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      Tissue Tropism and Target Cells of NSs-Deleted Rift Valley Fever Virus in Live Immunodeficient Mice

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          Abstract

          Background

          Rift Valley fever virus (RVFV) causes disease in livestock and humans. It can be transmitted by mosquitoes, inhalation or physical contact with the body fluids of infected animals. Severe clinical cases are characterized by acute hepatitis with hemorrhage, meningoencephalitis and/or retinitis. The dynamics of RVFV infection and the cell types infected in vivo are poorly understood.

          Methodology/Principal Findings

          RVFV strains expressing humanized Renilla luciferase (hRLuc) or green fluorescent protein (GFP) were generated and inoculated to susceptible Ifnar1-deficient mice. We investigated the tissue tropism in these mice and the nature of the target cells in vivo using whole-organ imaging and flow cytometry. After intraperitoneal inoculation, hRLuc signal was observed primarily in the thymus, spleen and liver. Macrophages infiltrating various tissues, in particular the adipose tissue surrounding the pancreas also expressed the virus. The liver rapidly turned into the major luminescent organ and the mice succumbed to severe hepatitis. The brain remained weakly luminescent throughout infection. FACS analysis in RVFV-GFP-infected mice showed that the macrophages, dendritic cells and granulocytes were main target cells for RVFV. The crucial role of cells of the monocyte/macrophage/dendritic lineage during RVFV infection was confirmed by the slower viral dissemination, decrease in RVFV titers in blood, and prolonged survival of macrophage- and dendritic cell-depleted mice following treatment with clodronate liposomes. Upon dermal and nasal inoculations, the viral dissemination was primarily observed in the lymph node draining the injected ear and in the lungs respectively, with a significant increase in survival time.

          Conclusions/Significance

          These findings reveal the high levels of phagocytic cells harboring RVFV during viral infection in Ifnar1-deficient mice. They demonstrate that bioluminescent and fluorescent viruses can shed new light into the pathogenesis of RVFV infection.

          Author Summary

          Rift Valley fever, caused by a member of the Bunyaviridae family, has spread during recent years to most sub-Saharan African countries, in Egypt and in the Arabian peninsula. The virus can be transmitted by insect vectors or by direct contacts with infectious tissues. The analysis of virus replication and dissemination in laboratory animals has been hampered by the need to euthanize sufficient numbers of animals and to assay appropriate organs at various time points after infection to evaluate the viral replication. By following the bioluminescence and fluorescence of Rift Valley fever viruses expressing light reporters, we were able to track the real-time dissemination of the viruses in live immunodeficient mice. We showed that the first infected organs were the thymus, spleen and liver, but the liver rapidly became the main location of viral replication. Phagocytes also appeared as important targets, and their systemic depletion by use of clodronate liposomes decreased the number of viruses in the blood, delayed the viral dissemination and prolonged the survival of the infected mice.

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          Most cited references56

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          Liposome mediated depletion of macrophages: mechanism of action, preparation of liposomes and applications.

          Selective depletion of macrophages from tissues in vivo can be used to investigate whether these cells are playing a role in defined biological processes. This question is particularly relevant to various host defense mechanisms. We have developed a macrophage 'suicide' technique, using the liposome mediated intracellular delivery of dichloromethylene-bisphosphonate (Cl2MBP or clodronate). The method is specific with respect to phagocytic cells of the mononuclear phagocyte system (MPS) for the following reasons: (1) The natural fate of liposomes is phagocytosis. (2) Once ingested by macrophages, the phospholipid bilayers of the liposomes are disrupted under the influence of lysosomal phospholipases. (3) Cl2MBP intracellularly released in this way does not easily escape from the cell by crossing the cell membranes. (4) Cl2MBP released in the circulation from dead macrophages or by leakage from liposomes, will not easily enter non-phagocytic cells and has an extremely short half life in the circulation and body fluids. In the present review, the preparation of Cl2MBP-liposomes has been described in detail. Furthermore, the mechanism of action of the new approach and its applicabilities are discussed.
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            Analysis of in vivo dynamics of influenza virus infection in mice using a GFP reporter virus.

            Influenza A virus is being extensively studied because of its major impact on human and animal health. However, the dynamics of influenza virus infection and the cell types infected in vivo are poorly understood. These characteristics are challenging to determine, partly because there is no efficient replication-competent virus expressing an easily traceable reporter gene. Here, we report the generation of a recombinant influenza virus carrying a GFP reporter gene in the NS segment (NS1-GFP virus). Although attenuated when compared with wild-type virus, the NS1-GFP virus replicates efficiently in murine lungs and shows pathogenicity in mice. Using whole-organ imaging and flow cytometry, we have tracked the dynamics of influenza virus infection progression in mice. Imaging of murine lungs shows that infection starts in the respiratory tract in areas close to large conducting airways and later spreads to deeper sections of the lungs. In addition to epithelial cells, we found GFP-positive antigen-presenting cells, such as CD11b(+)CD11c(-), CD11b(-)CD11c(+), and CD11b(+)CD11c(+), as early as 24 h after intranasal infection. In addition, a significant proportion of NK and B cells were GFP positive, suggesting active infection of these cells. We next tested the effects of the influenza virus inhibitors oseltamivir and amantadine on the kinetics of in vivo infection progression. Treatment with oseltamivir dramatically reduced influenza infection in all cell types, whereas, surprisingly, amantadine treatment more efficiently blocked infection in B and NK cells. Our results demonstrate high levels of immune cells harboring influenza virus antigen during viral infection and cell-type-specific effects upon treatment with antiviral agents, opening additional avenues of research in the influenza virus field.
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              Alpha/beta interferon protects against lethal West Nile virus infection by restricting cellular tropism and enhancing neuronal survival.

              West Nile virus (WNV) is a mosquito-borne flavivirus that is neurotropic in humans, birds, and other animals. While adaptive immunity plays an important role in preventing WNV spread to the central nervous system (CNS), little is known about how alpha/beta interferon (IFN-alpha/beta) protects against peripheral and CNS infection. In this study, we examine the virulence and tropism of WNV in IFN-alpha/beta receptor-deficient (IFN- alpha/betaR-/-) mice and primary neuronal cultures. IFN-alpha/betaR-/- mice were acutely susceptible to WNV infection through subcutaneous inoculation, with 100% mortality and a mean time to death (MTD) of 4.6 +/- 0.7 and 3.8+/- 0.5 days after infection with 10(0) and 10(2) PFU, respectively. In contrast, congenic wild-type 129Sv/Ev mice infected with 10(2) PFU showed 62% mortality and a MTD of 11.9 +/- 1.9 days. IFN-alpha/betaR-/- mice developed high viral loads by day 3 after infection in nearly all tissues assayed, including many that were not infected in wild-type mice. IFN-alpha/betaR-/- mice also demonstrated altered cellular tropism, with increased infection in macrophages, B cells, and T cells in the spleen. Additionally, treatment of primary wild-type neurons in vitro with IFN-beta either before or after infection increased neuronal survival independent of its effect on WNV replication. Collectively, our data suggest that IFN-alpha/beta controls WNV infection by restricting tropism and viral burden and by preventing death of infected neurons.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS Negl Trop Dis
                plos
                plosntds
                PLoS Neglected Tropical Diseases
                Public Library of Science (San Francisco, USA )
                1935-2727
                1935-2735
                December 2011
                6 December 2011
                : 5
                : 12
                : e1421
                Affiliations
                [1 ]Central Animal Facilities, Institut Pasteur, Paris, France
                [2 ]Molecular Genetics of Bunyaviruses, Institut Pasteur, Paris, France
                [3 ]Human Histopathology and Animal Models, Institut Pasteur, Paris, France
                [4 ]Center for Human Immunology, Institut Pasteur, Paris, France
                [5 ]Mouse Functional Genetics, Institut Pasteur, Paris, France
                [6 ]Centre National de la Recherche Scientifique, Unité de Recherche Associée 2578, Paris, France
                [7 ]Cytokines and Inflammation, Institut Pasteur, Paris, France
                [8 ]Department of Molecular Cell Biology, Vrije Universiteit Medical Center, Amsterdam, The Netherlands
                Centers for Disease Control and Prevention, United States of America
                Author notes

                Conceived and designed the experiments: CG AB MH TZdV XM MB JJP. Performed the experiments: CG AB GJ MH TZdV LG CB. Analyzed the data: CG AB GJ MH CB TZdV XM MB JJP. Contributed reagents/materials/analysis tools: NvR. Wrote the paper: CG MB JJP.

                Article
                PNTD-D-11-00535
                10.1371/journal.pntd.0001421
                3232203
                22163058
                6b2590ca-5a3c-490b-bf08-55a7fe0a5437
                Gommet et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
                History
                : 7 June 2011
                : 24 October 2011
                Page count
                Pages: 12
                Categories
                Research Article
                Biology
                Anatomy and Physiology
                Biotechnology
                Genetics
                Immunology
                Microbiology
                Model Organisms
                Medicine
                Infectious Diseases
                Veterinary Science
                Animal Types
                Veterinary Anatomy and Physiology
                Veterinary Diseases
                Veterinary Medicine
                Veterinary Microbiology
                Veterinary Pathology

                Infectious disease & Microbiology
                Infectious disease & Microbiology

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