Visna Virus

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Gordon D. Harkiss - One of the best experts on this subject based on the ideXlab platform.

  • granulocyte macrophage colony stimulating factor is elevated in alveolar macrophages from sheep naturally infected with maedi Visna Virus and stimulates maedi Visna Virus replication in macrophages in vitro
    Clinical and Experimental Immunology, 2002
    Co-Authors: Z Zhang, J Hopkins, Gordon D. Harkiss, C J Woodall
    Abstract:

    Infection by maedi-Visna Virus, a lentiVirus of sheep, leads to chronic inflammatory reactions of various tissues. In this report we have analysed the role of specific cytokines in the disease process. A significant increase in expression of interleukin-6, interleukin-10, granulocyte macrophage-colony stimulating factor (GM-CSF) and transforming growth factor-beta1 mRNA was observed in alveolar macrophages isolated from the lungs of naturally infected animals when compared with lungs of seronegative controls. Levels of GM-CSF mRNA expression in alveolar macrophages correlated with the presence of lung lesions, but there was no correlation of interleukin-10, interleukin-6, tumour necrosis factor-alpha and transforming growth factor-beta1 mRNA levels in alveolar macrophages from animals with pulmonary lesions. In vitro investigation showed that GM-CSF in the range 0.1-10 ng/ml induced a significant increase in viral p25 production after 7 days in acutely infected blood monocyte-derived macrophages. The production of p25 peaked between 7 and 14 days exposure to 10 ng/ml of GM-CSF. Quantitative polymerase chain reaction showed that the level of viral DNA in monocyte-derived macrophages was dose-dependent following GM-CSF treatment in the range 0.1-100 ng/ml after 7 days. Viral mRNA expression was also enhanced. These findings indicate a role for GM-CSF in the pathogenesis of lymphoid interstitial pneumonia in infected animals.

  • quantitative analysis of maedi Visna Virus dna load in peripheral blood monocytes and alveolar macrophages
    Journal of Virological Methods, 2000
    Co-Authors: Zhidong Zhang, N J Watt, Gordon D. Harkiss, C J Woodall
    Abstract:

    Viral load may be an important indicator of disease progression in sheep infected with maedi-Visna Virus (MVV). To assess this variable accurately in MVV-infected sheep, a quantitative competitive-polymerase chain reaction (QC-PCR) was developed. A conserved region of the MVV pol gene was selected. The RT-PCR MVV pol product was cloned and mutagenised in vitro by PCR to produce a competitor template reduced in length from 217 to 192 bp, but which retained the original flanking MVV pol PCR primers. The competitor template was quantified accurately and in an optimised QC-PCR protocol serial dilutions of this template were co-amplified with known amounts of sample DNA. MVV DNA levels in peripheral blood monocytes and alveolar macrophages from MVV-infected sheep (n=12) were assessed by QC-PCR. Viral DNA load in alveolar macrophages was significantly higher than that in peripheral blood monocytes when the animals were compared overall. A comparison was also made between alveolar macrophages from the lungs of seropositive animals with or without histopathological evidence of pulmonary lesions. The load of MVV DNA in alveolar macrophages was low in sheep without histopathological evidence of lesions in the lung. In contrast, in alveolar macrophages from sheep with histopathological lesions in the lung, there was a significantly higher level of MVV DNA. The correlation of MVV load with pulmonary lesions suggests that infected alveolar macrophages play a key role in the pathogenesis of this lymphoid interstitial pneumonia.

  • phenotypic characterisation and infection of ovine microglial cells with maedi Visna Virus
    Journal of NeuroVirology, 2000
    Co-Authors: Bahrain Ebrahimi, Timothy E Allsopp, John K Fazakerley, Gordon D. Harkiss
    Abstract:

    Maedi-Visna Virus (MVV) infection of the central nervous system (CNS) results in pathological changes, the mechanisms of which are poorly understood. MVV preferentially infects cell of the monocyte/macrophage lineage in vivo. The neuroparenchymal microglial cells are the resident tissue macrophages in the CNS and therefore likely targets for MVV infection. However, no information is currently available on the susceptibility of these cells to MVV infection or their contribution to neuropathological changes as a result of MVV infection. Highly enriched primary ovine microglial cell cultures were set up from brain tissues of lambs. These cells were amoeboid or bipolar with spikes, a morphology consistent with microglial cells of other species, and stained positive for CD1, CD11a, CD11c, CD14, MHC-class I, MHC-class II, and beta-N-acetyl galactose, but not with markers of astrocytes or oligodendrocytes. These sheep microglial cells were permissive for MVV infection. Productive MVV infection resulted in selective transcriptional up-regulation of the pro-inflammatory cytokines TNFalpha and IL-6. In contrast, there was no change in levels of transcripts for TGFbeta1, IL-1beta, GM-CSF, IL-10, or IL-12. These data provide the first evidence that ovine microglial cells can support productive infection with MVV, and that this leads to a selective up-regulation of proinflammatory cytokines. These may contribute to Visna neuropathology.

  • acute in vivo neurotoxicity of peptides from maedi Visna Virus transactivating protein tat
    Brain Research, 1999
    Co-Authors: Isabella Starling, Gordon W. Arbuthnott, Ann K. Wright, Gordon D. Harkiss
    Abstract:

    LentiViruses such as Maedi Visna Virus (MVV) in sheep, and human immunodeficiency Virus (HIV) in man often cause a variety of neurological syndromes in later stages of infection. Neuropathological investigations reveal damage to myelin and astrocytosis in both white and grey matter. MVV infection induces axonal damage with some areas of necrosis while neuronal loss, and synaptic damage have been reported in HIV-1 infection. It is not clear, at present, how this neurodegeneration is mediated but, as these Viruses do not directly infect neurons, an indirect neurotoxic action of the Viruses is indicated. Previous experiments have shown that the intra-striatal injection in rats of a synthetic peptide derived from the basic region of the MVV transactivating protein Tat causes considerable neurotoxicity 1 week post-operatively. By in vivo stereotaxic injections of the same synthetic peptide, and subsequent immunocytochemical detection of neurons, astrocytes and microglia, we show that this neurotoxicity displays a distinctive and unusual lesion profile and is evident as rapidly as 0.5 h post-operatively. Furthermore, neuroprotection studies suggest that the early effects of the MVV tat peptide may involve glutamate neurotoxicity via the N-methyl-D-aspartate (NMDA) receptors since the application of dizolcipine (MK801) reduces the volume of the lesion seen at 1 h after the injection of neurotoxic peptide, while L-NAME is ineffective. The mechanism of this early neurotoxicity is thus different from the longer term actions already described.

  • regulation of the long terminal repeat in Visna Virus by a transcription factor related to the aml pebp2 cbf superfamily
    Virology, 1997
    Co-Authors: Keith A Sutton, Ian Mcconnell, Gordon D. Harkiss, Chungtien Lin, David R Sargan
    Abstract:

    The long terminal repeats of maedi Visna Virus strain 1514 contain a consensus AP-1 binding site which has been shown to be important in controlling Virus transcription. However, this consensus site is absent in strain EV-1. Here, we have compared the ability of oligonucleotides corresponding to LTR sequences from EV-1 with those from 1514 to bind transcription factors in competitive gel retardation assays and activate reporter gene expression. The experiments demonstrated no observable binding of AP-1 to the EV-1-derived sequences and significant differences in the abilities of the 1514 and EV-1 sequences to activate transcription. However, both viral sequences interacted with a second, previously undetected, transcription factor. This factor gave specific gel shifts which were competed by an oligonucleotide containing the consensus sequence for the AML/PEBP2/CBF family of transcriptional factors, but not by control AP-1 or OCT-1 oligonucleotides. The factor was therefore denoted AML (vis). A second AML (vis) site, noted upstream of the TATA box proximal AP-1 site, gave single shifts which were competed by the downstream AML (vis) oligonucleotide. Both sites were functional in transfection assays. In gel shift retardation assays, polyclonal antisera directed against known runt domain proteins were able to supershift part of the AML (vis) binding activity in nuclear extracts from physiologically relevant cell types. The results thus suggest that the AML (vis) binding factor belongs to the AML/PEBP2/CBF family of transcription factors and may be important in controlling Virus replication in these and other strains of ruminant lentiViruses.

Janice E. Clements - One of the best experts on this subject based on the ideXlab platform.

  • Visna Virus induced activation of mapk is required for Virus replication and correlates with Virus induced neuropathology
    Journal of Virology, 2002
    Co-Authors: Sheila A Barber, Linda Bruett, Brian R Douglass, David S Herbst, Christine M Zink, Janice E. Clements
    Abstract:

    It is well accepted that Viruses require access to specific intracellular environments in order to proliferate or, minimally, to secure future proliferative potential as latent reservoirs. Hence, identification of essential Virus-cell interactions should both refine current models of Virus replication and proffer alternative targets for therapeutic intervention. In the present study, we examined the activation states of mitogen-activated protein kinases (MAPKs), ERK-1/2, in primary cells susceptible to Visna Virus and report that Virus infection induces and sustains activation of the ERK/MAPK pathway. Treatment of infected cells with PD98059, a specific inhibitor of the ERK/MAPK pathway, abolishes Visna Virus replication, as evidenced by extremely low levels of Gag protein expression and reverse transcriptase activity in culture supernatants. In addition, although Visna Virus-induced activation of MAPK is detectable within 15 min, early events of viral replication (i.e., reverse transcription, integration, and transcription) are largely unaffected by PD98059. Interestingly, further examination demonstrated that treatment with PD98059 results in decreased cytoplasmic expression of gag and env, but not rev, mRNA, highly suggestive of an ERK/MAPK-dependent defect in Rev function. In vivo analysis of ERK-1/2 activation in brains derived from Visna Virus-infected sheep demonstrates a strong correlation between ERK/MAPK activation and Virus-associated encephalitis. Moreover, double-labeling experiments revealed that activation of MAPK occurs not only in cells classically infected by Visna Virus (i.e., macrophages and microglia), but also in astrocytes, cells not considered to be major targets of Visna Virus replication, suggesting that activation of the ERK/MAPK pathway may contribute to the Virus-induced processes leading to neurodegenerative pathology.

  • Functional Murine Leukemia Virus Vectors Pseudotyped with the Visna Virus Envelope Show Expanded Visna Virus Cell Tropism
    Journal of virology, 2001
    Co-Authors: Linda Bruett, Janice E. Clements
    Abstract:

    Pseudotype Virus vectors serve as a powerful tool for the study of Virus receptor usage and entry. We describe the development of murine leukemia Virus (MuLV) particles pseudotyped with the Visna Virus envelope glycoprotein and encoding a green fluorescent protein reporter as a tool to study the expression of the Visna Virus receptor. Functional MuLV/Visna Virus pseudotypes were obtained when the cytoplasmic tail of the Visna Virus envelope TM protein was truncated to 3, 7, or 11 amino acids in length. MuLV/Visna Virus particles were used to transduce a panel of cell types from various organisms, including sheep, goat, human, hamster, mouse, monkey, and quail. The majority of the cells examined were susceptible to MuLV/Visna pseudotype Viruses, supporting the notion that the Visna Virus cellular receptor is a widely expressed protein found in many species. Of 16 different cell types tested, only mouse embryo fibroblast NIH 3T3 cells, hamster ovary CHO cells, and the human promonocyte cell line U937 cells were not susceptible to transduction by the pseudotyped Virus. The production of functional MuLV/Visna Virus pseudotypes has provided a sensitive, biologically relevant system to study Visna Virus cell entry and envelope-receptor interactions.

  • characterization of a membrane associated protein implicated in Visna Virus binding and infection
    Virology, 2000
    Co-Authors: Linda Bruett, Sheila A Barber, Janice E. Clements
    Abstract:

    Abstract The identity of the cellular receptor(s) for Visna Virus, an ovine lentiVirus, is currently unknown; however, previous studies from our laboratory have identified membrane-associated proteins expressed selectively in susceptible cells which bind Visna Virus. Moreover, a polyclonal antibody (2-23), raised against a 45-kDa Visna Virus binding protein, bound specifically to the surface of susceptible cells in immunofluorescence assays and significantly reduced binding of Visna Virus to cells (S. E. Crane et al., 1991, J. Virol., 65, 6137–6143). In this report we extend our studies of this antibody (2-23), showing both that 2-23 significantly reduces Visna Virus infection of susceptible cells and that 2-23 immunoprecipitates a putative protein complex consisting of a prominent 30-kDa protein, as well as the 45-kDa immunogen, specifically from radiolabeled Virus-susceptible sheep cells. Further, we demonstrate that the 30-kDa protein is a membrane-associated proteoglycan substituted with a chondroitin sulfate glycosaminoglycan (GAG) chain(s) and that treatment of susceptible cells with an inhibitor of GAG synthesis significantly reduces Visna Virus production. Collectively, these data support a role for a proteoglycan in Visna Virus cell binding and infection.

  • development of transgenic sheep that express the Visna Virus envelope gene
    Virology, 1994
    Co-Authors: Janice E. Clements, R Schoborg, Lucy M Carruth, Darlene Sheffer, Anne M. Powell, R J Wall, Mary Christine Zink, Debra Hauer, Opendra Narayan, Caird E. Rexroad
    Abstract:

    Abstract The ovine lentiViruses cause encephalitis, pneumonia, and arthritis in sheep worldwide. Visna Virus is a prototype of this family and the pathogenesis and molecular biology of the Virus has been well characterized. The envelope proteins of Visna Virus are responsible for binding of Virus to host cells and for causing cell fusion. The surface glycoprotein also elicits cellular and humoral immune responses to the Virus, the former being thought to be responsible for eliminating infected cells as well as causing inflammatory lesions. In this study, transgenic sheep were constructed that expressed the envelope genes of Visna Virus under the control of the Visna LTR to investigate the role of the env gene in the pathogenesis of lentiviral disease in its natural host. Three transgenic lambs were identified that contain the env transgene and express the envelope glycoproteins. These transgenic animals have remained healthy and expression of the viral gene has had no obvious deleterious effect. Expression of the Visna envelope protein was demonstrated by cell fusion mediated by the envelope gene as well as by immunoprecipitation of the envelope proteins with monoclonal antibodies and immunofluorescence analyses of Env protein in cells. The target cell for Visna Virus replication in infected animals is the monocyte/macrophage. In natural infection, the level of viral gene expression in these cells increases with cell maturation. In the transgenic sheep, monocytes did not express the envelope glycoproteins until they differentiated into macrophages in vitro . Expression of the env mRNA in macrophages was quantitated by an RNase protection assay. In addition to expression in macrophages, the transgene was expressed by fibroblasts isolated from skin of the transgenic sheep. Expression of both the Env and Rev proteins was detected by immunoprecipitation and immunofluorescence. Two of the three lambs responded immunologically to the expression of the transgene by producing binding antibodies to the envelope glycoproteins. Thus, these transgenic sheep provide a model to study whether a lentiVirus glycoprotein will prevent infection or modulate disease in its natural host after Virus challenge.

  • molecular mechanisms of Visna Virus tat identification of the targets for transcriptional activation and evidence for a post transcriptional effect
    Virology, 1992
    Co-Authors: S L Gdovin, Janice E. Clements
    Abstract:

    Visna Virus is a pathogenic lentiVirus of sheep that is distantly related to the primate lentiViruses, including the human immunodeficiency Virus type 1 (HIV-1). Replication of HIV-1 in cell culture requires the expression of a Virus-encoded protein, Tat, which is a potent trans-activator of viral gene expression. Visna Virus encodes an analogous Tat protein that greatly increases gene expression directed by the Visna viral LTR. This report uses a stable vero cell line that constitutively expresses Visna Virus Tat to investigate the molecular mechanism of action of Tat on viral gene expression. Transient expression assays, using the Visna Virus LTR to drive transcription of the bacterial gene for chloramphenicol acetyltransferase (CAT), demonstrate that Tat trans-activates gene expression by increasing steady-state mRNA levels. The increase in steady-state mRNA levels is sufficient to account for the increase in protein observed and is due, in part, to an increase in the rate of transcription initiation. Tat mediates the accumulation of mRNA through AP-4 and AP-1 binding sites located in the U3 region of the LTR. Deletion of the upstream AP-1 and AP-4 binding sites results in a residual low level of trans-activation by Tat. Further experiments, using LTRs with R-U5 sequences deleted to +10, demonstrate AP-1 and AP-4 mediated responses to TAT at the RNA level, but no increase was observed in CAT protein.

Lynn M Herrmannhoesing - One of the best experts on this subject based on the ideXlab platform.

Donald P Knowles - One of the best experts on this subject based on the ideXlab platform.

Ernst Peterhans - One of the best experts on this subject based on the ideXlab platform.

  • maedi Visna Virus infection in sheep a review
    Veterinary Research, 1998
    Co-Authors: Michel Pepin, Christian Vitu, Pierre Russo, Jeanfrancois Mornex, Ernst Peterhans
    Abstract:

    The maedi-Visna Virus (MVV) is classified as a lentiVirus of the retroviridae family. The genome of MVV includes three genes: gag, which encodes for group-specific antigens; pol, which encodes for reverse transcriptase, integrase, RNAse H, protease and dUTPase and env, the gene encoding for the surface glycoprotein responsible for receptor binding and entry of the Virus into its host cell. In addition, analogous to other lentiViruses, the genome contains genes for regulatory proteins, i.e. vif, rev and tat. The coding regions of the genome are flanked by long terminal repeats (LTR) which play a crucial role in the replication of the viral genome and provide binding sites for cellular transcription factors. The organs targeted by MVV are, in descending order of importance, the lungs, mammary glands, joints and the brain. In these organs, the Virus replicates in mature macrophages and induces slowly progressing inflammatory lesions containing B and T lymphocytes. The clinical signs of MVV infection, i.e. dyspnea, loss of weight, mastitis and arthritis, are related to the location of these lesions. Infection with MVV induces the formation of antibodies which can be detected by agar gel immunodiffusion, ELISA and the serum neutralization assay. As neither antiviral treatment nor vaccination is available, diagnostic tests are the backbone of most of the schemes implemented to prevent the spread of MVV. However, since current serological assays are still lacking in sensitivity and specificity, molecular biological methods are being developed permitting the detection of Virus in peripheral blood, milk and tissue samples. Future research will have to focus on both the development of new diagnostic tests and a better understanding of the pathogenesis of MVV infection.

  • expression in escherichia coli and sequencing of the coding region for the capsid protein of dutch maedi Visna Virus strain zzv 1050 application of recombinant protein in enzyme linked immunosorbent assay for the detection of caprine and ovine lentiv
    Journal of Clinical Microbiology, 1991
    Co-Authors: R Zanoni, I M Nauta, U Pauli, Ernst Peterhans
    Abstract:

    Maedi-Visna in sheep and caprine arthritis-encephalitis in goats are caused by two closely related and widespread lentiViruses. The infections are characterized by life-long Virus persistence and slow induction of antiviral antibodies. The diagnosis is based on the detection of antiviral antibodies. We have used the polymerase chain reaction (PCR) to amplify a part of the gag gene coding for the entire capsid protein and for parts of the matrix and nucleocapsid proteins. Sequencing of the PCR fragment of the Dutch maedi-Visna Virus strain ZZV 1050 revealed 85 and 92% homology to the DNA and deduced amino acid sequences, respectively, of the distantly related Icelandic Visna Virus strain 1514. The respective homologies with caprine arthritis-encephalitis Virus strain CO were 76 and 80%. The PCR fragment was cloned into pGEX-2T and expressed as a glutathione S-transferase fusion protein. The recombinant protein could be detected on immunoblots by using a monoclonal antibody and polyclonal antisera and was further purified by glutathione-based affinity chromatography. Enzyme-linked immunosorbent assay with purified recombinant fusion protein is shown to be a sensitive and specific diagnostic tool for the detection of lentiviral infection in goats and sheep.

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