RESUMO
Marek's disease (MD), caused by the Marek's disease virus (MDV), is a common infectious tumor disease in chickens and was the first neoplastic disease preventable by vaccination. However, the vaccine cannot completely prevent virulent MDV infections, allowing both the vaccine and virulent MDV to coexist in the same chicken for extended periods. This study aims to investigate the changes in viral load of the very virulent strain Md5 and the rHVT-IBD vaccine in different chicken tissues using a real-time PCR assay. The results showed that the rHVT-IBD vaccine significantly reduced the viral load of MDV-Md5 in different organs, while the load of rHVT-IBD was significantly increased when co-infected with Md5. Additionally, co-infection with Md5 and rHVT-IBD in chickens not only changed the original viral load of both viruses but also affected the positive rate of Md5 at 14 days post-vaccination. The positive rate decreased from 100% to 14.29% (feather tips), 0% (skin), 33.33% (liver), 16.67% (spleen), 28.57% (thymus), 33.33% (bursa), and 66.67% (PBL), respectively. This study enhances our understanding of the interactions between HVT vector vaccines and very virulent MDV in chickens and provides valuable insights for the future development of MD vaccines.
Assuntos
Galinhas , Coinfecção , Vacinas contra Doença de Marek , Doença de Marek , Doenças das Aves Domésticas , Carga Viral , Animais , Doença de Marek/virologia , Doença de Marek/prevenção & controle , Doença de Marek/imunologia , Galinhas/virologia , Coinfecção/virologia , Coinfecção/veterinária , Doenças das Aves Domésticas/virologia , Doenças das Aves Domésticas/prevenção & controle , Vacinas contra Doença de Marek/imunologia , Vacinas contra Doença de Marek/genética , Virulência , Herpesvirus Meleagrídeo 1/imunologia , Herpesvirus Meleagrídeo 1/genética , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/genética , Herpesvirus Galináceo 2/genética , Herpesvirus Galináceo 2/imunologia , Herpesvirus Galináceo 2/patogenicidade , Vacinação , Vetores Genéticos/genéticaRESUMO
Marek's disease (MD), caused by the Marek's disease virus, is a lymphoproliferative disease in chickens that can be controlled by vaccination. However, the current vaccines can limit tumor growth and death but not virus replication and transmission. The present study aimed to evaluate host responses following intramuscular injection of an mRNA vaccine encoding gB and pp38 proteins of the MDV within the first 36 h. The vaccine was injected in low and high doses using prime and prime-boost strategies. The expression of type I and II interferons (IFNs), a panel of interferon-stimulated genes, and two key antiviral cytokines, IL-1ß and IL-2, were measured in spleen and lungs after vaccination. The transcriptional analysis of the above genes showed significant increases in the expression of MDA5, Myd88, IFN-α, IFN-ß, IFN-γ, IRF7, OAS, Mx1, and IL-2 in both the spleen and lungs within the first 36 h of immunization. Secondary immunization increased expression of all the above genes in the lungs. In contrast, only IFN-γ, MDA5, MyD88, Mx1, and OAS showed significant upregulation in the spleen after the secondary immunization. This study shows that two doses of the MDV mRNA vaccine encoding gB and pp38 antigens activate innate and adaptive responses and induce an antiviral state in chickens.
Assuntos
Galinhas , Citocinas , Herpesvirus Galináceo 2 , Vacinas contra Doença de Marek , Doença de Marek , Animais , Galinhas/imunologia , Doença de Marek/prevenção & controle , Doença de Marek/imunologia , Doença de Marek/virologia , Vacinas contra Doença de Marek/imunologia , Vacinas contra Doença de Marek/administração & dosagem , Vacinas contra Doença de Marek/genética , Citocinas/metabolismo , Citocinas/imunologia , Herpesvirus Galináceo 2/imunologia , Herpesvirus Galináceo 2/genética , Pulmão/virologia , Pulmão/imunologia , Baço/imunologia , Baço/virologia , Doenças das Aves Domésticas/prevenção & controle , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Vacinas de mRNA/imunologia , Vacinação , RNA Mensageiro/genética , RNA Mensageiro/imunologia , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genéticaRESUMO
Marek's disease virus (MDV) is an oncogenic alphaherpesvirus that causes deadly lymphomas in chickens. In chickens, up to 50% of all peripheral T cells are gamma delta (γδ) T cells. Until now, their role in MDV pathogenesis and tumor formation remains poorly understood. To investigate the role of γδ T cells in MDV pathogenesis, we infected recently generated γδ T cell knockout chickens with very virulent MDV. Strikingly, disease and tumor incidence were highly increased in the absence of γδ T cells, indicating that γδ T cells play an important role in the immune response against MDV. In the absence of γδ T cells, virus replication was drastically increased in the thymus and spleen, which are potential sites of T cell transformation. Taken together, our data provide the first evidence that γδ T cells play an important role in the pathogenesis and tumor formation of this highly oncogenic herpesvirus.IMPORTANCEGamma delta (γδ) T cells are the most abundant T cells in chickens, but their role in fighting pathogens remains poorly understood. Marek's disease virus (MDV) is an important veterinary pathogen, that causes one of the most frequent cancers in animals and is used as a model for virus-induced tumor formation. Our study revealed that γδ T cells play a crucial role in combating MDV, as disease and tumor incidence drastically increased in the absence of these cells. γδ T cells restricted virus replication in the key lymphoid organs, thereby decreasing the likelihood of causing tumors and disease. This study provides novel insights into the role of γδ T cells in the pathogenesis of this highly oncogenic virus.
Assuntos
Galinhas , Herpesvirus Galináceo 2 , Doença de Marek , Replicação Viral , Animais , Galinhas/virologia , Doença de Marek/virologia , Doença de Marek/imunologia , Herpesvirus Galináceo 2/patogenicidade , Herpesvirus Galináceo 2/imunologia , Herpesvirus Galináceo 2/genética , Baço/imunologia , Baço/virologia , Baço/patologia , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/genética , Linfócitos Intraepiteliais/imunologia , Timo/imunologia , Timo/virologia , Timo/patologia , Linfócitos T/imunologia , Doenças das Aves Domésticas/virologia , Doenças das Aves Domésticas/imunologiaRESUMO
Marek's disease virus (MDV) is a significant tumorigenic virus that causes severe immunosuppression in chickens. Lentinan (LNT) is an immunomodulator containing ß-glucans and is widely used in areas such as antiviral, anticancer, and immune regulation. To investigate the immunomodulatory effects of LNT on specific pathogen-free (SPF) chicks and its potential to inhibit MDV infection, we conducted an MDV challenge experiment and observed the immune-enhancing effect of LNT on SPF chicks. The results showed that LNT promoted the growth and development of SPF chicks and induced the upregulation of cytokines such as Mx protein, interferon-γ (INF-γ), tumor necrosis factor-α (TNF-α), and interleukin-2 (IL-2). The specific gravity of CD4+ T-lymphocytes and CD8+ T-lymphocytes and their ratios were also significantly upregulated. Prophylactic use of LNT inhibited MDV replication in lymphocytes, liver, and spleen. It also alleviated MDV-induced weight loss and hepatosplenomegaly in SPF chicks. The present study confirms that LNT can enhance the levels of innate and cellular immunity in SPF chicks and contributes to the inhibition of MDV replication in vivo and mitigation of immune organ damage in chicks due to MDV infection. This provides an adjunctive measure for better control of MDV infection.
Assuntos
Galinhas , Herpesvirus Galináceo 2 , Lentinano , Doença de Marek , Doenças das Aves Domésticas , Animais , Doença de Marek/imunologia , Lentinano/farmacologia , Lentinano/administração & dosagem , Doenças das Aves Domésticas/virologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/tratamento farmacológico , Herpesvirus Galináceo 2/fisiologia , Organismos Livres de Patógenos Específicos , Ração Animal/análise , Fatores Imunológicos/farmacologia , Fatores Imunológicos/administração & dosagem , Dieta/veterinária , Distribuição AleatóriaRESUMO
Marek's disease virus (MDV) vaccines were the first vaccines that protected against cancer. The avirulent turkey herpesvirus (HVT) was widely employed and protected billions of chickens from a deadly MDV infection. It is also among the most common vaccine vectors providing protection against a plethora of pathogens. HVT establishes latency in T-cells, allowing the vaccine virus to persist in the host for life. Intriguingly, the HVT genome contains telomeric repeat arrays (TMRs) at both ends; however, their role in the HVT life cycle remains elusive. We have previously shown that similar TMRs in the MDV genome facilitate its integration into host telomeres, which ensures efficient maintenance of the virus genome during latency and tumorigenesis. In this study, we investigated the role of the TMRs in HVT genome integration, latency, and reactivation in vitro and in vivo. Additionally, we examined HVT infection of feather follicles. We generated an HVT mutant lacking both TMRs (vΔTMR) that efficiently replicated in cell culture. We could demonstrate that wild type HVT integrates at the ends of chromosomes containing the telomeres in T-cells, while integration was severely impaired in the absence of the TMRs. To assess the role of TMRs in vivo, we infected one-day-old chickens with HVT or vΔTMR. vΔTMR loads were significantly reduced in the blood and hardly any virus was transported to the feather follicle epithelium where the virus is commonly shed. Strikingly, latency in the spleen and reactivation of the virus were severely impaired in the absence of the TMRs, indicating that the TMRs are crucial for the establishment of latency and reactivation of HVT. Our findings revealed that the TMRs facilitate integration of the HVT genome into host chromosomes, which ensures efficient persistence in the host, reactivation, and transport of the virus to the skin.
Assuntos
Galinhas , Doença de Marek , Telômero , Integração Viral , Latência Viral , Animais , Galinhas/virologia , Telômero/genética , Telômero/virologia , Doença de Marek/virologia , Doença de Marek/imunologia , Doença de Marek/prevenção & controle , Vetores Genéticos , Herpesvirus Meleagrídeo 1/genética , Herpesvirus Meleagrídeo 1/imunologia , Vacinas contra Doença de Marek/imunologia , Vacinas contra Doença de Marek/genética , Genoma Viral , Herpesvirus Galináceo 2/genética , Herpesvirus Galináceo 2/imunologia , Sequências Repetitivas de Ácido Nucleico , Doenças das Aves Domésticas/virologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/prevenção & controleRESUMO
Marek's disease (MD) is a lymphoproliferative disease of chickens induced by Marek's disease virus (MDV), an oncogenic α-herpesvirus. MDV has increased in virulence, prompting continued efforts in both improved vaccines and enhanced genetic resistance. Model pairs of genetically MD-resistant and MD-susceptible chickens that were either MHC-matched or MHC-congenic allowed characterization of T cell receptor (TCR) repertoires associated with MDV infection. MD-resistant chickens showed higher usage of Vß-1 TCRs than susceptible chickens in both the CD8 and CD4 subsets in the MHC-matched model, and in the CD8 subset only in the MHC-congenic model, with a shift towards Vß-1+ CD8 cells during MDV infection. Long and short read sequencing identified divergent TCRß loci between MHC-matched MD-resistant and MD-susceptible chickens, with MD-resistant chickens having more TCR Vß1 genes. TCR Vß1 CDR1 haplotype usage in MD-resistant x MD-susceptible F1 birds by RNAseq indicated that the most commonly used CDR1 variant was unique to the MD-susceptible line, suggesting that selection for MD resistance in the MHC-matched model optimized the TCR repertoire away from dominant recognition of one or more B2 haplotype MHC molecules. Finally, TCR downregulation during MDV infection in the MHC-matched model was strongest in the MD-susceptible line, and MDV reactivation downregulated TCR expression in a tumor cell line.
Assuntos
Resistência à Doença , Herpesvirus Galináceo 2 , Doença de Marek , Receptores de Antígenos de Linfócitos T alfa-beta , Animais , Linfócitos T CD8-Positivos , Galinhas , Doença de Marek/genética , Doença de Marek/imunologia , Resistência à Doença/genéticaRESUMO
Marek's disease (MD) vaccines reduce the incidence of MD but cannot control virus shedding. To develop new vaccines, it is essential to elucidate mechanisms of immunity to Marek's disease virus (MDV) infection. In this regard, gamma delta (γδ) T cells may play a significant role in prevention of viral spread and tumor surveillance. Here we demonstrated that MDV vaccination induced interferon (IFN)-γ+CD8α+ γδ T cells and transforming growth factor (TGF)-ß+ γδ T cells in lungs. γδ T cells from MDV-infected chickens exhibited cytotoxic activity. Importantly, γδ T cells from the vaccinated/challenged group exhibited maximum cytotoxic activity following ex vivo stimulation. These results suggest that MDV vaccines activate effector γδ T cells which may be involved in the development of protective immune responses against MD. Further, it was demonstrated that MDV infection increases the frequency of a subpopulation of γδ T cells expressing membrane-bound TGF-ß in MDV-infected birds.
Assuntos
Galinhas/imunologia , Doença de Marek/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Animais , Biomarcadores , Galinhas/virologia , Citocinas , Sequenciamento de Nucleotídeos em Larga Escala , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Imunização , Imunofenotipagem , Ativação Linfocitária , Contagem de Linfócitos , Doença de Marek/prevenção & controle , Doença de Marek/virologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/prevenção & controle , Doenças das Aves Domésticas/virologia , Vacinas Virais/imunologia , Replicação Viral , Eliminação de Partículas ViraisRESUMO
Viral diseases pose major threats to humans and other animals, including the billions of chickens that are an important food source as well as a public health concern due to zoonotic pathogens. Unlike humans and other typical mammals, the major histocompatibility complex (MHC) of chickens can confer decisive resistance or susceptibility to many viral diseases. An iconic example is Marek's disease, caused by an oncogenic herpesvirus with over 100 genes. Classical MHC class I and class II molecules present antigenic peptides to T lymphocytes, and it has been hard to understand how such MHC molecules could be involved in susceptibility to Marek's disease, given the potential number of peptides from over 100 genes. We used a new in vitro infection system and immunopeptidomics to determine peptide motifs for the 2 class II molecules expressed by the MHC haplotype B2, which is known to confer resistance to Marek's disease. Surprisingly, we found that the vast majority of viral peptide epitopes presented by chicken class II molecules arise from only 4 viral genes, nearly all having the peptide motif for BL2*02, the dominantly expressed class II molecule in chickens. We expressed BL2*02 linked to several Marek's disease virus (MDV) peptides and determined one X-ray crystal structure, showing how a single small amino acid in the binding site causes a crinkle in the peptide, leading to a core binding peptide of 10 amino acids, compared to the 9 amino acids in all other reported class II molecules. The limited number of potential T cell epitopes from such a complex virus can explain the differential MHC-determined resistance to MDV, but raises questions of mechanism and opportunities for vaccine targets in this important food species, as well as providing a basis for understanding class II molecules in other species including humans.
Assuntos
Galinhas/imunologia , Herpesvirus Galináceo 2/imunologia , Antígenos de Histocompatibilidade Classe II , Doença de Marek/imunologia , Animais , Apresentação de Antígeno/genética , Apresentação de Antígeno/imunologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Bolsa de Fabricius/imunologia , Células Cultivadas , Galinhas/genética , Galinhas/virologia , Resistência à Doença/genética , Resistência à Doença/imunologia , Haplótipos , Herpesvirus Galináceo 2/química , Antígenos de Histocompatibilidade Classe II/química , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe II/imunologia , Antígenos de Histocompatibilidade Classe II/metabolismo , Epitopos Imunodominantes/química , Epitopos Imunodominantes/genética , Epitopos Imunodominantes/imunologia , Epitopos Imunodominantes/metabolismo , Doença de Marek/genética , Doença de Marek/virologia , Modelos Moleculares , Peptídeos/química , Peptídeos/genética , Peptídeos/imunologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Proteínas Virais/química , Proteínas Virais/genética , Proteínas Virais/imunologiaRESUMO
Marek's disease virus (MDV) is a highly oncogenic alphaherpesvirus that causes deadly T-cell lymphomas and serves as a natural virus-induced tumor model in chickens. The most efficacious vaccine, CVI988/Rispens (CVI988), against MD has been used for several decades. However, the mechanisms leading to protective immunity following vaccination are not fully understood. In this study, employing multi-parameter flow cytometry, we performed a comprehensive analysis of T cell responses in CVI988-vaccinated chickens. CVI988 vaccination induced significant expansion of γδ T cells and CD8α+ T cells but not CD4+ T cells in spleen, lung and blood at early time-points. The expansion of these cells was CVI988-specific as infection with very virulent MDV RB1B did not elicit expansion of either γδ or CD8α+ T cells. Phenotypic analysis showed that CVI988 vaccination elicited preferential proliferation of CD8α+ γδ T cells and CD8αα co-receptor expression was upregulated on γδ T cells and CD8α+ T cells after immunization. Additionally, cell sorting and quantitative RT-PCR showed that CVI988 vaccination activated γδ T cells and CD8α+ T cells which exhibited differential expression of cytotoxic and T cell-related cytokines. Lastly, secondary immunization with CVI988 induced the expansion of CD8+ T cells but not γδ T cells at higher magnitude, compared to primary immunization, suggesting CVI988 did induce memory CD8+ T cells but not γδ T cells in chickens. Our results, for the first time, reveal a potential role of γδ T cells in CVI988-induced immune protection and provide new insights into the mechanism of immune protection against oncogenic MDV.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Vacinas Anticâncer/farmacologia , Galinhas , Herpesvirus Galináceo 2/imunologia , Doenças das Aves Domésticas , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Vacinas Virais/farmacologia , Animais , Galinhas/imunologia , Galinhas/virologia , Doença de Marek/imunologia , Doença de Marek/prevenção & controle , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/prevenção & controle , Doenças das Aves Domésticas/virologia , VacinaçãoRESUMO
DEAD-box helicase 5 (DDX5) plays a significant role in tumorigenesis and regulates viral replication of several viruses. An avian oncogenic herpesvirus, Marek's disease virus (MDV), is widely known to cause immunosuppression and lymphoma in chickens. However, the underlying mechanisms of how DDX5 plays a role in viral replication remain unclear. In this study, we show that MDV inhibits the production of interferon beta (IFN-ß) in chicken embryo fibroblasts (CEFs) by increasing the expression level and promoting the nuclear aggregation of DDX5. We further reveal how DDX5 down-regulates melanoma differentiation-associated gene 5/toll-like receptor 3 signaling through the fundamental transcription factor, interferon regulatory factor 1. MDV replication is suppressed, and the production of IFN-ß is promoted in the DDX5 absented CEFs. Taken together, our investigations demonstrate that MDV inhibits IFN-ß production by targeting DDX5-mediated signaling to facilitate viral replication, which offers a novel insight into the mechanism by which an avian oncogenic herpesvirus replicates in chicken cells.
Assuntos
Proteínas Aviárias/imunologia , RNA Helicases DEAD-box/imunologia , Fibroblastos/imunologia , Herpesvirus Galináceo 2/imunologia , Interferon beta/imunologia , Replicação Viral/imunologia , Animais , Proteínas Aviárias/genética , Proteínas Aviárias/metabolismo , Western Blotting , Células Cultivadas , Embrião de Galinha , Galinhas/genética , Galinhas/imunologia , Galinhas/virologia , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Fibroblastos/metabolismo , Fibroblastos/virologia , Regulação da Expressão Gênica/imunologia , Herpesvirus Galináceo 2/fisiologia , Interações Hospedeiro-Patógeno/imunologia , Imunidade Inata/genética , Imunidade Inata/imunologia , Interferon beta/genética , Interferon beta/metabolismo , Doença de Marek/genética , Doença de Marek/imunologia , Doença de Marek/virologia , RNA-Seq/métodos , Transcriptoma/imunologiaRESUMO
Marek's disease virus (MDV), the etiologic agent for Marek's disease (MD), causes a deadly lymphoproliferative disease in chickens. Causes of the well-documented association between genetically defined lines of chicken and resistance to MD remain unknown. Here, the frequencies of IFN-gamma producing pp38 and MEQ-specific T cell responses were determined in line N (B21 haplotype; MD-resistant) and line P2a (B19 haplotype, MD-susceptible) chickens after infection with vaccine and/or virulent (RB1B) strains of MDV using both standard ex vivo and cultured chIFN-gamma ELISPOT assays. Notably, MDV infection of naïve and vaccinated MD-resistant chickens induced higher frequencies of IFN-gamma producing MDV-specific T cell responses using the cultured and ex vivo ELISPOT assay, respectively. Remarkably, vaccination did not induce or boost MEQ-specific effector T cells in the susceptible chickens, while it boosted both pp38-and MEQ-specific response in resistant line. Taken together, our results revealed that there is a direct association between the magnitude of T cell responses to pp38 and MEQ of MDV antigens and resistance to the disease.
Assuntos
Galinhas/imunologia , Haplótipos/imunologia , Antígenos de Histocompatibilidade/imunologia , Interferon gama/imunologia , Mardivirus/imunologia , Doença de Marek/imunologia , Linfócitos T/imunologia , Animais , Galinhas/virologia , Suscetibilidade a Doenças/imunologia , Suscetibilidade a Doenças/virologia , Doença de Marek/virologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Virulência/imunologiaRESUMO
Marek's Disease Virus (MDV) infects chickens via respiratory route and causes lymphomas in internal organs including gastrointestinal tract. MDV infection causes a shift in the gut microbiota composition. However, interactions between the gut microbiota and immune responses against MDV infection are not well understood. Therefore, the current study was performed to understand the effect of the gut microbiota on Marek's disease (MD) pathogenesis. The findings showed that depletion of gut microbiota increased the severity of MD in infected chickens. In addition, an increase in the transcription of interferon (IFN)-α, IFN-ß and IFN-γ in the bursa of Fabricius at 4 days post-infection (dpi) was observed in the gut microbiota depleted chickens. The observations in this study shed more light on the association between the gut microbiota and MDV infection in chickens. More research is needed to explore the mechanisms of involvement of the gut microbiota in immunity against MD in chickens.
Assuntos
Galinhas , Microbioma Gastrointestinal/fisiologia , Trato Gastrointestinal/microbiologia , Herpesvirus Galináceo 2/fisiologia , Doença de Marek/imunologia , Doença de Marek/microbiologia , Animais , Antibacterianos/farmacologia , Bolsa de Fabricius/imunologia , Bolsa de Fabricius/metabolismo , Ceco/metabolismo , Ceco/microbiologia , Plumas/virologia , Trato Gastrointestinal/imunologia , Trato Gastrointestinal/metabolismo , Expressão Gênica , Genoma Viral , Herpesvirus Galináceo 2/genética , Herpesvirus Galináceo 2/imunologia , Interferons/genética , Interleucinas/genética , Interleucinas/metabolismo , Doença de Marek/virologia , Índice de Gravidade de Doença , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo , Interleucina 22RESUMO
Modified-live herpesvirus vaccines are widely used in humans and animals, but field strains can emerge that have a higher virulence and break vaccinal protection. Since the introduction of the first vaccine in the 1970s, Marek's disease virus overcame the vaccine barrier by the acquisition of numerous genomic mutations. However, the evolutionary adaptations in the herpesvirus genome responsible for the vaccine breaks have remained elusive. Here, we demonstrate that point mutations in the multifunctional meq gene acquired during evolution can significantly alter virulence. Defined mutations found in highly virulent strains also allowed the virus to overcome innate cellular responses and vaccinal protection. Concomitantly, the adaptations in meq enhanced virus shedding into the environment, likely providing a selective advantage for the virus. Our study provides the first experimental evidence that few point mutations in a single herpesviral gene result in drastically increased virulence, enhanced shedding, and escape from vaccinal protection.
Assuntos
Vacinas contra Doença de Marek/imunologia , Doença de Marek/genética , Doença de Marek/imunologia , Proteínas Oncogênicas Virais/genética , Virulência/genética , Animais , Galinhas , Genes Virais/genética , Herpesvirus Galináceo 2/genética , Mutação PuntualRESUMO
Marek's disease (MD) is an oncogenic, lymphoproliferative, and highly contagious disease of chickens. Its etiologic agent is the alphaherpesvirus Marek's disease virus (MDV, Gallid alphaherpesvirus 2), and it is a chronic and ubiquitous problem for the poultry industry with significant economic impact in the United States and worldwide. We have previously demonstrated that MDV attenuated by dicodon deoptimization of the UL54 gene results in reduced gene product accumulation in vitro, with reduced viral genome copy number upon infection and reduced atrophy of bursa and thymus in vivo as well. In this report we detail our attempts to use the same attenuation strategy on a meq-deleted MDV mutant, rMd5B40ΔMeq. Unlike the wild-type rMd5B40 virus the rMd5B40ΔMeq is no longer oncogenic, but infected birds experience an unacceptable amount of bursa and thymus atrophy (BTA). We produced two meq-deleted MDV recombinants with a dicodon-deoptimized UL54 (rMd5B40ΔMeq/UL54deop1 and -deop2) and tested their tendency to cause BTA and to serve as a protective vaccine. We found that, although dicodon deoptimization of the UL54 gene results in a virus that spares the infected animal from atrophy of the bursa and thymus, the meq-deleted UL54-deoptimized recombinant is also less protective than the meq-deleted virus without UL54 deoptimization, the HVT + SB1 combination vaccine, or the Rispens (CVI988) vaccine.
Assuntos
Galinhas , Códon/genética , Proteínas Imediatamente Precoces/genética , Vacinas contra Doença de Marek/genética , Doença de Marek/imunologia , Proteínas Oncogênicas Virais/deficiência , Doenças das Aves Domésticas/imunologia , Proteínas Virais/genética , Animais , Atrofia/veterinária , Deleção de Genes , Linfócitos/patologia , Organismos Livres de Patógenos EspecíficosRESUMO
Marek's disease (MD) vaccines are unique in their capability to prevent MD lymphomas as early as a few days after vaccination, despite the fact that they do not eliminate virulent viruses from the host. To help understand the mechanism behind this unique MD vaccine effect, we compared the expression of MDV oncoprotein Meq among CD4+ T cells between vaccinated and unvaccinated birds. Chickens were vaccinated by an MD vaccine, herpesvirus of turkeys, and then challenged by a recombinant virulent MDV that expresses green fluorescent protein simultaneously with Meq. We found significantly fewer Meq-expressing CD4+ T cells appeared in peripheral blood mononuclear cells (PBMC) of the vaccinated birds compared to the unvaccinated birds as early as one week after the virulent virus challenge. In contrast, the quantity of virulent MDV genome remained similar in Meq- PBMC in both vaccinated and unvaccinated birds. Our results suggest that MD vaccination affects the dynamics of Meq-expressing, possibly transformed, cells while impact on the overall infection in the Meq- cells was not significant.
Assuntos
Linfócitos T CD4-Positivos/virologia , Herpesvirus Galináceo 2/genética , Vacinas contra Doença de Marek/imunologia , Doença de Marek/virologia , Proteínas Oncogênicas Virais/genética , Animais , Galinhas/virologia , Genoma Viral , Herpesvirus Galináceo 2/imunologia , Doença de Marek/imunologia , Vacinas contra Doença de Marek/administração & dosagem , Proteínas Oncogênicas Virais/imunologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Organismos Livres de Patógenos Específicos , Latência ViralRESUMO
In order to evaluate the influence of the vertical transmission of avian leukosis virus (ALV) from J subgroup (ALV-J) positive parents on the vaccine efficacy of Marek's disease virus (MDV), ALV-J positive male breeders × female breeders of Three-yellow chickens and the ALV negative male breeder × the negative female breeders were used respectively for crossbreeding to produce eggs and the hatching offspring. The commercial CVI988/Rispens vaccine was used to vaccinate the crossbred offspring at 1-day-old. At 7-days-old, the birds were inoculated with the inactivated oil-emulsion vaccines (OEVs) AIV-H5 monovalent and NDV + AIV-H9 bivalent, respectively. Then the birds were challenged with a Chinese very virulent (vv) MDV field strain GXY2 at 14-day-old. The results showed that the viral load of the challenged GXY2 in the offspring from the ALV-J positive breeders was significantly higher than that from the ALV-negative breeders' (P < 0.05), and the mortality and tumor incidence of offspring from the ALV-J positive breeders were higher than those of the ALV-negative breeders. Also the offspring of the ALV-J positive breeders exhibited a significant negative effect on the development of the immune organs (P < 0.05) and lower antibody responses to the vaccinations with the commercial OEVs (P<0.05). The MD vaccine protective index in the offspring from the ALV-J positive breeders was lower than that from the ALV-negative breeders. The results of the study demonstrated that the vertical transmission of ALV from the ALV-J positive parents caused severe immunosuppression and significantly reduced the Marek's disease vaccine efficacy in Three-yellow chickens.
Assuntos
Leucose Aviária/imunologia , Leucose Aviária/transmissão , Terapia de Imunossupressão/veterinária , Transmissão Vertical de Doenças Infecciosas , Vacinas contra Doença de Marek/imunologia , Animais , Vírus da Leucose Aviária/patogenicidade , Cruzamento , Galinhas/imunologia , Galinhas/virologia , Feminino , Masculino , Doença de Marek/imunologia , Doença de Marek/prevenção & controle , Aves Domésticas/imunologia , Aves Domésticas/virologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/prevenção & controle , Doenças das Aves Domésticas/virologia , Potência de Vacina , Carga ViralRESUMO
SC9-2 is a recombinant Marek's disease virus (MDV) strain lacking the meq oncogene. Previous study demonstrated that SC9-2 virus provides good protection against challenge with a very virulent MDV rMd5, but it induces immunosuppressive effects in specific pathogen-free (SPF) chickens. In the present study, SC9-2 was serially passaged on chicken embryo fibroblast (CEF) cell cultures. The pathogenicity and immune efficacy of SC9-2/10th and SC9-2/40th against rMd5 were evaluated. Animal experimental results showed that SC9-2/10th and SC9-2/40th showed no lethality or tumorigenicity in SPF chickens. Body weight of chickens inoculated with SC9-2/40th were significantly higher than that of the chickens inoculated with SC9-2/10th but lower than that of the uninoculated controls. The severity of bursa and thymus atrophy (BTA) and spleen enlargement in SC9-2/40th-inoculated chickens were also weaker than the SC9-2/10th-inoculated ones but stronger than the uninoculated controls. Chickens inoculated with SC9-2/40th and SC9-2/10th showed similar antibody levels induced by H9N2 subtype avian influenza virus/Newcastle disease virus inactivated vaccines, both of which were lower than the uninoculated controls. Replication of SC9-2/40th was significantly lower than SC9-2/10th in feather follicle epithelium (FFE) of infected chickens. The immune protection index of SC9-2/40th was also lower than that of SC9-2/10th, but the difference was not significantly, and both of which were significant higher than that of the commercial MDV vaccine CVI988/Rispens. The results of our studies demonstrated that SC9-2/40th showed weaker severity of BTA, spleen enlargement, and body weight loss and lower replication level in FFE than SC9-2/10th in SPF chickens. However, SC9-2/40th was able to confer better immune protection as compared with CVI988/Rispens vaccination in SPF chickens. In conclusion, serially attenuation of SC9-2 in CEFs reduced the lymphoid organ atrophy and replication in SPF chickens, and the immune protective efficacy of attenuated viruses was still superior than CVI988/Rispens.
Assuntos
Galinhas , Herpesvirus Galináceo 2/fisiologia , Vacinas contra Doença de Marek/imunologia , Doença de Marek/imunologia , Proteínas Oncogênicas Virais/deficiência , Doenças das Aves Domésticas/imunologia , Animais , Herpesvirus Galináceo 2/genética , Herpesvirus Galináceo 2/imunologia , Doença de Marek/virologia , Microrganismos Geneticamente Modificados/genética , Microrganismos Geneticamente Modificados/fisiologia , Doenças das Aves Domésticas/virologia , Organismos Livres de Patógenos EspecíficosRESUMO
Marek's disease virus (MDV), an alpha herpes virus, causes a lymphoproliferative state in chickens known as Marek's disease (MD), resulting in severe monetary losses to the poultry industry. Because lymphocytes of bursa of Fabricius and spleen are prime targets of MDV replication during the early cytolytic phase of infection, the immune response in bursa and spleen should be the foundation of late immunity induced by MDV. However, the mechanism of the MDV-mediated host immune response in lymphocytes in the early stage is poorly understood. The present study is primarily aimed at identifying the crucial genes and significant pathways involved in the immune response of chickens infected with MDV CVI988 and the very virulent RB1B (vvRB1B) strains. Using the RNA sequencing approach, we analyzed the generated transcriptomes from lymphocytes isolated from chicken bursa and spleen. Our findings validated the expression of previously characterized genes; however, they also revealed the expression of novel genes during the MDV-mediated immune response. The results showed that after challenge with CVI988 or vvRB1B strains, 634 and 313 differentially expressed genes (DEGs) were identified in splenic lymphocytes, respectively. However, 58 and 47 DEGs were observed in bursal lymphocytes infected with CVI988 and vvRB1B strains, respectively. Following MDV CVI988 or vvRB1B challenge, the bursal lymphocytes displayed changes in IL-6 and IL-4 gene expression. Surprisingly, splenic lymphocytes exhibited an overwhelming alteration in the expression of cytokines and cytokine receptors involved in immune response signaling. On the other hand, there was no distinct trend between infection with CVI988 and vvRB1B and the expression of cytokines and chemokines, such as IL-10, IFN-γ, STAT1, IRF1, CCL19, and CCL26. However, the expression profiles of IL-1ß, IL-6, IL8L1, CCL4 (GGCL1), and CCL5 were significantly upregulated in splenic lymphocytes from chickens infected with CVI988 compared with those of chickens infected with vvRB1B. Because these cytokines and chemokines are considered to be associated with B cell activation and antigenic signal transduction to T cells, they may indicate differences of immune responses initiated by vaccinal and virulent strains during the early phase of infection. Collectively, our study provides valuable data on the transcriptional landscape using high-throughput sequencing to understand the different mechanism between vaccine-mediated protection and pathogenesis of virulent MDV in vivo.
Assuntos
Herpesvirus Galináceo 2/fisiologia , Imunidade/genética , Linfócitos/metabolismo , Linfócitos/virologia , Doença de Marek/genética , Doença de Marek/virologia , Transcriptoma , Animais , Linfócitos B/metabolismo , Linfócitos B/virologia , Biomarcadores , Galinhas , Biologia Computacional/métodos , Citocinas/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Ontologia Genética , Redes Reguladoras de Genes , Doença de Marek/imunologia , Mapeamento de Interação de Proteínas , Mapas de Interação de Proteínas , Baço/imunologia , Baço/metabolismo , Baço/virologia , Replicação ViralRESUMO
Marek's disease virus (MDV) is a highly oncogenic alphaherpesvirus that causes deadly T-cell lymphomas and serves as a natural virus-induced tumor model in chickens. Although Marek's disease (MD) is well controlled by current vaccines, the evolution of MDV field viruses towards increasing virulence is concerning as a better vaccine to combat very virulent plus MDV is still lacking. Our understanding of molecular and cellular immunity to MDV and its immunopathogenesis has significantly improved, but those findings about cellular immunity to MDV are largely out-of-date, hampering the development of more effective vaccines against MD. T-cell-mediated cellular immunity was thought to be of paramount importance against MDV. However, MDV also infects macrophages, B cells and T cells, leading to immunosuppression and T-cell lymphoma. Additionally, there is limited information about how uninfected immune cells respond to MDV infection or vaccination, specifically, the mechanisms by which T cells are activated and recognize MDV antigens and how the function and properties of activated T cells correlate with immune protection against MDV or MD tumor. The current review revisits the roles of each immune cell subset and its effector mechanisms in the host immune response to MDV infection or vaccination from the point of view of comparative immunology. We particularly emphasize areas of research requiring further investigation and provide useful information for rational design and development of novel MDV vaccines.
Assuntos
Galinhas/imunologia , Galinhas/virologia , Imunidade Celular/imunologia , Doença de Marek/imunologia , Vírus Oncogênicos/imunologia , Linfócitos T/imunologia , Animais , Herpesvirus Galináceo 2/imunologia , Humanos , Doença de Marek/virologia , Linfócitos T/virologia , Virulência/imunologiaRESUMO
The zinc finger antiviral protein (ZAP), as a host restriction factor, inhibits the replication of certain viruses by binding viral mRNA or proteins for degradation. However, little is known about the role of ZAP in the antiviral immune response. We now show that ZAP participates in the antiviral immune response by activating T cells. Overexpression of ZAP significantly inhibited avian leukosis virus subgroup J (ALV-J) replication and reduced the associated inflammatory damage in vivo. In this study, we found that ZAP tended to be expressed in T lymphocytes, especially after ALV-J infection. T lymphocyte proliferation proceeded as usual in response to ALV-J infection in the presence of ZAP, indicating that ZAP endows T lymphocytes with resistance to the immunosuppression caused by ALV-J. Furthermore, ZAP activated cytokine secretion by T lymphocytes by contributing to nuclear translocation of nuclear factors of activated T cells and indirectly promoted anti-ALV-J antibody generation. Together, our findings show that ZAP, acting as an immunomodulatory factor, is involved in the antiviral immune response via T lymphocyte activation.