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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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Rectal body temperature and clinical signs of vaccinated sheep with recombinant adenovirus.(A) The rectal temperature was recorded daily for a total period of 14 days. Temperature for individual animals in each group is shown. Fever was considered when temperature was above 40°C (indicated with dotted line). The asterisks indicate statistically significant at peak of fever (D6 and D7 pi) (p = 0.0009 for Ad5-BTV-VP7 and p = 0.0036 for Ad-BTV-VP7+Ad5-BTV-VP2) by unpaired t test with Welch’s correction. (B) Clinical signs recorded in vaccinated and control sheep after challenge with BTV-8. Sheep were scored daily using a clinical index score that takes into account general symptoms, fever, respiratory signs or death as described in Material and Methods. Total scores are given for individual animals as the accumulative values for each symptom collected through the observation period. The asterisks indicate statistically significant (* p = 0.0194; *** p< 0.0001) by Wilcoxon Test.
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pone.0143273.g004: Rectal body temperature and clinical signs of vaccinated sheep with recombinant adenovirus.(A) The rectal temperature was recorded daily for a total period of 14 days. Temperature for individual animals in each group is shown. Fever was considered when temperature was above 40°C (indicated with dotted line). The asterisks indicate statistically significant at peak of fever (D6 and D7 pi) (p = 0.0009 for Ad5-BTV-VP7 and p = 0.0036 for Ad-BTV-VP7+Ad5-BTV-VP2) by unpaired t test with Welch’s correction. (B) Clinical signs recorded in vaccinated and control sheep after challenge with BTV-8. Sheep were scored daily using a clinical index score that takes into account general symptoms, fever, respiratory signs or death as described in Material and Methods. Total scores are given for individual animals as the accumulative values for each symptom collected through the observation period. The asterisks indicate statistically significant (* p = 0.0194; *** p< 0.0001) by Wilcoxon Test.

Mentions: The efficacy of the new generated adenoviruses as a BTV vaccine in IFNAR(-/-) mice encouraged us to investigate their efficiency in sheep. Ad5-BTV-VP2 and Ad5-BTV-VP7 were chosen based on their capacity to induce humoral and T cell responses as well as because the lack of NS3 directed antibodies by the Ad5-BTV-VP2 or Ad5-BTV-VP7 vaccine may enable differentiation of infected from vaccinated animals (DIVA principle). Four different vaccination groups were used. In group # 1, five sheep (numbered 1 to 5) were vaccinated with Ad5-BTV-VP7; in group # 2, five sheep (numbered 6 to 10) were vaccinated with Ad5-BTV-VP7 and Ad5-BTV-VP2; in group # 3, three sheep (numbered 11 to 13) were vaccinated with Ad5-DsRed; and in group # 4, three sheep (numbered 14 to 16) were administered PBS. All groups received a booster injection at day 15 post-immunization with the same dose and antigen composition. Finally, animals were challenged at 15 days post-booster with BTV-8. During the experiment, clinical signs and rectal temperature were recorded (Fig 4 and S2 Fig). Sheep in both control groups showed increased temperature of >40°C at D5pc (Fig 4A), declining to normal values by D9pc. Vaccinated sheep challenged with BTV-8 did not show high temperature but one animal in group #1 (vaccinated with Ad5-BTV-VP7) that however did not develop any additional clinical sign and one animal in group #2 (vaccinated with Ad5-BTV-VP7 + Ad5-BTV-VP2) that developed clinical signs. All animals in the control groups developed clinical signs associated to BTV infection (see Material and Methods), with an average clinical score of 28.7 ± 2.7 (Fig 4B). In stark contrast, the animals vaccinated with Ad5-BTV-VP7 developed very mild clinical signs of disease (average score of 3.8 ± 0.5) but significantly reduced when compared with control groups (p value < 0.0001, Student’s t-test). Four sheep of the group vaccinated with Ad5-BTV-VP7 + Ad5-BTV-VP2 showed also reduced clinical signs but one animal developed clinical signs similar to control groups (clinical score of 24) (average clinical score of the group 10 ± 8.3, significantly lower that the control groups, p value = 0.0194, Student’s t-test. In addition to the observed clinical signs recorded daily, 3 sheep per group (1 from each control group) were sacrificed at day D9pc to evaluate the lesions presented post-mortem. All control sheep showed gross changes such as haemorrhages, ulceration of the mucosal of the upper gastrointestinal tract; haemorrhages in lymph nodes and pulmonary artery and pulmonary oedema as main lesions. Vaccinated animals (group #1 and #2) did not show any of these lesions. Histological examination of the lungs showed hyperaemia, severe alveolar oedema with interstitial cellularity in control animals whereas normal lung histology is found in vaccinated sheep (Fig 4A). Comparison of the results of vaccinated sheep with control groups suggests that the new adenoviral BTV vaccine protects sheep against BTV-8 challenge.


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)

Rectal body temperature and clinical signs of vaccinated sheep with recombinant adenovirus.(A) The rectal temperature was recorded daily for a total period of 14 days. Temperature for individual animals in each group is shown. Fever was considered when temperature was above 40°C (indicated with dotted line). The asterisks indicate statistically significant at peak of fever (D6 and D7 pi) (p = 0.0009 for Ad5-BTV-VP7 and p = 0.0036 for Ad-BTV-VP7+Ad5-BTV-VP2) by unpaired t test with Welch’s correction. (B) Clinical signs recorded in vaccinated and control sheep after challenge with BTV-8. Sheep were scored daily using a clinical index score that takes into account general symptoms, fever, respiratory signs or death as described in Material and Methods. Total scores are given for individual animals as the accumulative values for each symptom collected through the observation period. The asterisks indicate statistically significant (* p = 0.0194; *** p< 0.0001) by Wilcoxon Test.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4664254&req=5

pone.0143273.g004: Rectal body temperature and clinical signs of vaccinated sheep with recombinant adenovirus.(A) The rectal temperature was recorded daily for a total period of 14 days. Temperature for individual animals in each group is shown. Fever was considered when temperature was above 40°C (indicated with dotted line). The asterisks indicate statistically significant at peak of fever (D6 and D7 pi) (p = 0.0009 for Ad5-BTV-VP7 and p = 0.0036 for Ad-BTV-VP7+Ad5-BTV-VP2) by unpaired t test with Welch’s correction. (B) Clinical signs recorded in vaccinated and control sheep after challenge with BTV-8. Sheep were scored daily using a clinical index score that takes into account general symptoms, fever, respiratory signs or death as described in Material and Methods. Total scores are given for individual animals as the accumulative values for each symptom collected through the observation period. The asterisks indicate statistically significant (* p = 0.0194; *** p< 0.0001) by Wilcoxon Test.
Mentions: The efficacy of the new generated adenoviruses as a BTV vaccine in IFNAR(-/-) mice encouraged us to investigate their efficiency in sheep. Ad5-BTV-VP2 and Ad5-BTV-VP7 were chosen based on their capacity to induce humoral and T cell responses as well as because the lack of NS3 directed antibodies by the Ad5-BTV-VP2 or Ad5-BTV-VP7 vaccine may enable differentiation of infected from vaccinated animals (DIVA principle). Four different vaccination groups were used. In group # 1, five sheep (numbered 1 to 5) were vaccinated with Ad5-BTV-VP7; in group # 2, five sheep (numbered 6 to 10) were vaccinated with Ad5-BTV-VP7 and Ad5-BTV-VP2; in group # 3, three sheep (numbered 11 to 13) were vaccinated with Ad5-DsRed; and in group # 4, three sheep (numbered 14 to 16) were administered PBS. All groups received a booster injection at day 15 post-immunization with the same dose and antigen composition. Finally, animals were challenged at 15 days post-booster with BTV-8. During the experiment, clinical signs and rectal temperature were recorded (Fig 4 and S2 Fig). Sheep in both control groups showed increased temperature of >40°C at D5pc (Fig 4A), declining to normal values by D9pc. Vaccinated sheep challenged with BTV-8 did not show high temperature but one animal in group #1 (vaccinated with Ad5-BTV-VP7) that however did not develop any additional clinical sign and one animal in group #2 (vaccinated with Ad5-BTV-VP7 + Ad5-BTV-VP2) that developed clinical signs. All animals in the control groups developed clinical signs associated to BTV infection (see Material and Methods), with an average clinical score of 28.7 ± 2.7 (Fig 4B). In stark contrast, the animals vaccinated with Ad5-BTV-VP7 developed very mild clinical signs of disease (average score of 3.8 ± 0.5) but significantly reduced when compared with control groups (p value < 0.0001, Student’s t-test). Four sheep of the group vaccinated with Ad5-BTV-VP7 + Ad5-BTV-VP2 showed also reduced clinical signs but one animal developed clinical signs similar to control groups (clinical score of 24) (average clinical score of the group 10 ± 8.3, significantly lower that the control groups, p value = 0.0194, Student’s t-test. In addition to the observed clinical signs recorded daily, 3 sheep per group (1 from each control group) were sacrificed at day D9pc to evaluate the lesions presented post-mortem. All control sheep showed gross changes such as haemorrhages, ulceration of the mucosal of the upper gastrointestinal tract; haemorrhages in lymph nodes and pulmonary artery and pulmonary oedema as main lesions. Vaccinated animals (group #1 and #2) did not show any of these lesions. Histological examination of the lungs showed hyperaemia, severe alveolar oedema with interstitial cellularity in control animals whereas normal lung histology is found in vaccinated sheep (Fig 4A). Comparison of the results of vaccinated sheep with control groups suggests that the new adenoviral BTV vaccine protects sheep against BTV-8 challenge.

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