RESUMO
Introduction: The critical early stages of infection and innate immune responses to porcine epidemic diarrhea virus (PEDV) at the intestinal epithelium remain underexplored due to the limitations of traditional cell culture and animal models. This study aims to establish a porcine enteroid culture model to investigate potential differences in susceptibility to infection across segments of the porcine small intestine (duodenum, jejunum, and ileum). Methods: Intestinal crypt cells from nursery pigs were cultured in Matrigel to differentiate into porcine enteroid monolayer cultures (PEMCs). Following characterization, PEMCs were enzymatically dissociated and subcultured on transwell inserts (PETCs) for apical surface exposure and infection studies. Characterization of region-specific PEMCs and PETCs included assessment of morphology, proliferation, viability, and cellular phenotyping via immunohistochemistry/immunocytochemistry and gene expression analysis. Subsequently, PETCs were inoculated with 105 TCID50 (50% tissue culture infectious dose)/mL of a high pathogenic PEDV non-S INDEL strain and incubated for 24 h. Infection outcomes were assessed by cytopathic effect, PEDV N protein expression (immunofluorescence assay, IFA), and PEDV N-gene detection (quantitative reverse transcription polymerase chain reaction, RT-qPCR). Results: No significant morphological and phenotypical differences were observed among PEMCs and PETCs across intestinal regions, resembling the porcine intestinal epithelium. Although PETCs established from different segments of the small intestine were susceptible to PEDV infection, jejunum-derived PETCs exhibited higher PEDV replication, confirmed by IFA and RT-qPCR. Discussion: This segment-specific enteroid culture model provides a reliable platform for virological studies, offering a controlled environment that overcomes the limitations of in vivo and traditional cell culture methods. Standardizing culture conditions and characterizing the model are essential for advancing enteroid-based infection models.
Assuntos
Infecções por Coronavirus , Intestino Delgado , Vírus da Diarreia Epidêmica Suína , Animais , Vírus da Diarreia Epidêmica Suína/fisiologia , Suínos , Intestino Delgado/imunologia , Intestino Delgado/virologia , Infecções por Coronavirus/virologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/imunologia , Laminina , Combinação de Medicamentos , Doenças dos Suínos/virologia , Doenças dos Suínos/imunologia , Suscetibilidade a Doenças , Colágeno/metabolismo , Organoides/virologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/virologia , Proteoglicanas , Células CultivadasRESUMO
The objective of this field trial was to determine the efficacy of a recombinant toxoid vaccine against Shiga toxin 2e (Stx2e) in piglets suffering from edema disease (ED). Three farms with confirmed ED cases were selected for the field trials. On each farm, a total of 40 4-day-old pigs were randomly allocated to either the vaccinated or unvaccinated group, with 20 pigs per group (10 males and 10 females). A 1.0-mL dose of the recombinant toxoid vaccine was administered intramuscularly to pigs in the vaccinated groups in accordance with the manufacturer's recommendations at 4 d of age. A single 2.0-mL dose of phosphate-buffered saline was administered to unvaccinated pigs at the same age. Clinical signs of ED were observed in 12 piglets in the unvaccinated groups and 7 unvaccinated pigs died as a result of ED out of the total number of piglets on Farms A, B, and C. Vaccination had a positive effect on pig growth performance compared to that of unvaccinated pigs on 2 of the 3 farms. Seroconversion of neutralizing antibodies against Stx2e occurred in 70% of piglets in the vaccinated group on Farm A, 75% of vaccinated piglets on Farm B, and 55% of vaccinated piglets on Farm C, when detectable antibodies were measured at 17 d post-vaccination (dpv). Detectable antibodies were measured in 85% of vaccinated piglets on Farms A and B and in 90% of these piglets on Farm C at 37 dpv. These field trials determined that the ED recombinant toxoid vaccine successfully reduced clinical signs and mortality, improved average daily weight gain, and resulted in the production of neutralizing antibodies against Stx2e in pigs.
L'objectif de cette étude sur le terrain était de déterminer l'efficacité d'un vaccin à base d'anatoxine recombinante contre la toxine Shiga 2e (Stx2e) chez les porcelets souffrant de la maladie de l'oedème (ED). Trois fermes ayant des cas confirmés de ED ont été sélectionnées pour les études sur le terrain. Dans chaque ferme, un total de 40 porcs âgés de 4 jours ont été répartis au hasard dans le groupe vacciné ou non vacciné, avec 20 porcs par groupe (10 mâles et 10 femelles). Une dose de 1,0 ml du vaccin à base d'anatoxine recombinante a été administrée par voie intramusculaire aux porcs des groupes vaccinés conformément aux recommandations du fabricant à l'âge de 4 jours. Une dose unique de 2,0 ml de solution saline tamponnée au phosphate a été administrée aux porcs non vaccinés au même âge. Des signes cliniques de ED ont été observés chez 12 porcelets des groupes non vaccinés et 7 porcs non vaccinés sont morts des suites d'une maladie de l'oedème sur le nombre total de porcelets des fermes A, B et C. La vaccination a eu un effet positif sur les performances de croissance des porcs par rapport à celles des porcs non vaccinés dans 2 des 3 fermes. La séroconversion des anticorps neutralisants contre Stx2e s'est produite chez 70 % des porcelets du groupe vacciné de la ferme A, 75 % des porcelets vaccinés de la ferme B et 55 % des porcelets vaccinés de la ferme C, lorsque des anticorps détectables ont été mesurés 17 jours après la vaccination (dpv). Des anticorps détectables ont été mesurés chez 85 % des porcelets vaccinés des fermes A et B et chez 90 % de ces porcelets de la ferme C à 37 dpv. Ces études sur le terrain ont déterminé que le vaccin toxoïde recombinant ED réduisait avec succès les signes cliniques et la mortalité, améliorait le gain de poids quotidien moyen et entraînait la production d'anticorps neutralisants contre Stx2e chez les porcs.(Traduit par Docteur Serge Messier).
Assuntos
Vacinas Sintéticas , Animais , Suínos , Feminino , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/administração & dosagem , Masculino , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/imunologia , Edematose Suína/prevenção & controle , Edematose Suína/imunologia , Infecções por Escherichia coli/veterinária , Infecções por Escherichia coli/prevenção & controle , Infecções por Escherichia coli/imunologia , Toxina Shiga II/imunologia , Vacinas contra Escherichia coli/imunologia , Vacinas contra Escherichia coli/administração & dosagem , Toxoides/imunologia , Vacinas Bacterianas/imunologia , Vacinas Bacterianas/administração & dosagemRESUMO
Foot-and-mouth disease (FMD) is one of the most infectious viral transboundary diseases of livestock, which causes devastating global economic losses. Different enzyme-linked immunosorbent assays (ELISAs) are used for sero-surveillance of the foot-and-mouth disease virus (FMDV). However, more sensitive, accurate, and convenient ELISAs are still required to detect antibodies against FMDV serotypes. The primary goal of this study was to establish serotype-specific monoclonal antibody (mAb)-based blocking ELISAs (mAb-bELISAs) that would provide better performance characteristics or be equivalent in performance characteristics compared with a conventional polyclonal antibody (pAb)-based competitive ELISA (pAb-cELISA). Four mAb-bELISAs were developed using FMDV serotype-specific mAbs for the detection of anti-FMDV/O/A/Asia1/SAT2 antibodies. Using a 50% cut-off, all four mAb-bELISAs exhibited species-independent 99.74%, 98.01%, 96.59%, and 98.55% diagnostic specificity (DSp) and 98.93%, 98.25%, 100%, and 87.50% diagnostic sensitivity (DSe) for FMDV serotypes O, A, Asia1, and SAT2, respectively. In addition, a 100% DSe of serotypes O- and SAT2-specific mAb-bELISAs was observed for porcine sera when the cut-off was 30%. All mAb-bELISAs developed in this study displayed high repeatability/reproducibility without cross-reactivity. Finally, the diagnostic performance of mAb-bELISAs was found to be better than or equivalent to compared with pAb-cELISAs, suggesting that mAb-bELISAs can be used to replace existing pAb-ELISAs for the detection of antibodies against these four FMDV serotypes.
Assuntos
Anticorpos Monoclonais , Anticorpos Antivirais , Ensaio de Imunoadsorção Enzimática , Vírus da Febre Aftosa , Febre Aftosa , Sensibilidade e Especificidade , Sorogrupo , Ensaio de Imunoadsorção Enzimática/métodos , Vírus da Febre Aftosa/imunologia , Vírus da Febre Aftosa/classificação , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Animais , Anticorpos Monoclonais/imunologia , Febre Aftosa/diagnóstico , Febre Aftosa/imunologia , Febre Aftosa/virologia , Suínos , Bovinos , Doenças dos Suínos/diagnóstico , Doenças dos Suínos/virologia , Doenças dos Suínos/imunologia , Camundongos , Reprodutibilidade dos TestesRESUMO
Streptococcus suis (S. suis) is one of the most important porcine pathogens, causing severe pathologies such as meningitis or polyarthritis. It is also a very successful colonizer of mucosal surfaces. The IgM-degrading enzyme of S. suis (IdeSsuis) specifically cleaves porcine IgM, which results in complement evasion. On the basis of our previous finding that IdeSsuis also cleaves the IgM B cell receptor in vitro, we verified IgM B cell receptor cleavage ex vivo in whole regional lymph nodes and investigated the working hypothesis that this IgM B cell receptor cleavage results in a long-lasting impaired B cell function. The number of IgM-secreting cells was determined via ELISpot analysis after porcine peripheral blood mononuclear cells had initially been treated with different recombinant S. suis proteins and subsequently stimulated with interleukin-2 and the toll-like receptor 7/8 ligand R848. Compared with treatment with medium or recombinant muramidase-released protein, treatment with rIdeSsuis but also with a cleavage-deficient variant led to a reduction in the number of IgM-secreting cells as well as the level of secreted IgM. Flow cytometry analysis confirmed that the IgM B cell receptor was cleaved only by rIdeSsuis, and the receptor recovered to pretreatment levels on day 2 after treatment. Flow cytometry analysis of B and T cells incubated with fluorescein-labelled recombinant proteins revealed that different rIdeSsuis variants bind specifically to B cells, most prominently the cleavage-deficient variant. Our results indicate that in vitro interference of rIdeSsuis with the IgM B cell receptor results in long-lasting impaired IgM secretion by B cells after toll-like receptor activation. Further studies are warranted to prove that the modulation of B cell function by IdeSsuis could play a role in vivo.
Assuntos
Linfócitos B , Imunoglobulina M , Streptococcus suis , Animais , Streptococcus suis/imunologia , Imunoglobulina M/imunologia , Imunoglobulina M/metabolismo , Linfócitos B/imunologia , Suínos , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Doenças dos Suínos/microbiologia , Doenças dos Suínos/imunologia , Infecções Estreptocócicas/veterinária , Infecções Estreptocócicas/imunologia , Infecções Estreptocócicas/microbiologiaRESUMO
Lawsonia intracellularis is the etiological agent of proliferative enteropathy (PE) in pigs, horses and wide range of mammals. Little is known about the role of innate immune response during L. intracellularis infection. In this study, we investigated the nuclear factor-κB (NF-κB)-regulated immune response against infection of a clinical strain Dkp23 and a live-attenuated Enterisol vaccine strain in PK-15 cells. We found that expression of NF-κB target genes TNF-α, IFN-γ, IL-6 and IL-8 were modulated during the course of infection. At 5 dpi, there was a significant increase in p65 NF-κB activation, including protein nuclear translocation and phosphorylation, synchronous with the induction of IL-6, IFN-γ and IL-8 expression in L. intracellularis infected cells, especially for Enterisol vaccine strain-infected cells. This result suggests that NF-κB signalling level is induced when L. intracellularis bacterial load peaks at 5 dpi. The induction of pro-inflammatory cytokines expression is consistent with the decreased viability of L. intracellularis-infected cells especially that of the vaccine strain. There were no significant changes in NF-κB signalling between vaccine and Dkp23 infection in PK-15 cells, except for moderate levels of differences in NF-κB target genes expression which might be a reflection of differences in intracellular bacterial load. Overall, the data presented here indicate a correlation between the induction of NF-κB signalling and the L. intracellularis bacterial load in PK-15 cells.
Assuntos
Infecções por Desulfovibrionaceae , Lawsonia (Bactéria) , NF-kappa B , Transdução de Sinais , Animais , Lawsonia (Bactéria)/imunologia , NF-kappa B/metabolismo , Suínos , Linhagem Celular , Infecções por Desulfovibrionaceae/microbiologia , Infecções por Desulfovibrionaceae/veterinária , Infecções por Desulfovibrionaceae/imunologia , Citocinas/metabolismo , Doenças dos Suínos/microbiologia , Doenças dos Suínos/imunologia , Doenças dos Suínos/metabolismo , Regulação da Expressão GênicaRESUMO
INTRODUCTION: Pigs without intestinal receptors for F4 fimbriae are congenitally resistant to F4 fimbriae-bearing enterotoxigenic Escherichia coli (ETEC F4). In general, 50 % and 100 % of piglets born to resistant (RR) sows crossed with hetero- or homozygous susceptible (SR, SS) boars, respectively, are susceptible but do not receive colostral antibodies against F4 fimbriae unless the sows have been vaccinated. The question arises as to whether resistant sows produce protective amounts of F4 antifimbrial antibodies after vaccination. The serum and colostrum antibody titres of 12 resistant and 12 susceptible vaccinated gilts were compared. The effect of the receptor status of the dam and sire on the preweaning performance of 5027 piglets was evaluated using Agroscope's recordings. The sows of the experimental herd, where ETEC F4 was circulating, were vaccinated against ETEC twice during the first pregnancy and once during each following pregnancy. The log2 transformed F4 antibody titres in the serum obtained after the second vaccine injection as well as in the colostrum of the 12 resistant animals were lower than the titres of the susceptible animals (serum: F4ab 11,19 ± 1,44 vs. 12,18 ± 1,33, P = 0,096; F4ac 10,03 ± 1,58 vs. 11,59 ± 1,43, P = 0,019; colostrum: F4ab 12,20 ± 2,41 vs. 14,02 ± 1,31, P = 0,033; F4ac 10,93 ± 2,46 vs. 13,03 ± 5,21, P = 0,006). The heat labile enterotoxin (LT) antibody titres after vaccination did not differ between susceptible and resistant animals (p > 0,10). Preweaning mortality in the offspring of RR sows × SS boars was slightly lower than in the offspring of SS sows × RR boars (P = 0,04), suggesting that the disease risk of susceptible piglets born to vaccinated resistant sows was not increased, even though they received colostrum with a slightly reduced content of antibody against F4 fimbriae.
INTRODUCTION: Les porcs dépourvus de récepteurs intestinaux pour les fimbriae F4 sont congénitalement résistants aux Escherichia coli entérotoxinogènes porteurs de fimbriae F4 (ETEC F4). En général, 50 % et 100 % des porcelets nés de truies résistantes (RR) croisées avec des verrats hétéro- ou homozygotes sensibles (SR, SS), respectivement, sont sensibles mais ne reçoivent pas d'anticorps colostraux contre les fimbriae F4, à moins que les truies n'aient été vaccinées. La question se pose de savoir si les truies résistantes produisent des quantités protectrices d'anticorps antifimbriae F4 après la vaccination. Les titres d'anticorps dans le sérum et le colostrum de 12 truies reproductrices vaccinées résistantes et de 12 truies reproductrices vaccinées sensibles ont été comparés et l'effet du statut récepteur de la mère et du père sur les performances avant sevrage de 5027 porcelets a été évalué. Les truies du troupeau expérimental, où circulait ETEC F4, ont été vaccinées deux fois au cours de la première gestation et une fois au cours de chaque gestation suivante contre ETEC. Les titres d'anticorps F4 transformés en log2 dans le sérum obtenu après la deuxième injection de vaccin ainsi que dans le colostrum des 12 animaux résistants étaient inférieurs aux titres des animaux sensibles (sérum : F4ab 11,19 ± 1,44 vs. 12,18 ± 1,33, P = 0,096 ; F4ac 10,03 ± 1,58 vs. 11,59 ± 1,43, P = 0,019 ; colostrum : F4ab 12,20 ± 2,41 vs. 14,02 ± 1,31, P = 0,033 ; F4ac 10,93 ± 2,46 vs. 13,03 ± 5,21, P = 0,006). Les titres d'anticorps contre l'entérotoxine thermolabile (LT) après la vaccination ne différaient pas entre les animaux sensibles et résistants (p > 0,10). La mortalité avant sevrage dans la progéniture des truies RR × verrats SS était légèrement inférieure à celle de la progéniture des truies SS × verrats RR (P = 0,04), ce qui suggère que le risque de maladie des porcelets sensibles nés de truies résistantes vaccinées n'a pas été augmenté, même s'ils ont reçu du colostrum avec une teneur légèrement réduite en anticorps contre les fimbriae F4.
Assuntos
Escherichia coli Enterotoxigênica , Infecções por Escherichia coli , Vacinas contra Escherichia coli , Fímbrias Bacterianas , Doenças dos Suínos , Animais , Suínos , Infecções por Escherichia coli/veterinária , Infecções por Escherichia coli/prevenção & controle , Infecções por Escherichia coli/imunologia , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/imunologia , Doenças dos Suínos/microbiologia , Feminino , Vacinas contra Escherichia coli/imunologia , Vacinas contra Escherichia coli/administração & dosagem , Escherichia coli Enterotoxigênica/imunologia , Fímbrias Bacterianas/imunologia , Fímbrias Bacterianas/genética , Gravidez , Colostro/imunologia , Anticorpos Antibacterianos/sangue , DesmameRESUMO
INTRODUCTION: Clostridioides difficile is the main cause of antibiotic-associated diarrhea in humans and is a major enteropathogen in several animal species. In newborn piglets, colonic lesions caused by C. difficile A and B toxins (TcdA and TcdB, respectively) cause diarrhea and significant production losses. OBJECTIVE: The present study aimed to develop two recombinant vaccines from immunogenic C-terminal fragments of TcdA and TcdB and evaluate the immune response in rabbits and in breeding sows. Two vaccines were produced: bivalent (rAB), consisting of recombinant fragments of TcdA and TcdB, and chimeric (rQAB), corresponding to the synthesis of the same fragments in a single protein. Groups of rabbits were inoculated with 10 or 50 µg of proteins adjuvanted with aluminum or 0.85 % sterile saline in a final volume of 1 mL/dose. Anti-TcdA and anti-TcdB IgG antibodies were detected in rabbits and sows immunized with both rAB and rQAB vaccines by ELISA. The vaccinated sows were inoculated intramuscularly with 20 µg/dose using a prime-boost approach. RESULTS: Different antibody titers (p ≤ 0.05) were observed among the vaccinated groups of sows (rAB and rQAB) and control. Additionally, newborn piglets from vaccinated sows were also positive for anti-TcdA and anti-TcdB IgGs, in contrast to control piglets (p ≤ 0.05). Immunization of sows with the rQAB vaccine conferred higher anti-TcdA and anti-TcdB responses in piglets, suggesting the superiority of this compound over rAB. CONCLUSION: The synthesized recombinant proteins were capable of inducing antibody titers against C. difficile toxins A and B in sows, and were passively transferred to piglets through colostrum.
Assuntos
Animais Recém-Nascidos , Anticorpos Antibacterianos , Toxinas Bacterianas , Vacinas Bacterianas , Clostridioides difficile , Infecções por Clostridium , Doenças dos Suínos , Vacinas Sintéticas , Animais , Feminino , Suínos , Coelhos , Infecções por Clostridium/prevenção & controle , Infecções por Clostridium/veterinária , Infecções por Clostridium/imunologia , Vacinas Bacterianas/imunologia , Vacinas Bacterianas/administração & dosagem , Vacinas Bacterianas/genética , Gravidez , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/administração & dosagem , Clostridioides difficile/imunologia , Clostridioides difficile/genética , Anticorpos Antibacterianos/sangue , Toxinas Bacterianas/imunologia , Toxinas Bacterianas/genética , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/imunologia , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/genética , Enterotoxinas/imunologia , Enterotoxinas/genéticaRESUMO
Lawsonia intracellularis is the causative agent of ileitis in swine that manifests as slower weight gain, mild or hemorrhagic diarrhea and/or death in severe cases. As an economically important swine pathogen, development of effective vaccines is important to the swine industry. In developing a subunit vaccine with three recombinant antigens - FliC, GroEL and YopN - we wanted to identify a formulation that would produce robust immune responses that reduce disease parameters associated with Lawsonia intracellularis infection. We formulated these three antigens with four adjuvants: Montanide ISA 660 VG, Montanide Gel 02 PR, Montanide IMS 1313 VG NST, and Montanide ISA 61 VG in an immunogenicity study. Groups vaccinated with formulations including Montanide ISA 660 VG or Montanide ISA 61 VG had significantly more robust immune responses than groups vaccinated with formulations including Montanide Gel 02 PR or Montanide IMS 1313 VG NST. In the challenge study, animals vaccinated with these antigens and Montanide ISA 61 VG had reduced lesion scores, reduced lesion lengths, and increased average daily gain, but no reduction in shedding relative to the control animals. This work shows that this vaccine formulation should be considered for future study in a field and performance trial.
Assuntos
Infecções por Desulfovibrionaceae , Lawsonia (Bactéria) , Doenças dos Suínos , Vacinas de Subunidades Antigênicas , Animais , Suínos , Lawsonia (Bactéria)/imunologia , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/imunologia , Doenças dos Suínos/microbiologia , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/administração & dosagem , Infecções por Desulfovibrionaceae/prevenção & controle , Infecções por Desulfovibrionaceae/imunologia , Infecções por Desulfovibrionaceae/veterinária , Vacinação/métodos , Vacinas Bacterianas/imunologia , Vacinas Bacterianas/administração & dosagem , Adjuvantes Imunológicos/administração & dosagem , Adjuvantes Imunológicos/farmacologia , Emulsões , Derrame de BactériasRESUMO
Coronaviruses (CoVs) are important pathogens for humans and other vertebrates, causing severe respiratory and intestinal infections that have become a threat to public health because of the potential for interspecies transmission between animals and humans. Therefore, the development of safe, effective vaccines remains a top priority for the control of CoV infection. The unique immunological characteristics of vaccines featuring messenger RNA (mRNA) present an advantageous tool for coronavirus vaccine development. Here, we designed two lipid nanoparticle (LNP)-encapsulated mRNA (mRNA-LNP) vaccines: one encoding full-length spike (S) protein and the other encoding the spike ectodomain (Se) from porcine deltacoronavirus (PDCoV). Fourteen days after primary immunization, both mRNA vaccines induced high levels of immunoglobulin G and neutralizing antibodies in mice, with the S vaccine showing better performance than the Se vaccine. Passive immune protection of the S mRNA vaccine in suckling piglets was confirmed by the induction of robust PDCoV-specific humoral and cellular immune responses. The S mRNA vaccine also showed better protective effects than the inactivated vaccine. Our results suggest that the novel PDCoV-S mRNA-LNP vaccine may have the potential to combat PDCoV infection. IMPORTANCE: As an emerging porcine enteropathogenic coronavirus, porcine deltacoronavirus (PDCoV) has the potential for cross-species transmission, attracting extensive attention. Messenger RNA (mRNA) vaccines are a promising option for combating emerging and re-emerging infectious diseases, as evidenced by the demonstrated efficacy of the COVID-19 mRNA vaccine. Here, we first demonstrated that PDCoV-S mRNA-lipid nanoparticle (LNP) vaccines could induce potent humoral and cellular immune responses in mice. An evaluation of passive immune protection of S mRNA vaccines in suckling piglets confirmed that the protective effect of mRNA vaccine was better than that of inactivated vaccine. This study suggests that the PDCoV-S mRNA-LNP vaccine may serve as a potential and novel vaccine candidate for combating PDCoV infection.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , Infecções por Coronavirus , Glicoproteína da Espícula de Coronavírus , Doenças dos Suínos , Vacinas Virais , Animais , Suínos , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/genética , Camundongos , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/virologia , Doenças dos Suínos/imunologia , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/sangue , Vacinas Virais/imunologia , Vacinas Virais/administração & dosagem , Vacinas de mRNA , Deltacoronavirus/imunologia , Deltacoronavirus/genética , Nanopartículas , RNA Mensageiro/genética , RNA Mensageiro/imunologia , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Camundongos Endogâmicos BALB C , Feminino , Imunidade Humoral , LipossomosRESUMO
Senecavirus A (SVA) is an emerging virus that poses a threat to swine herds worldwide. To date, the role of tripartite motif 5 (TRIM5) in the replication of viruses has not been evaluated. Here, TRIM5 was reported to inhibit SVA replication by promoting the type I interferon (IFN) antiviral response mediated by retinoic acid-inducible gene I (RIG-I). TRIM5 expression was significantly upregulated in SVA-infected cells, and TRIM5 overexpression inhibited viral replication and promoted IFN-α, IFN-ß, interleukin-1beta (IL-1ß), IL-6, and IL-18 expression. Conversely, interfering with the expression of TRIM5 had the opposite effect. Viral adsorption and entry assays showed that TRIM5 did not affect the adsorption of SVA but inhibited its entry. In addition, TRIM5 promoted the expression of RIG-I and RIG-I-mediated IFNs and proinflammatory cytokines, and this effect was also proven by inhibiting the expression of TRIM5. These findings expand the scope of knowledge on host factors inhibiting the replication of SVA and indicate that targeting TRIM5 may aid in the development of new agents against SVA.
Assuntos
Interferon Tipo I , Picornaviridae , Replicação Viral , Animais , Interferon Tipo I/metabolismo , Suínos , Picornaviridae/fisiologia , Picornaviridae/imunologia , Proteínas com Motivo Tripartido/metabolismo , Proteínas com Motivo Tripartido/genética , Doenças dos Suínos/virologia , Doenças dos Suínos/imunologiaRESUMO
In 2009, a novel swine-origin H1N1 virus emerged, causing a pandemic. The virus, known as H1N1pdm09, quickly displaced the circulating H1 lineage and became the dominant seasonal influenza A virus subtype infecting humans. Human-to-swine spillovers of the H1N1pdm09 have occurred frequently, and each occurrence has led to sustained transmission of the human-origin H1N1pdm09 within swine populations. In the present study, we developed a lipid nanoparticle-based DNA vaccine (LNP-DNA) containing the hemagglutinin gene of a swine-origin H1N1pdm09. In pigs, this LNP-DNA vaccine induced a robust antibody response after a single intramuscular immunization and protected the pigs against challenge infection with the homologous swine-origin H1N1pdm09 virus. In a mouse model, the LNP-DNA vaccine induced antibody and T-cell responses and protected mice against lethal challenge with a mouse-adapted human-origin H1N1pdm09 virus. These findings demonstrate the potential of the LNP-DNA vaccine to protect against both swine- and human-origin H1N1pdm09 viruses. IMPORTANCE: Swine influenza A virus (IAV) is widespread and causes significant economic losses to the swine industry. Moreover, bidirectional transmission of IAV between swine and humans commonly occurs. Once introduced into the swine population, human-origin IAV often reassorts with endemic swine IAV, resulting in reassortant viruses. Thus, it is imperative to develop a vaccine that is not only effective against IAV strains endemic in swine but also capable of preventing the spillover of human-origin IAV. In this study, we developed a lipid nanoparticle-encapsulated DNA plasmid vaccine (LNP-DNA) that demonstrates efficacy against both swine- and human-origin H1N1 viruses. The LNP-DNA vaccines are non-infectious and non-viable, meeting the criteria to serve as a vaccine platform for rapidly updating vaccines. Collectively, this LNP-DNA vaccine approach holds great potential for alleviating the impact of IAV on the swine industry and preventing the emergence of reassortant IAV strains.
Assuntos
Anticorpos Antivirais , Vírus da Influenza A Subtipo H1N1 , Vacinas contra Influenza , Influenza Humana , Nanopartículas , Infecções por Orthomyxoviridae , Doenças dos Suínos , Vacinas de DNA , Animais , Vírus da Influenza A Subtipo H1N1/imunologia , Vírus da Influenza A Subtipo H1N1/genética , Vacinas de DNA/imunologia , Vacinas de DNA/administração & dosagem , Suínos , Vacinas contra Influenza/imunologia , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/genética , Infecções por Orthomyxoviridae/prevenção & controle , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/veterinária , Nanopartículas/administração & dosagem , Humanos , Camundongos , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/virologia , Doenças dos Suínos/imunologia , Anticorpos Antivirais/sangue , Influenza Humana/prevenção & controle , Influenza Humana/imunologia , Influenza Humana/virologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Feminino , Camundongos Endogâmicos BALB C , Lipossomos/administração & dosagemRESUMO
Porcine epidemic diarrhea virus (PEDV) is a serious disease that poses a significant threat to the pig industry. This study focused on analyzing the Spike protein of PEDV, which harbors crucial antigenic determinants, in identifying dominant epitopes. Immunoinformatics tools were used to screen for B-cell, CD4+ and CD8+ predominance epitopes. These epitopes were then connected to the N-terminal of ferritin to form a self-assembled nanoparticle vaccine. Various physical and chemical properties of the candidate vaccine were analyzed, including secondary structure prediction, tertiary structure modeling, molecular docking, immune response simulation and computer cloning. The results demonstrated that the candidate vaccine was antigenic, soluble, stable, non-allergic, and formed a stable complex with the target receptor TLR-3. Immune simulation analysis showed that the candidate vaccine effectively stimulated both cellular and humoral reactions, leading to increased related cytokines production. Furthermore, efficient and stable expression of the candidate vaccine was achieved through reverse translation in the Escherichia coli K12 expression system following codon optimization and in silico cloning. The developed nanoparticle candidate vaccine in this study holds promise as an effective PEDV vaccine candidate, offering a new approach for the research, development and improvement of vaccines targeting porcine enteric diarrhea coronavirus.
Assuntos
Infecções por Coronavirus , Imunoinformática , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos , Vacinas Virais , Animais , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Epitopos/imunologia , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito T/imunologia , Imunoinformática/métodos , Simulação de Acoplamento Molecular , Vírus da Diarreia Epidêmica Suína/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/química , Suínos , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/imunologia , Doenças dos Suínos/virologia , Vacinas Virais/imunologiaRESUMO
As one of the most important swine enteropathogenic coronavirus, porcine epidemic diarrhea virus (PEDV) is the causative agent of an acute and devastating enteric disease that causes lethal watery diarrhea in suckling piglets. Recent progress in studying PEDV has revealed many intriguing findings on its prevalence and genetic evolution, rapid diagnosis, suppression of host gene expression, and suppression of the host innate immune system. Due to the continuous mutation of the PEDV genome, viral evasions from innate immune defenses and mixed infection with other coronaviruses, the spread of the virus is becoming wider and faster, making it even more necessary to prevent the infections caused by wild-type PEDV variants. It has also been reported that PEDV nsp1 is an essential virulence determinant and is critical for inhibiting host gene expression by structural and biochemical analyses. The inhibition of host protein synthesis employed by PEDV nsp1 may contribute to the regulation of host cell proliferation and immune evasion-related biological functions. In this review, we critically evaluate the recent studies on these aspects of PEDV and assess prospects in understanding the function of PEDV proteins in regulating host innate immune response and viral virulence.
Assuntos
Infecções por Coronavirus , Evasão da Resposta Imune , Imunidade Inata , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos , Vírus da Diarreia Epidêmica Suína/genética , Vírus da Diarreia Epidêmica Suína/imunologia , Vírus da Diarreia Epidêmica Suína/patogenicidade , Animais , Suínos , Doenças dos Suínos/virologia , Doenças dos Suínos/imunologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Virulência/genética , Proteínas não Estruturais Virais/genética , Proteínas não Estruturais Virais/imunologia , Proteínas não Estruturais Virais/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Fatores de Virulência/genéticaRESUMO
As one genotype of porcine circovirus (PCV) identified in 2016, PCV3 has brought huge hidden dangers to the global swine industry together with PCV2. Virus-like particles (VLPs) of capsid protein (Cap) of PCV2 serve as an alternative nano-antigen delivery strategy to efficiently induce antiviral immune response against PCV2 and/or other covalently displayed swine pathogens. However, the current understanding is limited on the capability of PCV3 as a nano-vaccine vehicle. Here we systematically compared the characteristics and the immunogenic efficacy of PCV3 Cap (Cap3) and PCV2 Cap (Cap2) in a VLP form. Cap3 VLPs presented higher internalization efficiency into cells and cytokines production compared to those of Cap2. Meanwhile, cross-reactive immunity between Cap3 VLPs and Cap2 VLPs was detected. Furthermore, to evaluate the function of Cap3 VLPs and Cap2 VLPs as vaccine vehicles carrying foreign proteins, the non-structural protein 6 of porcine reproductive and respiratory syndrome virus (PRRSV) was fused to C-terminus of Cap. Cap3-based chimeric particles induced a higher level of nsp6-specific immune response and PRRSV inhibition. Collectively, these self-assembling, Cap-based VLPs offer a compelling platform for enhancing the effectiveness of subunit vaccinations against newly emerging diseases and hold great promise for the development of Cap3-based chimeric subunit vaccines.
Assuntos
Proteínas do Capsídeo , Circovirus , Vacinas de Partículas Semelhantes a Vírus , Vacinas Virais , Animais , Circovirus/imunologia , Circovirus/genética , Suínos , Proteínas do Capsídeo/imunologia , Proteínas do Capsídeo/genética , Vacinas de Partículas Semelhantes a Vírus/imunologia , Vacinas Virais/imunologia , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/imunologia , Doenças dos Suínos/virologia , Anticorpos Antivirais/imunologia , Infecções por Circoviridae/prevenção & controle , Infecções por Circoviridae/imunologiaRESUMO
Atypical porcine pestivirus (APPV) is a novel member of the Pestivirus genus detected in association with congenital tremor (CT) type A-II outbreaks and from apparently healthy pigs, both as singular infection and as part of multi-pathogen infections. 'Classical' pestiviruses are known to cause immunosuppression of their host, which can increase susceptibility to secondary infections, severely impacting health, welfare, and production. To investigate APPV's effect on the host's immune system and characterise disease outcomes, 12 piglets from a natural APPV CT type A-II outbreak were experimentally infected with porcine reproductive and respiratory syndrome virus (PRRSV), a significant porcine pathogen. Rectal temperatures indicating febrile responses, viremia and viral-specific humoral and cellular responses were assessed throughout the study. Pathological assessment of the lungs and APPV-PRRSV co-localisation within the lungs was performed at necropsy. Viral co-localisation and pathological assessment of the lungs (Immunohistochemistry, BaseScope in situ hybridisation) were performed post-mortem. APPV status did not impact virological or immunological differences in PRRSV-infected groups. However, significantly higher rectal temperatures were observed in the APPV+ve/PRRSV+ve group over four days, indicating APPV increased the febrile response. Significant differences in the lung consolidation of the apical and intermediate lobes were also present, suggesting that APPV co-infection may augment lung pathology.
Assuntos
Coinfecção , Pulmão , Infecções por Pestivirus , Pestivirus , Síndrome Respiratória e Reprodutiva Suína , Vírus da Síndrome Respiratória e Reprodutiva Suína , Animais , Suínos , Infecções por Pestivirus/veterinária , Infecções por Pestivirus/virologia , Infecções por Pestivirus/patologia , Pestivirus/patogenicidade , Pestivirus/genética , Coinfecção/virologia , Coinfecção/veterinária , Síndrome Respiratória e Reprodutiva Suína/virologia , Síndrome Respiratória e Reprodutiva Suína/patologia , Síndrome Respiratória e Reprodutiva Suína/imunologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/patogenicidade , Pulmão/virologia , Pulmão/patologia , Viremia , Doenças dos Suínos/virologia , Doenças dos Suínos/patologia , Doenças dos Suínos/imunologia , Anticorpos Antivirais/sangueRESUMO
Ascaris is one of the most widespread helminth infections, leading to chronic morbidity in humans and considerable economic losses in pig farming. In addition, pigs are an important reservoir for the zoonotic salmonellosis, where pigs can serve as asymptomatic carriers. Here, we investigated the impact of an ongoing Ascaris infection on the immune response to Salmonella in pigs. We observed higher bacterial burdens in experimentally coinfected pigs compared to pigs infected with Salmonella alone. The impaired control of Salmonella in the coinfected pigs was associated with repressed interferon gamma responses in the small intestine and with the alternative activation of gut macrophages evident in elevated CD206 expression. Ascaris single and coinfection were associated with a rise of CD4-CD8α+FoxP3+ Treg in the lymph nodes draining the small intestine and liver. In addition, macrophages from coinfected pigs showed enhanced susceptibility to Salmonella infection in vitro and the Salmonella-induced monocytosis and tumor necrosis factor alpha production by myeloid cells was repressed in pigs coinfected with Ascaris. Hence, our data indicate that acute Ascaris infection modulates different immune effector functions with important consequences for the control of tissue-invasive coinfecting pathogens.IMPORTANCEIn experimentally infected pigs, we show that an ongoing infection with the parasitic worm Ascaris suum modulates host immunity, and coinfected pigs have higher Salmonella burdens compared to pigs infected with Salmonella alone. Both infections are widespread in pig production and the prevalence of Salmonella is high in endemic regions of human Ascariasis, indicating that this is a clinically meaningful coinfection. We observed the type 2/regulatory immune response to be induced during an Ascaris infection correlates with increased susceptibility of pigs to the concurrent bacterial infection.
Assuntos
Ascaríase , Ascaris suum , Coinfecção , Salmonelose Animal , Doenças dos Suínos , Animais , Ascaríase/imunologia , Ascaríase/veterinária , Suínos , Coinfecção/imunologia , Coinfecção/microbiologia , Coinfecção/parasitologia , Doenças dos Suínos/imunologia , Doenças dos Suínos/microbiologia , Doenças dos Suínos/parasitologia , Ascaris suum/imunologia , Salmonelose Animal/imunologia , Salmonelose Animal/microbiologia , Macrófagos/imunologia , Linfócitos T Reguladores/imunologia , Linfonodos/imunologia , Intestino Delgado/imunologia , Intestino Delgado/microbiologia , Intestino Delgado/parasitologia , Interferon gama/imunologia , Interferon gama/metabolismo , Fígado/imunologia , Fígado/parasitologiaRESUMO
We previously showed that several polymorphisms in genes encoding pattern recognition receptors that cause amino acid substitutions alter pathogen recognition ability and disease susceptibility in pigs. In this study, we expanded our analysis to a wide range of immune-related genes and investigated polymorphism distribution and its influence on pneumonia in multiple commercial pig populations. Among the polymorphisms in 42 genes causing 634 amino acid substitutions extracted from the swine genome database, 80 in 24 genes were found to have a minor allele frequency of at least 10% in Japanese breeding stock pigs via targeted resequencing. Of these, 62 single nucleotide polymorphisms (SNPs) in 23 genes were successfully genotyped in 862 pigs belonging to four populations with data on pneumonia severity. Association analysis using a generalized linear mixed model revealed that 12 SNPs in nine genes were associated with pneumonia severity. In particular, SNPs in the cellular receptor for immunoglobulin G FCGR2B and the intracellular nucleic acid sensors IFI16 and LRRFIP1 were found to be associated with mycoplasmal pneumonia of swine or porcine pleuropneumonia in multiple populations and may therefore have wide applications in the improvement of disease resistance in pigs. Functional analyses at the cellular and animal levels are required to clarify the mechanisms underlying the effects of these SNPs on disease susceptibility.
Assuntos
Pneumonia , Polimorfismo de Nucleotídeo Único , Doenças dos Suínos , Suínos , Pneumonia/genética , Pneumonia/imunologia , Pneumonia/microbiologia , Pneumonia/veterinária , Doenças dos Suínos/genética , Doenças dos Suínos/imunologia , Doenças dos Suínos/microbiologia , Receptores de Reconhecimento de Padrão/genética , Receptores de Reconhecimento de Padrão/imunologia , Masculino , Feminino , Genótipo , Alelos , Índice de Gravidade de DoençaRESUMO
Dihydromyricetin (DMY), a natural flavonoid compound, are believed to prevent inflammatory response, dealing with pathogens and repairing the intestinal barrier. The objective of this study was to investigate whether DMY supplementation could attenuate intestinal damage in the context of enterotoxigenic Escherichia coli K88 (ETEC F4+) infection. After weaning, different litters of pigs were randomly assigned to one of the following treatments: (1) non-challenged control (CON, fed with basal diet); (2) ETEC-challenged control (ECON, fed with basal diet); and (3) ETEC challenge + DMY treatment (EDMY, fed with basal diet plus 300 mg kg-1 DMY). We observed a significant reduction in fecal Escherichia coli shedding and diarrhea incidence, but an increase in ADG in pigs of EDMY group compared to the pigs of ECON group. Relative to the pigs of ECON group, dietary DMY treatment decreased (P < 0.05) concentrations of the serum D-xylose, D-lactate and diamine oxidase (DAO), but increased the abundance of zonula occludens-1 (ZO-1) in the jejunum of pigs. In addition, DMY also decreased (P < 0.05) the number of S-phase cells and the percentage of total apoptotic epithelial cells of jejunal epithelium in pigs of the EDMY group compared to the pigs of the ECON group. Furthermore, DMY decreased the mRNA expression levels of critical immune-associated genes TLR4, NFκB, Caspase3, Caspase9, IL-1ß, IL-6, TNF-α and the protein p-NFκB and p-IκBα expressions of intestinal epithelium in pigs of the EDMY group compared to the pigs of the ECON group. Compared to the ECON group, DMY elevated (P < 0.05) the expression levels of ß-defensins PBD1, PBD2, PBD3, PBD129, as well as the abundance of secreted IgA in intestinal mucosae of the EDMY group. Thus, our results indicate that DMY may relieve intestinal integrity damage due to Escherichia coli F4.
Assuntos
Escherichia coli Enterotoxigênica , Infecções por Escherichia coli , Flavonóis , Mucosa Intestinal , Doenças dos Suínos , Animais , Escherichia coli Enterotoxigênica/efeitos dos fármacos , Suínos , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/patologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Mucosa Intestinal/imunologia , Flavonóis/farmacologia , Flavonóis/uso terapêutico , Infecções por Escherichia coli/tratamento farmacológico , Infecções por Escherichia coli/veterinária , Infecções por Escherichia coli/imunologia , Doenças dos Suínos/tratamento farmacológico , Doenças dos Suínos/microbiologia , Doenças dos Suínos/imunologia , Desmame , Citocinas/metabolismo , Diarreia/tratamento farmacológico , Diarreia/veterinária , Apoptose/efeitos dos fármacos , Proteína da Zônula de Oclusão-1/metabolismo , Proteína da Zônula de Oclusão-1/genéticaRESUMO
Glaesserella parasuis (GPS) can cause severe systemic inflammation in pigs, resulting in huge economic losses to the pig industry. At present, no effective method is available for the prevention and control of GPS infection. Molecular breeding for disease resistance is imminent, but disease-resistance genes have not been identified. To study the mechanism of systemic acute inflammation caused by GPS, we established three in vitro infection models (3D4/21 cells, PK15 cells, and PAVEC cells) according to its infection path. There was no significant difference in apoptosis among the three kinds of cells after 12 h of continuous GPS stimulation, while inflammatory factors were significantly upregulated. Subsequent transcriptome analysis revealed 1969, 1207, and 3564 differentially expressed genes (DEGs) in 3D4/21 cells, PK15 cells, and PAVEC cells, respectively, after GPS infection. Many of the DEGs were predicted to be associated with inflammatory responses (C3, CD44, etc.); cell proliferation, growth and apoptosis; gene expression; and protein phosphorylation. Key signaling pathways, including S100 family signaling, bacteria and virus recognition, and pathogen-induced cytokine storm signaling, were enriched based on Ingenuity Pathway Analysis (IPA). Furthermore, a total of three putative transmembrane receptors and two putative G-protein-coupled receptors, namely F3, ICAM1, PLAUR, ACKR3, and GPRC5A, were identified by IPA among the three types of cells. ACKR3 and GPRC5A play pivotal roles in bacterial adhesion, invasion, host immune response and inflammatory response through the S100 family signaling pathway. Our findings provide new insights into the pathological mechanisms underlying systemic inflammation caused by GPS infection in pigs, and they lay a foundation for further research on disease-resistance breeding to GPS.
Assuntos
Haemophilus parasuis , Inflamação , Transdução de Sinais , Doenças dos Suínos , Animais , Suínos , Haemophilus parasuis/genética , Haemophilus parasuis/patogenicidade , Transdução de Sinais/genética , Inflamação/genética , Inflamação/microbiologia , Doenças dos Suínos/microbiologia , Doenças dos Suínos/genética , Doenças dos Suínos/imunologia , Infecções por Haemophilus/veterinária , Infecções por Haemophilus/genética , Infecções por Haemophilus/microbiologia , Infecções por Haemophilus/imunologia , Transcriptoma/genética , Perfilação da Expressão Gênica , Linhagem Celular , Apoptose/genéticaRESUMO
BACKGROUND: Porcine deltacoronavirus (PDCoV) is a swine enteropathogenic coronavirus that affects young pigs, causing vomiting, acute diarrhea, dehydration, and even death. There is growing evidence that PDCoV can undergo cross-species as well as zoonotic transmissions. Due to the frequent outbreaks of this deadly virus, early detection is essential for effective prevention and control. Therefore, developing a more convenient and reliable method for PDCoV detection is the need of the hour. RESULTS: This study utilized a high-affinity monoclonal antibody as the capture antibody and a horseradish peroxidase labeled polyclonal antibody as the detection antibody to develop an enzyme-linked immunosorbent assay (DAS-ELSA) for PDCoV detection.Both antibodies target the PDCoV nucleocapsid (N) protein. The findings of this study revealed that DAS-ELISA was highly specific to PDCoV and did not cross-react with other viruses to cause swine diarrhea. The limit of detection of the virus titer using this method was 103 TCID50/mL of PDCoV particles. The results of a parallel analysis of 239 known pig samples revealed a coincidence rate of 97.07% (κ = 0.922) using DAS-ELISA and reverse transcriptase PCR (RT-PCR). The DAS-ELISA was used to measure the one-step growth curve of PDCoV in LLC-PK cells and the tissue distribution of PDCoV in infected piglets. The study found that the DAS-ELISA was comparable in accuracy to the TCID50 method while measuring the one-step growth curve. Furthermore, the tissue distribution measured by DAS-ELISA was also consistent with the qRT-PCR method. CONCLUSION: The developed DAS-ELISA method can be conveniently used for the early clinical detection of PDCoV infection in pigs, and it may also serve as an alternative method for laboratory testing of PDCoV.