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Protective Efficacy in Sheep of Adenovirus-Vectored Vaccines against Bluetongue Virus Is Associated with Specific T Cell Responses.

Martín V, Pascual E, Avia M, Peña L, Valcárcel F, Sevilla N - PLoS ONE (2015)

Bottom Line: Sheep vaccinated with Ad5-BTV-VP2 and Ad5-BTV-VP7 or only with Ad5-BTV-VP7 and challenged with BTV showed mild disease symptoms and reduced viremia.This partial protection was achieved in the absence of neutralizing antibodies but strong BTV-specific CD8+ T cell responses in those sheep vaccinated with Ad5-BTV-VP7.These data indicate that rAd5 is a suitable vaccine vector to induce T cell immunity during BTV vaccination and provide new data regarding the relevance of T cell responses in protection during BTV infection.

View Article: PubMed Central - PubMed

Affiliation: Centro de Investigación en Sanidad Animal (CISA-INIA), Instituto Nacional de Investigación Agraria y Alimentaria, Valdeolmos, Madrid, Spain.

ABSTRACT
Bluetongue virus (BTV) is an economically important Orbivirus of the Reoviridae family that causes a hemorrhagic disease in ruminants. Its control has been achieved by inactivated-vaccines that have proven to protect against homologous BTV challenge although unable to induce long-term immunity. Therefore, a more efficient control strategy needs to be developed. Recombinant adenovirus vectors are lead vaccine candidates for protection of several diseases, mainly because of their potency to induce potent T cell immunity. Here we report the induction of humoral and T-cell mediated responses able to protect animals against BTV challenge by recombinant replication-defective human adenovirus serotype 5 (Ad5) expressing either VP7, VP2 or NS3 BTV proteins. First we used the IFNAR(-/-) mouse model system to establish a proof of principle, and afterwards we assayed the protective efficacy in sheep, the natural host of BTV. Mice were completely protected against BTV challenge, developing humoral and BTV-specific CD8+- and CD4+-T cell responses by vaccination with the different rAd5. Sheep vaccinated with Ad5-BTV-VP2 and Ad5-BTV-VP7 or only with Ad5-BTV-VP7 and challenged with BTV showed mild disease symptoms and reduced viremia. This partial protection was achieved in the absence of neutralizing antibodies but strong BTV-specific CD8+ T cell responses in those sheep vaccinated with Ad5-BTV-VP7. These data indicate that rAd5 is a suitable vaccine vector to induce T cell immunity during BTV vaccination and provide new data regarding the relevance of T cell responses in protection during BTV infection.

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Immunogenicity of recombinant adenoviruses Ad5-BTV-NS3, Ad5-BTV-VP7 and Ad5-BTV-VP2 in IFNAR(-/-) mice.Mice were immunized twice with recombinant adenoviruses (group#1: Ad5-BTV-NS3 + Ad5-BTV-VP7 + Ad5-BTV-VP2; group #2: Ad5-BTV-VP2; and group#3: Ad5-BTV-VP7). At D30 post-vaccination, 15 days after the booster injection, mice were challenged with 103 PFUs of BTV-8. (A) Survival curves after challenge. (B) BTV-IgG titers in serum of mice immunized with recombinant adenovirus and after challenge (indicated with an arrow the day of challenge). (C) BTV-specific T cell responses in vaccinated sheep. Splenocytes from 3 mice from group #1 (vaccinated) and 3 mice from the control group inoculated with Ad5-DsRed were obtained 7 days after the booster vaccination and stimulated with BEI-inactivated BTV-8. Data show the percentage of CD8+ and CD4+ T cells producing IFN-γ detected by intracellular cytokine staining. * indicates statistically significant (p< 0.005, Wilcoxon test).
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pone.0143273.g003: Immunogenicity of recombinant adenoviruses Ad5-BTV-NS3, Ad5-BTV-VP7 and Ad5-BTV-VP2 in IFNAR(-/-) mice.Mice were immunized twice with recombinant adenoviruses (group#1: Ad5-BTV-NS3 + Ad5-BTV-VP7 + Ad5-BTV-VP2; group #2: Ad5-BTV-VP2; and group#3: Ad5-BTV-VP7). At D30 post-vaccination, 15 days after the booster injection, mice were challenged with 103 PFUs of BTV-8. (A) Survival curves after challenge. (B) BTV-IgG titers in serum of mice immunized with recombinant adenovirus and after challenge (indicated with an arrow the day of challenge). (C) BTV-specific T cell responses in vaccinated sheep. Splenocytes from 3 mice from group #1 (vaccinated) and 3 mice from the control group inoculated with Ad5-DsRed were obtained 7 days after the booster vaccination and stimulated with BEI-inactivated BTV-8. Data show the percentage of CD8+ and CD4+ T cells producing IFN-γ detected by intracellular cytokine staining. * indicates statistically significant (p< 0.005, Wilcoxon test).

Mentions: Adult IFNAR(-/-) mice are a valid surrogate model to study the effectiveness of BTV vaccines [34]. Therefore, we used IFNAR(-/-) mice as a model to evaluate the immunogenicity and capacity to confer protection against infection with BTV-8 of the Ad5-BTV-NS3, Ad5-BTV-VP2 and Ad5-BTV-VP7. Five groups of mice were inoculated with either Ad5-BTV-VP2 + Ad5-BTV-VP7 + Ad5-BTV-NS3 (group #1, 7 mice), Ad5-BTV-VP2 (group #2, 5 mice), Ad5-BTV-VP7 (group #3, 5 mice), Ad-DsRed (6 mice), or PBS (3 mice). Animals received a booster dose at day 15 post-priming and were challenged at day 15 post-booster with 103 PFUs of BTV-8. Each mouse was bled before the booster vaccination and before challenge and at days 3, 5, 7, 10 and 12 post-challenge (pc) to determine levels of BTV-specific antibody titer and T-cell responses. All the control animals (mice receiving Ad-DsRed or PBS) died at day 4 post-challenge (pc) (Fig 3A), as expected [34]. In sharp contrast, none of the mice vaccinated died or developed any signs associated with BTV replication (Fig 3A) but one mouse of group #2 (vaccinated with Ad5-BTV-VP2) that died at D7 pc. BTV-specific IgG levels increased following immunization in vaccinated groups (Fig 3B) to get high levels before challenge. To evaluate the induction of BTV-specific T-cell responses as a result of vaccination, 3 mice of group #1 were chosen because they were inoculated with the 3 rAds, and 3 mice from the control group, inoculated with Ad-DsRed. These were sacrificed at 7 days after the booster injection and splenocytes were cultured in the presence of BEI-inactivated BTV-8. T-cell responses were measured using flow cytometry analyses for intracellular IFN-γ staining. As shown in Fig 3C, a significant percentage of CD8+ and CD4+ T cells from vaccinated mice were producing IFN-γ (p<0.005, Wilcoxon test). These data indicate that mice immunized with Ad5-BTV-VP2 + Ad5-BTV-VP7 + Ad5-BTV-NS3 (group #1) were able to elicit both humoral and cellular immune responses against BTV which was potent enough to confer protection.


Protective Efficacy in Sheep of Adenovirus-Vectored Vaccines against Bluetongue Virus Is Associated with Specific T Cell Responses.

Martín V, Pascual E, Avia M, Peña L, Valcárcel F, Sevilla N - PLoS ONE (2015)

Immunogenicity of recombinant adenoviruses Ad5-BTV-NS3, Ad5-BTV-VP7 and Ad5-BTV-VP2 in IFNAR(-/-) mice.Mice were immunized twice with recombinant adenoviruses (group#1: Ad5-BTV-NS3 + Ad5-BTV-VP7 + Ad5-BTV-VP2; group #2: Ad5-BTV-VP2; and group#3: Ad5-BTV-VP7). At D30 post-vaccination, 15 days after the booster injection, mice were challenged with 103 PFUs of BTV-8. (A) Survival curves after challenge. (B) BTV-IgG titers in serum of mice immunized with recombinant adenovirus and after challenge (indicated with an arrow the day of challenge). (C) BTV-specific T cell responses in vaccinated sheep. Splenocytes from 3 mice from group #1 (vaccinated) and 3 mice from the control group inoculated with Ad5-DsRed were obtained 7 days after the booster vaccination and stimulated with BEI-inactivated BTV-8. Data show the percentage of CD8+ and CD4+ T cells producing IFN-γ detected by intracellular cytokine staining. * indicates statistically significant (p< 0.005, Wilcoxon test).
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pone.0143273.g003: Immunogenicity of recombinant adenoviruses Ad5-BTV-NS3, Ad5-BTV-VP7 and Ad5-BTV-VP2 in IFNAR(-/-) mice.Mice were immunized twice with recombinant adenoviruses (group#1: Ad5-BTV-NS3 + Ad5-BTV-VP7 + Ad5-BTV-VP2; group #2: Ad5-BTV-VP2; and group#3: Ad5-BTV-VP7). At D30 post-vaccination, 15 days after the booster injection, mice were challenged with 103 PFUs of BTV-8. (A) Survival curves after challenge. (B) BTV-IgG titers in serum of mice immunized with recombinant adenovirus and after challenge (indicated with an arrow the day of challenge). (C) BTV-specific T cell responses in vaccinated sheep. Splenocytes from 3 mice from group #1 (vaccinated) and 3 mice from the control group inoculated with Ad5-DsRed were obtained 7 days after the booster vaccination and stimulated with BEI-inactivated BTV-8. Data show the percentage of CD8+ and CD4+ T cells producing IFN-γ detected by intracellular cytokine staining. * indicates statistically significant (p< 0.005, Wilcoxon test).
Mentions: Adult IFNAR(-/-) mice are a valid surrogate model to study the effectiveness of BTV vaccines [34]. Therefore, we used IFNAR(-/-) mice as a model to evaluate the immunogenicity and capacity to confer protection against infection with BTV-8 of the Ad5-BTV-NS3, Ad5-BTV-VP2 and Ad5-BTV-VP7. Five groups of mice were inoculated with either Ad5-BTV-VP2 + Ad5-BTV-VP7 + Ad5-BTV-NS3 (group #1, 7 mice), Ad5-BTV-VP2 (group #2, 5 mice), Ad5-BTV-VP7 (group #3, 5 mice), Ad-DsRed (6 mice), or PBS (3 mice). Animals received a booster dose at day 15 post-priming and were challenged at day 15 post-booster with 103 PFUs of BTV-8. Each mouse was bled before the booster vaccination and before challenge and at days 3, 5, 7, 10 and 12 post-challenge (pc) to determine levels of BTV-specific antibody titer and T-cell responses. All the control animals (mice receiving Ad-DsRed or PBS) died at day 4 post-challenge (pc) (Fig 3A), as expected [34]. In sharp contrast, none of the mice vaccinated died or developed any signs associated with BTV replication (Fig 3A) but one mouse of group #2 (vaccinated with Ad5-BTV-VP2) that died at D7 pc. BTV-specific IgG levels increased following immunization in vaccinated groups (Fig 3B) to get high levels before challenge. To evaluate the induction of BTV-specific T-cell responses as a result of vaccination, 3 mice of group #1 were chosen because they were inoculated with the 3 rAds, and 3 mice from the control group, inoculated with Ad-DsRed. These were sacrificed at 7 days after the booster injection and splenocytes were cultured in the presence of BEI-inactivated BTV-8. T-cell responses were measured using flow cytometry analyses for intracellular IFN-γ staining. As shown in Fig 3C, a significant percentage of CD8+ and CD4+ T cells from vaccinated mice were producing IFN-γ (p<0.005, Wilcoxon test). These data indicate that mice immunized with Ad5-BTV-VP2 + Ad5-BTV-VP7 + Ad5-BTV-NS3 (group #1) were able to elicit both humoral and cellular immune responses against BTV which was potent enough to confer protection.

Bottom Line: Sheep vaccinated with Ad5-BTV-VP2 and Ad5-BTV-VP7 or only with Ad5-BTV-VP7 and challenged with BTV showed mild disease symptoms and reduced viremia.This partial protection was achieved in the absence of neutralizing antibodies but strong BTV-specific CD8+ T cell responses in those sheep vaccinated with Ad5-BTV-VP7.These data indicate that rAd5 is a suitable vaccine vector to induce T cell immunity during BTV vaccination and provide new data regarding the relevance of T cell responses in protection during BTV infection.

View Article: PubMed Central - PubMed

Affiliation: Centro de Investigación en Sanidad Animal (CISA-INIA), Instituto Nacional de Investigación Agraria y Alimentaria, Valdeolmos, Madrid, Spain.

ABSTRACT
Bluetongue virus (BTV) is an economically important Orbivirus of the Reoviridae family that causes a hemorrhagic disease in ruminants. Its control has been achieved by inactivated-vaccines that have proven to protect against homologous BTV challenge although unable to induce long-term immunity. Therefore, a more efficient control strategy needs to be developed. Recombinant adenovirus vectors are lead vaccine candidates for protection of several diseases, mainly because of their potency to induce potent T cell immunity. Here we report the induction of humoral and T-cell mediated responses able to protect animals against BTV challenge by recombinant replication-defective human adenovirus serotype 5 (Ad5) expressing either VP7, VP2 or NS3 BTV proteins. First we used the IFNAR(-/-) mouse model system to establish a proof of principle, and afterwards we assayed the protective efficacy in sheep, the natural host of BTV. Mice were completely protected against BTV challenge, developing humoral and BTV-specific CD8+- and CD4+-T cell responses by vaccination with the different rAd5. Sheep vaccinated with Ad5-BTV-VP2 and Ad5-BTV-VP7 or only with Ad5-BTV-VP7 and challenged with BTV showed mild disease symptoms and reduced viremia. This partial protection was achieved in the absence of neutralizing antibodies but strong BTV-specific CD8+ T cell responses in those sheep vaccinated with Ad5-BTV-VP7. These data indicate that rAd5 is a suitable vaccine vector to induce T cell immunity during BTV vaccination and provide new data regarding the relevance of T cell responses in protection during BTV infection.

Show MeSH
Related in: MedlinePlus