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1.
Acta Virol ; 63(3): 301-308, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31507196

RESUMO

Transmissible gastroenteritis virus (TGEV) causes great economic loss to swine industry worldwide. Vaccination is an important method to control the TGEV infection. In this study, a TGEV oral vaccine was generated by transferring a eukaryotic expression recombinant plasmid carrying the SAD (A and D antigenic sites of the S protein) epitope of TGEV into a swine-origin Lactobacillus acidophilus (L. acidophilus). In orally immunized BALB/c mice, the TGEV L. acidophilus oral vaccine induced significantly higher level of SIgA antibodies specific to TGEV compared with the mice immunized with a commercial inactivated TGEV vaccine and similar levels of IgG specific to TGEV as the inactivated vaccine. Furthermore, the TGEV L. acidophilus oral vaccine induced higher levels of IFN-γ, which suggested that the vaccine was able to induce immune response. In brief, this novel TGEV L. acidophilus oral vaccine could induce high levels of both mucosal and humoral immune responses, which has a potential to be used in the pig industries in the future. Keywords: transmissible gastroenteritis virus (TGEV); live L. acidophilus oral vaccine; SIgA antibody; IgG antibody; IFN-γ; IL-4.


Assuntos
Anticorpos Antivirais , Epitopos , Gastroenterite Suína Transmissível , Lactobacillus acidophilus , Vírus da Gastroenterite Transmissível , Vacinas Virais , Administração Oral , Animais , Anticorpos Antivirais/sangue , Epitopos/genética , Epitopos/imunologia , Gastroenterite Suína Transmissível/imunologia , Gastroenterite Suína Transmissível/patologia , Imunogenicidade da Vacina/imunologia , Lactobacillus acidophilus/genética , Lactobacillus acidophilus/virologia , Camundongos , Camundongos Endogâmicos BALB C , Plasmídeos/genética , Suínos , Proteínas Virais/genética , Proteínas Virais/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/imunologia
2.
Arch Virol ; 164(11): 2659-2669, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31385116

RESUMO

Interferon gamma (IFN-γ) is best known for its ability to regulate host immune responses; however, its direct antiviral activity is less well studied. Transmissible gastroenteritis virus (TGEV) is an economically important swine enteric coronavirus and causes acute diarrhea in piglets. At present, little is known about the function of IFN-γ in the control of TGEV infection. In this study, we demonstrated that IFN-γ inhibited TGEV infection directly in ST cells and intestine epithelial IPEC-J2 cells and that the anti-TGEV activity of IFN-γ was independent of IFN-α/ß. Moreover, IFN-γ suppressed TGEV infection in ST cells more efficiently than did IFN-α, and the combination of IFN-γ and IFN-α displayed a synergistic effect against TGEV. Mechanistically, using overexpression and functional knockdown experiments, we demonstrated that porcine interferon regulatory factor 1 (poIRF1) elicited by IFN-γ primarily mediated IFN-γ signaling cascades and the inhibition of TGEV infection by IFN-γ. Importantly, we found that TGEV elevated the expression of poIRF1 and IFN-γ in infected small intestines and peripheral blood mononuclear cells. Thus, IFN-γ plays a crucial role in curtailing enteric coronavirus infection and may serve as an effective prophylactic and/or therapeutic agent against TGEV infection.


Assuntos
Gastroenterite Suína Transmissível/imunologia , Fator Regulador 1 de Interferon/metabolismo , Interferon gama/imunologia , Vírus da Gastroenterite Transmissível/imunologia , Animais , Linhagem Celular , Cercopithecus aethiops , Fator Regulador 1 de Interferon/genética , Interferon-alfa/imunologia , Interferência de RNA , RNA Interferente Pequeno/genética , Transdução de Sinais/imunologia , Suínos , Células Vero
3.
Arch Virol ; 164(4): 983-994, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30729994

RESUMO

Transmissible gastroenteritis virus (TGEV) infection causes severe diarrhea in piglets and imposes a significant economic burden on pig farms. Single-chain fragment variable (scFv) antibodies effectively inhibit virus infection and could be a potential therapeutic reagent for preventing disease. In this study, a recombinant scFv antibody phage display library was constructed from peripheral blood lymphocytes of piglets infected with TGEV. The library was screened with four rounds of biopanning using purified TGEV antigen, and scFv antibodies that bound to TGEV were obtained. The scFv gene was subcloned into the pET-28a(+), and the constituted plasmid was introduced into Escherichia coli BL21 (DE3) for protein expression. All three scFv clones identified had neutralizing activity against TGEV. An immunofluorescence assay and western blot analysis demonstrated that two scFv antibodies reacted with the spike protein of TGEV. These results indicate that scFv antibodies provide protection against viral infection in vitro and may be a therapeutic candidate for both prevention and treatment of TGEV infection in swine.


Assuntos
Anticorpos Antivirais/imunologia , Gastroenterite Suína Transmissível/virologia , Anticorpos de Cadeia Única/imunologia , Vírus da Gastroenterite Transmissível/imunologia , Animais , Anticorpos Antivirais/genética , Gastroenterite Suína Transmissível/imunologia , Testes de Neutralização , Anticorpos de Cadeia Única/genética , Suínos , Vírus da Gastroenterite Transmissível/genética
4.
Appl Microbiol Biotechnol ; 102(19): 8403-8417, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30022263

RESUMO

Transmissible gastroenteritis coronavirus (TGEV) is one of the most severe threats to the swine industry. In this study, we constructed a suite of recombinant Lactobacillus plantarum with surface displaying the spike (S) protein coming from TGEV and fused with DC cells targeting peptides (DCpep) to develop an effective, safe, and convenient vaccine against transmissible gastroenteritis. Our research results found that the recombinant Lactobacillus plantarum (NC8-pSIP409-pgsA-S-DCpep) group expressing S fused with DCpep could not only significantly increase the percentages of MHC-II+CD80+ B cells and CD3+CD4+ T cells but also the number of IgA+ B cells and CD3+CD4+ T cells of ileum lamina propria, which elevated the specific secretory immunoglobulin A (SIgA) titers in feces and IgG titers in serum. Taken together, these results suggest that NC8-pSIP409-pgsA-S-DCpep expressing the S of TGEV fused with DCpep could effectively induce immune responses and provide a feasible original strategy and approach for the design of TGEV vaccines.


Assuntos
Proteínas de Bactérias/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Lactobacillus plantarum/imunologia , Vírus da Gastroenterite Transmissível/imunologia , Animais , Anticorpos Antivirais/imunologia , Linfócitos B/imunologia , Gastroenterite Suína Transmissível/imunologia , Imunoglobulina A Secretora/imunologia , Imunoglobulina G/imunologia , Suínos , Linfócitos T/imunologia , Vacinas Virais/imunologia
5.
Virology ; 507: 170-178, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28448848

RESUMO

Transmissible gastroenteritis virus (TGEV) is a porcine enteric coronavirus which causes lethal severe watery diarrhea in piglets. The pathogenesis of TGEV is strongly associated with inflammation. In this study, we found that TGEV infection activates transcription factors NF-κB, IRF3 and AP-1 in a time- and dose-dependent manner in porcine kidney cells. Treatment with the NF-κB-specific inhibitor BAY11-7082 significantly decreased TGEV-induced proinflammatory cytokine production, but did not affect virus replication. Phosphorylation of NF-κB subunit p65 and proinflammatory cytokine production were greatly decreased after knockdown of retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs) or its adaptors MAVS and STING, while only slight reduction was observed in cells following silencing of Toll-like receptor adaptors, MyD88 and TRIF. Furthermore, TGEV infection significantly upregulated mRNA expression of RIG-I and MDA5. Taken together, our results indicate that the RLR signaling pathway is involved in TGEV-induced inflammatory responses.


Assuntos
Gastroenterite Suína Transmissível/imunologia , Gastroenterite Suína Transmissível/virologia , NF-kappa B/imunologia , Vírus da Gastroenterite Transmissível/fisiologia , Animais , Citocinas/genética , Citocinas/imunologia , Proteína DEAD-box 58/genética , Proteína DEAD-box 58/imunologia , Gastroenterite Suína Transmissível/genética , Helicase IFIH1 Induzida por Interferon/genética , Helicase IFIH1 Induzida por Interferon/imunologia , NF-kappa B/genética , Fosforilação , Transdução de Sinais , Suínos , Fator de Transcrição AP-1/genética , Fator de Transcrição AP-1/imunologia , Vírus da Gastroenterite Transmissível/genética
6.
Vet Microbiol ; 199: 128-134, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28110779

RESUMO

Coronaviruses tend to efficiently evade innate immune sensing. Alpha-coronaviruses interfere with the type I interferon (IFN) response in various ways, ensuring the limited activation of IFN responses. Transmissible gastroenteritis virus (TGEV), an Alphacoronavirus genera virus, is an important pathogen that mainly infects piglet, but little is known about the activation of the host immune response. We show that TGEV induces a delayed activation of the IFN response in intestinal epithelial cells. Briefly, IFN-ß expression induced by TGEV infection is delayed with respect to that induced by poly(I:C) transfection. In addition, some of the IFN-stimulated genes (ISGs) were up-regulated in the early infection stage without obvious expression of IFN-ß. Moreover, we show that activation of IFN responses induced by poly(I:C) could inhibit viral replication in the early infection stage, but failed in the late infection stage in IPEC-J2 cells. Finally, the activation of IFN responses induced by TGEV infection cannot inhibit viral replication. Taken together, this study provides a preliminary analysis of an interaction between TGEV and IFN-ß responses of intestinal epithelial cells.


Assuntos
Células Epiteliais/imunologia , Células Epiteliais/virologia , Gastroenterite Suína Transmissível/imunologia , Regulação da Expressão Gênica/imunologia , Interações Hospedeiro-Patógeno/imunologia , Vírus da Gastroenterite Transmissível/imunologia , Animais , Linhagem Celular , Gastroenterite Suína Transmissível/virologia , Técnicas de Inativação de Genes , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Fatores Imunológicos/farmacologia , Interferon Tipo I/imunologia , Interferon beta/genética , Poli I-C/farmacologia , Receptores de Interferon/genética , Suínos , Replicação Viral/efeitos dos fármacos
7.
Sci Rep ; 6: 32154, 2016 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-27555521

RESUMO

It has been well characterized that the neonatal Fc receptor (FcRn) transports maternal IgG to a fetus or newborn and protects IgG from degradation. We previously reported that FcRn is expressed in a model of normal porcine intestinal epithelial cells (IPEC-J2). Transmissible gastroenteritis is an acute enteric disease of swine that is caused by transmissible gastroenteritis virus (TGEV). How porcine FcRn (pFcRn) expression is regulated by pathogenic infection remains unknown. Our research shows that IPEC-J2 cells infected with TGEV had up-regulated pFcRn expression. In addition, the NF-κB signaling pathway was activated in IPEC-J2 cells by TGEV infection. Furthermore, treatment of TGEV-infected IPEC-J2 cells with the NF-κB-specific inhibitor BAY 11-7082 resulted in down-regulation of pFcRn expression. Transient transfection of pFcRn promoter luciferase report plasmids with overexpression of NF-κB p65 transcription factor enhanced the activation of the luciferase report plasmids. We identified four NF-κB transcription factor binding sites in the promoter region of this gene using luciferase reporter system, chromatin immunoprecipitation, electromobility shift assay, and supershift analysis. Together, the data provide the first evidence that TGEV infection up-regulates pFcRn expression via activation of NF-κB signaling.


Assuntos
Gastroenterite Suína Transmissível/metabolismo , Antígenos de Histocompatibilidade Classe I/genética , NF-kappa B/metabolismo , Receptores Fc/genética , Animais , Sítios de Ligação , Linhagem Celular , Gastroenterite Suína Transmissível/genética , Gastroenterite Suína Transmissível/imunologia , Regulação da Expressão Gênica , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Interações Hospedeiro-Patógeno , Imunidade Inata , NF-kappa B/antagonistas & inibidores , Nitrilos/farmacologia , Regiões Promotoras Genéticas , Receptores Fc/imunologia , Receptores Fc/metabolismo , Transdução de Sinais , Sulfonas/farmacologia , Suínos , Vírus da Gastroenterite Transmissível/patogenicidade
8.
Virus Res ; 226: 128-141, 2016 12 02.
Artigo em Inglês | MEDLINE | ID: mdl-27212682

RESUMO

Porcine epidemic diarrhea virus (PEDV) and porcine deltacoronavirus (PDCoV) are emerged and reemerging viruses in pigs, and together with transmissible gastroenteritis virus (TGEV), pose significant economic concerns to the swine industry. These viruses infect epithelial cells of the small intestine and cause watery diarrhea, dehydration, and a high mortality in neonatal piglets. Type I interferons (IFN-α/ß) are major antiviral cytokines forming host innate immunity, and in turn, these enteric coronaviruses have evolved to modulate the host innate immune signaling during infection. Accumulating evidence however suggests that IFN induction and signaling in the intestinal epithelial cells differ from other epithelial cells, largely due to distinct features of the gut epithelial mucosal surface and commensal microflora, and it appears that type III interferon (IFN-λ) plays a key role to maintain the antiviral state in the gut. This review describes the recent understanding on the immune evasion strategies of porcine enteric coronaviruses and the role of different types of IFNs for intestinal antiviral innate immunity.


Assuntos
Gastroenterite Suína Transmissível/imunologia , Gastroenterite Suína Transmissível/virologia , Interações Hospedeiro-Patógeno/imunologia , Evasão da Resposta Imune , Imunidade Inata , Vírus da Gastroenterite Transmissível/imunologia , Animais , Gastroenterite Suína Transmissível/metabolismo , Imunomodulação , Interferon Tipo I/metabolismo , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/virologia , Transdução de Sinais , Suínos , Vírus da Gastroenterite Transmissível/genética
9.
Vet Immunol Immunopathol ; 172: 1-8, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-27032496

RESUMO

Transmissible gastroenteritis virus (TGEV) replicates in the small intestine and induces enteritis and watery diarrhea. Establishment of local immunity in the intestine would thus prevent TGEV transmission. CpG DNA has been reported as a promising mucosal adjuvant in some animals. The effects of oral immunization of CpG DNA together with inactivated TGEV (ITGEV) were investigated in this study. Pigs (6 weeks old) were orally immunized with ITGEV plus CpG DNA. The TGEV-specific IgA level in the intestinal tract and the TGEV-specific IgG level in serum significantly increased following immunization with ITGEV plus CpG DNA (P ≤ 0.05). Moreover, populations of IgA-secreting cells, CD3+ T lymphocytes and intraepithelial lymphocytes (IELs), in the intestine increased significantly after immunization with ITGEV plus CpG DNA (P ≤ 0.05). Furthermore, the expression of IL-6, IL-12 and interferon-γ (IFN-γ) in ligated intestine segments increased significantly after injection with ITGEV plus CpG DNA (P ≤ 0.05). Taken together, these data suggest that oral immunization of ITGEV plus CpG DNA elicits a local immune response. Further studies are required to determine whether this immunity provides protection against TGEV in pigs.


Assuntos
Adjuvantes Imunológicos , Suínos , Vírus da Gastroenterite Transmissível/imunologia , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Adjuvantes Imunológicos/administração & dosagem , Administração Oftálmica , Animais , Anticorpos Antivirais/imunologia , Nucleotídeos de Citosina/administração & dosagem , Nucleotídeos de Citosina/imunologia , Sequência Rica em GC/imunologia , Gastroenterite Suína Transmissível/imunologia , Gastroenterite Suína Transmissível/prevenção & controle , Nucleotídeos de Guanina/administração & dosagem , Nucleotídeos de Guanina/imunologia , Imunoglobulina A/imunologia , Imunoglobulina A/metabolismo , Interferon gama/biossíntese , Interleucina-12/biossíntese , Interleucina-6/biossíntese , Intestinos/imunologia , Doenças dos Suínos/imunologia , Doenças dos Suínos/prevenção & controle , Linfócitos T/imunologia , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia , Vacinas Virais/administração & dosagem
10.
Virus Genes ; 52(3): 354-64, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26980672

RESUMO

Porcine transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV) can cause severe diarrhea in newborn piglets and led to significant economic losses. The S proteins are the main structural proteins of PEDV and TGEV capable of inducing neutralizing antibodies in vivo. In this study, a DNA vaccine SL7207 (pVAXD-PS1-TS) co-expressing S proteins of TGEV and PEDV delivered by attenuated Salmonella typhimurium was constructed and its immunogenicity in piglets was investigated. Twenty-day-old piglets were orally immunized with SL7207 (pVAXD-PS1-TS) at a dosage of 1.6 × 10(11) CFU per piglet and then booster immunized with 2.0 × 10(11) CFU after 2 weeks. Humoral immune responses, as reflected by virus neutralizing antibodies and specific IgG and sIgA, and cellular immune responses, as reflected by IFN-γ, IL-4, and lymphocyte proliferation, were evaluated. SL7207 (pVAXD-PS1-TS) simultaneously elicited immune responses against TGEV and PEDV after oral immunization. The immune levels started to increase at 2 weeks after immunization and increased to levels statistically significantly different than controls at 4 weeks post-immunization, peaking at 6 weeks and declined at 8 weeks. The humoral, mucosal, and cellular immune responses induced by SL7207 (pAXD-PS1-TS) were significantly higher than those of the PBS and SL7207 (pVAXD) (p < 0.01). In particular, the levels of IFN-γ and IL-4 were higher than those induced by the single-gene vaccine SL7207 (pVAXD-PS1) (p < 0.05). These results demonstrated that SL7207 (pVAXD-PS1-TS) possess the immunological functions of the two S proteins of TGEV and PEDV, indicating that SL7207 (pVAXD-PS1-TS) is a candidate oral vaccine for TGE and PED.


Assuntos
Vírus da Diarreia Epidêmica Suína/imunologia , Salmonella typhimurium/genética , Salmonella typhimurium/imunologia , Vírus da Gastroenterite Transmissível/imunologia , Vacinas de DNA/imunologia , Proteínas Virais/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Células COS , DNA Recombinante/genética , DNA Recombinante/imunologia , DNA Viral/genética , DNA Viral/imunologia , Gastroenterite Suína Transmissível/imunologia , Gastroenterite Suína Transmissível/prevenção & controle , Genes Virais , Imunoglobulina G/sangue , Interferon gama/sangue , Interleucina-4/sangue , Vírus da Diarreia Epidêmica Suína/genética , Suínos , Vírus da Gastroenterite Transmissível/genética , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Proteínas Virais/genética , Vacinas Virais/administração & dosagem , Vacinas Virais/genética , Vacinas Virais/imunologia
11.
Antiviral Res ; 131: 74-84, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-26988122

RESUMO

Transmissible gastroenteritis (TGE) causes severe diarrhea in suckling piglets, results in enormous economic loss in swine-producing areas of the world. To develop an effective, safe, and convenient vaccine for the prevention of TGE, we have constructed a recombinant Bacillus subtilis strain (B. subtilis CotGSG) displaying the transmissible gastroenteritis virus (TGEV) spike (S) protein and discussed its immune function to intestinal submucosal dendritic cells (DCs). Our results showed that the recombinant B. subtilis had the ability to recruit more DCs to sample B. subtilis CotGSG, migrate to MLNs, and induce immune responses. Immunized piglets with B. subtilis CotGSG could significantly elevate the specific SIgA titers in feces, IgG titers and neutralizing antibodies in serum. Collectively, our results suggested that recombinant B. subtilis CotGSG expressing the TGEV S protein could effectively induce immune responses via DCs, and provided a perspective on potential novel strategy and approach that may be applicable to the development of the next generation of TGEV vaccines.


Assuntos
Anticorpos Antivirais/sangue , Bacillus subtilis/imunologia , Gastroenterite Suína Transmissível/imunologia , Vírus da Gastroenterite Transmissível/química , Proteínas Virais/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Neutralizantes/sangue , Antígenos Virais/imunologia , Bacillus subtilis/genética , Movimento Celular , Células Dendríticas/imunologia , Gastroenterite Suína Transmissível/prevenção & controle , Imunoglobulina A Secretora/análise , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/virologia , Suínos , Vírus da Gastroenterite Transmissível/genética , Vírus da Gastroenterite Transmissível/imunologia , Vacinas Sintéticas/imunologia , Proteínas Virais/administração & dosagem , Proteínas Virais/genética , Vacinas Virais/administração & dosagem , Vacinas Virais/genética
12.
DNA Cell Biol ; 35(6): 301-9, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26982157

RESUMO

It has been well characterized that piglets can absorb colostrum IgG across the intestine to neonatal bloodstream and a certain level of IgG has been found in the mucosal secretions of the porcine intestinal tract. However, little is known about how the maternal IgG transport across the intestinal barrier and how IgG enter the lumen of intestinal tract. In this study, we demonstrated that the porcine neonatal Fc receptor (pFcRn) was expressed in a model of normal porcine intestinal epithelial cells (IPEC-J2) as well as in kidney cells (PK-15), and pFcRn was mainly distributed in the apical side of the polarized IPEC-J2 cells. Analyzing the phylogenetic relatedness of this gene we found that swine and human neonatal Fc receptor (FcRn) amino acid sequence are closer than rodents. We also showed that pFcRn mediated bidirectional IgG transport across polarized IPEC-J2 cells and bound to IgG in a pH-dependent manner. Furthermore, pFcRn-transcytosed viral-specific IgG reduced the transmissible gastroenteritis virus (TGEV) yield from the luminal direction by a 50% tissue culture infective dose (TCID50) assay. Our results indicate that pFcRn-dependent bidirectional IgG transport across the intestinal epithelium plays critical role in the acquisition of humoral immunity in early life and in host defense at mucosal surfaces.


Assuntos
Gastroenterite Suína Transmissível/imunologia , Imunidade nas Mucosas , Imunoglobulina G/imunologia , Mucosa Intestinal/imunologia , Receptores Fc/imunologia , Vírus da Gastroenterite Transmissível/fisiologia , Animais , Linhagem Celular , Células Epiteliais/imunologia , Concentração de Íons de Hidrogênio , Mucosa Intestinal/virologia , Jejuno/imunologia , Modelos Animais , Suínos
13.
Vaccine ; 33(32): 3900-6, 2015 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-26116254

RESUMO

To explore the possibility of developing a vaccine against transmissible gastroenteritis virus (TGEV) infection, a recombinant swinepox virus (rSPV-SA) expressing a TGEV protective antigen has been constructed. Immune responses and protection efficacy of the vaccination vector were assessed in both mice and pig models. An indirect ELISA assay suggested that when mice were vaccinated with rSPV-SA, the level of IgG against TGEV was enhanced dramatically. The cytokine assays were employed and the results indicated that both the Th1-type and Th2-type cytokine levels raised after vaccination with rSPV-SA in mice models. Results from the passive immunity protection test of new born piglets demonstrated that the recombinant live-vector vaccine, rSPV-SA, could 100% protect piglets from the SPV infection, and there was no significant clinical symptom in the rSPV-SA treatment group during this experiment. The data suggest that the novel recombinant swinepox virus is a potential vaccine against TGEV infection.


Assuntos
Portadores de Fármacos , Epitopos/imunologia , Gastroenterite Suína Transmissível/prevenção & controle , Suipoxvirus/genética , Vírus da Gastroenterite Transmissível/imunologia , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/sangue , Citocinas/metabolismo , Epitopos/genética , Gastroenterite Suína Transmissível/imunologia , Vetores Genéticos , Imunização Passiva , Imunoglobulina G/sangue , Camundongos Endogâmicos BALB C , Suínos , Linfócitos T/imunologia , Vírus da Gastroenterite Transmissível/genética , Resultado do Tratamento , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/genética
14.
J Virol ; 89(6): 3332-42, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25589635

RESUMO

UNLABELLED: Porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV) are economically important swine enteropathogenic coronaviruses. These two viruses belong to two distinct species of the Alphacoronavirus genus within Coronaviridae and induce similar clinical signs and pathological lesions in newborn piglets, but they are presumed to be antigenically distinct. In the present study, two-way antigenic cross-reactivity examinations between the prototype PEDV CV777 strain, three distinct U.S. PEDV strains (the original highly virulent PC22A, S indel Iowa106, and S 197del PC177), and two representative U.S. TGEV strains (Miller and Purdue) were conducted by cell culture immunofluorescent (CCIF) and viral neutralization (VN) assays. None of the pig TGEV antisera neutralized PEDV and vice versa. One-way cross-reactions were observed by CCIF between TGEV Miller hyperimmune pig antisera and all PEDV strains. Enzyme-linked immunosorbent assays, immunoblotting using monoclonal antibodies and Escherichia coli-expressed recombinant PEDV and TGEV nucleocapsid (N) proteins, and sequence analysis suggested at least one epitope on the N-terminal region of PEDV/TGEV N protein that contributed to this cross-reactivity. Biologically, PEDV strain CV777 induced greater cell fusion in Vero cells than did U.S. PEDV strains. Consistent with the reported genetic differences, the results of CCIF and VN assays also revealed higher antigenic variation between PEDV CV777 and U.S. strains. IMPORTANCE: Evidence of antigenic cross-reactivity between porcine enteric coronaviruses, PEDV and TGEV, in CCIF assays supports the idea that these two species are evolutionarily related, but they are distinct species defined by VN assays. Identification of PEDV- or TGEV-specific antigenic regions allows the development of more specific immunoassays for each virus. Antigenic and biologic variations between the prototype and current PEDV strains could explain, at least partially, the recurrence of PEDV epidemics. Information on the conserved antigenicity among PEDV strains is important for the development of PEDV vaccines to protect swine from current highly virulent PEDV infections.


Assuntos
Anticorpos Antivirais/imunologia , Infecções por Coronavirus/veterinária , Vírus da Diarreia Epidêmica Suína/imunologia , Doenças dos Suínos/imunologia , Vírus da Gastroenterite Transmissível/imunologia , Sequência de Aminoácidos , Animais , Variação Antigênica , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Reações Cruzadas , Gastroenterite Suína Transmissível/imunologia , Gastroenterite Suína Transmissível/virologia , Dados de Sequência Molecular , Vírus da Diarreia Epidêmica Suína/química , Vírus da Diarreia Epidêmica Suína/classificação , Vírus da Diarreia Epidêmica Suína/genética , Alinhamento de Sequência , Suínos , Doenças dos Suínos/virologia , Vírus da Gastroenterite Transmissível/química , Vírus da Gastroenterite Transmissível/classificação , Vírus da Gastroenterite Transmissível/genética
15.
Res Vet Sci ; 97(3): 623-30, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25307113

RESUMO

Exposure of piglets less than 2 weeks of age to virulent transmissible gastroenteritis virus (TGEV) gives rise to mortality as high as 100%, and adult pigs recovering from its infection often become TGEV carriers. These facts suggest an evasion of the immune system by virulent TGEV. In this study, we showed that a virulent TGEV SHXB strain could infect porcine immature monocyte-derived dendritic cells (Mo-DCs), and down-regulate cell surface markers (SLA-II-DR, CD1a and CD80/86). Moreover, SHXB-infected immature Mo-DCs showed low expression of IL-12 and IFN-γ, and also lost the ability to stimulate T cell proliferation. Finally, SHXB inhibited the activation of nuclear factor kappa B (NF-κB) in these cells. Instead, UV-inactivated SHXB (UV-SHXB) had the opposite effects in immature Mo-DCs. In conclusion, the virulent SHXB could severely impair immature Mo-DCs, which might be involved in the pathogenesis of virulent TGEV in vivo.


Assuntos
Células Dendríticas/imunologia , Gastroenterite Suína Transmissível/imunologia , Regulação Viral da Expressão Gênica/fisiologia , Evasão da Resposta Imune/imunologia , Monócitos/citologia , Vírus da Gastroenterite Transmissível/imunologia , Vírus da Gastroenterite Transmissível/patogenicidade , Análise de Variância , Animais , Western Blotting , Células Dendríticas/virologia , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Interleucina-12/metabolismo , Ativação Linfocitária/imunologia , Monócitos/imunologia , NF-kappa B/metabolismo , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Suínos , Linfócitos T/imunologia , Virulência
16.
Appl Microbiol Biotechnol ; 98(19): 8301-12, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-24993357

RESUMO

The role of muramyl dipeptide (MDP) and tuftsin in oral immune adjustment remains unclear, particularly in a Lactobacillus casei (L. casei) vaccine. To address this, we investigated the effects of different repetitive peptides expressed by L. casei, specifically the MDP and tuftsin fusion protein (MT) repeated 20 and 40 times (20MT and 40MT), in mice also expressing the D antigenic site of the spike (S) protein of transmissible gastroenteritis virus (TGEV) on intestinal and systemic immune responses and confirmed the immunoregulation of these peptides. Treatment of mice with a different vaccine consisting of L. casei expressing MDP and tuftsin stimulated humoral and cellular immune responses. Both 20MT and 40MT induced an increase in IgG and IgA levels against TGEV, as determined using enzyme-linked immunosorbent assay. Increased IgG and IgA resulted in the activation of TGEV-neutralising antibody activity in vitro. In addition, 20MT and 40MT stimulated the differentiation of innate immune cells, including T helper cell subclasses and regulatory T (Treg) cells, which induced robust T helper type 1 and T helper type 17 (Th17) responses and reduced Treg T cell immune responses in the 20MT and 40MT groups, respectively. Notably, treatment of mice with L. casei expressing 20MT and 40MT enhanced the anti-TGEV antibody immune responses of both the humoral and mucosal immune systems. These findings suggest that L. casei expressing MDP and tuftsin possesses substantial immunopotentiating properties, as it can induce humoral and T cell-mediated immune responses upon oral administration, and it may be useful in oral vaccines against TGEV challenge.


Assuntos
Acetilmuramil-Alanil-Isoglutamina/genética , Gastroenterite Suína Transmissível/imunologia , Lactobacillus casei/genética , Células Th1/imunologia , Células Th17/imunologia , Vírus da Gastroenterite Transmissível/imunologia , Tuftsina/genética , Vacinas Virais/imunologia , Acetilmuramil-Alanil-Isoglutamina/administração & dosagem , Acetilmuramil-Alanil-Isoglutamina/imunologia , Administração Oral , Animais , Feminino , Gastroenterite Suína Transmissível/prevenção & controle , Gastroenterite Suína Transmissível/virologia , Lactobacillus casei/imunologia , Masculino , Camundongos , Glicoproteína da Espícula de Coronavírus/administração & dosagem , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Suínos , Vírus da Gastroenterite Transmissível/genética , Tuftsina/administração & dosagem , Tuftsina/imunologia , Regulação para Cima , Vacinas Virais/administração & dosagem , Vacinas Virais/genética
17.
Vet Microbiol ; 171(1-2): 74-86, 2014 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-24742951

RESUMO

Virulent transmissible gastroenteritis virus (TGEV) results in an acute, severe pathology and high mortality in piglets, while attenuated TGEV only causes moderate clinical reactions. Dendritic cells (DCs), through uptake and presentation of antigens to T cells, initiate distinct immune responses to different infections. In this study, an attenuated TGEV (STC3) and a virulent TGEV (SHXB) were used to determine whether porcine DCs play an important role in pathogenetic differences between these two TGEVs. Our results showed that immature and mature monocyte-derived dendritic cells (Mo-DCs) were susceptible to infection with SHXB and STC3. However, only SHXB inhibited Mo-DCs to activate T-cell proliferation by down-regulating the expression of cell-surface markers and the secretion of cytokines in vitro. In addition, after 48 h of SHXB infection, there was the impairment in the ability of porcine intestinal DCs to sample the antigen, to migrate from the villi to the lamina propria and to activate T-cell proliferation in vivo. In contrast, these abilities of intestinal DCs were enhanced in STC3-infected piglets. In conclusion, our results show that SHXB significantly impaired the functions of Mo-DCs and intestinal DCs in vitro and in vivo, while STC3 had the opposite effect. These differences may underlie the pathogenesis of virulent and attenuated TGEV in piglets, and could help us to develop a better strategy to prevent virulent TGEV infection.


Assuntos
Antígenos Virais/imunologia , Células Dendríticas/virologia , Gastroenterite Suína Transmissível/virologia , Vírus da Gastroenterite Transmissível , Animais , Proliferação de Células , Citocinas/metabolismo , Células Dendríticas/citologia , Células Dendríticas/imunologia , Gastroenterite Suína Transmissível/imunologia , Gastroenterite Suína Transmissível/patologia , Regulação da Expressão Gênica/imunologia , Intestinos/patologia , Suínos , Linfócitos T/citologia , Linfócitos T/imunologia , Vírus da Gastroenterite Transmissível/imunologia , Vírus da Gastroenterite Transmissível/patogenicidade
18.
PLoS One ; 8(3): e57468, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23526943

RESUMO

Porcine transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PDEV) can cause severe diarrhea in pigs. Development of effective vaccines against TGEV and PEDV is one of important prevention measures. The spike (S) protein is the surface glycoprotein of TGEV and PEDV, which can induce specific neutralization antibodies and is a candidate antigen for vaccination attempts. In this study, the open reading frames of the TGEV S1 protein and in addition of the S or S1 proteins of PEDV were inserted into the eukaryotic expression vector, pIRES, resulting in recombinant plasmids, pIRES-(TGEV-S1-PEDV-S1) and pIRES-(TGEV-S1-PEDV-S). Subsequently, 6-8 weeks old Kunming mice were inoculated with both DNA plasmids. Lymphocyte proliferation assay, virus neutralization assay, IFN-γ assay and CTL activity assay were performed. TGEV/PEDV specific antibody responses as well as kinetic changes of T lymphocyte subgroups of the immunized mice were analyzed. The results showed that the recombinant DNA plasmids increased the proliferation of T lymphocytes and the number of CD4+ and CD8+ T lymphocyte subgroups. In addition, the DNA vaccines induced a high level of IFN-γ in the immunized mice. The specific CTL activity in the pIRES-(TGEV-S1-PEDV-S) group became significant at 42 days post-immunization. At 35 days post-immunization, the recombinant DNA plasmids bearing full-length S genes of TGEV and PEDV stimulated higher levels of specific antibodies and neutralizing antibodies in immunized mice.


Assuntos
Vírus da Diarreia Epidêmica Suína/genética , Vírus da Diarreia Epidêmica Suína/imunologia , Vírus da Gastroenterite Transmissível/genética , Vírus da Gastroenterite Transmissível/imunologia , Vacinas de DNA/genética , Vacinas de DNA/imunologia , Proteínas Virais/genética , Proteínas Virais/imunologia , Vacinas Virais/genética , Vacinas Virais/imunologia , Animais , Anticorpos Antivirais/sangue , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/veterinária , DNA Recombinante/genética , DNA Recombinante/imunologia , DNA Viral/genética , DNA Viral/imunologia , Gastroenterite Suína Transmissível/imunologia , Gastroenterite Suína Transmissível/prevenção & controle , Genes Virais , Interferon gama/sangue , Interleucina-4/sangue , Camundongos , Plasmídeos/genética , Plasmídeos/imunologia , Suínos , Doenças dos Suínos/imunologia , Doenças dos Suínos/prevenção & controle , Linfócitos T/imunologia
19.
Comp Immunol Microbiol Infect Dis ; 34(4): 369-80, 2011 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-21719106

RESUMO

The enhanced effect of cytokine combinations has been assessed empirically, based on their immunobiological mechanisms. However, far less is known of the enhanced protection of practical cytokine combinations against viral infection in the livestock industry, due to cost and production issues associated with mass administration. This study demonstrates the enhanced protection of oral co-administration of swine interferon-α (swIFN-α) and interleukin-18 (swIL-18) against infection with transmissible gastroenteritis virus (TGEV) in piglets using attenuated Salmonella enterica serovar Typhimurium as carrier of cytokine proteins. A single oral co-administration of S. enterica serovar Typhimurium expressing swIFN-α and swIL-18 induced enhanced alleviation of the severity of diarrhea caused by TGEV infection, compared to piglets administered S. enterica serovar Typhimurium expressing swIFN-α or swIL-18 alone. This enhancement was further observed by the reduction of TGEV shedding and replication, and the expression of IFN-stimulated gene products in the intestinal tract. The results suggest that the combined administration of the swIFN-α and swIL-18 cytokines using attenuated S. enterica serovar Typhimurium as an oral carrier provides enhanced protection against intestinal tract infection with TGEV.


Assuntos
Gastroenterite Suína Transmissível/prevenção & controle , Imunidade Ativa , Interferon-alfa/imunologia , Interleucina-18/imunologia , Intestinos/imunologia , Salmonella typhimurium/genética , Vírus da Gastroenterite Transmissível/efeitos dos fármacos , Vacinação , Vacinas Atenuadas/administração & dosagem , Administração Oral , Animais , Feminino , Gastroenterite Suína Transmissível/imunologia , Gastroenterite Suína Transmissível/metabolismo , Gastroenterite Suína Transmissível/virologia , Interferon-alfa/genética , Interleucina-18/genética , Intestinos/virologia , Gado , Camundongos , Plasmídeos , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Salmonella typhimurium/química , Salmonella typhimurium/imunologia , Suínos , Transfecção , Vírus da Gastroenterite Transmissível/crescimento & desenvolvimento , Vírus da Gastroenterite Transmissível/imunologia , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Carga Viral/efeitos dos fármacos , Carga Viral/imunologia , Replicação Viral/efeitos dos fármacos , Replicação Viral/imunologia
20.
Hybridoma (Larchmt) ; 29(4): 345-50, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20715993

RESUMO

Transmissible gastroenteritis virus (TGEV) is a member of coronaviruses. The viral spike (S) protein mediates the interaction between TGEV and its susceptible cells. Here, we expressed a truncated gene encoding the N terminal half of TGEV S gene (designated S1 gene) in a prokaryotic system. The resulting S1 protein was used to immunize BALB/c mice followed by the generation of a monoclonal antibody (MAb). A generated MAb (7F9) was identified by ELISA and the chromosome number of the hybridoma cell was analyzed. The immunoreactivity of the MAb to TGEV S protein was confirmed by Western blot analysis. Moreover, immunofluorescence assays showed that the MAb is able to detect cell infection by TGEV. The MAb achieved in this study can be used as a specific diagnostic reagent for detecting TGEV S protein.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/isolamento & purificação , Gastroenterite Suína Transmissível/imunologia , Glicoproteínas de Membrana/imunologia , Proteínas Recombinantes/imunologia , Vírus da Gastroenterite Transmissível/imunologia , Proteínas do Envelope Viral/imunologia , Animais , Especificidade de Anticorpos , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Imunofluorescência , Gastroenterite Suína Transmissível/genética , Gastroenterite Suína Transmissível/metabolismo , Hibridomas , Immunoblotting , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Glicoproteína da Espícula de Coronavírus , Suínos , Testículo/citologia , Testículo/metabolismo , Vírus da Gastroenterite Transmissível/genética
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