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2.
PLoS Pathog ; 16(8): e1008763, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32834002

RESUMO

The various sub-species of Salmonella enterica cause a range of disease in human hosts. The human-adapted Salmonella enterica serovar Typhi enters the gastrointestinal tract and invades systemic sites to cause enteric (typhoid) fever. In contrast, most non-typhoidal serovars of Salmonella are primarily restricted to gut tissues. Across Africa, invasive non-typhoidal Salmonella (iNTS) have emerged with an ability to spread beyond the gastrointestinal tract and cause systemic bloodstream infections with increased morbidity and mortality. To investigate this evolution in pathogenesis, we compared the genomes of African iNTS isolates with other Salmonella enterica serovar Typhimurium and identified several macA and macB gene variants unique to African iNTS. MacAB forms a tripartite efflux pump with TolC and is implicated in Salmonella pathogenesis. We show that macAB transcription is upregulated during macrophage infection and after antimicrobial peptide exposure, with macAB transcription being supported by the PhoP/Q two-component system. Constitutive expression of macAB improves survival of Salmonella in the presence of the antimicrobial peptide C18G. Furthermore, these macAB variants affect replication in macrophages and influence fitness during colonization of the murine gastrointestinal tract. Importantly, the infection outcome resulting from these macAB variants depends upon both the Salmonella Typhimurium genetic background and the host gene Nramp1, an important determinant of innate resistance to intracellular bacterial infection. The variations we have identified in the MacAB-TolC efflux pump in African iNTS may reflect evolution within human host populations that are compromised in their ability to clear intracellular Salmonella infections.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Proteínas de Bactérias/genética , Colite/patologia , Variação Genética , Macrófagos/imunologia , Salmonelose Animal/patologia , Salmonella typhimurium/imunologia , Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Proteínas de Bactérias/metabolismo , Linhagem da Célula , Colite/induzido quimicamente , Colite/imunologia , Colite/microbiologia , Análise Mutacional de DNA , Modelos Animais de Doenças , Macrófagos/metabolismo , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Salmonelose Animal/imunologia , Salmonelose Animal/microbiologia , Replicação Viral
3.
PLoS Pathog ; 16(8): e1008766, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32857822

RESUMO

Pathogens commonly disrupt the intestinal epithelial barrier; however, how the epithelial immune system senses the loss of intestinal barrier as a danger signal to activate self-defense is unclear. Through an unbiased approach in the model nematode Caenorhabditis elegans, we found that the EGL-44/TEAD transcription factor and its transcriptional activator YAP-1/YAP (Yes-associated protein) were activated when the intestinal barrier was disrupted by infections with the pathogenic bacterium Pseudomonas aeruginosa PA14. Gene Ontology enrichment analysis of the genes containing the TEAD-binding sites revealed that "innate immune response" and "defense response to Gram-negative bacterium" were two top significantly overrepresented terms. Genetic inactivation of yap-1 and egl-44 significantly reduced the survival rate and promoted bacterial accumulation in worms after bacterial infections. Furthermore, we found that disturbance of the E-cadherin-based adherens junction triggered the nuclear translocation and activation of YAP-1/YAP in the gut of worms. Although YAP is a major downstream effector of the Hippo signaling, our study revealed that the activation of YAP-1/YAP was independent of the Hippo pathway during disruption of intestinal barrier. After screening 10 serine/threonine phosphatases, we identified that PP2A phosphatase was involved in the activation of YAP-1/YAP after intestinal barrier loss induced by bacterial infections. Additionally, our study demonstrated that the function of YAP was evolutionarily conserved in mice. Our study highlights how the intestinal epithelium recognizes the loss of the epithelial barrier as a danger signal to deploy defenses against pathogens, uncovering an immune surveillance program in the intestinal epithelium.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Permeabilidade da Membrana Celular , Células Epiteliais/imunologia , Microbioma Gastrointestinal/imunologia , Salmonelose Animal/imunologia , Salmonella typhimurium/imunologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Células Epiteliais/metabolismo , Células Epiteliais/microbiologia , Células Epiteliais/patologia , Camundongos , Salmonelose Animal/metabolismo , Salmonelose Animal/microbiologia , Salmonelose Animal/patologia , Transdução de Sinais
4.
Appl Environ Microbiol ; 86(21)2020 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-32859592

RESUMO

Salmonella enterica serovar Typhimurium is a prevalent incitant of enteritis in human beings and nonhuman animals. It has been proposed that host defense responses incited by Salmonella allow the bacterium to overcome colonization resistance. Piglets (n = 24) were orally inoculated with S. enterica serovar Typhimurium DT104 or buffer alone, and the host and microbial responses were temporally examined at the acute (2 days postinoculation [dpi]), subacute (6 dpi), and recovery (10 dpi) stages of salmonellosis. At the acute stage of disease, body temperatures were elevated, and feed consumption and weight gain were reduced. The densities of Salmonella associated with the gut mucosa decreased over time, with higher densities of the bacterium in the ileum and the large intestine. Moreover, substantive histopathological changes were observed as a function of time, with prominent epithelial injury and neutrophil infiltration observed at 2 dpi. Correspondingly, a variety of host metrics were temporally affected in piglets with salmonellosis (e.g., TNFα, IFNγ, PR39, ßD2, iNOS, IL8, REGIIIγ). The enteric microbiota was characterized using culture-independent and -dependent methods in concert, and taxon- and location-specific changes to the microbiota were observed in infected piglets. Bacteroides spp. (e.g., Bacteroides uniformis, Bacteroides fragilis), Streptococcus spp. (e.g., Streptococcus gallolyticus), and various Gammaproteobacteria were highly associated with inflamed tissues, while bacteria within the Ruminococcaceae and Veillonellaceae families were mainly associated with healthy mucosae. In conclusion, the study findings showed that S Typhimurium incited temporal and spatial modifications to the swine autochthonous microbiota, and to host defense responses, that were consistent with overcoming colonization resistance to incite salmonellosis in swine.IMPORTANCE Limited information is available on host and enteric microbiota responses incited by Salmonella enterica serovar Typhimurium in swine and on possible mechanisms by which the bacterium overcomes colonization resistance to incite salmonellosis. Temporal characterization of a variety of host metrics in piglets (e.g., physiological, histopathological, and immunological) showed the importance of studying the progression of salmonellosis. A number of host responses integrally associated with disease development were identified. Utilization of next-generation sequence analysis to characterize the enteric microbiota was found to lack sufficient resolution; however, culture-dependent and -independent methods in combination identified taxon- and location-specific changes to bacterial communities in infected piglets. The study identified bacterial and host responses associated with salmonellosis, which will be beneficial in understanding colonization resistance and in the development of effective alternatives to antibiotics to mitigate salmonellosis.


Assuntos
Ceco/microbiologia , Colo/microbiologia , Microbioma Gastrointestinal , Interações entre Hospedeiro e Microrganismos/imunologia , Íleo/microbiologia , Salmonella typhimurium/fisiologia , Animais , Ceco/imunologia , Colo/imunologia , Íleo/imunologia , Masculino , Distribuição Aleatória , Salmonelose Animal/imunologia , Salmonelose Animal/microbiologia , Suínos , Doenças dos Suínos/imunologia , Doenças dos Suínos/microbiologia , Fatores de Tempo
5.
Nat Cell Biol ; 22(6): 663-673, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32393887

RESUMO

The linear ubiquitin chain assembly complex (LUBAC), which consists of HOIP, SHARPIN and HOIL-1L, promotes NF-κB activation and protects against cell death by generating linear ubiquitin chains. LUBAC contains two RING-IBR-RING (RBR) ubiquitin ligases (E3), and the HOIP RBR is responsible for catalysing linear ubiquitination. We found that HOIL-1L RBR plays a crucial role in regulating LUBAC. HOIL-1L RBR conjugates monoubiquitin onto all LUBAC subunits, followed by HOIP-mediated conjugation of linear chains onto monoubiquitin, and these linear chains attenuate the functions of LUBAC. The introduction of E3-defective HOIL-1L mutants into cells augmented linear ubiquitination, which protected the cells against Salmonella infection and cured dermatitis caused by reduced LUBAC levels due to SHARPIN loss. Our results reveal a regulatory mode of E3 ligases in which the accessory E3 in LUBAC downregulates the main E3 by providing preferred substrates for autolinear ubiquitination. Thus, inhibition of HOIL-1L E3 represents a promising strategy for treating severe infections or immunodeficiency.


Assuntos
Proteínas de Transporte/fisiologia , Morte Celular , Doença Hepática Induzida por Substâncias e Drogas/imunologia , Dermatite de Contato/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Salmonelose Animal/imunologia , Ubiquitina-Proteína Ligases/fisiologia , Ubiquitina/metabolismo , Animais , Células Cultivadas , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/patologia , Dermatite de Contato/metabolismo , Dermatite de Contato/patologia , Embrião de Mamíferos/imunologia , Embrião de Mamíferos/metabolismo , Embrião de Mamíferos/patologia , Fibroblastos/imunologia , Fibroblastos/metabolismo , Fibroblastos/patologia , Regulação da Expressão Gênica , Camundongos Knockout , NF-kappa B/genética , NF-kappa B/metabolismo , Salmonella/patogenicidade , Salmonelose Animal/metabolismo , Salmonelose Animal/patologia , Índice de Gravidade de Doença , Transdução de Sinais , Ubiquitinação
6.
PLoS One ; 15(4): e0231942, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32339193

RESUMO

Salmonella enterica serovar Typhimurium is an animal welfare and public health concern due to its ability to parasite livestock and potentially contaminate pork products. To reduce Salmonella shedding and the risk of pork contamination, antibiotic therapy is used and can contribute to antimicrobial resistance. Here we hypothesized that immune system education by the microbiota can play a role in intestinal resilience to infection. We used amoxicillin (15mg/Kg) to modulate the intestinal microbiome of 10 piglets, paired with same age pigs that received a placebo (n = 10) from 0 to 14 days of age. Animals were euthanized at 4-weeks old. Each pig donated colon sections for ex vivo culture (n = 20 explants/pig). Explants were inoculated with S. Typhimurium, PBS or LPS (n = 6 explants/pig/group, plus technical controls). The gut bacteriome was characterized by sequencing of the 16S rRNA at 7, 21 days of age, and upon in vitro culture. Explants response to infection was profiled through high-throughput mRNA sequencing. In vivo antibiotic treatment led to ß-diversity differences between groups at all times (P<0.05), while α-diversity did not differ between amoxicillin and placebo groups on day 21 and at euthanasia (P<0.03 on day 7). Explant microbiomes were not different from in vivo. In vitro challenge with S. Typhimurium led to lower necrosis scores in explants from amoxicillin-treated pigs, when compared to explants placebo-treated pigs (P<0.05). This was coupled with the activation of immune-related pathways in explants from amoxicillin-treated pigs (IL-2 production, NO production, BCR activation), when compared to placebo-treated pigs. In addition, several DNA repair and intestinal wound healing pathways were also only activated in explants from amoxicillin-treated pigs. Taken together, these findings suggest that immune education by the amoxicillin-disturbed microbiota may have contributed to mitigate intestinal lesions following pathogen exposure.


Assuntos
Antibacterianos/farmacologia , Epigênese Genética/efeitos dos fármacos , Microbioma Gastrointestinal/efeitos dos fármacos , Salmonella typhimurium/patogenicidade , Amoxicilina/farmacologia , Animais , Animais Recém-Nascidos , Bactérias/genética , Bactérias/isolamento & purificação , Colo/citologia , Colo/microbiologia , Colo/patologia , Regulação para Baixo/efeitos dos fármacos , Fezes/microbiologia , Análise de Componente Principal , RNA Ribossômico 16S/química , RNA Ribossômico 16S/metabolismo , Salmonelose Animal/imunologia , Suínos , Doenças dos Suínos/imunologia , Regulação para Cima/efeitos dos fármacos
7.
PLoS One ; 15(4): e0231998, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32330169

RESUMO

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


Assuntos
Galinhas/imunologia , Quitosana/farmacologia , Vacinas contra Salmonella/imunologia , Animais , Proteínas da Membrana Bacteriana Externa/imunologia , Ceco/metabolismo , Galinhas/microbiologia , Quitosana/imunologia , Flagelina/imunologia , Antígenos de Histocompatibilidade Classe II , Nanopartículas/uso terapêutico , Doenças das Aves Domésticas/imunologia , Salmonella/imunologia , Salmonella/patogenicidade , Salmonelose Animal/imunologia , Salmonella enterica/metabolismo , Salmonella enteritidis/imunologia , Vacinas/imunologia , Vacinas Atenuadas/imunologia
8.
Vet Microbiol ; 243: 108633, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32273012

RESUMO

Salmonella Enteritidis (SE) is one of the most common culprits of foodborne disease in humans due to its horizontal transmission from infected animals to humans. The development of a safe vaccine against Salmonella would be important for both farm animals and humans concerning disease containment. The SE ghosts carrying FliC were genetically constructed using a special ghost plasmid pJHL184 that co-expressed FliC and the phage lysis gene E. These SE ghosts were characterized by ghost generation efficacy by increasing the culture temperature to "42 °C" in the absence of L-arabinose. This temperature change led to an ghost generation with almost complete lysis of the SE host strain in 48 hs. The expression of FliC was confirmed by Western blot analysis. Also, indirect ELISA was used to prove FliC specific antibody generation in immunized mice. The parenteral adjuvant effect of the FliC antigen was demonstrated by immunizing mice with pJHL184::flC, pJHL184 alone, or PBS alone. The mice were intramuscularly immunized at six weeks of age (n = 8) and boosted after three weeks of primary inoculation. A total of 32 mice were equally divided into four groups. Each group was treated with pJHL-ghosts alone, ghost surface displaying FliC adjuvant, and compared to the PBS and naïve control groups. The immunized mice demonstrated greater IgG and IgA antibody responses than did the PBS control group. Furthermore, the addition of the ghosts to the FliC led to a significant increase in both the humoral and cell-mediated immune responses compared to those in the ghost alone group. Besides, the in vitro antigen uptake and presentation studies revealed efficient antigen presentation on the mouse macrophage cell surfaces. This finding further corroborated the potential efficacy of immune stimulation by SE ghosts. After the virulent challenge, we observed a significant reduction in the bacterial load in the spleen and liver tissues in SE ghosts surface, displaying FliC adjuvant. Our results demonstrate a safe and effective strategy to prevent salmonellosis. They also suggest that the surface expression of flagellin (FliC) significantly enhances antigen-specific humoral and cell-mediated immune responses. This FliC expression can also enhance the protective efficacy of the bacterial ghosts-based vaccine against virulent challenge.


Assuntos
Anticorpos Antibacterianos/sangue , Flagelina/imunologia , Doenças das Aves Domésticas/prevenção & controle , Salmonelose Animal/prevenção & controle , Salmonella enteritidis/imunologia , Adjuvantes Imunológicos , Animais , Feminino , Flagelina/genética , Injeções Intramusculares , Camundongos , Camundongos Endogâmicos BALB C , Plasmídeos/genética , Plasmídeos/imunologia , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/microbiologia , Salmonelose Animal/imunologia , Vacinas contra Salmonella/administração & dosagem , Vacinas contra Salmonella/imunologia , Salmonella enteritidis/genética
9.
Int J Nanomedicine ; 15: 761-777, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32099364

RESUMO

Purpose: Salmonellosis in poultry is a serious economic burden. A major concern is the public health hazard caused by consumption of Salmonella-contaminated poultry products. Currently used Salmonella vaccines are ineffective in combating poultry Salmonellosis warranting the need of a potent vaccine, especially an oral vaccine that can elicit robust local intestinal immunity. Materials and Methods: A Salmonella subunit chitosan nanoparticles (NPs)-based vaccine was prepared that contained immunogenic outer membrane proteins (OMPs) and -flagellin (F) protein (OMPs-F-CS NPs). OMPs-F-CS NPs were administered as an oral vaccine in layer chickens and the resultant humoral and cell-mediated immune responses and localization of NPs were examined using standard detection methods. Results: We demonstrated targeting of surface F-protein coated chitosan NPs to immune cells when delivered orally to layer chickens, the particles were localized in ileal Peyer's patches. The OMPs-F-CS NPs vaccinated layer chickens had significantly higher OMPs-specific mucosal IgA production and lymphocyte proliferation response. The candidate vaccine increased the expression of toll-like receptor (TLR)-2, TLR-4, IFN-γ, TGF-ß and IL-4 mRNA expression in chicken cecal tonsils. Conclusion: Our study demonstrated that the chitosan-based oral Salmonella nanovaccine targets immune cells of chickens and induced antigen-specific B and T cell responses. This candidate oral Salmonella nanovaccine has the potential to mitigate Salmonellosis in poultry.


Assuntos
Galinhas , Salmonelose Animal/prevenção & controle , Vacinas contra Salmonella/administração & dosagem , Vacinas contra Salmonella/imunologia , Administração Oral , Animais , Galinhas/imunologia , Quitosana/química , Feminino , Imunidade Celular , Interleucina-4/genética , Mucosa Intestinal/efeitos dos fármacos , Nanopartículas/administração & dosagem , Nanopartículas/química , Porinas/química , Salmonelose Animal/imunologia , Salmonella enteritidis/imunologia , Receptores Toll-Like/genética , Fator de Crescimento Transformador beta/genética , Vacinas de Subunidades/administração & dosagem
10.
Vet Res ; 51(1): 26, 2020 Feb 24.
Artigo em Inglês | MEDLINE | ID: mdl-32093767

RESUMO

Probiotic pretreatment is an effective non-antibiotic strategy for preventing or controlling Salmonella infections. We found that Lactobacillus johnsonii L531, isolated from the colon of a clinically healthy weaned piglet, effectively prevented infection with Salmonella enterica serovar Infantis in a pig model. Newly weaned piglets were intragastrically administered Lactobacillus johnsonii L531 at 1.0 × 1010 CFU/day for 1 week before S. Infantis challenge. Pretreatment with L. johnsonii L531 lessened the severity of diarrhea and ileal inflammation in S. Infantis-infected piglets. Lactobacilli were more abundant in the ileum than jejunum after L. johnsonii L531 pretreatment. Treatment with L. johnsonii L531 reduced the abundance of total bacteria in the ileal mucosa and the production of lipocalin 2 in the jejunum of piglets challenged with Salmonella. Both intestinal morphology and transmission electron microscopy results indicated that L. johnsonii L531 alleviated intestinal tissue damage following S. Infantis challenge, especially in the villus and endoplasmic reticulum (ER). ER stress induced by S. Infantis was attenuated by L. johnsonii L531 treatment. The number of CD4- CCR6+ T cells decreased following S. Infantis challenge, but the percentage of CCR6- IFNγ+ T cells in peripheral blood increased. In intestinal mesenteric lymph nodes, S. Infantis increased the proportion of CCR6+ IFNγ+ T cells, whereas L. johnsonii L531 induced an increase in the proportion of CD4+ CCR6+ T cells in response to S. Infantis infection. Our data thus suggest that L. johnsonii L531 contributes to the maintenance of intestinal homeostasis by modulating T-cell responses and ER stress.


Assuntos
Anti-Inflamatórios/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Lactobacillus johnsonii/química , Probióticos/farmacologia , Salmonelose Animal/prevenção & controle , Doenças dos Suínos/prevenção & controle , Linfócitos T/imunologia , Animais , Salmonelose Animal/imunologia , Suínos , Doenças dos Suínos/imunologia
11.
Nat Commun ; 11(1): 851, 2020 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-32051408

RESUMO

Lipopolysaccharide (LPS) O-antigen (O-Ag) is known to limit antibody binding to surface antigens, although the relationship between antibody, O-Ag and other outer-membrane antigens is poorly understood. Here we report, immunization with the trimeric porin OmpD from Salmonella Typhimurium (STmOmpD) protects against infection. Atomistic molecular dynamics simulations indicate this is because OmpD trimers generate footprints within the O-Ag layer sufficiently sized for a single IgG Fab to access. While STmOmpD differs from its orthologue in S. Enteritidis (SEn) by a single amino-acid residue, immunization with STmOmpD confers minimal protection to SEn. This is due to the OmpD-O-Ag interplay restricting IgG binding, with the pairing of OmpD with its native O-Ag being essential for optimal protection after immunization. Thus, both the chemical and physical structure of O-Ag are key for the presentation of specific epitopes within proteinaceous surface-antigens. This enhances combinatorial antigenic diversity in Gram-negative bacteria, while reducing associated fitness costs.


Assuntos
Anticorpos Antibacterianos/imunologia , Proteínas da Membrana Bacteriana Externa/química , Proteínas da Membrana Bacteriana Externa/imunologia , Imunização , Antígenos O/imunologia , Salmonella typhimurium/imunologia , Animais , Anticorpos Antibacterianos/sangue , Formação de Anticorpos , Especificidade de Anticorpos , Antígenos de Bactérias/imunologia , Proteínas da Membrana Bacteriana Externa/genética , Proteção Cruzada , Modelos Animais de Doenças , Epitopos/química , Epitopos/imunologia , Imunoglobulina G/sangue , Camundongos , Modelos Moleculares , Antígenos O/química , Antígenos O/genética , Porinas/química , Porinas/genética , Porinas/imunologia , Conformação Proteica , Salmonelose Animal/imunologia , Salmonelose Animal/prevenção & controle , Análise de Sequência de Proteína
12.
Microbes Infect ; 22(1): 40-45, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31430539

RESUMO

Receptor interacting protein kinase 3 (Ripk3) is a signal relay protein involved in initiation of programmed cell death (necroptosis) and modulation of inflammasome activation. While caspase 1 and 11 are pro-inflammatory caspases responsible for unleashing inflammation and cell death by enzymatic activation of the executioners of inflammation and cell death (pyroptosis). Upon Salmonella infection, the host mounts a pro-inflammatory response which require Ripk3 and Caspase1/11. Here we show that bone marrow derived macrophages with combined deficiency of Ripk3 and Casp1/11 are highly resistant to Salmonella induced cell death, and that these macrophages show an anti-inflammatory cytokine profile. We confirm what was previously known that mice deficient in Casp1/11 have impaired ability to control Salmonella burden, and that the absence of Ripk3 alone does not influence the innate immune responses to Salmonella infection. However, we describe a synergistic role of Ripk3 and Casp1/11 in regulating Salmonella in vivo burden and that Ripk3-dependent host protection in the absence of Casp1/11 is evident during infection by sifA-expressing Salmonella. In summary, we show that the Ripk3 protection to Salmonella infection is obscured by presence of Caspase 1/11 and that the Ripk3-dependent protection requires a phagosome-bound Salmonella.


Assuntos
Caspase 1/metabolismo , Caspases Iniciadoras/metabolismo , Inflamassomos/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Animais , Carga Bacteriana , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Caspase 1/deficiência , Caspases Iniciadoras/deficiência , Morte Celular , Citocinas/metabolismo , Macrófagos/metabolismo , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína Serina-Treonina Quinases de Interação com Receptores/deficiência , Salmonella/fisiologia , Salmonelose Animal/imunologia , Salmonelose Animal/microbiologia
13.
Microb Pathog ; 139: 103854, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31704463

RESUMO

Salmonella enteric serovar Typhimurium (S. Typhimurium) is a zoonotic pathogen causing public health hazards. Identification of genes related to macrophages resistance to S. Typhimurium and their immune mechanisms can provide a theoretical basis for disease resistance. In this study, sixty significant differentially expressed genes were screened between susceptible and resistant sheep macrophages by transcriptome RNA-seq. Eight significantly enriched GO terms and six canonical pathways were involved by GO and KEGG enrichment analysis. Furthermore, knockdown of HMOX1 and SLPI increased remarkably the clearance of S. typhimurium, but SPP1 had little effect on the clearance of S. Typhimurium within sheep macrophages. Altogether, these results suggest that many genes of macrophages were reprogrammed via S. Typhimurium infection, some of which may facilitate host defence against Salmonella, while others allow Salmonella to escape.


Assuntos
Resistência à Doença/genética , Macrófagos/imunologia , Salmonelose Animal/genética , Salmonella typhimurium , Animais , Feminino , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Macrófagos/metabolismo , Macrófagos/microbiologia , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Osteopontina/genética , Osteopontina/metabolismo , Salmonelose Animal/imunologia , Salmonelose Animal/microbiologia , Inibidor Secretado de Peptidases Leucocitárias/genética , Inibidor Secretado de Peptidases Leucocitárias/metabolismo , Ovinos , Doenças dos Ovinos , Transcriptoma
14.
Infect Immun ; 88(3)2020 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-31843963

RESUMO

Salmonella enterica serovar Pullorum is the pathogen of pullorum disease, which leads to severe economic losses in many developing countries. In contrast to the strong inflammatory response induced by Salmonella enterica serovar Typhimurium and Salmonella enterica serovar Enteritidis, S Pullorum causes systemic infection with little inflammation. The effector proteins secreted by Salmonella often play a crucial role in modulating host signal transduction and cellular processes to the pathogen's advantage. In the present study, the invasion plasmid antigen J (IpaJ) protein specifically identified in S Pullorum was found to significantly inhibit activation of the key proinflammatory transcription factor, NF-κB, which was induced by tumor necrosis factor alpha (TNF-α), interleukin-1ß (IL-1ß), and lipopolysaccharide (LPS). IpaJ inhibited the NF-κB pathway in cells infected with S Pullorum through the stabilization of IκBα. Deletion of ipaJ in S Pullorum caused a significantly increased level of ubiquitinated IκBα that was subsequently degraded by the proteasome in HeLa cells. Moreover, IpaJ was efficient in the prevention of NF-κB translocation to the nucleus and ultimately interfered with the secretion of the proinflammatory cytokines IL-1ß, IL-6, and IL-8 in infected HeLa cells. Additionally, the transformation of ipaJ into S Enteritidis decreased the secretion of proinflammatory cytokines in HeLa cells through suppression of the NF-κB pathway. The infection of chicken peripheral blood monocyte-derived macrophages (chMDM) confirmed that ipaJ-deleted S Pullorum induced a stronger expression of proinflammatory cytokines than the wild-type and complementary strains. In summary, the present study revealed that IpaJ functions as an important anti-inflammatory protein involved in S Pullorum infection through inhibition of the NF-κB pathway and the subsequent inflammatory response.


Assuntos
Antígenos de Bactérias/imunologia , NF-kappa B/imunologia , Salmonelose Animal/imunologia , Salmonella enterica/patogenicidade , Ubiquitinação/fisiologia , Animais , Galinhas , Células HeLa/metabolismo , Humanos , Interleucinas/metabolismo
15.
Avian Dis ; 63(4): 559-567, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31865669

RESUMO

Intestinal inflammation may provide a growth advantage for Salmonella and enhance its systemic spread in chickens. Salmonella triggers intestinal inflammation in the host by using type III secretion systems (T3SS) and produces the inflammatory end product tetrathionate. In mice, tetrathionate respiration confers a growth advantage for Salmonella Typhimurium over the competitive microbiome in the inflamed intestine. Coccidia also promote intestinal inflammation and enhance Salmonella intestinal growth and systemic spread in chickens. The objective of this study was to evaluate the contribution of inflammation, induced by Eimeria spp. or Salmonella Typhimurium, to Salmonella colonization and dissemination in chickens. In addition, the fitness costs associated with defects in tetrathionate reductase and T3SS associated with Salmonella Pathogenicity Island 1 or 2 (SPI-1 or SPI-2) were evaluated in in vivo competition experiments with wild-type Salmonella strain, with or without Eimeria coinfection. One-day-old specific-pathogen-free chickens were orally inoculated with a sham inoculum or with 4 × 102 Eimeria oocysts cocktail of Eimeria tenella, Eimeria acervulina, Eimeria maxima, and Eimeria mitis. At 6 days of age, birds were orally administered a 1:1 ratio of Salmonella Typhimurium wild-type and mutant deficient in tetrathionate reductase, SPI-1, or SPI-2 (108 colony forming units/bird). Ceca, livers, and drumsticks were collected at 3, 7, 14, and 42 days after Salmonella infection, for bacteriology. Intestinal inflammation was scored by histology. Significantly higher intestinal inflammation was observed in challenge groups compared with the control. However, there were no significant differences in intestinal inflammation scores between groups coinfected with both Eimeria spp. and Salmonella Typhimurium and birds infected with Salmonella alone, and Eimeria coinfection did not increase Salmonella prevalence or abundance. Contrary to mouse studies, tetrathionate reductase did not enhance Salmonella Typhimurium cecal colonization or systemic spread in chickens. SPI-1 and SPI-2 played a significant role in Salmonella dissemination and cecal colonization in chickens, respectively.


Assuntos
Proteínas de Bactérias/genética , Galinhas , Coccidiose/veterinária , Coinfecção/veterinária , Doenças das Aves Domésticas/microbiologia , Salmonelose Animal/microbiologia , Animais , Proteínas de Bactérias/metabolismo , Ceco/microbiologia , Coccidiose/imunologia , Coccidiose/parasitologia , Coinfecção/microbiologia , Coinfecção/parasitologia , Eimeria/fisiologia , Inflamação/imunologia , Inflamação/microbiologia , Inflamação/parasitologia , Inflamação/veterinária , Enteropatias/imunologia , Enteropatias/microbiologia , Enteropatias/parasitologia , Enteropatias/veterinária , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Oxirredutases/genética , Oxirredutases/metabolismo , Doenças das Aves Domésticas/imunologia , Doenças das Aves Domésticas/parasitologia , Salmonelose Animal/imunologia , Salmonella typhimurium/fisiologia , Organismos Livres de Patógenos Específicos , Sistemas de Secreção Tipo III/genética , Sistemas de Secreção Tipo III/metabolismo
16.
Sci Rep ; 9(1): 18017, 2019 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-31784576

RESUMO

Routine use of antibiotics in livestock animals strongly contributed to the creation of multidrug-resistant Salmonella Typhimurium strains (STM). Vaccination is an alternative to the use of antibiotics but often suffers from low efficacy. The present study investigated whether long-chain inulin (lcITF) and Lactobacillus acidophilus W37 (LaW37) can support vaccination efficacy against STM and if the interventions influence possible gut microbiota changes. Piglets received daily supplementation until sacrifice. Animals were vaccinated on day 25 after birth, one day after weaning, and were challenged with STM on days 52-54. Dietary intervention with lcITF/LaW37 enhanced vaccination efficacy by 2-fold during challenge and resulted in higher relative abundance of Prevotellaceae and lower relative abundance of Lactobacillaceae in faeces. Although strongest microbial effects were observed post STM challenge on day 55, transient effects of the lcITF/LaW37 intervention were also detected on day 10 after birth, and post-weaning on day 30 where increased relative abundance of faecal lactobacilli was correlated with higher faecal consistency. LcITF treatment increased post-weaning feed efficiency and faecal consistency but did not support vaccination efficacy. Vaccination in immune-immature young animals can be enhanced with functional additives which can simultaneously promote health in an ingredient-dependent fashion.


Assuntos
Ração Animal/microbiologia , Probióticos/administração & dosagem , Salmonelose Animal/prevenção & controle , Vacinas contra Salmonella/administração & dosagem , Vacinação/veterinária , Administração Oral , Criação de Animais Domésticos/métodos , Animais , Fezes/microbiologia , Feminino , Microbioma Gastrointestinal/imunologia , Imunogenicidade da Vacina , Inulina/administração & dosagem , Lactobacillus acidophilus/imunologia , Lactobacillus acidophilus/isolamento & purificação , Salmonelose Animal/imunologia , Salmonelose Animal/microbiologia , Vacinas contra Salmonella/imunologia , Salmonella typhimurium/imunologia , Suínos , Vacinação/métodos , Desmame
17.
Infect Immun ; 87(12)2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31548317

RESUMO

Most Salmonella serovars cause disease in many host species, while a few serovars have evolved to be host specific. Very little is known about the mechanisms that contribute to Salmonella host specificity. We compared the interactions between chicken primary macrophages (CDPM) and host-generalist serovar Salmonella enterica serovar Typhimurium, host-adapted Salmonella enterica serovar Dublin, and avian host-specific Salmonella enterica serovar Gallinarum. S Gallinarum was taken up in lower numbers by CDPM than S Typhimurium and S Dublin; however, a higher survival rate was observed for this serovar. In addition, S Typhimurium and S Dublin caused substantially higher levels of cell death to the CDPM, while significantly higher concentrations of NO were produced by S Gallinarum-infected cells. Global transcriptome analysis performed 2 h postinfection showed that S Gallinarum infection triggered a more comprehensive response in CDPM with 1,114 differentially expressed genes (DEGs) compared to the responses of S Typhimurium (625 DEGs) and S Dublin (656 DEGs). Comparable levels of proinflammation responses were observed in CDPM infected by these three different serovars at the initial infection phase, but a substantially quicker reduction in levels of interleukin-1ß (IL-1ß), CXCLi1, and CXCLi2 gene expression was detected in the S Gallinarum-infected macrophages than that of two other groups as infections proceeded. KEGG cluster analysis for unique DEGs after S Gallinarum infection showed that the JAK-STAT signaling pathway was top enriched, indicating a specific role for this pathway in response to S Gallinarum infection of CDPM. Together, these findings provide new insights into the interaction between Salmonella and the host and increase our understanding of S Gallinarum host specificity.


Assuntos
Especificidade de Hospedeiro/imunologia , Macrófagos/imunologia , Doenças das Aves Domésticas/imunologia , Salmonelose Animal/imunologia , Salmonella typhimurium/imunologia , Animais , Bovinos , Células Cultivadas , Galinhas , Interleucina-1beta/imunologia , Óxido Nítrico/metabolismo , Doenças das Aves Domésticas/microbiologia , Salmonella typhimurium/classificação
18.
Front Immunol ; 10: 2124, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31555302

RESUMO

Non-typhoidal Salmonella (NTS) are a leading cause of pediatric invasive bacterial infections in sub-Saharan Africa with high associated case fatality rates in children under 5 years old. We have developed glycoconjugate vaccines consisting of the lipid A-removed surface polysaccharide of NTS, core and O-polysaccharide (COPS), and the flagellar monomer protein (FliC) from the homologous serovar as the carrier. We previously established that COPS:FliC was immunogenic and protective in mice immunized as adults or infants; however, the brief period of murine infancy precluded the evaluation of protection against invasive NTS (iNTS) disease in early life. In the present study, we used a mouse model of maternal immunization to investigate transmission of S. Typhimurium COPS:FliC-induced maternal antibodies and protection against lethal iNTS challenge in infant mice. We found that vaccinated dams developed high levels of COPS- and FliC-specific IgG, which were transferred to their offspring. Sera from both vaccinated mothers and their litters mediated complement-dependent bactericidal activity in-vitro. Passively immunized 2-week old infant mice born to vaccinated mothers were fully protected from challenge with an S. Typhimurium blood isolate from sub-Saharan Africa. The pre-clinical findings reported herein demonstrate that anti-COPS:FliC antibodies induced by vaccination are sufficient for protection of murine infants against experimental S. Typhimurium infection. By underscoring the protective role of antibody, our results suggest that maintaining an adequate titer of protective anti-Salmonella antibodies during early life, either through pediatric or maternal COPS:FliC vaccination, may reduce iNTS disease in young children in sub-Saharan Africa.


Assuntos
Anticorpos Antibacterianos/imunologia , Troca Materno-Fetal , Salmonelose Animal/imunologia , Vacinas contra Salmonella/imunologia , Animais , Animais Recém-Nascidos , Feminino , Glicoconjugados/imunologia , Imunoconjugados , Camundongos , Antígenos O/imunologia , Gravidez , Salmonelose Animal/prevenção & controle
19.
Biomedica ; 39(Supl. 2): 172-181, 2019 08 01.
Artigo em Inglês, Espanhol | MEDLINE | ID: mdl-31529843

RESUMO

Introduction: The immunological role of plasmacytoid dendritic cells (pDC) in bacterial infections such as Salmonella has been poorly documented. Therefore, we analyzed the effector function of these cells by presenting cytotoxic T lymphocytes (CTL) with Salmonella Typhimurium antigens. Objective: To analyze the Salmonella-specific CTL response evoked by pDCs. Materials and methods: We used plasmacytoid dendritic cells stained with carboxyfluorescein succinimidyl ester (CFSE) and pulsed with OmpC73, Salmonella Kb restricted epitopes or S. Typhimurium as targets for cytotoxicity assays. Results: Specific lysis was shown to be statistically significant in pDC + OmpC73 for all effector:target ratios (p≤0.05). For pDC + S. Typhimurium, statistical significance was only observed at a 1:100 ratio (p≤0.05) using OmpC73. Conclusion: Plasmacytoid dendritic cells evoke CTL response during S. Typhimurium infection.


Assuntos
Células Dendríticas/imunologia , Salmonelose Animal/imunologia , Linfócitos T Citotóxicos/imunologia , Animais , Ilhas de CpG , Feminino , Antígeno de Histocompatibilidade H-2D/imunologia , Humanos , Imunidade Celular , Imunização , Camundongos , Camundongos Endogâmicos C57BL , Salmonella typhimurium
20.
Int J Med Microbiol ; 309(8): 151337, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31477487

RESUMO

Salmonella Enteritidis (SE) is a highly adapted pathogen causing severe economic losses in the poultry industry worldwide. Chickens infected by SE are a major source of human food poisoning. Vaccination is an effective approach to control SE infections. This study evaluated the immunogenicity and protective efficacy of a SE sptP deletion mutant (C50336ΔsptP) as a live attenuated vaccine (LAV) candidate in chickens. 14 day-old specific pathogen-free (SPF) chickens were intramuscularly immunized with various doses of C50336ΔsptP. Several groups of chickens were challenged with the virulent wild-type SE strain Z-11 via the same route at 14 days post vaccination. Compared to the control group, the groups vaccinated with 1 × 106, 1 × 107 and 1 × 108 colony-forming units (CFU) of C50336ΔsptP exhibited no clinical symptoms after immunization. Only slight pathological changes occurred in the organs of the 1 × 109 CFU vaccinated group. C50336ΔsptP bacteria were cleared from the organs of immunized chickens within 14 days after vaccination. Lymphocyte proliferation and serum cytokine analyses indicated that significant cellular immune responses were induced after the vaccination of C50336ΔsptP. Compared to the control group, specific IgG antibody levels increased significantly in vaccinated chickens, and the levels increased markedly after the challenge. The 1 × 107, 1 × 108, and 1 × 109 CFU vaccinated chickens groups showed no clinical symptoms or pathological changes, and no death after the lethal challenge. Whereas severe clinical signs of disease and pathological changes were observed in the control group chickens after the challenge. These results suggest that a single dose of C50336ΔsptP could be an effective LAV candidate to against SE infection in chickens.


Assuntos
Proteínas de Bactérias/genética , Imunogenicidade da Vacina , Doenças das Aves Domésticas/prevenção & controle , Salmonelose Animal/prevenção & controle , Vacinas contra Salmonella/imunologia , Deleção de Sequência , Animais , Anticorpos Antibacterianos/sangue , Proteínas de Bactérias/imunologia , Galinhas , Citocinas/sangue , Imunoglobulina G/sangue , Doenças das Aves Domésticas/microbiologia , Salmonelose Animal/imunologia , Salmonella enteritidis/genética , Salmonella enteritidis/imunologia , Organismos Livres de Patógenos Específicos , Vacinação/veterinária , Vacinas Atenuadas/imunologia
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