Recovery of Virulent and RNase-Negative Attenuated Type 2 Bovine Viral Diarrhea Viruses from Infectious cDNA Clones

Meyer, Christiane; Freyburg, Martina von; Elbers, Knut; Meyers, Gregor

doi:10.1128/jvi.76.16.8494-8503.2002

Cited by 53 publications

(76 citation statements)

References 59 publications

(38 reference statements)

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“…A single-or double-codon mutation abrogating E rns RNase activity of the CSFV Alfort/Tü-bingen strain attenuated the virus for pigs (19). Notably, a similar result was obtained by mutating the same E rns domain of BVDV (18). More recently, N pro has also been shown to be involved in CSFV virulence, since deletion of N pro in the virulent Eystrup strain abrogated its virulence for swine (17).…”

Section: Discussionsupporting

confidence: 58%

The E2 Glycoprotein of Classical Swine Fever Virus Is a Virulence Determinant in Swine

Risatti

Borca

Kutish

et al. 2005

J Virol

133

108

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To identify genetic determinants of classical swine fever virus (CSFV) virulence and host range, chimeras of the highly pathogenic Brescia strain and the attenuated vaccine strain CS were constructed and evaluated for viral virulence in swine. Upon initial screening, only chimeras 138.8v and 337.14v, the only chimeras containing the E2 glycoprotein of CS, were attenuated in swine despite exhibiting unaltered growth characteristics in primary porcine macrophage cell cultures. Additional viral chimeras were constructed to confirm the role of E2 in virulence. Chimeric virus 319.1v, which contained only the CS E2 glycoprotein in the Brescia background, was markedly attenuated in pigs, exhibiting significantly decreased virus replication in tonsils, a transient viremia, limited generalization of infection, and decreased virus shedding. Chimeras encoding all Brescia structural proteins in a CS genetic background remained attenuated, indicating that additional mutations outside the structural region are important for CS vaccine virus attenuation. These results demonstrate that CS E2 alone is sufficient for attenuating Brescia, indicating a significant role for the CSFV E2 glycoprotein in swine virulence.Classical swine fever (CSF) is a highly contagious and often fatal hemorrhagic disease of swine and is caused by Classical swine fever virus (CSFV), a member of the genus Pestivirus of the family Flaviviridae (5). Intermittent CSF outbreaks in Europe and other parts of the world result in significant economic losses. While infection with highly virulent CSFV strains can cause acute CSF characterized by high morbidity and mortality, isolates of moderate to low virulence induce a prolonged, chronic disease (35).CSFV is a small enveloped virus with a single-stranded, 12.5-kb RNA genome of positive polarity. The CSFV genome contains a single open reading frame encoding an approximately 4,000-amino-acid polyprotein which through cellular and viral protease-mediated co-and posttranslational processing gives rise to the following 11 to 12 final cleavage products: NH 2 -Npro-C-E rns -E1-E2-p7-NS2-NS3-NS4A-NS4B-NS5A-NS5B-COOH (23). Protein C and the glycoproteins E rns , E1, and E2 represent structural components of the virion (28). E1 and E2 are anchored to the envelope by their carboxy termini, with E rns loosely associated with the envelope. E rns and E2 are present as homodimers linked by disulfide bridges on the surfaces of CSFV virions, whereas E2 is also found dimerized with E1 (28, 37, 38). The genetic basis of CSFV virulence and host range remains poorly understood (35).Development of infectious CSFV cDNA clones has enabled genetic approaches for defining mechanisms of viral replication and pathogenesis. Infectious clones (IC) of the attenuated CSFV C-strain and the pathogenic Alfort/Tübingen and Eystrup strains have been constructed, enabling identification of E rns and N pro as virulence factors in swine and of the role of different E rns mutations in virus attenuation (17, 19). Additionally, attempts to mutate and in...

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Section: Discussionsupporting

confidence: 58%

The E2 Glycoprotein of Classical Swine Fever Virus Is a Virulence Determinant in Swine

Risatti

Borca

Kutish

et al. 2005

J Virol

133

108

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“…pDCs produce type III IFN in response to BVDV. pDC-enriched cell populations were isolated from three animals and infected with rBVDV [XIKE A (42)] at an MOI of 0.1 for 48 h. Supernatants were either left untreated or acid treated for 24 h prior to assay by IL-28B ELISA (Biotang). The IL-28B ELISA results ( Fig.…”

Section: Resultsmentioning

confidence: 99%

Type I and III IFNs Produced by Plasmacytoid Dendritic Cells in Response to a Member of theFlaviviridaeSuppress Cellular Immune Responses

Reid

Juleff

Windsor

et al. 2016

The Journal of Immunology

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The pestivirus noncytopathic bovine viral diarrhea virus (BVDV) can suppress IFN production in the majority of cell types in vitro. However, IFN is detectable in serum during acute infection in vivo for ∼5–7 d, which correlates with a period of leucopoenia and immunosuppression. In this study, we demonstrate that a highly enriched population of bovine plasmacytoid dendritic cells (DCs) produced IFN in response to BVDV in vitro. We further show that the majority of the IFN produced in response to infection both in vitro and in vivo is type III IFN and acid labile. Further, we show IL-28B (IFN-λ3) mRNA is induced in this cell population in vitro. Supernatant from plasmacytoid DCs harvested postinfection with BVDV or recombinant bovine IFN-α or human IL-28B significantly reduced CD4+ T cell proliferation induced by tubercle bacillus Ag 85–stimulated monocyte-derived DCs. Furthermore, these IFNs induced IFN-stimulated gene expression predominantly in monocyte-derived DCs. IFN-treated immature DCs derived from murine bone marrow also had a reduced capacity to stimulate T cell proliferative responses to tubercle bacillus Ag 85. Immature DCs derived from either source had a reduced capacity for Ag uptake following IFN treatment that is dose dependent. Immunosuppression is a feature of a number of pestivirus infections; our studies suggest type III IFN production plays a key role in the pathogenesis of this family of viruses. Overall, in a natural host, we have demonstrated a link between the induction of type I and III IFN after acute viral infection and transient immunosuppression.

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“…On the one hand, it is a glycoprotein associated with the virion and essential for the production of infectious particles. On the other hand, this protein is secreted from infected cells and seems to play an important role as a virulence factor in the host via its RNase activity (27,28). The latter activity is also important for the establishment of persistent pestivirus infections (30).…”

Section: Discussionmentioning

confidence: 99%

“…The protein is essential for virus growth (26), but the RNase activity is dispensable. Viruses in which the RNase activity had been knocked out were shown to be clinically attenuated (27,28).…”

mentioning

confidence: 99%

The Pestivirus Glycoprotein Erns Is Anchored in Plane in the Membrane via an Amphipathic Helix

Tews

Meyers

2007

Journal of Biological Chemistry

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100

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Erns is a structural glycoprotein of pestiviruses found to be attached to the virion and to membranes within infected cells via its COOH terminus, although it lacks a hydrophobic anchor sequence. The COOH-terminal sequence was hypothesized to fold into an amphipathic ␣-helix. Alanine insertion scanning revealed that the ability of the E rns COOH terminus to bind membranes is considerably reduced by the insertion of a single amino acid at a wide variety of positions. Mutations decreasing the hydrophobicity of the apolar face of the putative helix led to reduction of membrane association. Proteinase K protection assays showed that E rns translated in vitro in the presence of microsomal membranes was protected, whereas a mutant with an artificial transmembrane region and a short cytosolic tag was shortened by the protease treatment. A tag fused to the COOH terminus of wild type E rns was not accessible for antibodies within digitonin-permeabilized cells, but the variant with the tag located downstream of the artificial transmembrane region was detected under the same conditions. These results are in accordance with the model that the COOH-terminal membrane anchor of E rns represents an amphipathic helix embedded in plane into the membrane. The integrity of the membrane anchor was found to be important for recovery of infectious virus.

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Recovery of Virulent and RNase-Negative Attenuated Type 2 Bovine Viral Diarrhea Viruses from Infectious cDNA Clones

Cited by 53 publications

References 59 publications

The E2 Glycoprotein of Classical Swine Fever Virus Is a Virulence Determinant in Swine

The E2 Glycoprotein of Classical Swine Fever Virus Is a Virulence Determinant in Swine

Type I and III IFNs Produced by Plasmacytoid Dendritic Cells in Response to a Member of theFlaviviridaeSuppress Cellular Immune Responses

The Pestivirus Glycoprotein Erns Is Anchored in Plane in the Membrane via an Amphipathic Helix

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