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Multiple genome segments determine virulence of bluetongue virus serotype 8.

Janowicz A, Caporale M, Shaw A, Gulletta S, Di Gialleonardo L, Ratinier M, Palmarini M - J. Virol. (2015)

Bottom Line: We found that partial attenuation of BTV8L in IFNAR(-/-) mice was achieved by simply replacing genomic segment 2 (Seg2, encoding VP2) or Seg10 (encoding NS3) with the BTV8H homologous segments.Replication of BTV8H was relatively poor in interferon (IFN)-competent primary ovine endothelial cells compared to replication of BTV8L, and this phenotype was determined by several viral genomic segments, including Seg4 and Seg9.The possibility to determine the pathogenicity of virus isolates on the basis of their genome sequences will help in the design of control strategies that fit the risk posed by new emerging BTV strains.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Glasgow, United Kingdom.

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IFN production and gene expression induced by infection of OvEC by rgBTV8L, rgBTV8H, and BTV8L/BTV8H monoreassortants. OvEC were infected with rgBTV8L, rgBTV8H, and monoreassortants within the BTV8L backbone (MOI of 1). (A) IFN protection assay. Supernatants were collected at 18 h p.i., inactivated by UV treatment, and used in a biological assay to estimate the amount of IFN present, as described in Materials and Methods. The only major differences were observed in cells infected with BTV8L+S9H, where the amount of IFN released was significantly lower than what was found in cells infected with rgBTV8L (P < 0.05; one-way analysis of variance followed by Dunnett's multiple-comparison test to dissect individual interactions). (B) IFN-β, ActB, RSAD2, and Mx1 gene expression. mRNA was measured by qPCR in OvEC at 18 h p.i. with parental and reassortant viruses (MOI of 1) as described in Materials and Methods. Mock-treated and UIFN-treated cells were used as controls. Panels show gene expression relative to rgBTV8L and normalized to GAPDH gene levels.
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Figure 7: IFN production and gene expression induced by infection of OvEC by rgBTV8L, rgBTV8H, and BTV8L/BTV8H monoreassortants. OvEC were infected with rgBTV8L, rgBTV8H, and monoreassortants within the BTV8L backbone (MOI of 1). (A) IFN protection assay. Supernatants were collected at 18 h p.i., inactivated by UV treatment, and used in a biological assay to estimate the amount of IFN present, as described in Materials and Methods. The only major differences were observed in cells infected with BTV8L+S9H, where the amount of IFN released was significantly lower than what was found in cells infected with rgBTV8L (P < 0.05; one-way analysis of variance followed by Dunnett's multiple-comparison test to dissect individual interactions). (B) IFN-β, ActB, RSAD2, and Mx1 gene expression. mRNA was measured by qPCR in OvEC at 18 h p.i. with parental and reassortant viruses (MOI of 1) as described in Materials and Methods. Mock-treated and UIFN-treated cells were used as controls. Panels show gene expression relative to rgBTV8L and normalized to GAPDH gene levels.

Mentions: Next, we wanted to establish whether BTV8H was a more potent IFN inducer than the parental BTV8L. To this end, we measured IFN production in the supernatants of OvEC infected with the same set of viruses as above. rgBTV8H and most of the monoreassortants induced similar amounts of IFN. However, statistically significant differences (P < 0.05) were observed between rgBTV8L and BTV8L+S9H (Fig. 7A).


Multiple genome segments determine virulence of bluetongue virus serotype 8.

Janowicz A, Caporale M, Shaw A, Gulletta S, Di Gialleonardo L, Ratinier M, Palmarini M - J. Virol. (2015)

IFN production and gene expression induced by infection of OvEC by rgBTV8L, rgBTV8H, and BTV8L/BTV8H monoreassortants. OvEC were infected with rgBTV8L, rgBTV8H, and monoreassortants within the BTV8L backbone (MOI of 1). (A) IFN protection assay. Supernatants were collected at 18 h p.i., inactivated by UV treatment, and used in a biological assay to estimate the amount of IFN present, as described in Materials and Methods. The only major differences were observed in cells infected with BTV8L+S9H, where the amount of IFN released was significantly lower than what was found in cells infected with rgBTV8L (P < 0.05; one-way analysis of variance followed by Dunnett's multiple-comparison test to dissect individual interactions). (B) IFN-β, ActB, RSAD2, and Mx1 gene expression. mRNA was measured by qPCR in OvEC at 18 h p.i. with parental and reassortant viruses (MOI of 1) as described in Materials and Methods. Mock-treated and UIFN-treated cells were used as controls. Panels show gene expression relative to rgBTV8L and normalized to GAPDH gene levels.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 7: IFN production and gene expression induced by infection of OvEC by rgBTV8L, rgBTV8H, and BTV8L/BTV8H monoreassortants. OvEC were infected with rgBTV8L, rgBTV8H, and monoreassortants within the BTV8L backbone (MOI of 1). (A) IFN protection assay. Supernatants were collected at 18 h p.i., inactivated by UV treatment, and used in a biological assay to estimate the amount of IFN present, as described in Materials and Methods. The only major differences were observed in cells infected with BTV8L+S9H, where the amount of IFN released was significantly lower than what was found in cells infected with rgBTV8L (P < 0.05; one-way analysis of variance followed by Dunnett's multiple-comparison test to dissect individual interactions). (B) IFN-β, ActB, RSAD2, and Mx1 gene expression. mRNA was measured by qPCR in OvEC at 18 h p.i. with parental and reassortant viruses (MOI of 1) as described in Materials and Methods. Mock-treated and UIFN-treated cells were used as controls. Panels show gene expression relative to rgBTV8L and normalized to GAPDH gene levels.
Mentions: Next, we wanted to establish whether BTV8H was a more potent IFN inducer than the parental BTV8L. To this end, we measured IFN production in the supernatants of OvEC infected with the same set of viruses as above. rgBTV8H and most of the monoreassortants induced similar amounts of IFN. However, statistically significant differences (P < 0.05) were observed between rgBTV8L and BTV8L+S9H (Fig. 7A).

Bottom Line: We found that partial attenuation of BTV8L in IFNAR(-/-) mice was achieved by simply replacing genomic segment 2 (Seg2, encoding VP2) or Seg10 (encoding NS3) with the BTV8H homologous segments.Replication of BTV8H was relatively poor in interferon (IFN)-competent primary ovine endothelial cells compared to replication of BTV8L, and this phenotype was determined by several viral genomic segments, including Seg4 and Seg9.The possibility to determine the pathogenicity of virus isolates on the basis of their genome sequences will help in the design of control strategies that fit the risk posed by new emerging BTV strains.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Glasgow, United Kingdom.

Show MeSH
Related in: MedlinePlus