kwang Sung; Won, Ji Young [Department of Physiology, Dankook University School of Medicine, Cheonan (Korea, Republic of); Park, Jin-Ki [Animal Biotechnology Division, National Institute of Animal Science, Suwon (Korea, Republic of); Sorrell, Alice M. [Department of Physiology, Dankook University School of Medicine, Cheonan (Korea, Republic of); Heo, Soon Young; Kang, Jee Hyun [Department of Nanobiomedical Science, Dankook University, Cheonan (Korea, Republic of); Woo, Jae-Seok [Animal Biotechnology Division, National Institute of Animal Science, Suwon (Korea, Republic of); Choi, Bong-Hwan [Genomics and Bioinformatics Division, National Institute of Animal Science, Suwon (Korea, Republic of); Chang, Won-Kyong [Animal Biotechnology Division, National Institute of Animal Science, Suwon (Korea, Republic of); Shim, Hosup, E-mail: shim@dku.edu [Department of Nanobiomedical Science, Dankook University, Cheonan (Korea, Republic of); Institute of Tissue Regeneration Engineering, Dankook University, Cheonan (Korea, Republic of)

    2010-10-01

    Research highlights: {yields} Human CD59 (hCD59) gene was introduced into porcine embryonic germ (EG) cells. {yields} hCD59-transgenic EG cells were resistant to hyperacute rejection in cytolytic assay. {yields} hCD59-transgenic pigs were produced by EG cell nuclear transfer. -- Abstract: This study was performed to produce transgenic pigs expressing the human complement regulatory protein CD59 (hCD59) using the nuclear transfer (NT) of embryonic germ (EG) cells, which are undifferentiated stem cells derived from primordial germ cells. Because EG cells can be cultured indefinitely in an undifferentiated state, they may provide an inexhaustible source of nuclear donor cells for NT to produce transgenic pigs. A total of 1980 NT embryos derived from hCD59-transgenic EG cells were transferred to ten recipients, resulting in the birth of fifteen piglets from three pregnancies. Among these offspring, ten were alive without overt health problems. Based on PCR analysis, all fifteen piglets were confirmed as hCD59 transgenic. The expression of the hCD59 transgene in the ten living piglets was verified by RT-PCR. Western analysis showed the expression of the hCD59 protein in four of the ten RT-PCR-positive piglets. These results demonstrate that hCD59-transgenic pigs could effectively be produced by EG cell NT and that such transgenic pigs may be used as organ donors in pig-to-human xenotransplantation.

  3. Measuring Human Performance in Simulated Nuclear Power Plant Control Rooms Using Eye Tracking

    Energy Technology Data Exchange (ETDEWEB)

    Kovesdi, Casey Robert [Idaho National Lab. (INL), Idaho Falls, ID (United States); Rice, Brandon Charles [Idaho National Lab. (INL), Idaho Falls, ID (United States); Bower, Gordon Ross [Idaho National Lab. (INL), Idaho Falls, ID (United States); Spielman, Zachary Alexander [Idaho National Lab. (INL), Idaho Falls, ID (United States); Hill, Rachael Ann [Idaho National Lab. (INL), Idaho Falls, ID (United States); LeBlanc, Katya Lee [Idaho National Lab. (INL), Idaho Falls, ID (United States)

    2015-11-01

    Control room modernization will be an important part of life extension for the existing light water reactor fleet. As part of modernization efforts, personnel will need to gain a full understanding of how control room technologies affect performance of human operators. Recent advances in technology enables the use of eye tracking technology to continuously measure an operator’s eye movement, which correlates with a variety of human performance constructs such as situation awareness and workload. This report describes eye tracking metrics in the context of how they will be used in nuclear power plant control room simulator studies.

  4. Spontaneous calcium oscillations and nuclear PLC-beta1 in human GV oocytes.

    Science.gov (United States)

    Miyara, F; Pesty, A; Migne, C; Djediat, C; Huang, X B; Dumont-Hassan, M; Debey, P; Lefèvre, B

    2008-02-01

    Our aim was to investigate if human oocytes, like mouse oocytes, exhibit spontaneous Ca(2+) oscillations and nuclear translocation of PLC-beta1 prior to germinal vesicle breakdown (GVBD), and to correlate these events with the evolution of chromatin configuration as a landmark for the meiosis resumption kinetics. Human germinal vesicle (GV) oocytes were either loaded with Fluo-3 probe to record Ca(2+) signals or fixed for subsequent fluorescent labeling of both chromatin and PLC-beta1, and immunogold labeling of PLC-beta1. Here for the first time, we show that human oocytes at the GV-stage exhibit spontaneous Ca(2+) oscillations. Interestingly, only oocytes with a large diameter and characterized by a compact chromatin surrounding the nucleolus of the GV could reveal these kind of oscillations. We also observed a translocation of PLC-beta1 from the cytoplasm towards the nucleus during in vitro maturation of human oocytes. Spontaneous calcium oscillations and nuclear translocation of PLC-beta1 may reflect some degree of oocyte maturity. The impact of our results may be very helpful to understand and resolve many enigmatic problems usually encountered during the in vitro meiotic maturation of human GV oocytes.

  5. Review of Methods Related to Assessing Human Performance in Nuclear Power Plant Control Room Simulations

    Energy Technology Data Exchange (ETDEWEB)

    Katya L Le Blanc; Ronald L Boring; David I Gertman

    2001-11-01

    With the increased use of digital systems in Nuclear Power Plant (NPP) control rooms comes a need to thoroughly understand the human performance issues associated with digital systems. A common way to evaluate human performance is to test operators and crews in NPP control room simulators. However, it is often challenging to characterize human performance in meaningful ways when measuring performance in NPP control room simulations. A review of the literature in NPP simulator studies reveals a variety of ways to measure human performance in NPP control room simulations including direct observation, automated computer logging, recordings from physiological equipment, self-report techniques, protocol analysis and structured debriefs, and application of model-based evaluation. These methods and the particular measures used are summarized and evaluated.

  6. Crystal structure of cytomegalovirus IE1 protein reveals targeting of TRIM family member PML via coiled-coil interactions.

    Directory of Open Access Journals (Sweden)

    Myriam Scherer

    2014-11-01

    Full Text Available PML nuclear bodies (PML-NBs are enigmatic structures of the cell nucleus that act as key mediators of intrinsic immunity against viral pathogens. PML itself is a member of the E3-ligase TRIM family of proteins that regulates a variety of innate immune signaling pathways. Consequently, viruses have evolved effector proteins to modify PML-NBs; however, little is known concerning structure-function relationships of viral antagonists. The herpesvirus human cytomegalovirus (HCMV expresses the abundant immediate-early protein IE1 that colocalizes with PML-NBs and induces their dispersal, which correlates with the antagonization of NB-mediated intrinsic immunity. Here, we delineate the molecular basis for this antagonization by presenting the first crystal structure for the evolutionary conserved primate cytomegalovirus IE1 proteins. We show that IE1 consists of a globular core (IE1CORE flanked by intrinsically disordered regions. The 2.3 Å crystal structure of IE1CORE displays an all α-helical, femur-shaped fold, which lacks overall fold similarity with known protein structures, but shares secondary structure features recently observed in the coiled-coil domain of TRIM proteins. Yeast two-hybrid and coimmunoprecipitation experiments demonstrate that IE1CORE binds efficiently to the TRIM family member PML, and is able to induce PML deSUMOylation. Intriguingly, this results in the release of NB-associated proteins into the nucleoplasm, but not of PML itself. Importantly, we show that PML deSUMOylation by IE1CORE is sufficient to antagonize PML-NB-instituted intrinsic immunity. Moreover, co-immunoprecipitation experiments demonstrate that IE1CORE binds via the coiled-coil domain to PML and also interacts with TRIM5α We propose that IE1CORE sequesters PML and possibly other TRIM family members via structural mimicry using an extended binding surface formed by the coiled-coil region. This mode of interaction might render the antagonizing activity less

  7. Neurochemical dynamics of acute orofacial pain in the human trigeminal brainstem nuclear complex.

    Science.gov (United States)

    de Matos, Nuno M P; Hock, Andreas; Wyss, Michael; Ettlin, Dominik A; Brügger, Mike

    2017-09-04

    The trigeminal brainstem sensory nuclear complex is the first central relay structure mediating orofacial somatosensory and nociceptive perception. Animal studies suggest a substantial involvement of neurochemical alterations at such basal CNS levels in acute and chronic pain processing. Translating this animal based knowledge to humans is challenging. Human related examining of brainstem functions are challenged by MR related peculiarities as well as applicability aspects of experimentally standardized paradigms. Based on our experience with an MR compatible human orofacial pain model, the aims of the present study were twofold: 1) from a technical perspective, the evaluation of proton magnetic resonance spectroscopy at 3 T regarding measurement accuracy of neurochemical profiles in this small brainstem nuclear complex and 2) the examination of possible neurochemical alterations induced by an experimental orofacial pain model. Data from 13 healthy volunteers aged 19-46 years were analyzed and revealed high quality spectra with significant reductions in total N-acetylaspartate (N-acetylaspartate + N-acetylaspartylglutamate) (-3.7%, p = 0.009) and GABA (-10.88%, p = 0.041) during the pain condition. These results might reflect contributions of N-acetylaspartate and N-acetylaspartylglutamate in neuronal activity-dependent physiologic processes and/or excitatory neurotransmission, whereas changes in GABA might indicate towards a reduction in tonic GABAergic functioning during nociceptive signaling. Summarized, the present study indicates the applicability of (1)H-MRS to obtain neurochemical dynamics within the human trigeminal brainstem sensory nuclear complex. Further developments are needed to pave the way towards bridging important animal based knowledge with human research to understand the neurochemistry of orofacial nociception and pain. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Human Resource Managements as a part of the Human Factors Management Program(HFMP) for Nuclear Power Plants

    Energy Technology Data Exchange (ETDEWEB)

    Kim, DaeHo; Lee, YongHee; Lee, JungWoon [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of); Kim, Younggab [Korea Hydro and Nuclear Power Co. LTD, Daejeon (Korea, Republic of)

    2007-10-15

    Programs for the effective implementation and management of human factor issues in nuclear power plants (NPPs) should contain technical criteria, an establishment of a job process, and activities for job improvements and be a system through which human factors can be managed in an integrated way. Human factors to be managed should include those related to an operation of plants as well as those related to a plant design as mentioned in NUREG-0800(2004), NUREG- 0711(2004), and NUREG-0700(2002). Human factor items to be managed for an operation of plants are listed in the PSR (Periodic Safety Review) items defined in the Enforcement of Regulation of the Atomic Energy Act. They are procedures, a work management system including a shift work management, a qualification management of plant personnel, training, a work amount assessment, a MMI (Man Machine Interface), and the use of experience. Among these factors, factors related to a human resource management include work management systems and the status of a work management including shift work, a qualification management ensuring qualified workers on duty at all times, and the systems for and the status of training and education. This paper addresses the scope of a human resource management, guidelines and procedures to be developed for a human resource management, and considerations critical in the development of guidelines and procedures.

  9. Cytomegalovirus as a cause of anterior uveitis in immunocompetent patients

    NARCIS (Netherlands)

    van Boxtel, Lonneke A. A.; van der Lelij, Allegonda; van der Meer, Johannes; Los, Leonoor I.

    Purpose: To describe 7 cases of unilateral, chronic and/or recurrent anterior uveitis caused by cytomegalovirus (CMV) in immunocompetent patients; to identify specific ophthalmologic characteristics; and to evaluate the clinical effect of valganciclovir treatment. Design: Retrospective observational

  10. Cytomegalovirus, inflammatory bowel disease, and anti-TNFα.

    Science.gov (United States)

    Campos, Sara T; Portela, Francisco A; Tomé, Luís

    2017-05-01

    Anti-TNFα agents emerged in inflammatory bowel disease (IBD) as an effective option in situations that, otherwise, would be refractory to medical therapy. Cytomegalovirus infection may present with a high spectrum of manifestations and lead to high morbidity and mortality. However, its clinical significance in IBD course remains unknown and data on its association with anti-TNFα are limited. This study aims to evaluate cytomegalovirus (CMV) infection/disease in patients with IBD treated with anti-TNFα; if possible, possible risk factors associated with CMV infection/disease in IBD patients under anti-TNFα as well as the influence of CMV infection/disease in IBD course would be determined. During three consecutive years, all IBD patients starting infliximab in our department were included. Cytomegalovirus status before anti-TNFα was evaluated. Data regarding IBD, therapeutic and IBD course after infliximab, were recorded. CMV analysis was performed with polymerase chain reaction (PCR)-cytomegalovirus in peripheral blood and colonoscopy with biopsies (histopathology/immunohistochemistry). We included 29 patients: female-83%; Crohn's disease-51.8%, ulcerative colitis-44.8%, non-classified colitis-3.4%; 23 cytomegalovirus seropositive. Median follow-up: 19 months (3-36). During follow-up, 14 patients were under combination therapy with azathioprine and 5 did at least 1 cycle of corticosteroids. Twenty-one patients responded to infliximab. We registered 8 exacerbations of IBD. Four patients discontinued infliximab: none had CMV infection. We documented 1 case of intestinal cytomegalovirus infection-detected in biopsies performed per protocol in an asymptomatic UC patient, who responded to valganciclovir without infliximab discontinuation. Infliximab, with/without immunosuppression, does not confer an increased risk of (re)activation of cytomegalovirus. Cytomegalovirus was not responsible neither for significant morbidity nor mortality in IBD.

  11. Symptomatic Congenital Cytomegalovirus Infection Is Underdiagnosed in British Columbia.

    Science.gov (United States)

    Sorichetti, Brendan; Goshen, Oran; Pauwels, Julie; Kozak, Frederick K; Tilley, Peter; Krajden, Mel; Gantt, Soren

    2016-02-01

    Records were reviewed from all infants tested for congenital cytomegalovirus infection in British Columbia, Canada from 2006 to June 2014. Fourteen of 701 infants, or approximately 4.2 per 100,000 live births, had a positive test, indicating that >90% of expected symptomatic congenital cytomegalovirus infection cases were not diagnosed using clinician-initiated testing. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. Controlling Cytomegalovirus: Helping the Immune System Take the Lead

    Directory of Open Access Journals (Sweden)

    Patrick J. Hanley

    2014-05-01

    Full Text Available Cytomegalovirus, of the Herpesviridae family, has evolved alongside humans for thousands of years with an intricate balance of latency, immune evasion, and transmission. While upwards of 70% of humans have evidence of CMV infection, the majority of healthy people show little to no clinical symptoms of primary infection and CMV disease is rarely observed during persistent infection in immunocompetent hosts. Despite the fact that the majority of infected individuals are asymptomatic, immunologically, CMV hijacks the immune system by infecting and remaining latent in antigen-presenting cells that occasionally reactivate subclinically and present antigen to T cells, eventually causing the inflation of CMV-specific T cells until they can compromise up to 10% of the entire T cell repertoire. Because of this impact on the immune system, as well as its importance in fields such as stem cell and organ transplant, the relationship between CMV and the immune response has been studied in depth. Here we provide a review of many of these studies and insights into how CMV-specific T cells are currently being used therapeutically.

  13. Controlling Cytomegalovirus: Helping the Immune System Take the Lead

    Science.gov (United States)

    Hanley, Patrick J.; Bollard, Catherine M.

    2014-01-01

    Cytomegalovirus, of the Herpesviridae family, has evolved alongside humans for thousands of years with an intricate balance of latency, immune evasion, and transmission. While upwards of 70% of humans have evidence of CMV infection, the majority of healthy people show little to no clinical symptoms of primary infection and CMV disease is rarely observed during persistent infection in immunocompetent hosts. Despite the fact that the majority of infected individuals are asymptomatic, immunologically, CMV hijacks the immune system by infecting and remaining latent in antigen-presenting cells that occasionally reactivate subclinically and present antigen to T cells, eventually causing the inflation of CMV-specific T cells until they can compromise up to 10% of the entire T cell repertoire. Because of this impact on the immune system, as well as its importance in fields such as stem cell and organ transplant, the relationship between CMV and the immune response has been studied in depth. Here we provide a review of many of these studies and insights into how CMV-specific T cells are currently being used therapeutically. PMID:24872114

  14. Results of a nuclear power plant application of A New Technique for Human Error Analysis (ATHEANA)

    Energy Technology Data Exchange (ETDEWEB)

    Whitehead, D.W.; Forester, J.A. [Sandia National Labs., Albuquerque, NM (United States); Bley, D.C. [Buttonwood Consulting, Inc. (United States)] [and others

    1998-03-01

    A new method to analyze human errors has been demonstrated at a pressurized water reactor (PWR) nuclear power plant. This was the first application of the new method referred to as A Technique for Human Error Analysis (ATHEANA). The main goals of the demonstration were to test the ATHEANA process as described in the frame-of-reference manual and the implementation guideline, test a training package developed for the method, test the hypothesis that plant operators and trainers have significant insight into the error-forcing-contexts (EFCs) that can make unsafe actions (UAs) more likely, and to identify ways to improve the method and its documentation. A set of criteria to evaluate the success of the ATHEANA method as used in the demonstration was identified. A human reliability analysis (HRA) team was formed that consisted of an expert in probabilistic risk assessment (PRA) with some background in HRA (not ATHEANA) and four personnel from the nuclear power plant. Personnel from the plant included two individuals from their PRA staff and two individuals from their training staff. Both individuals from training are currently licensed operators and one of them was a senior reactor operator on shift until a few months before the demonstration. The demonstration was conducted over a 5-month period and was observed by members of the Nuclear Regulatory Commission`s ATHEANA development team, who also served as consultants to the HRA team when necessary. Example results of the demonstration to date, including identified human failure events (HFEs), UAs, and EFCs are discussed. Also addressed is how simulator exercises are used in the ATHEANA demonstration project.

  15. Acetylation dynamics of human nuclear proteins during the ionizing radiation-induced DNA damage response

    DEFF Research Database (Denmark)

    Bennetzen, Martin V; Larsen, Dorthe Helena; Dinant, Christoffel

    2013-01-01

    -dependent posttranslational modifications (PTMs). To complement our previous analysis of IR-induced temporal dynamics of nuclear phosphoproteome, we now identify a range of human nuclear proteins that are dynamically regulated by acetylation, and predominantly deacetylation, during IR-induced DDR by using mass spectrometry...... to genotoxic insults. Overall, these results present a resource of temporal profiles of a spectrum of protein acetylation sites during DDR and provide further insights into the highly dynamic nature of regulatory PTMs that help orchestrate the maintenance of genome integrity.......Genotoxic insults, such as ionizing radiation (IR), cause DNA damage that evokes a multifaceted cellular DNA damage response (DDR). DNA damage signaling events that control protein activity, subcellular localization, DNA binding, protein-protein interactions, etc. rely heavily on time...

  16. The history of vaccination against cytomegalovirus.

    Science.gov (United States)

    Plotkin, Stanley

    2015-06-01

    Cytomegalovirus vaccine development started in the 1970s with attenuated strains. In the 1980s, one of the strains was shown to be safe and effective in renal transplant patients. Then, attention switched to glycoprotein gB, which was shown to give moderate but transient protection against acquisition of the virus by women. The identification of the pp65 tegument protein as the principal target of cellular immune responses resulted in new approaches, particularly DNA, plasmids to protect hematogenous stem cell recipients. The subsequent discovery of the pentameric protein complex that generates most neutralizing antibodies led to efforts to incorporate that complex into vaccines. At this point, there are many candidate CMV vaccines, including live recombinants, replication-defective virus, DNA plasmids, soluble pentameric proteins, peptides, virus-like particles and vectored envelope proteins.

  17. [Respiratory infections associated with a cytomegalovirus].

    Science.gov (United States)

    Calicó, I; Moraga Llop, F A; Español, T; Bertrán Sangués, J M; Fernández Pérez, F

    1985-11-15

    We report 27 children with respiratory tract disease in whom cytomegalovirus was isolated. These group excludes transplant patients and those on hemodialysis and mononucleosis. At the time of virus studies 7 had pneumonia, 3 chronic bronchopneumopathy, 2 bronchopneumonia, 8 pertussoid syndrome, 6 bronchitis or bronchiolitis and 1 laryngitis with glottic oedema. Virus studies consisted in cell cultures of biological products (pharyngeal exudates and urine). They were positive in 18 pharyngeal exudates, 24 urines, 2 bronchial brushings and 1 bronchoaspiration. In only 5 patients a complete serologic study was performed, with 3 seroconversions and one case of persistent high titers. Three patients had severe immune disease (2 hipogammaglobulinemias) and 1 dysgammaglobulinemia. These findings are discussed.

  18. Pathological aggression in "fierce" mice corrected by human nuclear receptor 2E1.

    Science.gov (United States)

    Abrahams, Brett S; Kwok, Melvin C H; Trinh, Eric; Budaghzadeh, Saeed; Hossain, Sazzad M; Simpson, Elizabeth M

    2005-07-06

    "Fierce" mice, homozygous for the deletion of nuclear receptor 2E1 (NR2E1), show abnormal brain-eye development and pathological aggression. To evaluate functional equivalency between mouse and human NR2E1, we generated mice transgenic for a genomic clone spanning the human NR2E1 locus and bred these animals to fierce mice deleted for the corresponding mouse gene. In fierce mutants carrying human NR2E1, structural brain defects were eliminated and eye abnormalities ameliorated. Excitingly, behavior in these "rescue" mice was indistinguishable from controls. Because no artificial promoter was used to drive transgene expression, promoter and regulatory elements within the human NR2E1 clone are functional in mouse. Normal behavior in rescue animals suggests that mechanisms underlying the behavioral abnormalities in fierce mice may also be conserved in humans. Our data support the hypothesis that variation at NR2E1 may contribute to human behavioral disorders. Use of this rescue paradigm with other genes will permit the direct evaluation of human genes hypothesized to play a causal role in psychiatric disease but for which evidence is lacking or equivocal.

  19. The Relationship between Human Operators' Psycho-physiological Condition and Human Errors in Nuclear Power Plants

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Arryum; Jang, Inseok; Kang, Hyungook; Seong, Poonghyun [Korea Advanced Institute of Science and Technology, Daejeon (Korea, Republic of)

    2013-05-15

    The safe operation of nuclear power plants (NPPs) is substantially dependent on the performance of the human operators who operate the systems. In this environment, human errors caused by inappropriate performance of operator have been considered to be critical since it may lead serious problems in the safety-critical plants. In order to provide meaningful insights to prevent human errors and enhance the human performance, operators' physiological conditions such as stress and workload have been investigated. Physiological measurements were considered as reliable tools to assess the stress and workload. T. Q. Tran et al. and J. B. Brooking et al pointed out that operators' workload can be assessed using eye tracking, galvanic skin response, electroencephalograms (EEGs), heart rate, respiration and other measurements. The purpose of this study is to investigate the effect of the human operators' tense level and knowledge level to the number of human errors. For this study, the experiments were conducted in the mimic of the main control rooms (MCR) in NPP. It utilized the compact nuclear simulator (CNS) which is modeled based on the three loop Pressurized Water Reactor, 993MWe, Kori unit 3 and 4 in Korea and the subjects were asked to follow the tasks described in the emergency operating procedures (EOP). During the simulation, three kinds of physiological measurement were utilized; Electrocardiogram (ECG), EEG and nose temperature. Also, subjects were divided into three groups based on their knowledge of the plant operation. The result shows that subjects who are tense make fewer errors. In addition, subjects who are in higher knowledge level tend to be tense and make fewer errors. For the ECG data, subjects who make fewer human errors tend to be located in higher tense level area of high SNS activity and low PSNS activity. The results of EEG data are also similar to ECG result. Beta power ratio of subjects who make fewer errors was higher. Since beta

  20. Structure of a herpesvirus nuclear egress complex subunit reveals an interaction groove that is essential for viral replication

    Science.gov (United States)

    Leigh, Kendra E.; Sharma, Mayuri; Mansueto, My Sam; Boeszoermenyi, Andras; Filman, David J.; Hogle, James M.; Wagner, Gerhard; Coen, Donald M.; Arthanari, Haribabu

    2015-01-01

    Herpesviruses require a nuclear egress complex (NEC) for efficient transit of nucleocapsids from the nucleus to the cytoplasm. The NEC orchestrates multiple steps during herpesvirus nuclear egress, including disruption of nuclear lamina and particle budding through the inner nuclear membrane. In the important human pathogen human cytomegalovirus (HCMV), this complex consists of nuclear membrane protein UL50, and nucleoplasmic protein UL53, which is recruited to the nuclear membrane through its interaction with UL50. Here, we present an NMR-determined solution-state structure of the murine CMV homolog of UL50 (M50; residues 1–168) with a strikingly intricate protein fold that is matched by no other known protein folds in its entirety. Using NMR methods, we mapped the interaction of M50 with a highly conserved UL53-derived peptide, corresponding to a segment that is required for heterodimerization. The UL53 peptide binding site mapped onto an M50 surface groove, which harbors a large cavity. Point mutations of UL50 residues corresponding to surface residues in the characterized M50 heterodimerization interface substantially decreased UL50–UL53 binding in vitro, eliminated UL50–UL53 colocalization, prevented disruption of nuclear lamina, and halted productive virus replication in HCMV-infected cells. Our results provide detailed structural information on a key protein–protein interaction involved in nuclear egress and suggest that NEC subunit interactions can be an attractive drug target. PMID:26150520

  1. Spent nuclear fuel project, Cold Vacuum Drying Facility human factors engineering (HFE) analysis: Results and findings

    Energy Technology Data Exchange (ETDEWEB)

    Garvin, L.J.

    1998-07-17

    This report presents the background, methodology, and findings of a human factors engineering (HFE) analysis performed in May, 1998, of the Spent Nuclear Fuels (SNF) Project Cold Vacuum Drying Facility (CVDF), to support its Preliminary Safety Analysis Report (PSAR), in responding to the requirements of Department of Energy (DOE) Order 5480.23 (DOE 1992a) and drafted to DOE-STD-3009-94 format. This HFE analysis focused on general environment, physical and computer workstations, and handling devices involved in or directly supporting the technical operations of the facility. This report makes no attempt to interpret or evaluate the safety significance of the HFE analysis findings. The HFE findings presented in this report, along with the results of the CVDF PSAR Chapter 3, Hazards and Accident Analyses, provide the technical basis for preparing the CVDF PSAR Chapter 13, Human Factors Engineering, including interpretation and disposition of findings. The findings presented in this report allow the PSAR Chapter 13 to fully respond to HFE requirements established in DOE Order 5480.23. DOE 5480.23, Nuclear Safety Analysis Reports, Section 8b(3)(n) and Attachment 1, Section-M, require that HFE be analyzed in the PSAR for the adequacy of the current design and planned construction for internal and external communications, operational aids, instrumentation and controls, environmental factors such as heat, light, and noise and that an assessment of human performance under abnormal and emergency conditions be performed (DOE 1992a).

  2. 7SK small nuclear RNA transcription level down-regulates in human tumors and stem cells.

    Science.gov (United States)

    Abasi, Mozhgan; Bazi, Zahra; Mohammadi-Yeganeh, Samira; Soleimani, Masoud; Haghpanah, Vahid; Zargami, Nosratollah; Ghanbarian, Hossein

    2016-11-01

    The small nuclear noncoding RNA (snRNA) 7SK is a highly conserved noncoding RNA of 331 nucleotides in animals, which is present in a nuclear ribonucleoprotein complex with proteins such as methylphosphate capping enzyme (MePCE), hexamethylene bisacetamide-inducible proteins 1 and 2 (HEXIM1 and HEXIM2) and La-related protein 7 (Larp7). Regulating the activity of the positive transcription elongation factor b (P-TEFb) is the key function of 7SK noncoding RNA. Recently, we have shown that 7SK snRNA over-expression reduces human embryonic kidney 293T cell line viability. Here, we attempt to monitor the expression level of 7SK snRNA in different human cell lines and cancer tissues. Examination of 7SK transcription either in cell lines or in different malignant tissues including blood (CML), breast and colon showed that 7SK expression significantly down-regulated in cancer. Similar to human cancer tissues and cell lines, 7SK transcriptional level decreased in stem cells in comparison with differentiated cell types. In this regard, over-expression of 7SK snRNA might be a powerful tool for blocking cancer progression by controlling the activity of P-TEFb.

  3. A Function-Behavior-State Approach to Designing Human Machine Interface for Nuclear Power Plant Operators

    Science.gov (United States)

    Lin, Y.; Zhang, W. J.

    2005-02-01

    This paper presents an approach to human-machine interface design for control room operators of nuclear power plants. The first step in designing an interface for a particular application is to determine information content that needs to be displayed. The design methodology for this step is called the interface design framework (called framework ). Several frameworks have been proposed for applications at varying levels, including process plants. However, none is based on the design and manufacture of a plant system for which the interface is designed. This paper presents an interface design framework which originates from design theory and methodology for general technical systems. Specifically, the framework is based on a set of core concepts of a function-behavior-state model originally proposed by the artificial intelligence research community and widely applied in the design research community. Benefits of this new framework include the provision of a model-based fault diagnosis facility, and the seamless integration of the design (manufacture, maintenance) of plants and the design of human-machine interfaces. The missing linkage between design and operation of a plant was one of the causes of the Three Mile Island nuclear reactor incident. A simulated plant system is presented to explain how to apply this framework in designing an interface. The resulting human-machine interface is discussed; specifically, several fault diagnosis examples are elaborated to demonstrate how this interface could support operators' fault diagnosis in an unanticipated situation.

  4. Updating Human Factors Engineering Guidelines for Conducting Safety Reviews of Nuclear Power Plants

    Energy Technology Data Exchange (ETDEWEB)

    O, J.M.; Higgins, J.; Stephen Fleger - NRC

    2011-09-19

    The U.S. Nuclear Regulatory Commission (NRC) reviews the human factors engineering (HFE) programs of applicants for nuclear power plant construction permits, operating licenses, standard design certifications, and combined operating licenses. The purpose of these safety reviews is to help ensure that personnel performance and reliability are appropriately supported. Detailed design review procedures and guidance for the evaluations is provided in three key documents: the Standard Review Plan (NUREG-0800), the HFE Program Review Model (NUREG-0711), and the Human-System Interface Design Review Guidelines (NUREG-0700). These documents were last revised in 2007, 2004 and 2002, respectively. The NRC is committed to the periodic update and improvement of the guidance to ensure that it remains a state-of-the-art design evaluation tool. To this end, the NRC is updating its guidance to stay current with recent research on human performance, advances in HFE methods and tools, and new technology being employed in plant and control room design. This paper describes the role of HFE guidelines in the safety review process and the content of the key HFE guidelines used. Then we will present the methodology used to develop HFE guidance and update these documents, and describe the current status of the update program.

  5. Visual field loss among patients with cytomegalovirus retinitis

    Science.gov (United States)

    Thorne, Jennifer E.; Van Natta, Mark L.; Jabs, Douglas A.; Duncan, Jacque L.; Srivastava, Sunil K.

    2010-01-01

    Purpose To describe visual field (VF) loss among patients with cytomegalovirus (CMV) retinitis and the risk factors for such loss. Design Multicenter, prospective, observational study Participants 476 patients with AIDS and CMV retinitis and VF data Methods Follow-up every 3 months with medical history, ophthalmological examination, Goldman visual fields, and laboratory testing Main outcome measures Incidence of visual field loss in eyes affected with CMV retinitis and characteristics associated with such VF loss Results Over a median follow up of 4 years (range = 0.5 to 9 years), the incidence rates of VF loss to 75% and 50% of normal were 0.22/eye-year (EY, 95% confidence interval [CI]: 0.20, 0.25) and 0.08/EY (95% CI: 0.06, 0.10), respectively. The observed rates were 6- to 7-fold less than those observed rates of VF loss in the era prior to highly active antiretroviral therapy (HAART). Decreased CD4+ T cell count, whether measured at enrollment or over follow up time, was associated with increased rates of VF loss for all VF outcomes in a dose-dependent fashion. Risk factors for VF loss included lower CD4+ T cell count, CMV lesion size >25% of the total retinal area, and active CMV retinitis after controlling for potential confounding. HAART use and immune recovery (CD4+ T cell count >100 cells/μL) were associated with reduced risk of VF loss in multiple regression models. Immune recovery was statistically significantly associated with a lower risk of VF loss to 75% of normal (relative risk [RR] = 0.63 95% CI: 0.49, 0.86 P = 0.003) and to 50% of normal (RR = 0.60 95% CI: 0.44, 0.82 P = 0.001) after controlling for demographic characteristics, human immunodeficiency virus (HIV) viral load, HAART use, CMV lesion location and size, and retinitis activity. Conclusions Cytomegalovirus retinitis was associated with a substantial risk of incident VF loss, but the incidence is approximately 6- fold lower than that observed in the pre-HAART era. Those who have HAART

  6. Application of laboratory data from small-scale simulators to human performance issues in the nuclear industry

    Energy Technology Data Exchange (ETDEWEB)

    Spettell, C.M.

    1986-01-01

    Laboratory analogs of nuclear power plant tasks were simulated on personal computers in two experimental studies. Human performance data were collected during each experimental study. The goal of the first experiment was to validate a quantitative model of dependence among human errors during testing, calibration, and maintenance activities. This model, the Multiple Sequential Failure (MSF) model (NUREG/CR-2211) has been used to quantify dependent human error failure probabilities for human reliability analyses in Probabilistic Risk Assessments (PRAs). The goal of the second experiment was to examine the relationship among psychological and behavioral characteristics of individuals and their performance at controlling a simulated nuclear power plant. These studies demonstrated the usefulness of the experimental psychology approach for validating models of human performance at nuclear power plant tasks.

  7. Cytomegalovirus (CMV) Infection: A Guide for Patients and Families After Stem Cell Transplant

    Science.gov (United States)

    ... Infection: A Guide for Patients and Families after Stem Cell Transplant What is cytomegalovirus (CMV)? Cytomegalovirus (CMV), a ... weakened by medicines that you must take after stem cell transplant and by the transplant itself. Your body ...

  8. Applying Human Factors Evaluation and Design Guidance to a Nuclear Power Plant Digital Control System

    Energy Technology Data Exchange (ETDEWEB)

    Thomas Ulrich; Ronald Boring; William Phoenix; Emily Dehority; Tim Whiting; Jonathan Morrell; Rhett Backstrom

    2012-08-01

    The United States (U.S.) nuclear industry, like similar process control industries, has moved toward upgrading its control rooms. The upgraded control rooms typically feature digital control system (DCS) displays embedded in the panels. These displays gather information from the system and represent that information on a single display surface. In this manner, the DCS combines many previously separate analog indicators and controls into a single digital display, whereby the operators can toggle between multiple windows to monitor and control different aspects of the plant. The design of the DCS depends on the function of the system it monitors, but revolves around presenting the information most germane to an operator at any point in time. DCSs require a carefully designed human system interface. This report centers on redesigning existing DCS displays for an example chemical volume control system (CVCS) at a U.S. nuclear power plant. The crucial nature of the CVCS, which controls coolant levels and boration in the primary system, requires a thorough human factors evaluation of its supporting DCS. The initial digital controls being developed for the DCSs tend to directly mimic the former analog controls. There are, however, unique operator interactions with a digital vs. analog interface, and the differences have not always been carefully factored in the translation of an analog interface to a replacement DCS. To ensure safety, efficiency, and usability of the emerging DCSs, a human factors usability evaluation was conducted on a CVCS DCS currently being used and refined at an existing U.S. nuclear power plant. Subject matter experts from process control engineering, software development, and human factors evaluated the DCS displays to document potential usability issues and propose design recommendations. The evaluation yielded 167 potential usability issues with the DCS. These issues should not be considered operator performance problems but rather opportunities

  9. Can active proton interrogation find shielded nuclear threats at human-safe radiation levels?

    Energy Technology Data Exchange (ETDEWEB)

    Liew, Seth Van, E-mail: vanliew@gmail.com

    2017-05-21

    A new method of low-dose proton radiography is presented. The system is composed of an 800 MeV proton source, bending magnets, and compact detectors, and is designed for drive-through cargo scanning. The system has been simulated using GEANT4. Material identification algorithms and pixel sorting methods are presented that allow the system to perform imaging at doses low enough to scan passenger vehicles and people. Results are presented on imaging efficacy of various materials and cluttered cargoes. The identification of shielded nuclear materials at human-safe doses has been demonstrated.