RESUMEN
Marek's disease virus (MDV) can cause severe immunosuppression in chickens. Our previous study showed that infection with very virulent plus (vv+) MDV strains of one-day-old commercial meat-type chickens possessing maternal antibodies against MDV resulted in severe depletion of splenocytes at 28-30 days of age. In the present study, we have investigated the effect of vv+MDV strain 686 on splenic immunophenotypes at 6, 20, and 30 days post-infection (dpi). Both live and dead cells were analyzed, and the data were statistically compared to the uninfected control. The results revealed a decrease in the total live cell population starting on day 20, primarily affecting B cells, CD8ß+, and gamma delta (γδ) T cells, while the frequencies of both live and dead CD3+ and CD4+ T cells were increased. The MHC-I expression of CD3+ and CD4+ T cells was higher at 20 and 30 dpi, while the expression of MHC-II on these cells was downregulated at 6 dpi but was upregulated at 30 dpi. Collectively, these results suggest that maternal antibodies seem to delay the negative effects of vv+MDV on the splenic lymphoid populations, albeit being non-protective. Our results emphasize the importance of MD vaccination in vv+MDV endemic areas.
Asunto(s)
Pollos , Enfermedad de Marek , Enfermedades de las Aves de Corral , Bazo , Animales , Bazo/inmunología , Bazo/virología , Enfermedad de Marek/inmunología , Enfermedad de Marek/virología , Enfermedades de las Aves de Corral/virología , Enfermedades de las Aves de Corral/inmunología , Inmunofenotipificación , Virulencia , Linfocitos B/inmunología , Herpesvirus Gallináceo 2/inmunología , Herpesvirus Gallináceo 2/genéticaRESUMEN
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.
Asunto(s)
Pollos , Citocinas , Herpesvirus Gallináceo 2 , Vacunas contra la Enfermedad de Marek , Enfermedad de Marek , Animales , Pollos/inmunología , Enfermedad de Marek/prevención & control , Enfermedad de Marek/inmunología , Enfermedad de Marek/virología , Vacunas contra la Enfermedad de Marek/inmunología , Vacunas contra la Enfermedad de Marek/administración & dosificación , Vacunas contra la Enfermedad de Marek/genética , Citocinas/metabolismo , Citocinas/inmunología , Herpesvirus Gallináceo 2/inmunología , Herpesvirus Gallináceo 2/genética , Pulmón/virología , Pulmón/inmunología , Bazo/inmunología , Bazo/virología , Enfermedades de las Aves de Corral/prevención & control , Enfermedades de las Aves de Corral/inmunología , Enfermedades de las Aves de Corral/virología , Vacunas de ARNm/inmunología , Vacunación , ARN Mensajero/genética , ARN Mensajero/inmunología , Vacunas Sintéticas/inmunología , Vacunas Sintéticas/administración & dosificación , Vacunas Sintéticas/genéticaRESUMEN
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.
Asunto(s)
Pollos , Herpesvirus Gallináceo 2 , Enfermedad de Marek , Replicación Viral , Animales , Pollos/virología , Enfermedad de Marek/virología , Enfermedad de Marek/inmunología , Herpesvirus Gallináceo 2/patogenicidad , Herpesvirus Gallináceo 2/inmunología , Herpesvirus Gallináceo 2/genética , Bazo/inmunología , Bazo/virología , Bazo/patología , Receptores de Antígenos de Linfocitos T gamma-delta/inmunología , Receptores de Antígenos de Linfocitos T gamma-delta/genética , Linfocitos Intraepiteliales/inmunología , Timo/inmunología , Timo/virología , Timo/patología , Linfocitos T/inmunología , Enfermedades de las Aves de Corral/virología , Enfermedades de las Aves de Corral/inmunologíaRESUMEN
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.
Asunto(s)
Pollos , Coinfección , Vacunas contra la Enfermedad de Marek , Enfermedad de Marek , Enfermedades de las Aves de Corral , Carga Viral , Animales , Enfermedad de Marek/virología , Enfermedad de Marek/prevención & control , Enfermedad de Marek/inmunología , Pollos/virología , Coinfección/virología , Coinfección/veterinaria , Enfermedades de las Aves de Corral/virología , Enfermedades de las Aves de Corral/prevención & control , Vacunas contra la Enfermedad de Marek/inmunología , Vacunas contra la Enfermedad de Marek/genética , Virulencia , Herpesvirus Meleágrido 1/inmunología , Herpesvirus Meleágrido 1/genética , Vacunas Sintéticas/inmunología , Vacunas Sintéticas/genética , Herpesvirus Gallináceo 2/genética , Herpesvirus Gallináceo 2/inmunología , Herpesvirus Gallináceo 2/patogenicidad , Vacunación , Vectores Genéticos/genéticaRESUMEN
Cytokines are co-administrated with vaccines or co-expressed in the vaccine virus genome to improve protective efficacy by stimulating immune responses. Using glycosylphosphatidylinositol (GPI) anchoring by attachment to the target cytokine, we constructed recombinant Marek's disease virus (MDV) vaccine strain 301B/1 (v301B/1-rtg-IL-15) that expresses chicken interleukin-15 (IL-15) as the membrane-bound form at the cell surface. We evaluated the vaccine efficacy of v301B/1-rtg-IL-15 given as a bivalent Marek's disease (MD) vaccine in combination with turkey herpesvirus (HVT) against a very virulent plus MDV strain 648A challenge. The efficacy was compared with that of conventional bivalent MD vaccine, as a mixture with HVT plus parental v301B/1 or v301B/1-IL-15, which expresses a natural form of IL-15. The membrane-bound IL-15 expression did not interfere with the virus growth of recombinant v301B/1-rtg-IL-15. However, the MD incidence in birds vaccinated with v301B/1-rtg-IL-15 was higher than that of birds given the conventional bivalent MD vaccine containing parental v301B/1 virus, although the v301B/1-rtg-IL-15 vaccinated group showed increased natural killer cell activation at day 5 postvaccination, the same day as challenge. Overall, the protection of v301B/1-rtg-IL-15 was not improved from that of v301B/1 against very virulent plus MDV challenge.
Eficacia de una vacuna contra el virus de la enfermedad de Marek cepa 301B/1 recombinante que expresa la interleucina-15 de pollo anclada a la membrana. Las citocinas se administran junto con vacunas o se co-expresan en el genoma del virus de la vacuna para mejorar la eficacia protectora mediante la estimulación de respuestas inmunitarias. Utilizando el anclaje de glicosilfosfatidilinositol (GPI) mediante unión a la citoquina objetivo, se construyó una cepa de vacuna recombinante del virus de la enfermedad de Marek (MDV) 301B/1 (v301B/1-rtg-IL-15) que expresa la interleucina-15 de pollo (IL-15) como la forma unida a la membrana en la superficie celular. Se evaluó la eficacia de la vacuna v301B/1-rtg-IL-15 administrada como vacuna bivalente en combinación con el herpesvirus del pavo (HVT) contra el desafío con un virus muy virulento cepa 648A de la enfermedad de Marek (MD). La eficacia se comparó con la de la vacuna bivalente convencional contra la enfermedad de Marek, como una mezcla con HVT más la cepa v301B/1 parental o con el virus recombinante v301B/1-IL-15, que expresa una forma natural de IL-15. La expresión de IL-15 unida a membrana no interfirió con el crecimiento del virus de v301B/1-rtg-IL-15 recombinante. Sin embargo, la incidencia de la enfermedad de Marek en aves vacunadas con v301B/1-rtg-IL-15 fue mayor que la de las aves que recibieron la vacuna de Marek bivalente convencional que contenía el virus v301B/1 parental, aunque el grupo vacunado con v301B/1-rtg-IL-15 mostró una mayor activación de las células asesinas naturales en el día 5 después de la vacunación, que fue el mismo día del desafío. En general, la protección por la vacuna v301B/1-rtg-IL-15 no mejoró con respecto a la conferida por v301B/1 contra un desafío muy virulento de la enfermedad de Marek.
Asunto(s)
Pollos , Herpesvirus Gallináceo 2 , Interleucina-15 , Vacunas contra la Enfermedad de Marek , Enfermedad de Marek , Vacunas Sintéticas , Animales , Interleucina-15/genética , Interleucina-15/inmunología , Interleucina-15/metabolismo , Enfermedad de Marek/prevención & control , Enfermedad de Marek/inmunología , Vacunas contra la Enfermedad de Marek/inmunología , Vacunas contra la Enfermedad de Marek/genética , Vacunas Sintéticas/inmunología , Herpesvirus Gallináceo 2/genética , Herpesvirus Gallináceo 2/inmunología , Enfermedades de las Aves de Corral/prevención & control , Enfermedades de las Aves de Corral/virología , Enfermedades de las Aves de Corral/inmunología , Herpesvirus Meleágrido 1/inmunología , Herpesvirus Meleágrido 1/genética , Herpesvirus Meleágrido 1/metabolismoRESUMEN
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.
Asunto(s)
Pollos , Enfermedad de Marek , Telómero , Integración Viral , Latencia del Virus , Animales , Pollos/virología , Telómero/genética , Telómero/virología , Enfermedad de Marek/virología , Enfermedad de Marek/inmunología , Enfermedad de Marek/prevención & control , Vectores Genéticos , Herpesvirus Meleágrido 1/genética , Herpesvirus Meleágrido 1/inmunología , Vacunas contra la Enfermedad de Marek/inmunología , Vacunas contra la Enfermedad de Marek/genética , Genoma Viral , Herpesvirus Gallináceo 2/genética , Herpesvirus Gallináceo 2/inmunología , Secuencias Repetitivas de Ácidos Nucleicos , Enfermedades de las Aves de Corral/virología , Enfermedades de las Aves de Corral/inmunología , Enfermedades de las Aves de Corral/prevención & controlRESUMEN
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.
Asunto(s)
Pollos , Herpesvirus Gallináceo 2 , Lentinano , Enfermedad de Marek , Enfermedades de las Aves de Corral , Animales , Enfermedad de Marek/inmunología , Lentinano/farmacología , Lentinano/administración & dosificación , Enfermedades de las Aves de Corral/virología , Enfermedades de las Aves de Corral/inmunología , Enfermedades de las Aves de Corral/tratamiento farmacológico , Herpesvirus Gallináceo 2/fisiología , Organismos Libres de Patógenos Específicos , Alimentación Animal/análisis , Factores Inmunológicos/farmacología , Factores Inmunológicos/administración & dosificación , Dieta/veterinaria , Distribución AleatoriaRESUMEN
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.
Asunto(s)
Resistencia a la Enfermedad , Herpesvirus Gallináceo 2 , Enfermedad de Marek , Receptores de Antígenos de Linfocitos T alfa-beta , Animales , Linfocitos T CD8-positivos , Pollos , Enfermedad de Marek/genética , Enfermedad de Marek/inmunología , Resistencia a la Enfermedad/genéticaRESUMEN
BACKGROUND: Duck plague virus (DPV), belonging to herpesviruses, is a linear double-stranded DNA virus. There are many reports about the outbreak of the duck plague in a variety of countries, which caused huge economic losses. Recently, increasing reports revealed that multiple long non-coding RNAs (lncRNAs) can possess great potential in the regulation of host antiviral immune response. Furthermore, it remains to be determined which specific molecular mechanisms are responsible for the DPV-host interaction in host immunity. Here, lncRNAs and mRNAs in DPV infected duck embryonic fibroblast (DEF) cells were identified by high-throughput RNA-sequencing (RNA-seq). And we predicted target genes of differentially expressed genes (DEGs) and formed a complex regulatory network depending on in-silico analysis and prediction. RESULT: RNA-seq analysis results showed that 2921 lncRNAs were found at 30 h post-infection (hpi). In our study, 218 DE lncRNAs and 2840 DE mRNAs were obtained in DEF after DPV infection. Among these DEGs and target genes, some have been authenticated as immune-related molecules, such as a Macrophage mannose receptor (MR), Anas platyrhynchos toll-like receptor 2 (TLR2), leukocyte differentiation antigen, interleukin family, and their related regulatory factors. Furthermore, according to the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analysis, we found that the target genes may have important effects on biological development, biosynthesis, signal transduction, cell biological regulation, and cell process. Also, we obtained, the potential targeting relationship existing in DEF cells between host lncRNAs and DPV-encoded miRNAs by software. CONCLUSIONS: This study revealed not only expression changes, but also the possible biological regulatory relationship of lncRNAs and mRNAs in DPV infected DEF cells. Together, these data and analyses provide additional insight into the role of lncRNAs and mRNAs in the host's immune response to DPV infection.
Asunto(s)
Patos/embriología , Fibroblastos/virología , Enfermedad de Marek/virología , Enfermedades de las Aves de Corral/virología , ARN Largo no Codificante/metabolismo , ARN Mensajero/metabolismo , Animales , Brotes de Enfermedades/veterinaria , Patos/genética , Patos/virología , Fibroblastos/metabolismo , Perfilación de la Expresión Génica , Infecciones por Herpesviridae/metabolismo , Mardivirus , Enfermedad de Marek/epidemiología , Enfermedad de Marek/inmunología , Enfermedades de las Aves de Corral/epidemiología , Enfermedades de las Aves de Corral/inmunología , ARN Largo no Codificante/análisis , ARN Largo no Codificante/genética , ARN Mensajero/análisis , ARN Mensajero/genéticaRESUMEN
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.
Asunto(s)
Pollos/inmunología , Enfermedad de Marek/inmunología , Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Animales , Biomarcadores , Pollos/virología , Citocinas , Secuenciación de Nucleótidos de Alto Rendimiento , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/inmunología , Inmunización , Inmunofenotipificación , Activación de Linfocitos , Recuento de Linfocitos , Enfermedad de Marek/prevención & control , Enfermedad de Marek/virología , Enfermedades de las Aves de Corral/inmunología , Enfermedades de las Aves de Corral/prevención & control , Enfermedades de las Aves de Corral/virología , Vacunas Virales/inmunología , Replicación Viral , Esparcimiento de VirusRESUMEN
Vaccines against Marek's disease can protect chickens against clinical disease; however, infected chickens continue to propagate the Marek's disease virus (MDV) in feather follicles and can shed the virus into the environment. Therefore, the present study investigated if MDV could induce an immunoregulatory microenvironment in feathers of chickens and whether vaccines can overcome the immune evasive mechanisms of MDV. The results showed an abundance of CD4+CD25+ and CD4+ transforming growth factor-beta (TGF-ß)+ T regulatory cells in the feathers of MDV-infected chickens at 21 days post-infection. In contrast, vaccinated chickens had a lower number of regulatory T cells. Furthermore, the expression of TGF-ß and programmed cell death receptor (PD)-1 increased considerably in the feathers of Marek's disease virus-infected chickens. The results of the present study raise the possibility of an immunoregulatory environment in the feather pulp of MDV-infected chickens, which may in turn favor replication of infectious MDV in this tissue. Exploring the evasive strategies employed by MDV will facilitate the development of control measures to prevent viral replication and transmission.
Asunto(s)
Pollos/virología , Plumas/virología , Enfermedad de Marek/inmunología , Animales , Linfocitos T CD4-Positivos , Linfocitos T CD8-positivos , Expresión Génica , Herpesvirus Gallináceo 2/inmunología , Enfermedad de Marek/virología , Vacunas contra la Enfermedad de Marek/inmunología , Bazo/inmunología , Factor de Crecimiento Transformador beta/genética , Factor de Crecimiento Transformador beta/metabolismo , Vacunación , Carga Viral/veterinaria , Replicación Viral/fisiologíaRESUMEN
Cytokine transcripts were evaluated chronologically in the brain and in the eye of chickens infected with the very virulent plus Marek's disease virus (vv + MDV) strain 648A. Brain and eye samples were collected from chickens that were either suffering from transient paralysis (TP) (11 days post inoculation, dpi) or had completely recovered from TP but started developing clinical signs of persistent neurological disease (PND) (18-31 dpi). Results obtained from samples collected at 11 dpi are referred as EL (early lesions) and results obtained from samples collected at later times (18-31 dpi) are referred as LL (late lesions). Marked differences were found in the cytokine transcripts in brain and eye. While proinflammatory cytokines (IL-1ß, IL-8, IL-18), iNOS, IFN-α, IFN-γ, and IL-15 were upregulated in the brain during EL and LL, only IL-8 and IFN-γ were upregulated in the eye at both times (EL and LL). The two evaluated viral transcripts (gB and meq) were found in both eye and brain during EL and LL. Levels of the two viral transcripts evaluated were higher at LL than at EL in both brain and eye. No differences were found in any of the viral transcripts between eye and brain during EL. However, during the LL, the levels of meq transcripts were higher in the eye than in the brain. Our results suggest that MDV elicits different immune responses in the brain and in the eye of infected chickens. Because immune responses in the eye of chickens have been poorly studied, further studies on the pathogenesis of MDV in the eye could greatly contribute to our knowledge on the chicken eye immunity.
Asunto(s)
Encéfalo/inmunología , Pollos , Citocinas/biosíntesis , Ojo/inmunología , Herpesvirus Gallináceo 2/patogenicidad , Enfermedad de Marek/inmunología , Enfermedades del Sistema Nervioso/veterinaria , Animales , Encéfalo/patología , Ojo/patología , Enfermedad de Marek/patología , Enfermedades del Sistema Nervioso/inmunología , Enfermedades del Sistema Nervioso/patología , Enfermedades del Sistema Nervioso/virología , Transcriptoma , VirulenciaRESUMEN
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.
Asunto(s)
Pollos/inmunología , Herpesvirus Gallináceo 2/inmunología , Antígenos de Histocompatibilidad Clase II , Enfermedad de Marek/inmunología , Animales , Presentación de Antígeno/genética , Presentación de Antígeno/inmunología , Linfocitos B/inmunología , Linfocitos B/metabolismo , Bolsa de Fabricio/inmunología , Células Cultivadas , Pollos/genética , Pollos/virología , Resistencia a la Enfermedad/genética , Resistencia a la Enfermedad/inmunología , Haplotipos , Herpesvirus Gallináceo 2/química , Antígenos de Histocompatibilidad Clase II/química , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/inmunología , Antígenos de Histocompatibilidad Clase II/metabolismo , Epítopos Inmunodominantes/química , Epítopos Inmunodominantes/genética , Epítopos Inmunodominantes/inmunología , Epítopos Inmunodominantes/metabolismo , Enfermedad de Marek/genética , Enfermedad de Marek/virología , Modelos Moleculares , Péptidos/química , Péptidos/genética , Péptidos/inmunología , Enfermedades de las Aves de Corral/inmunología , Enfermedades de las Aves de Corral/virología , Proteínas Virales/química , Proteínas Virales/genética , Proteínas Virales/inmunologíaRESUMEN
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.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Vacunas contra el Cáncer/farmacología , Pollos , Herpesvirus Gallináceo 2/inmunología , Enfermedades de las Aves de Corral , Receptores de Antígenos de Linfocitos T gamma-delta/inmunología , Vacunas Virales/farmacología , Animales , Pollos/inmunología , Pollos/virología , Enfermedad de Marek/inmunología , Enfermedad de Marek/prevención & control , Enfermedades de las Aves de Corral/inmunología , Enfermedades de las Aves de Corral/prevención & control , Enfermedades de las Aves de Corral/virología , VacunaciónRESUMEN
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.
Asunto(s)
Proteínas Aviares/inmunología , ARN Helicasas DEAD-box/inmunología , Fibroblastos/inmunología , Herpesvirus Gallináceo 2/inmunología , Interferón beta/inmunología , Replicación Viral/inmunología , Animales , Proteínas Aviares/genética , Proteínas Aviares/metabolismo , Western Blotting , Células Cultivadas , Embrión de Pollo , Pollos/genética , Pollos/inmunología , Pollos/virología , ARN Helicasas DEAD-box/genética , ARN Helicasas DEAD-box/metabolismo , Fibroblastos/metabolismo , Fibroblastos/virología , Regulación de la Expresión Génica/inmunología , Herpesvirus Gallináceo 2/fisiología , Interacciones Huésped-Patógeno/inmunología , Inmunidad Innata/genética , Inmunidad Innata/inmunología , Interferón beta/genética , Interferón beta/metabolismo , Enfermedad de Marek/genética , Enfermedad de Marek/inmunología , Enfermedad de Marek/virología , RNA-Seq/métodos , Transcriptoma/inmunologíaRESUMEN
Marek's disease virus (MDV), an avian alphaherpesvirus, infects chickens, transforms CD4+ T cells, and induces immunosuppression early during infection. However, the exact mechanisms involved in MDV-induced immunosuppression are yet to be identified. Here, our results demonstrate that MDV infection in vitro and in vivo induces activation of cyclooxygenase-2 (COX-2) and production of prostaglandin E2 (PGE2). This exerts its inhibitory effects on T cell proliferation at day 21 post infection via PGE2 receptor 2 (EP2) and receptor 4 (EP4). Impairment of the MDV-induced T cell proliferation was associated with downregulation of IL-2 and transferrin uptake in a COX-2/PGE2 dependent manner in vitro. Interestingly, oral administration of a COX-2 inhibitor, meloxicam, during MDV infection inhibited COX-2 activation and rescued T cell proliferation at day 21 post infection. Taken together, our results reveal a novel mechanism that contributes to immunosuppression in the MDV-infected chickens.
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Ciclooxigenasa 2/metabolismo , Dinoprostona/metabolismo , Tolerancia Inmunológica/inmunología , Enfermedad de Marek/inmunología , Linfocitos T/inmunología , Animales , Proliferación Celular/fisiología , Pollos , Activación Enzimática/inmunología , Herpesvirus Gallináceo 2 , Activación de Linfocitos/inmunología , Enfermedad de Marek/metabolismo , Enfermedad de Marek/virologíaRESUMEN
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.
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Pollos/inmunología , Haplotipos/inmunología , Antígenos de Histocompatibilidad/inmunología , Interferón gamma/inmunología , Mardivirus/inmunología , Enfermedad de Marek/inmunología , Linfocitos T/inmunología , Animales , Pollos/virología , Susceptibilidad a Enfermedades/inmunología , Susceptibilidad a Enfermedades/virología , Enfermedad de Marek/virología , Enfermedades de las Aves de Corral/inmunología , Enfermedades de las Aves de Corral/virología , Virulencia/inmunologíaRESUMEN
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.
Asunto(s)
Pollos , Microbioma Gastrointestinal/fisiología , Tracto Gastrointestinal/microbiología , Herpesvirus Gallináceo 2/fisiología , Enfermedad de Marek/inmunología , Enfermedad de Marek/microbiología , Animales , Antibacterianos/farmacología , Bolsa de Fabricio/inmunología , Bolsa de Fabricio/metabolismo , Ciego/metabolismo , Ciego/microbiología , Plumas/virología , Tracto Gastrointestinal/inmunología , Tracto Gastrointestinal/metabolismo , Expresión Génica , Genoma Viral , Herpesvirus Gallináceo 2/genética , Herpesvirus Gallináceo 2/inmunología , Interferones/genética , Interleucinas/genética , Interleucinas/metabolismo , Enfermedad de Marek/virología , Índice de Severidad de la Enfermedad , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo , Interleucina-22RESUMEN
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.
Asunto(s)
Vacunas contra la Enfermedad de Marek/inmunología , Enfermedad de Marek/genética , Enfermedad de Marek/inmunología , Proteínas Oncogénicas Virales/genética , Virulencia/genética , Animales , Pollos , Genes Virales/genética , Herpesvirus Gallináceo 2/genética , Mutación PuntualRESUMEN
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.