RESUMO
Vaccination is the best strategy to control Paratuberculosis (PTB), which is a significant disease in cattle and sheep. Previously we showed the humoral and cellular immune response induced by a novel vaccine candidate against PTB based on the Argentinian Mycobacterium avium subspecies paratuberculosis (Map) 6611 strain. To improve 6611 immunogenicity and efficacy, we evaluated this vaccine candidate in mice with two different adjuvants and a heterologous boost with a recombinant modified vaccinia Ankara virus (MVA) expressing the antigen 85A (MVA85A). We observed that boosting with MVA85A did not improve total IgG or specific isotypes in serum induced by one or two doses of 6611 formulated with incomplete Freund's adjuvant (IFA). However, when 6611 was formulated with ISA201 adjuvant, MVA85A boost enhanced the production of IFNγ, Th1/Th17 cytokines (IL-2, TNF, IL-17A) and IL-6, IL-4 and IL-10. Also, this group showed the highest levels of IgG2b and IgG3 isotypes, both important for better protection against Map infection in the murine model. Finally, the heterologous scheme elicited the highest levels of protection after Map challenge (lowest CFU count and liver lesion score). In conclusion, our results encourage further evaluation of 6611 strain + ISA201 prime and MVA85A boost in bovines.
Assuntos
Adjuvantes Imunológicos , Anticorpos Antibacterianos , Citocinas , Modelos Animais de Doenças , Imunização Secundária , Imunoglobulina G , Mycobacterium avium subsp. paratuberculosis , Paratuberculose , Animais , Mycobacterium avium subsp. paratuberculosis/imunologia , Imunização Secundária/métodos , Camundongos , Paratuberculose/prevenção & controle , Paratuberculose/imunologia , Imunoglobulina G/sangue , Citocinas/metabolismo , Feminino , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Adjuvantes Imunológicos/administração & dosagem , Vacinas Bacterianas/imunologia , Vacinas Bacterianas/administração & dosagem , Camundongos Endogâmicos BALB C , Vaccinia virus/imunologia , Vaccinia virus/genética , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/genética , Imunidade Celular/imunologia , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/administração & dosagem , Adjuvante de Freund/administração & dosagem , Adjuvante de Freund/imunologiaRESUMO
Bovine respiratory syncytial virus (BRSV) affects both beef and dairy cattle, reaching morbidity and mortality rates of 60-80% and 20%, respectively. The aim of this study was to obtain a recombinant MVA expressing the BRSV F protein (MVA-F) as a vaccine against BRSV and to evaluate the immune response induced by MVA-F after systemic immunization in homologous and heterologous vaccination (MVA-F alone or combined with a subunit vaccine), and after intranasal immunization of mice. MVA-F administered by intraperitoneal route in a homologous scheme elicited levels of neutralizing antibodies similar to those obtained with inactivated BRSV as well as better levels of IFN-γ secretion. In addition, nasal administration of MVA-F elicited local and systemic immunity with a Th1 profile. This study suggests that MVA-F is a good candidate for further evaluations combining intranasal and intramuscular routes, in order to induce local and systemic immune responses, to improve the vaccine efficacy against BRSV infection.
Assuntos
Administração Intranasal , Camundongos Endogâmicos BALB C , Vírus Sincicial Respiratório Bovino , Animais , Vírus Sincicial Respiratório Bovino/imunologia , Camundongos , Feminino , Bovinos , Proteínas Virais de Fusão/imunologia , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/administração & dosagem , Vacinas contra Vírus Sincicial Respiratório/imunologia , Vacinas contra Vírus Sincicial Respiratório/administração & dosagem , Vetores Genéticos , Infecções por Vírus Respiratório Sincicial/prevenção & controle , Infecções por Vírus Respiratório Sincicial/imunologia , Infecções por Vírus Respiratório Sincicial/veterinária , Vaccinia virus/imunologia , Vaccinia virus/genética , Anticorpos Antivirais/sangue , Imunidade nas Mucosas , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Imunização/métodos , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/administração & dosagemRESUMO
Mycobacterium tuberculosis (Mtb) infection is one of the leading causes of death worldwide. The Modified Vaccinia Ankara (MVA) vaccine vector expressing the mycobacterial antigen 85A (MVA85A) was demonstrated to be safe, although it did not improve BCG efficacy, denoting the need to search for improved tuberculosis vaccines. In this work, we investigated the effect of IL-12 DNA -as an adjuvant- on an Ag85A DNA prime/MVA85A boost vaccination regimen. We evaluated the immune response profile elicited in mice and the protection conferred against intratracheal Mtb H37Rv challenge. We observed that the immunization scheme including DNA-A85A+DNA-IL-12/MVA85A induced a strong IFN-γ production to Ag85A in vitro, with a significant expansion of IFN-γ+CD4+ and IFN-γ+CD8+ anti-Ag85A lymphocytes. Furthermore, we also detected a significant increase in the proportion of specific CD8+CD107+ T cells against Ag85A. Additionally, inclusion of IL-12 DNA in the DNA-A85A/MVA85A vaccine scheme induced a marked augment in anti-Ag85A IgG levels. Interestingly, after 30 days of infection with Mtb H37Rv, DNA-A85A+DNA-IL-12/MVA85A vaccinated mice displayed a significant reduction in lung bacterial burden. Together, our findings suggest that IL-12 DNA might be useful as a molecular adjuvant in an Ag85A DNA/MVA prime-boost vaccine against Mtb infection.
Assuntos
Mycobacterium tuberculosis , Vacinas contra a Tuberculose , Tuberculose , Vacinas de DNA , Aciltransferases/genética , Animais , Antígenos de Bactérias/genética , Vacina BCG , DNA , Imunização Secundária , Interleucina-12/genética , Camundongos , Mycobacterium tuberculosis/genética , Tuberculose/prevenção & controle , Vacinas de DNA/genéticaRESUMO
The baculovirus Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) infects lepidopteran invertebrates as natural hosts, although it also has been used as display vector for vaccine development. In this work, we evaluated the effectiveness of repetitive doses of AcMNPV-based vectors on the cytotoxic immune response specific to the capsid-displayed heterologous antigen ovalbumin (OVA). Our results demonstrate that baculovirus vectors induce a boosting effect in the cytotoxic immune response to OVA, making possible to recover the levels obtained in the primary response. Moreover, mice preimmunized with wild-type baculovirus showed a complete lack of antigen-specific CD8 cytotoxic T lymphocytes (CTLs) that may be related to the presence of antibodies directed to baculoviral surface proteins, particularly to GP64. However, baculovirus was able to induce the innate immune response in spite of a previous response against this vector, although some quantitative differences reflect a distinct activation of the immune cells in prime and boost. This is the first report in which the novel capsid display strategy is evaluated in prime-boost schemes to improve efficient CTL responses.
Assuntos
Proteínas do Capsídeo/imunologia , Capsídeo/imunologia , Nucleopoliedrovírus/imunologia , Vacinação , Animais , Antígenos Virais/imunologia , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular , Citocinas/sangue , Feminino , Interações entre Hospedeiro e Microrganismos/imunologia , Imunidade Inata , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/imunologia , Spodoptera/imunologia , Spodoptera/virologia , Linfócitos T Citotóxicos/imunologiaRESUMO
Canarypox viruses (CNPV) are excellent candidates to develop recombinant vector vaccines due to both their capability to induce protective immune responses and their incompetence to replicate in mammalian cells (safety profile). In addition, CNPV and the derived recombinants can be manipulated under biosafety level 1 conditions. There is no commercially available system to obtain recombinant CNPV; however, the methodology and tools required to develop recombinant vaccinia virus (VV), prototype of the Poxviridae family, can be easily adapted. This chapter provides protocols for the generation, plaque isolation, molecular characterization, amplification and purification of recombinant CNPV.
Assuntos
Vírus da Varíola dos Canários/crescimento & desenvolvimento , Fibroblastos/virologia , Vacinas Sintéticas/imunologia , Animais , Vírus da Varíola dos Canários/genética , Vírus da Varíola dos Canários/imunologia , Linhagem Celular , Galinhas , Fibroblastos/imunologia , Vacinas Virais/imunologiaRESUMO
In this study, we evaluated the immunogenicity and efficacy of mucosal delivery of a recombinant modified vaccinia Ankara virus (MVA) expressing the secreted version of bovine herpesvirus type 1 (BoHV-1) glycoprotein D (MVA-gDs) without addition of adjuvant in two animal models. First, we demonstrated the capability of MVA-gDs of inducing both local and systemic anti-gD humoral immune response after intranasal immunization of mice. Then, we confirmed that two doses of MVA-gDs administered intranasally to rabbits induced systemic anti-gD antibodies and conferred protection against BoHV-1 challenge. Our results show the potential of using MVA as a vector for the rational design of veterinary vaccines capable of inducing specific and protective immune responses both at local and systemic level.
Assuntos
Portadores de Fármacos , Infecções por Herpesviridae/prevenção & controle , Herpesvirus Bovino 1/imunologia , Vacinas contra Herpesvirus/imunologia , Vaccinia virus/genética , Proteínas Virais/imunologia , Administração Intranasal , Animais , Anticorpos Antivirais/sangue , Modelos Animais de Doenças , Feminino , Infecções por Herpesviridae/imunologia , Vacinas contra Herpesvirus/administração & dosagem , Vacinas contra Herpesvirus/genética , Camundongos Endogâmicos BALB C , Coelhos , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia , Proteínas Virais/genéticaRESUMO
MVA is an attenuated vector that still retains immunomodulatory genes. We have previously reported its optimization after deleting the C12L gene, coding for the IL-18 binding-protein. Here, we analyzed the immunogenicity of MVA vectors harboring the simultaneous deletion of A44L, related to steroid synthesis and A46R, a TLR-signaling inhibitor (MVAΔA44L-A46R); or also including a deletion of C12L (MVAΔC12L/ΔA44L-A46R). The absence of biological activities of the deleted genes in the MVA vectors was demonstrated. Adaptive T-cell responses against VACV epitopes, evaluated in spleen and draining lymph-nodes of C57Bl/6 mice at acute/memory phases, were of higher magnitude in those animals that received deleted MVAs compared to MVAwt. MVAΔC12L/ΔA44L-A46R generated cellular specific memory responses of higher quality characterized by bifunctionality (CD107a/bâº/IFN-γâº) and proliferation capacity. Deletion of selected genes from MVA generated innate immune responses with higher levels of determining cytokines related to T-cell response generation, such as IL-12, IFN-γ, as well as IL-1ß and IFN-ß. This study describes for the first time that simultaneous deletion of the A44L, A46R and C12L genes from MVA improved its immunogenicity by enhancing the host adaptive and innate immune responses, suggesting that this approach comprises an appropriate strategy to increase the MVA vaccine potential.
Assuntos
Imunidade Inata , Deleção de Sequência , Linfócitos T/imunologia , Vaccinia virus/genética , Vaccinia virus/imunologia , Proteínas Virais/genética , Animais , Antígenos Virais/imunologia , Citocinas/metabolismo , Epitopos/imunologia , Linfonodos/imunologia , Camundongos Endogâmicos C57BL , Baço/imunologiaRESUMO
A recombinant modified vaccinia Ankara (MVA) virus expressing mature viral protein 2 (VP2) of the infectious bursal disease virus (IBDV) was constructed to develop MVA-based vaccines for poultry. We demonstrated that this recombinant virus was able to induce a specific immune response by observing the production of anti-IBDV-seroneutralizing antibodies in specific pathogen-free chickens. Besides, as the epitopes of VP2 responsible to induce IBDV-neutralizing antibodies are discontinuous, our results suggest that VP2 protein expressed from MVA-VP2 maintained the correct conformational structure. To our knowledge, this is the first report on the usefulness of MVA-based vectors for developing recombinant vaccines for poultry.
Assuntos
Infecções por Birnaviridae/veterinária , Vírus da Doença Infecciosa da Bursa/imunologia , Doenças das Aves Domésticas/prevenção & controle , Vaccinia virus/genética , Proteínas Estruturais Virais/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais , Infecções por Birnaviridae/prevenção & controle , Células Cultivadas , Embrião de Galinha , Galinhas , Fibroblastos/metabolismo , Doenças das Aves Domésticas/virologia , Organismos Livres de Patógenos Específicos , Vaccinia virus/imunologia , Vaccinia virus/metabolismo , Proteínas Estruturais Virais/genética , Proteínas Estruturais Virais/metabolismoRESUMO
BACKGROUND: Modified Vaccinia Ankara (MVA) is an attenuated strain of Vaccinia virus (VACV) currently employed in many clinical trials against HIV/AIDS and other diseases. MVA still retains genes involved in host immune response evasion, enabling its optimization by removing some of them. The aim of this study was to evaluate cellular immune responses (CIR) induced by an IL-18 binding protein gene (C12L) deleted vector (MVAΔC12L). METHODOLOGY/PRINCIPAL FINDINGS: BALB/c and C57BL/6 mice were immunized with different doses of MVAΔC12L or MVA wild type (MVAwt), then CIR to VACV epitopes in immunogenic proteins were evaluated in spleen and draining lymph nodes at acute and memory phases (7 and 40 days post-immunization respectively). Compared with parental MVAwt, MVAΔC12L immunization induced a significant increase of two to three-fold in CD8(+) and CD4(+) T-cell responses to different VACV epitopes, with increased percentage of anti-VACV cytotoxic CD8(+) T-cells (CD107a/b(+)) during the acute phase of the response. Importantly, the immunogenicity enhancement was also observed after MVAΔC12L inoculation with different viral doses and by distinct routes (systemic and mucosal). Potentiation of MVA's CIR was also observed during the memory phase, in correlation with a higher protection against an intranasal challenge with VACV WR. Of note, we could also show a significant increase in the CIR against HIV antigens such as Env, Gag, Pol and Nef from different subtypes expressed from two recombinants of MVAΔC12L during heterologous DNA prime/MVA boost vaccination regimens. CONCLUSIONS/SIGNIFICANCE: This study demonstrates the relevance of IL-18 bp contribution in the immune response evasion during MVA infection. Our findings clearly show that the deletion of the viral IL-18 bp gene is an effective approach to increase MVA vaccine efficacy, as immunogenicity improvements were observed against vector antigens and more importantly to HIV antigens.