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1.
Poult Sci ; 99(9): 4351-4359, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32867979

RESUMO

The vaccines currently available to control infectious bursal disease (IBD) include live-attenuated and inactivated vaccines, immune-complex vaccines, and vaccines consisting of viral constructs of herpesvirus of turkeys genetically engineered to express VP2 surface protein. To evaluate the impact of vaccines on the chicken immune system, 2 animal trials were performed in specific pathogen-free broiler chickens. In trial 1, birds were either vaccinated when they are one-day old with a dual recombinant herpes virus of turkey construct vaccine, expressing VP2 protein of (IBDV) and F protein of Newcastle disease virus, or an immune-complex IBDV vaccine or birds were not vaccinated. At 14, 28, and 35 D, the bursa of Fabricius was collected for bursa:body weight (B:BW) ratio calculation. In trial 2, birds were vaccinated when they were 1-day old according to the same protocol as trial 1, but at day 14, all groups also received a live infectious bronchitis (IB) vaccine. At 0, 7, 14, 21, and 28 days after IB vaccination, birds were tested by ELISA for IB serology and, soon after the last blood sampling, they were euthanized for collection of Harderian glands, trachea, and spleen and testing by flow cytometry for characterization of mononuclear cells. The immune-complex vaccine groups showed significantly lower B:BW ratio, lower IBV antibody titers, and higher mean percentage of CD8+ T cells in the spleen, trachea, and Harderian glands than those in the other experimental groups. The results of the in vivo trials coupled with a depth analysis of the repertoire of parameters involved in the immune response to IBD and IB vaccinations show one vaccine may influence the immune response of other vaccines included in the vaccination program.


Assuntos
Infecções por Birnaviridae/veterinária , Doenças das Aves Domésticas/imunologia , Vacinas Virais/imunologia , Animais , Infecções por Birnaviridae/imunologia , Bolsa de Fabricius/patologia , Galinhas , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/veterinária , Vírus da Bronquite Infecciosa/imunologia , Vírus da Doença Infecciosa da Bursa/imunologia , Vírus da Doença de Newcastle/imunologia , Doenças das Aves Domésticas/prevenção & controle , Doenças das Aves Domésticas/virologia , Organismos Livres de Patógenos Específicos , Vacinação/veterinária , Vacinas Atenuadas/imunologia
2.
PLoS One ; 15(8): e0237118, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32764795

RESUMO

The objective of this study was to evaluate the effect of supplementation with 100ppm sodium monensin or 0.15% of a blend of functional oils (cashew nut oil + castor oil) on the intestinal microbiota of broilers challenged with three different Eimeria spp. The challenge was accomplished by inoculating broiler chicks with sporulated oocysts of Eimeria tenella, Eimeria acervulina, and Eimeria maxima via oral gavage. A total of 864, day-old male broiler chicks (Cobb) were randomly assigned to six treatments (eight pens/treatment; 18 broilers/pen) in a 3 × 2 factorial arrangement, composed of three additives (control, monensin or blend), with or without Eimeria challenge. Intestinal contents was collected at 28 days of age for microbiota analysis by sequencing 16s rRNA in V3 and V4 regions using the Illumina MiSeq platform. Taxonomy was assigned through the SILVA database version 132, using the QIIME 2 software version 2019.1. No treatment effects (p > 0.05) were observed in the microbial richness at the family level estimated by Chao1 and the biodiversity assessed by Simpson's index, except for Shannon's index (p < 0.05). The intestinal microbiota was dominated by members of the order Clostridiales and Lactobacillales, followed by the families Ruminococcaceae, Bacteroidaceae, and Lactobacillaceae, regardless of treatment. When the controls were compared, in the challenged control group there was an increase in Erysipelotrichaceae, Lactobacillaceae, Bacteroidaceae, Streptococcaceae, and Peptostreptococcaceae, and a decrease in Ruminococcaceae. Similar results were found for a challenged group that received monensin, while the blend partially mitigated this variation. Therefore, the blend alleviated the impact of coccidiosis challenge on the microbiome of broilers compared to monensin.


Assuntos
Coccidiose/veterinária , Eimeria/isolamento & purificação , Microbioma Gastrointestinal/efeitos dos fármacos , Monensin/administração & dosagem , Óleos Vegetais/administração & dosagem , Doenças das Aves Domésticas/dietoterapia , Anacardium/química , Ração Animal , Animais , Galinhas/parasitologia , Coccidiose/dietoterapia , Coccidiose/imunologia , Coccidiose/parasitologia , DNA de Protozoário/isolamento & purificação , Eimeria/genética , Eimeria/imunologia , Eimeria/patogenicidade , Microbioma Gastrointestinal/imunologia , Masculino , Oocistos/patogenicidade , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/parasitologia , RNA Ribossômico 16S/genética , Ricinus/química
3.
J Virol ; 94(19)2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32669327

RESUMO

Infectious bronchitis (IB) caused by infectious bronchitis virus (IBV) is currently a major threat to chicken health, with multiple outbreaks being reported in the United States over the past decade. Modified live virus (MLV) vaccines used in the field can persist and provide the genetic material needed for recombination and emergence of novel IBV serotypes. Inactivated and subunit vaccines overcome some of the limitations of MLV with no risk of virulence reversion and emergence of new virulent serotypes. However, these vaccines are weakly immunogenic and poorly protective. There is an urgent need to develop more effective vaccines that can elicit a robust, long-lasting immune response. In this study, we evaluate a novel adjuvant system developed from Quil-A and chitosan (QAC) for the intranasal delivery of nucleic acid immunogens to improve protective efficacy. The QAC adjuvant system forms nanocarriers (<100 nm) that efficiently encapsulate nucleic acid cargo, exhibit sustained release of payload, and can stably transfect cells. Encapsulation of plasmid DNA vaccine expressing IBV nucleocapsid (N) protein by the QAC adjuvant system (pQAC-N) enhanced immunogenicity, as evidenced by robust induction of adaptive humoral and cellular immune responses postvaccination and postchallenge. Birds immunized with pQAC-N showed reduced clinical severity and viral shedding postchallenge on par with protection observed with current commercial vaccines without the associated safety concerns. Presented results indicate that the QAC adjuvant system can offer a safer alternative to the use of live vaccines against avian and other emerging coronaviruses.IMPORTANCE According to 2017 U.S. agriculture statistics, the combined value of production and sales from broilers, eggs, turkeys, and chicks was $42.8 billion. Of this number, broiler sales comprised 67% of the industry value, with the production of >50 billion pounds of chicken meat. The economic success of the poultry industry in the United States hinges on the extensive use of vaccines to control infectious bronchitis virus (IBV) and other poultry pathogens. The majority of vaccines currently licensed for poultry health include both modified live vaccine and inactivated pathogens. Despite their proven efficacy, modified live vaccine constructs take time to produce and could revert to virulence, which limits their safety. The significance of our research stems from the development of a safer and potent alternative mucosal vaccine to replace live vaccines against IBV and other emerging coronaviruses.


Assuntos
Bronquite/prevenção & controle , Infecções por Coronavirus/veterinária , Gammacoronavirus/imunologia , Membrana Mucosa/imunologia , Doenças das Aves Domésticas/prevenção & controle , Vacinas Virais/imunologia , Adjuvantes Imunológicos/farmacologia , Animais , Bronquite/virologia , Galinhas , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Modelos Animais de Doenças , Imunidade Celular , Imunização , Vírus da Bronquite Infecciosa/imunologia , Nucleocapsídeo/imunologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Proteínas Recombinantes/imunologia , Vacinas de DNA/imunologia , Carga Viral
4.
Arch Virol ; 165(10): 2249-2258, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32696270

RESUMO

While infectious bursal disease virus (IBDV) mainly targets immature B cells and causes T cell infiltration in the bursa of Fabricius (BF) of chickens, the effect of IBDV infection on the properties of T cells and relevant cytokine production in avian gut-associated lymphoid tissues (GALTs) remains unknown. Here, we show that while the CD8+ T cell subset is not affected, IBDV infection decreases the percentage of CD4+ T cells in the cecal tonsil (CT), but not in esophagus tonsil, pylorus tonsil, and Meckel's diverticulum of GALTs, in contrast to BF and spleen, in which the proportion of CD4+ cells increases upon IBDV infection. Further, IBDV infection upregulates IFN-γ, IL-10, and the T cell checkpoint receptor LAG-3 mRNA expression in BF. In contrast, in CTs, IBDV infection significantly increases the production of IFN-ß and CTLA-4 mRNA, while no significant effect is seen in the case of IFN-γ, IL-10 and LAG-3. Together, our data reveal differential modulation of T cell subsets and proinflammatory cytokine production in different lymphoid tissues during the course of IBDV infection.


Assuntos
Subpopulações de Linfócitos B/imunologia , Infecções por Birnaviridae/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Regulação da Expressão Gênica/imunologia , Doenças das Aves Domésticas/imunologia , Animais , Antígenos CD/genética , Antígenos CD/imunologia , Subpopulações de Linfócitos B/virologia , Infecções por Birnaviridae/genética , Infecções por Birnaviridae/patologia , Infecções por Birnaviridae/virologia , Bolsa de Fabricius/imunologia , Bolsa de Fabricius/virologia , Linfócitos T CD4-Positivos/virologia , Linfócitos T CD8-Positivos/virologia , Antígeno CTLA-4/genética , Antígeno CTLA-4/imunologia , Galinhas/virologia , Vírus da Doença Infecciosa da Bursa/crescimento & desenvolvimento , Vírus da Doença Infecciosa da Bursa/imunologia , Vírus da Doença Infecciosa da Bursa/patogenicidade , Interferon beta/genética , Interferon beta/imunologia , Interferon gama/genética , Interferon gama/imunologia , Interleucina-10/genética , Interleucina-10/imunologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/virologia , Tonsila Palatina/imunologia , Tonsila Palatina/virologia , Doenças das Aves Domésticas/genética , Doenças das Aves Domésticas/patologia , Doenças das Aves Domésticas/virologia
5.
Exp Parasitol ; 216: 107945, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32615133

RESUMO

Despite decades of investigation to clarify protective mechanisms of anticoccidial responses, one crucial field is neglected, that is, protective memory responses in primed birds. Protective memory immunity is critical for host resistance to reinfection and is the basis of modern vaccinology, especially in developing successful subunit vaccines. There are important differences between the immune responses induced by infections and antigens delivered either as killed, recombinant proteins or as live, replicating vector vaccines or as DNA vaccines. Animals immunized with these vaccines may fail to develop protective memory immunity, and is still naïve to Eimeria infection. This may explain why limited success is achieved in developing next-generation anticoccidial vaccines. In this review, we try to decipher the protective memory responses against Eimeria infection, assess immune responses elicited by various anticoccidial vaccine candidates, and propose possible approaches to develop rational vaccines that can induce a protective memory response to chicken coccidiosis.


Assuntos
Galinhas/parasitologia , Coccidiose/veterinária , Eimeria/imunologia , Memória Imunológica/fisiologia , Doenças das Aves Domésticas/imunologia , Vacinas Protozoárias , Animais , Galinhas/imunologia , Coccidiose/imunologia , Coccidiose/prevenção & controle , Intestinos/imunologia , Intestinos/parasitologia , Doenças das Aves Domésticas/prevenção & controle , Vacinas Protozoárias/imunologia , Recidiva , Vacinação/veterinária , Vacinas de Subunidades/imunologia
6.
Arch Virol ; 165(9): 1959-1968, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32519007

RESUMO

Newcastle disease (ND), caused by virulent Newcastle disease virus (NDV) strains, has been one of the most problematic diseases affecting the poultry industry worldwide. Conventional vaccines provide effective protection for birds to survive ND outbreaks, but they may not completely suppress NDV shedding. NDV strains circulate on farms for a long time after the initial infection and cause potential risks. A new vaccine with fast clearance ability and low viral shedding is needed. In this study, we used interleukin-12 (IL-12) as an adjuvant and electroporation (EP) as an advanced delivery system to improve a DNA vaccine candidate. The fusion (F) protein gene from an NDV strain of the prevalent genotype VII.1.1 was cloned to prepare the vaccine. Chickens immunized with the F gene DNA vaccine co-delivered with an IL-12-expressing plasmid DNA showed higher neutralizing antibody levels and stronger concanavalin-A-induced lymphocyte proliferation than those treated with the F gene DNA vaccine alone. The co-delivered vaccine provided 100% protection, and less viral shedding and a shorter release time were observed in challenged chickens than when the F gene DNA vaccine was administered alone. The use of F gene DNA combined with IL-12 delivered by electroporation is a promising approach for vaccination against ND.


Assuntos
Adjuvantes Imunológicos/administração & dosagem , Interleucina-12/imunologia , Doença de Newcastle/prevenção & controle , Vírus da Doença de Newcastle/imunologia , Doenças das Aves Domésticas/prevenção & controle , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/imunologia , Galinhas , Eletroporação , Interleucina-12/administração & dosagem , Doença de Newcastle/imunologia , Doença de Newcastle/virologia , Vírus da Doença de Newcastle/genética , Vírus da Doença de Newcastle/fisiologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Vacinação , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Proteínas Virais de Fusão/administração & dosagem , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/genética , Eliminação de Partículas Virais
7.
Vet Immunol Immunopathol ; 224: 110059, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32408182

RESUMO

There are currently no licensed vaccines against Clostridium perfringens which causes necrotic enteritis in poultry. Chitosan nanoparticles were formulated with native (CN) or toxoids (CT) of extracellular proteins (ECP) of C. perfringens, both surface-tagged with Salmonella flagellar proteins. In a pH stability assay, CN and CT nanoparticles released 6% and 0% of their protein at 8.0 pH. In a protein release assay, CN and CT nanoparticles released 16% and 10% of their protein respectively at 7.4 pH after 24 h. CN and CT nanoparticles incubated at 100 µg/mL PBS with Chicken RBCs released 1% and 0% hemoglobin respectively. Ninety broilers were randomly assigned to treatments; sham-vaccinated (Control), CN-vaccinated (CN), and CT-vaccinated (CT). Each bird was orally gavaged with 50 µg vaccine in 0.5 mL PBS or 0.5 mL PBS only on d 0, 3, 7 and 14 of age. At 21 d of age, the CN group had higher anti-ECP IgA than control (P < 0.05). At 21 d of age, the CN and CT group had higher anti-ECP IgA than control (P < 0.05). At 17 d of age, the CN group had higher anti-flagellar IgG than control (P < 0.05). At 10 d of age, the CN group had higher anti-flagellar IgA than control (P < 0.05). Splenic T cells from chickens in the CN and CT group ex-vivo stimulated with 0.05 mg/mL ECP, had higher proliferation control (P < 0.05, P < 0.01 respectively). Splenic T cells from chickens in the CN and CT groups ex-vivo stimulated with 0.1 mg/mL ECP had proliferation than control (P < 0.05). Pooled serum from 17 d of age CN and CT-vaccinated birds partially neutralized toxins in 50 µg of ECP (P < 0.05). Pooled serum from 28 d of age CN-vaccinated birds also partially neutralized toxins in 50 µg of ECP. The result from this study indicates the potential for chitosan loaded with Clostridium perfringens extracellular proteins to be applied to necrotic enteritis challenge studies.


Assuntos
Vacinas Bacterianas/imunologia , Quitosana/química , Infecções por Clostridium/veterinária , Enterocolite Necrosante/veterinária , Nanopartículas/química , Administração Oral , Animais , Anticorpos Antibacterianos/sangue , Anticorpos Neutralizantes/sangue , Antígenos de Bactérias/imunologia , Toxinas Bacterianas/imunologia , Galinhas/imunologia , Galinhas/microbiologia , Infecções por Clostridium/imunologia , Infecções por Clostridium/prevenção & controle , Clostridium perfringens , Enterocolite Necrosante/imunologia , Enterocolite Necrosante/prevenção & controle , Flagelos/imunologia , Imunogenicidade da Vacina , Imunoglobulina A/análise , Imunoglobulina G/sangue , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/microbiologia , Doenças das Aves Domésticas/prevenção & controle , Salmonella , Vacinas Atenuadas/imunologia
8.
PLoS One ; 15(4): e0231998, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32330169

RESUMO

Two experiments were conducted to evaluate the immune response of broilers vaccinated with Salmonella chitosan-nanoparticle (CNP) vaccine and challenged with Salmonella. The Salmonella CNP vaccine was synthesized with Salmonella enterica outer membrane proteins (OMPs) and flagellin proteins. In Experiment I, birds were orally gavaged with PBS or 500, 1000, or 2000µg of CNP vaccine 1 and 7d-of-age. At 14d-of-age, birds were orally challenged with 1 X 105 CFU/bird of live S. Enteritidis (SE). Macrophage-nitrite production 11d-post-challenge was higher (P<0.05) in the 500µg group when compared to the control. At d14 (8h-post-challenge), broilers vaccinated with 1000µg CNP had higher (P<0.05) serum anti-OMPs IgG and IgA and cloacal anti-OMP IgA amounts. At 11d-post-challenge, birds vaccinated with 1000µg CNP vaccine had greater (P<0.05) bile anti-OMP and anti-flagellin IgA amounts. At 11d-post-challenge, birds administered 1000µg CNP vaccine has increased (P<0.05) IL-1ß and IL-10 mRNA in cecal tonsils. In Experiment II, birds were orally gavaged with PBS or 1000µg CNP or a live commercial vaccine at 1 and 7d-of-age. At 14d-of-age, birds were orally challenged with 1 X 105 CFU/bird of live SE or S. Heidelberg (SH). Birds vaccinated with CNP showed higher (P<0.05) serum anti-OMPs IgG amounts at 8h-post-challenge. At 4d-post-SH challenge, birds vaccinated with CNP had higher (P<0.05) bile anti-flagellin IgA amounts. CNP decreased (P<0.05) anti-OMPs IgG levels in serum at 2d-post-SE challenge and 4d-post-SH or SE challenge. Salmonella Enteritidis loads in cecal content at 2d-post-challenge was decreased (P<0.05) by 65.9% in birds vaccinated with CNP, when compared to the control. Chitosan-nanovaccine had no adverse effects on bird's production performance. In conclusion, 1000µg CNP vaccine can induce a specific immune response against Salmonella and has the potential to mitigate SE cecal colonization in broiler birds.


Assuntos
Galinhas/imunologia , Quitosana/farmacologia , Vacinas contra Salmonella/imunologia , Animais , Proteínas da Membrana Bacteriana Externa/imunologia , Ceco/metabolismo , Galinhas/microbiologia , Quitosana/imunologia , Flagelina/imunologia , Antígenos de Histocompatibilidade Classe II , Nanopartículas/uso terapêutico , Doenças das Aves Domésticas/imunologia , Salmonella/imunologia , Salmonella/patogenicidade , Salmonelose Animal/imunologia , Salmonella enterica/metabolismo , Salmonella enteritidis/imunologia , Vacinas/imunologia , Vacinas Atenuadas/imunologia
9.
Avian Dis ; 64(1): 53-59, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32267125

RESUMO

Recombinant Newcastle disease virus (NDV) LaSota (LS) expressing secreted trimeric spike (S)-ectodomain (Se) of infectious bronchitis virus (IBV) (rLS/IBV.Se) was developed and evaluated for protection conferred against IBV challenge. The IBV S-ectodomain protein, which is S excluding the transmembrane anchor and short cytoplasmic domain of S2, expressed from recombinant LS corresponds to an Arkansas (Ark)-type IBV. In a first experiment, chickens were primed at 1 day of age or primed at 1 day of age and boosted at 14 days of age with 104 50% embryo infectious doses (EID50)/bird of rLS/IBV.Se and challenged with a virulent Ark strain. A single vaccination proved completely ineffective at protecting chickens against challenge, whereas priming and boosting reduced clinical signs and tracheal lesions but did not reduce viral load in lachrymal fluids. In experiment 2, the vaccine dose was increased to 107 EID50/bird and a different virulent Ark strain was used for challenge. In addition, chickens were singly immunized on either day 1 or day 10 after hatch. NDV antibody levels detected in vaccinated chickens were moderate, with hemagglutination inhibition titers varying between 4 and 5 log2. Slightly higher antibody levels to NDV were observed in chickens vaccinated on day 10 versus day 1 but without the difference achieving statistical significance. In contrast, antibody responses measured using recombinant IBV S1 protein-coated ELISA plates were significantly greater in chickens vaccinated on day 10 than on day 1. The use of a higher rLS/IBV.Se dose substantially enhanced the success of a single vaccination compared to experiment 1. Signs and tracheal lesions were reduced more effectively in chickens vaccinated at day 10 after hatch. However, as in experiment 1, vaccination did not reduce the viral loads in tear fluids of challenged chickens. Similar results, in which no reduction in viral load in the trachea was apparent from rLS/IBV.S vaccination, have been obtained by others. Further work is needed to understand the immune responses induced by this recombinant virus that seems to provide some protection against the disease but does not reduce viral loads in the upper respiratory tract.


Assuntos
Galinhas , Infecções por Coronavirus/veterinária , Vírus da Bronquite Infecciosa/imunologia , Vírus da Doença de Newcastle/imunologia , Doenças das Aves Domésticas/prevenção & controle , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinas Virais/imunologia , Animais , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/virologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Vacinas Sintéticas/imunologia
10.
Avian Dis ; 64(1): 60-68, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32267126

RESUMO

A commercial Arkansas (Ark) Delmarva Poultry Industry (DPI)-type vaccine and a more homogeneous population of that vaccine obtained previously through adaptation to chicken embryo kidney (CEK) cells (CEK-ArkDPI) were used as a model to further understand the impact of population genetic structure on generation of immune responses and protection. In a first experiment, vaccinated chickens were challenged with an IBV Ark99-type virulent strain (AL/4614/98). Despite extensive sequence similarity between the vaccines, the more heterogeneous commercial ArkDPI was more efficient at reducing viral loads in challenged chickens, while respiratory signs and tracheal lesions were reduced similarly by either vaccine. A distinct subpopulation of the Ark challenge virus showing asparagine at S1 position 56 was consistently negatively selected by immune pressure originating from vaccination with either vaccine. Antibody levels and antibody avidity to Ark-type S1 protein were greater in CEK-ArkDPI-vaccinated chickens compared to chickens vaccinated with the more diverse commercial ArkDPI vaccine. Synchronous replication of a homogeneous virus population likely elicits clonal expansion and affinity maturation of a greater number of responding B cells compared to a diverse virus population continuously changing its proportion of phenotypes during replication. The results of a second experiment showed that during initial vaccine virus replication (24 and 48 hr postvaccination), the virus population showing increased diversity (commercial ArkDPI) achieved higher concentrations of IBV RNA in the trachea compared to the more homogenous virus. mRNA expression of genes associated with innate immune responses in the trachea 48 hr postvaccination generally showed greater upregulation in chickens vaccinated with the heterogeneous commercial ArkDPI vaccine compared to the CEK-adapted virus. The greater upregulation of these genes is likely associated with higher virus replication achieved by the heterogeneous commercial vaccine. Thus, while the adaptive antibody response was favored by the more homogenous structure of the CEK-ArkDPI vaccine population (higher antibody levels and antibody avidity), the innate immune response was favored by the more diverse viral population of the commercial ArkDPI. We confirmed previous results that distinct subpopulations in wild Ark challenge virus become selected by immune pressure originating from vaccination, and we concluded that the population structure of IBV vaccines impacts innate immune response, antibody avidity, and protection.


Assuntos
Galinhas , Infecções por Coronavirus/veterinária , Imunidade Inata/fisiologia , Vírus da Bronquite Infecciosa/fisiologia , Doenças das Aves Domésticas/imunologia , Vacinas Virais/imunologia , Animais , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Vírus da Bronquite Infecciosa/imunologia , Doenças das Aves Domésticas/virologia , Vacinas Atenuadas/imunologia
11.
Avian Dis ; 64(1): 92-95, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32267130

RESUMO

In recent years, Arkansas Delmarva Poultry Industry (ArkDPI)-derived infectious bronchitis (IB) virus (IBV) vaccines have been used to characterize the immune responses of chickens subsequent to vaccination on day of hatch or beyond. Perhaps because ArkDPI vaccines display increased heterogeneity, the results on cell immune responses have shown ambiguity. In the current study, we investigated the effects of vaccination with a highly stable and homogeneous Massachusetts (Mass)-type vaccine on days 1 or 7 of age on Harderian gland (HG) responses. Confirming previous studies, both IBV serum antibodies and lachrymal IgA levels were greater upon vaccination on day 7 compared with vaccination on day 1 of age. Unlike results with ArkDPI viruses, a clear trend was detected for both B and T cells in the HG after Mass-type vaccination. Consistent with antibody responses, B- and T-helper (CD3+CD4+) cell frequencies were higher in birds vaccinated on day 7 of age. Cytotoxic T cells (CD3+CD8+) were also increased compared with chickens vaccinated on day 1 of age. Depending on the most likely age of IB outbreaks to occur in a particular region, postponing the first IBV vaccination may optimize immune responses.


Assuntos
Galinhas , Infecções por Coronavirus/veterinária , Imunidade Inata , Vírus da Bronquite Infecciosa/imunologia , Doenças das Aves Domésticas/imunologia , Vacinação/veterinária , Vacinas Virais/imunologia , Animais , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Glândula de Harder/imunologia , Glândula de Harder/virologia , Doenças das Aves Domésticas/virologia
12.
Arch Razi Inst ; 75(1): 31-37, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32292000

RESUMO

Newcastle disease (ND) is a major threat to poultry industry production throughout developing countries. The Newcastle disease viruses (NDVs) infecting industrialized and indigenous poultry in Iran are velogenic strains and responsible for the frequent outbreaks of ND in poultry farms even in vaccinated flocks causing serious economic losses in the commercial and indigenous poultry. However, vaccination is the only way to protect against endemic ND, and the conventional vaccines are not heat stable and consequently require complex cold-chains to be transferred to users leading to not much resistance. The present study aimed to evaluate the efficacy of thermostable NDV strain I-2 in broiler chickens vaccinated via drinking water and coated on oiled wheat grain. The horizontal transmission of I-2 strain and transmission of disease from vaccinated to unvaccinated chickens were also evaluated in this study. The obtained results showed that both routes of administration, following primary and/or secondary dose, provoked the production of necessary antibody titer and adequate protective immunity in broiler chickens. Moreover, the horizontal transmission of I-2 strain from vaccinated to unvaccinated chickens housed together induced an antibody response and protected unvaccinated chickens against a local field isolate of a virulent strain of NDV (The intravenous pathogenicity index 2.46, mean death time 59 h). Nevertheless, all unvaccinated and Newcastle challenged broilers chickens against the NDV died in this study. It is noteworthy that the transmission of the virus from challenged broiler chickens was very low to induce clinical signs in susceptible chickens. The obtained results of this study revealed the efficacy of NDV strain I-2 coated on the oiled wheat and via drinking water as it protects broiler chickens from highly virulent NDV.


Assuntos
Galinhas , Doença de Newcastle/prevenção & controle , Vírus da Doença de Newcastle/imunologia , Doenças das Aves Domésticas/prevenção & controle , Vacinação/veterinária , Vacinas Virais/farmacologia , Animais , Anticorpos Antivirais/sangue , Formação de Anticorpos , Irã (Geográfico) , Doença de Newcastle/imunologia , Doença de Newcastle/virologia , Vírus da Doença de Newcastle/patogenicidade , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/virologia , Vacinas Virais/administração & dosagem , Vacinas Virais/química
13.
Infect Immun ; 88(5)2020 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-32122943

RESUMO

Mycoplasma gallisepticum is the primary etiological agent of chronic respiratory disease in chickens. Live attenuated vaccines are most commonly used in the field to control the disease, but current vaccines have some limitations. Vaxsafe MG (strain ts-304) is a new vaccine candidate that is efficacious at a lower dose than the current commercial vaccine strain ts-11, from which it is derived. In this study, the transcriptional profiles of the trachea of unvaccinated chickens and chickens vaccinated with strain ts-304 were compared 2 weeks after challenge with M. gallisepticum strain Ap3AS during the chronic stage of infection. After challenge, genes, gene ontologies, pathways, and protein classes involved in inflammation, cytokine production and signaling, and cell proliferation were upregulated, while those involved in formation and motor movement of cilia, formation of intercellular junctional complexes, and formation of the cytoskeleton were downregulated in the unvaccinated birds compared to the vaccinated birds, reflecting immune dysregulation and the pathological changes induced in the trachea by infection with M. gallisepticum Vaccination appears to protect the structural and functional integrity of the tracheal mucosa 2 weeks after infection with M. gallisepticum.


Assuntos
Galinhas/imunologia , Galinhas/microbiologia , Mycoplasma gallisepticum/imunologia , Traqueia/imunologia , Traqueia/microbiologia , Transcrição Genética/imunologia , Animais , Anticorpos Antibacterianos/imunologia , Vacinas Bacterianas/imunologia , Proliferação de Células/fisiologia , Membrana Mucosa/imunologia , Membrana Mucosa/microbiologia , Infecções por Mycoplasma/imunologia , Infecções por Mycoplasma/microbiologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/microbiologia , Regulação para Cima/imunologia , Vacinação/métodos , Vacinas Atenuadas/imunologia
14.
Mol Immunol ; 120: 52-60, 2020 04.
Artigo em Inglês | MEDLINE | ID: covidwho-2404

RESUMO

Avian infectious bronchitis (IB) is an acute, highly infectious and contagious viral disease of chickens caused by avian infectious bronchitis virus (IBV) belonging to the genus Coronavirus and family Coronaviridae. It can affect all age groups of birds. The toll-like receptors (TLRs) are a major class of innate immune pattern recognition receptors that have a key role in immune response and defense against various infections.The TLRs are essential for initiation of innate immune responses and in the development of adaptive immune responses. An in ovo model was employed to study the antiviral activity of TLR ligands (Pam3CSK4, LPS and CpG ODN) on replication of IBV. It was hypothesized that optimum dose and specific timing of TLR ligands may reduce viral load of IBV in specific pathogen free (SPF) embryonated chicken eggs (ECEs). Further, the mechanism involved in the TLR-mediated antiviral response in chorioallantoic membrane (CAM) of ECEs was investigated. The ECEs of 9-11 days old were treated with different doses (high, intermediate and low) of TLR-2 (Pam3CSK4), TLR-4 (LPS) and TLR-21 (CpG ODN) ligands. In addition, to know the timing of TLR ligand treatment, six time intervals were analyzed viz. 36, 24 and 12 h prior to infection, time of infection (co-administration of TLR ligands and avian IBV) and 12 and 24 h post-IBV infection. For studying the relative expression of immuno-stimulatory genes (IFN-α, IFN-ß, IFN-γ, IL-1ß, iNOS and OAS) in CAM, TLR ligands were administered through intra-allantoicroute and CAM were collected at 4, 8 and 16 h post treatment. The results demonstrated that intermediate dose of all the three TLR ligands significantly reduced virus titers and used in the present study. However, the LPS reduced virus titer pre- and post-IBV infection but Pam3CSK4 and CpG ODN reduced only pre-IBV infection. Further analysis showed that TLR ligands induced IFN-γ, IL-1ß and IFN stimulated genes viz. iNOS and OAS genes in CAM. The present study pointed towards the novel opportunities for rational design of LPS as immuno-stimulatory agent in chickens with reference to IBV. It may be speculated that in ovo administration of these TLR ligands may enhance resistance against viral infection in neonatal chicken and may contribute towards the development of more effective and safer vaccines including in ovo vaccines.


Assuntos
Vírus da Bronquite Infecciosa/imunologia , Receptores Toll-Like/agonistas , Adjuvantes Imunológicos/farmacologia , Animais , Antivirais/farmacologia , Proteínas Aviárias/agonistas , Proteínas Aviárias/imunologia , Embrião de Galinha , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Citocinas/metabolismo , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/imunologia , Imunidade Inata , Vírus da Bronquite Infecciosa/patogenicidade , Vírus da Bronquite Infecciosa/fisiologia , Ligantes , Lipopeptídeos/farmacologia , Lipopolissacarídeos/farmacologia , Oligodesoxirribonucleotídeos/farmacologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/prevenção & controle , Doenças das Aves Domésticas/virologia , Receptores Toll-Like/imunologia , Carga Viral/efeitos dos fármacos , Carga Viral/imunologia , Replicação Viral/efeitos dos fármacos , Replicação Viral/imunologia
15.
Arch Virol ; 165(4): 835-843, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: covidwho-71756

RESUMO

Avian infectious bronchitis virus (IBV) is a coronavirus with great economic impact on the poultry industry, causing an acute and highly contagious disease in chickens that primarily affects the respiratory and reproductive systems. The cellular regulation of IBV pathogenesis and the host immune responses involved remain to be fully elucidated. MicroRNAs (miRNAs) have emerged as a class of crucial regulators of numerous cellular processes, including responses to viral infections. Here, we employed a high-throughput sequencing approach to analyze the miRNA composition of the spleen and the lungs of chicken embryos upon IBV infection. Compared to healthy chicken embryos, 13 and six miRNAs were upregulated in the spleen and the lungs, respectively, all predicted to influence viral transcription, cytokine production, and lymphocyte functioning. Subsequent downregulation of NFATC3, NFAT5, SPPL3, and TGFB2 genes in particular was observed only in the spleen, demonstrating the biological functionality of the miRNAs in this lymphoid organ. This is the first study that describes the modulation of miRNAs and the related host immune factors by IBV in chicken embryos. Our data provide novel insight into complex virus-host interactions and specifically highlight components that could affect the host's immune response to IBV infection.


Assuntos
Infecções por Coronavirus/veterinária , Gammacoronavirus/fisiologia , MicroRNAs/imunologia , Óvulo/virologia , Doenças das Aves Domésticas/imunologia , Animais , Galinhas , Infecções por Coronavirus/genética , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Citocinas/genética , Citocinas/imunologia , Gammacoronavirus/genética , Pulmão/imunologia , Pulmão/patologia , MicroRNAs/genética , Óvulo/imunologia , Doenças das Aves Domésticas/genética , Doenças das Aves Domésticas/patologia , Doenças das Aves Domésticas/virologia , Baço/imunologia , Baço/patologia
16.
Vet Res ; 51(1): 8, 2020 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-32014061

RESUMO

MicroRNAs (miRNAs) are small non-coding RNAs that contribute to host immune response as post-transcriptional regulation. The current study investigated the biological role of the chicken (Gallus gallus) microRNA-200a-3p (gga-miR-200a-3p), using 2 necrotic enteritis (NE) afflicted genetically disparate chicken lines, 6.3 and 7.2, as well as the mechanisms underlying the fundamental signaling pathways in chicken. The expression of gga-miR-200a-3p in the intestinal mucosal layer of NE-induced chickens, was found to be upregulated during NE infection in the disease-susceptible chicken line 7.2. To validate the target genes, we performed an overexpression analysis of gga-miR-200a-3p using chemically synthesized oligonucleotides identical to gga-miR-200a-3p, reporter gene analysis including luciferase reporter assay, and a dual fluorescence reporter assay in cultured HD11 chicken macrophage cell lines. Gga-miR-200a-3p was observed to be a direct transcriptional repressor of ZAK, MAP2K4, and TGFß2 that are involved in mitogen-activated protein kinase (MAPK) pathway by targeting the 3'-UTR of their transcripts. Besides, gga-miR-200a-3p may indirectly affect the expression of protein kinases including p38 and ERK1/2 at both transcriptional and translational levels, suggesting that this miRNA may function as an important regulator of the MAPK signaling pathway. Proinflammatory cytokines consisting of IL-1ß, IFN-γ, IL-12p40, IL-17A, and LITAF belonging to Th1 and Th17-type cytokines, were upregulated upon gga-miR-200a-3p overexpression. These findings have enhanced our knowledge of the immune function of gga-miR-200a-3p mediating the chicken immune response via regulation of the MAPK signaling pathway and indicate that this miRNA may serve as an important biomarker of diseases in domestic animals.


Assuntos
Galinhas , Enterite/veterinária , Imunidade Inata/genética , MicroRNAs/imunologia , Necrose/veterinária , Doenças das Aves Domésticas/imunologia , Animais , Enterite/genética , Enterite/imunologia , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/imunologia , Necrose/genética , Necrose/imunologia , Doenças das Aves Domésticas/genética , Transdução de Sinais
17.
Poult Sci ; 99(2): 725-733, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32036975

RESUMO

The objective of this study was to investigate the effects of dietary Bacillus subtilis supplementation on growth performance, jejunal lesion scores, oocyst shedding, and cytokine and tight junction protein expression in broiler chickens infected with Eimeria maxima. A total of 196 male day-old Ross 708 broilers were given a nonexperimental diet until 14 D of age. Then, all chickens were randomly assigned to one of seven dietary treatments: 2 basal diets (CON and NC); CON + virginiamycin (AB1); CON + bacitracin methylene disalicylate (BMD; AB2); CON + B. subtilis 1781 (PB1); CON + B. subtilis 747 (PB2); or CON + B. subtilis 1781 + 747 (PB3). At day 21, all chickens except those in the CON group were orally inoculated with E. maxima oocysts. At 7 D after E. maxima infection, the body weight gains of chickens fed PB2 and PB3 increased (P = 0.032) as much as those in chickens fed AB2. The body weight gain and feed efficiency of chickens fed PB2 were significantly increased (P < 0.001), and PB2 chickens showed (P = 0.005) the lowest lesion scores after E. maxima infection. Chickens fed PB2 showed (P < 0.05) lower mRNA expression of IL-1ß in infected chicken groups. Chickens in the AB1, AB2, PB1, PB2, and PB3 groups showed (P < 0.05) greater mRNA expression of junctional adhesion molecule 2 in jejunal tissue, whereas occludin expression increased (P < 0.05) in the jejunal tissue of chickens fed AB2 or PB2. Dietary B. subtilis supplementation significantly improved the growth performance of young chickens to a level comparable with that induced by virginiamycin or BMD without E. maxima infection. After infection with E. maxima, dietary virginiamycin and BMD significantly enhanced the epithelial barrier integrity, and the dietary B. subtilis 747 showed significantly enhanced growth performance, intestinal immunity, and epithelial barrier integrity. Together our results indicated that certain strains of B. subtilis provide beneficial effects on the growth of young broiler chickens and have the potential to replace antibiotic growth promoters.


Assuntos
Bacillus subtilis/química , Galinhas , Coccidiose/veterinária , Eimeria/fisiologia , Imunidade Inata/efeitos dos fármacos , Mucosa Intestinal/imunologia , Doenças das Aves Domésticas/tratamento farmacológico , Probióticos/administração & dosagem , Ração Animal/análise , Animais , Galinhas/crescimento & desenvolvimento , Coccidiose/tratamento farmacológico , Coccidiose/imunologia , Coccidiose/parasitologia , Dieta/veterinária , Masculino , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/parasitologia
18.
Mol Immunol ; 120: 52-60, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32065987

RESUMO

Avian infectious bronchitis (IB) is an acute, highly infectious and contagious viral disease of chickens caused by avian infectious bronchitis virus (IBV) belonging to the genus Coronavirus and family Coronaviridae. It can affect all age groups of birds. The toll-like receptors (TLRs) are a major class of innate immune pattern recognition receptors that have a key role in immune response and defense against various infections.The TLRs are essential for initiation of innate immune responses and in the development of adaptive immune responses. An in ovo model was employed to study the antiviral activity of TLR ligands (Pam3CSK4, LPS and CpG ODN) on replication of IBV. It was hypothesized that optimum dose and specific timing of TLR ligands may reduce viral load of IBV in specific pathogen free (SPF) embryonated chicken eggs (ECEs). Further, the mechanism involved in the TLR-mediated antiviral response in chorioallantoic membrane (CAM) of ECEs was investigated. The ECEs of 9-11 days old were treated with different doses (high, intermediate and low) of TLR-2 (Pam3CSK4), TLR-4 (LPS) and TLR-21 (CpG ODN) ligands. In addition, to know the timing of TLR ligand treatment, six time intervals were analyzed viz. 36, 24 and 12 h prior to infection, time of infection (co-administration of TLR ligands and avian IBV) and 12 and 24 h post-IBV infection. For studying the relative expression of immuno-stimulatory genes (IFN-α, IFN-ß, IFN-γ, IL-1ß, iNOS and OAS) in CAM, TLR ligands were administered through intra-allantoicroute and CAM were collected at 4, 8 and 16 h post treatment. The results demonstrated that intermediate dose of all the three TLR ligands significantly reduced virus titers and used in the present study. However, the LPS reduced virus titer pre- and post-IBV infection but Pam3CSK4 and CpG ODN reduced only pre-IBV infection. Further analysis showed that TLR ligands induced IFN-γ, IL-1ß and IFN stimulated genes viz. iNOS and OAS genes in CAM. The present study pointed towards the novel opportunities for rational design of LPS as immuno-stimulatory agent in chickens with reference to IBV. It may be speculated that in ovo administration of these TLR ligands may enhance resistance against viral infection in neonatal chicken and may contribute towards the development of more effective and safer vaccines including in ovo vaccines.


Assuntos
Vírus da Bronquite Infecciosa/imunologia , Receptores Toll-Like/agonistas , Adjuvantes Imunológicos/farmacologia , Animais , Antivirais/farmacologia , Proteínas Aviárias/agonistas , Proteínas Aviárias/imunologia , Embrião de Galinha , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Citocinas/metabolismo , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/imunologia , Imunidade Inata , Vírus da Bronquite Infecciosa/patogenicidade , Vírus da Bronquite Infecciosa/fisiologia , Ligantes , Lipopeptídeos/farmacologia , Lipopolissacarídeos/farmacologia , Oligodesoxirribonucleotídeos/farmacologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/prevenção & controle , Doenças das Aves Domésticas/virologia , Receptores Toll-Like/imunologia , Carga Viral/efeitos dos fármacos , Carga Viral/imunologia , Replicação Viral/efeitos dos fármacos , Replicação Viral/imunologia
19.
J Virol ; 94(8)2020 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-32024774

RESUMO

Tembusu virus (TMUV) is a flavivirus responsible for panzootic outbreaks of severe egg-drop and fatal encephalitis of domestic waterfowl in China. Although TMUV can be attenuated by in vitro passaging, experimental evidence supporting the role of specific genetic changes in virulence attenuation is currently lacking. Here, we performed site-directed mutagenesis on five envelope (E) protein amino acid residues in accordance with the attenuated TMUV generated in our recent study. Our results showed that the Thr-to-Lys mutation of residue 367 in E protein (E367) plays a predominant role in viral cell adaptation and virulence attenuation in ducks compared with mutations in other residues. We further demonstrated that the positively charged basic amino acid substitution at E367 enhanced the viral binding affinity for glycosaminoglycans (GAGs) and reduced viremia levels and the efficiency of replication in major target organs in subcutaneously inoculated ducks. Interestingly, the T367K mutation increased viral neutralization sensitivity to the early immune sera. Together, our findings provide the first evidence that a basic amino acid substitution at E367 strongly impacts the in vitro and in vivo infection of TMUV.IMPORTANCE Outbreaks of Tembusu virus (TMUV) infection have caused huge economic losses in the production of domestic waterfowl since the virus was first recognized in China in 2010. To control TMUV infection, a live-attenuated vaccine candidate of TMUV was developed in our previous study, but the mechanisms of virulence attenuation are not fully understood. Here, we found that the Thr-to-Lys substitution at E367 is a crucial determinant of TMUV virulence attenuation in ducks. We demonstrated that the T367K mutation attenuates TMUV through reducing viral replication in the blood, brain, heart (ducklings), and ovaries. These data provide new insights into understanding the pathogenesis of TMUV and the rational development of novel TMUV vaccines.


Assuntos
Substituição de Aminoácidos , Infecções por Flavivirus/imunologia , Infecções por Flavivirus/virologia , Flavivirus/genética , Proteínas do Envelope Viral/genética , Substituição de Aminoácidos/imunologia , Animais , Anticorpos Neutralizantes , Linhagem Celular , China/epidemiologia , Patos/virologia , Feminino , Infecções por Flavivirus/epidemiologia , Infecções por Flavivirus/patologia , Mutagênese Sítio-Dirigida , Mutação , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/mortalidade , Doenças das Aves Domésticas/patologia , Doenças das Aves Domésticas/virologia , Carga Viral , Virulência , Replicação Viral
20.
BMC Vet Res ; 16(1): 47, 2020 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-32028947

RESUMO

BACKGROUND: Necrotic enteritis, which is caused by Clostridium perfringens, has resulted in more than $2 billion losses in the poultry industry every year. Due to the ban of antibiotics in feed industry, alternatives like environment improvement and probiotics have been found to be effective as well. In our study, we aim to explore the protective effect of Lactobacillus plantarum supplementation on CP infected chickens in two environments. RESULTS: The results showed that the Clostridium perfringens administration led to visible and histomorphological gut lesions. In the specific pathogen free or free-range system environment, dietary supplementation with LP obvious increased the ratio of intestinal villus height to crypt depth and the expression of MUC2 mRNA in ileum mucosa, then reduced the mRNA expression level of TNF-α gene in the ileum mucosa. LP treatment significantly reduced the contents of total protein, total superoxide dismutase and glutamic oxaloacetic transaminase in serum of the chickens. CONCLUSIONS: The specific pathogen free environment contributed to the recovery of pre-inflammation of the chickens, and free-range system environment contributed to the repair of damage in the later stages of chicken inflammation. Supplementation of LP in FRS environment was more conducive to the recovery of CP infected in chickens.


Assuntos
Infecções por Clostridium/veterinária , Clostridium perfringens , Lactobacillus plantarum , Probióticos/administração & dosagem , Animais , Galinhas , Infecções por Clostridium/imunologia , Infecções por Clostridium/patologia , Infecções por Clostridium/prevenção & controle , Dieta/veterinária , Inflamação , Mucosa Intestinal/patologia , Mucina-2/genética , Mucina-2/metabolismo , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/microbiologia , Doenças das Aves Domésticas/prevenção & controle , RNA Mensageiro , Organismos Livres de Patógenos Específicos , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
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