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1.
J Cell Sci ; 134(5)2021 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-33589501

RESUMO

Staphylococcus aureus infects ∼30% of the human population and causes a spectrum of pathologies ranging from mild skin infections to life-threatening invasive diseases. The strict host specificity of its virulence factors has severely limited the accuracy of in vivo models for the development of vaccines and therapeutics. To resolve this, we generated a humanised zebrafish model and determined that neutrophil-specific expression of the human C5a receptor conferred susceptibility to the S. aureus toxins PVL and HlgCB, leading to reduced neutrophil numbers at the site of infection and increased infection-associated mortality. These results show that humanised zebrafish provide a valuable platform to study the contribution of human-specific S. aureus virulence factors to infection in vivo that could facilitate the development of novel therapeutic approaches and essential vaccines.


Assuntos
Staphylococcus aureus , Fatores de Virulência , Animais , Humanos , Receptor da Anafilatoxina C5a/genética , Staphylococcus aureus/genética , Virulência , Fatores de Virulência/genética , Peixe-Zebra
2.
PLoS One ; 14(4): e0215592, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31002727

RESUMO

The neutrophil enzyme myeloperoxidase (MPO) is a major enzyme made by neutrophils to generate antimicrobial and immunomodulatory compounds, notably hypochlorous acid (HOCl), amplifying their capacity for destroying pathogens and regulating inflammation. Despite its roles in innate immunity, the importance of MPO in preventing infection is unclear, as individuals with MPO deficiency are asymptomatic with the exception of an increased risk of candidiasis. Dysregulation of MPO activity is also linked with inflammatory conditions such as atherosclerosis, emphasising a need to understand the roles of the enzyme in greater detail. Consequently, new tools for investigating granular dynamics in vivo can provide useful insights into how MPO localises within neutrophils, aiding understanding of its role in preventing and exacerbating disease. The zebrafish is a powerful model for investigating the immune system in vivo, as it is genetically tractable, and optically transparent. To visualise MPO activity within zebrafish neutrophils, we created a genetic construct that expresses human MPO as a fusion protein with a C-terminal fluorescent tag, driven by the neutrophil-specific promoter lyz. After introducing the construct into the zebrafish genome by Tol2 transgenesis, we established the Tg(lyz:Hsa.MPO-mEmerald,cmlc2:EGFP)sh496 line, and confirmed transgene expression in zebrafish neutrophils. We observed localisation of MPO-mEmerald within a subcellular location resembling neutrophil granules, mirroring MPO in human neutrophils. In Spotless (mpxNL144) larvae-which express a non-functional zebrafish myeloperoxidase-the MPO-mEmerald transgene does not disrupt neutrophil migration to sites of infection or inflammation, suggesting that it is a suitable line for the study of neutrophil granule function. We present a new transgenic line that can be used to investigate neutrophil granule dynamics in vivo without disrupting neutrophil behaviour, with potential applications in studying processing and maturation of MPO during development.


Assuntos
Proteínas de Fluorescência Verde/metabolismo , Proteínas Luminescentes/metabolismo , Neutrófilos/enzimologia , Peroxidase/metabolismo , Peixe-Zebra/metabolismo , Animais , Animais Geneticamente Modificados , Proteínas de Fluorescência Verde/genética , Humanos , Larva/genética , Larva/metabolismo , Proteínas Luminescentes/genética , Microscopia Confocal , Microscopia de Fluorescência , Peroxidase/genética , Transgenes/genética , Peixe-Zebra/genética , Proteína Vermelha Fluorescente
3.
Cell Rep ; 26(7): 1828-1840.e4, 2019 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-30759393

RESUMO

Infection by rapidly growing Mycobacterium abscessus is increasingly prevalent in cystic fibrosis (CF), a genetic disease caused by a defective CF transmembrane conductance regulator (CFTR). However, the potential link between a dysfunctional CFTR and vulnerability to M. abscessus infection remains unknown. Herein, we exploit a CFTR-depleted zebrafish model, recapitulating CF immuno-pathogenesis, to study the contribution of CFTR in innate immunity against M. abscessus infection. Loss of CFTR increases susceptibility to infection through impaired NADPH oxidase-dependent restriction of intracellular growth and reduced neutrophil chemotaxis, which together compromise granuloma formation and integrity. As a consequence, extracellular multiplication of M. abscessus expands rapidly, inducing abscess formation and causing lethal infections. Because these phenotypes are not observed with other mycobacteria, our findings highlight the crucial and specific role of CFTR in the immune control of M. abscessus by mounting effective oxidative responses.


Assuntos
Regulador de Condutância Transmembrana em Fibrose Cística/imunologia , Infecções por Mycobacterium não Tuberculosas/imunologia , Mycobacterium abscessus/imunologia , Estresse Oxidativo/imunologia , Proteínas de Peixe-Zebra/imunologia , Animais , Animais Geneticamente Modificados , Modelos Animais de Doenças , Infecções por Mycobacterium não Tuberculosas/microbiologia , Infecções por Mycobacterium não Tuberculosas/prevenção & controle , Mycobacterium abscessus/isolamento & purificação , Espécies Reativas de Oxigênio/imunologia , Peixe-Zebra
4.
J Immunol ; 202(2): 494-502, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30552162

RESUMO

Drug-resistant mycobacteria are a rising problem worldwide. There is an urgent need to understand the immune response to tuberculosis to identify host targets that, if targeted therapeutically, could be used to tackle these currently untreatable infections. In this study we use an Il-1ß fluorescent transgenic line to show that there is an early innate immune proinflammatory response to well-established zebrafish models of inflammation and Mycobacterium marinum infection. We demonstrate that host-derived hypoxia signaling, mediated by the Hif-1α transcription factor, can prime macrophages with increased levels of Il-1ß in the absence of infection, upregulating neutrophil antimicrobial NO production, leading to greater protection against infection. Our data link Hif-1α to proinflammatory macrophage Il-1ß transcription in vivo during early mycobacterial infection and importantly highlight a host protective mechanism, via antimicrobial NO, that decreases disease outcomes and that could be targeted therapeutically to stimulate the innate immune response to better deal with infections.


Assuntos
Hipóxia/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/imunologia , Infecções por Mycobacterium não Tuberculosas/imunologia , Mycobacterium marinum/fisiologia , Mycobacterium tuberculosis/fisiologia , Tuberculose/imunologia , Animais , Animais Geneticamente Modificados , Antituberculosos/metabolismo , Células Cultivadas , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Modelos Animais de Doenças , Técnicas de Inativação de Genes , Proteínas de Fluorescência Verde/genética , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Imunidade Inata , Interleucina-1beta/genética , Óxido Nítrico/metabolismo , Peixe-Zebra
5.
Blood ; 130(8): 1014-1025, 2017 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-28637666

RESUMO

The lifespan of neutrophils is plastic and highly responsive to factors that regulate cellular survival. Defects in neutrophil number and survival are common to both hematologic disorders and chronic inflammatory diseases. At sites of inflammation, neutrophils respond to multiple signals that activate protein kinase A (PKA) signaling, which positively regulates neutrophil survival. The aim of this study was to define transcriptional responses to PKA activation and to delineate the roles of these factors in neutrophil function and survival. In human neutrophil gene array studies, we show that PKA activation upregulates a significant number of apoptosis-related genes, the most highly regulated of these being NR4A2 and NR4A3 Direct PKA activation by the site-selective PKA agonist pair N6/8-AHA (8-AHA-cAMP and N6-MB-cAMP) and treatment with endogenous activators of PKA, including adenosine and prostaglandin E2, results in a profound delay of neutrophil apoptosis and concomitant upregulation of NR4A2/3 in a PKA-dependent manner. NR4A3 expression is also increased at sites of neutrophilic inflammation in a human model of intradermal inflammation. PKA activation also promotes survival of murine neutrophil progenitor cells, and small interfering RNA to NR4A2 decreases neutrophil production in this model. Antisense knockdown of NR4A2 and NR4A3 homologs in zebrafish larvae significantly reduces the absolute neutrophil number without affecting cellular migration. In summary, we show that NR4A2 and NR4A3 are components of a downstream transcriptional response to PKA activation in the neutrophil, and that they positively regulate neutrophil survival and homeostasis.


Assuntos
Neutrófilos/citologia , Neutrófilos/metabolismo , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Membro 3 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Peixe-Zebra/metabolismo , Animais , Contagem de Células , Proliferação de Células , Sobrevivência Celular , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Dinoprostona/metabolismo , Ativação Enzimática , Deleção de Genes , Técnicas de Silenciamento de Genes , Humanos , Inflamação/patologia , Larva/metabolismo , Camundongos , Família Multigênica , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Reprodutibilidade dos Testes , Transdução de Sinais , Transcrição Gênica
6.
Brain ; 140(4): 1128-1146, 2017 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-28334843

RESUMO

Mutations in the gene encoding tau (MAPT) cause frontotemporal dementia spectrum disorders. A rare tau variant p.A152T was reported as a risk factor for frontotemporal dementia spectrum and Alzheimer's disease in an initial case-control study. Such findings need replication in an independent cohort. We analysed an independent multinational cohort comprising 3100 patients with neurodegenerative disease and 4351 healthy control subjects and found p.A152T associated with significantly higher risk for clinically defined frontotemporal dementia and progressive supranuclear palsy syndrome. To assess the functional and biochemical consequences of this variant, we generated transgenic zebrafish models expressing wild-type or A152T-tau, where A152T caused neurodegeneration and proteasome compromise. Impaired proteasome activity may also enhance accumulation of other proteins associated with this variant. We increased A152T clearance kinetics by both pharmacological and genetic upregulation of autophagy and ameliorated the disease pathology observed in A152T-tau fish. Thus, autophagy-upregulating therapies may be a strategy for the treatment for tauopathies.


Assuntos
Autofagia , Transtornos Heredodegenerativos do Sistema Nervoso/genética , Transtornos Heredodegenerativos do Sistema Nervoso/terapia , Paralisia Supranuclear Progressiva/genética , Paralisia Supranuclear Progressiva/terapia , Tauopatias/genética , Tauopatias/terapia , Peixe-Zebra , Proteínas tau/genética , Alelos , Animais , Proteína 5 Relacionada à Autofagia , Comportamento Animal , Modelos Animais de Doenças , Embrião não Mamífero , Demência Frontotemporal/genética , Humanos , Cinética , Polimorfismo de Nucleotídeo Único , Complexo de Endopeptidases do Proteassoma/genética , RNA/biossíntese , RNA/genética , Tauopatias/psicologia , Proteínas de Peixe-Zebra , Proteínas tau/metabolismo
7.
Dis Model Mech ; 9(6): 621-32, 2016 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-27079522

RESUMO

Neutrophils are essential for host defence and are recruited to sites of inflammation in response to tissue injury or infection. For inflammation to resolve, these cells must be cleared efficiently and in a controlled manner, either by apoptosis or reverse migration. If the inflammatory response is not well-regulated, persistent neutrophils can cause damage to host tissues and contribute to the pathogenesis of chronic inflammatory diseases, which respond poorly to current treatments. It is therefore important to develop drug discovery strategies that can identify new therapeutics specifically targeting neutrophils, either by promoting their clearance or by preventing their recruitment. Our recent in vivo chemical genetic screen for accelerators of inflammation resolution identified a subset of compounds sharing a common chemical signature, the bicyclic benzopyrone rings. Here, we further investigate the mechanisms of action of the most active of this chemical series, isopimpinellin, in our zebrafish model of neutrophilic inflammation. We found that this compound targets both the recruitment and resolution phases of the inflammatory response. Neutrophil migration towards a site of injury is reduced by isopimpinellin and this occurs as a result of PI3K inhibition. We also show that isopimpinellin induces neutrophil apoptosis to drive inflammation resolution in vivo using a new zebrafish reporter line detecting in vivo neutrophil caspase-3 activity and allowing quantification of flux through the apoptotic pathway in real time. Finally, our studies reveal that clinically available 'cromones' are structurally related to isopimpinellin and have previously undescribed pro-resolution activity in vivo These findings could have implications for the therapeutic use of benzopyrones in inflammatory disease.


Assuntos
Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Cumarínicos/química , Cumarínicos/farmacologia , Avaliação Pré-Clínica de Medicamentos , Peixe-Zebra/metabolismo , Animais , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Cromolina Sódica/química , Cromolina Sódica/farmacologia , Furocumarinas/química , Furocumarinas/farmacologia , Inflamação/patologia , Infiltração de Neutrófilos/efeitos dos fármacos , Neutrófilos/citologia , Neutrófilos/efeitos dos fármacos , Fenótipo , Fosfatidilinositol 3-Quinases/metabolismo , Relação Estrutura-Atividade
8.
Sci Rep ; 6: 21489, 2016 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-26887656

RESUMO

Cryptococcus neoformans is a significant fungal pathogen of immunocompromised patients. Many questions remain regarding the function of macrophages in normal clearance of cryptococcal infection and the defects present in uncontrolled cryptococcosis. Two current limitations are: 1) The difficulties in interpreting studies using isolated macrophages in the context of the progression of infection, and 2) The use of high resolution imaging in understanding immune cell behavior during animal infection. Here we describe a high-content imaging method in a zebrafish model of cryptococcosis that permits the detailed analysis of macrophage interactions with C. neoformans during infection. Using this approach we demonstrate that, while macrophages are critical for control of C. neoformans, a failure of macrophage response is not the limiting defect in fatal infections. We find phagocytosis is restrained very early in infection and that increases in cryptococcal number are driven by intracellular proliferation. We show that macrophages preferentially phagocytose cryptococci with smaller polysaccharide capsules and that capsule size is greatly increased over twenty-four hours of infection, a change that is sufficient to severely limit further phagocytosis. Thus, high-content imaging of cryptococcal infection in vivo demonstrates how very early interactions between macrophages and cryptococci are critical in the outcome of cryptococcosis.


Assuntos
Proliferação de Células , Criptococose/metabolismo , Cryptococcus neoformans/metabolismo , Doenças dos Peixes/metabolismo , Macrófagos/metabolismo , Peixe-Zebra/metabolismo , Animais , Criptococose/patologia , Doenças dos Peixes/microbiologia , Macrófagos/microbiologia , Organismos Geneticamente Modificados/metabolismo , Peixe-Zebra/microbiologia
9.
J Leukoc Biol ; 98(6): 975-81, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26071567

RESUMO

Stimulation of neutrophil reverse migration presents an attractive, alternative therapeutic pathway to driving inflammation resolution. However, little is known about whether the activity of wound-experienced neutrophils is altered and whether encouraging dispersal of such neutrophils back into the body may have undesirable consequences. This study used a zebrafish tail transection inflammation model, in combination with a photoconvertible neutrophil transgenic line, to allow internally controlled, simultaneous comparison of reverse-migrated neutrophils with naïve neutrophils in the presence and absence of secondary insult. Detailed microscopy revealed that reverse-migrated neutrophils exhibited an activated morphology but responded normally to secondary insult and are able to mount an effective antimicrobial response to Staphylococcus aureus. These results support a model in which reverse-migrated neutrophils exhibit no long-term behavioral alterations and encourage the notion of enhanced reverse migration as a viable target for pharmaceutical manipulation.


Assuntos
Movimento Celular/imunologia , Neutrófilos/imunologia , Peixe-Zebra/imunologia , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/imunologia , Movimento Celular/genética , Inflamação/genética , Inflamação/imunologia , Staphylococcus aureus/imunologia , Peixe-Zebra/genética
10.
Science ; 346(6209): 641-646, 2014 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-25359976

RESUMO

Many key components of innate immunity to infection are shared between Drosophila and humans. However, the fly Toll ligand Spaetzle is not thought to have a vertebrate equivalent. We have found that the structurally related cystine-knot protein, nerve growth factor ß (NGFß), plays an unexpected Spaetzle-like role in immunity to Staphylococcus aureus infection in chordates. Deleterious mutations of either human NGFß or its high-affinity receptor tropomyosin-related kinase receptor A (TRKA) were associated with severe S. aureus infections. NGFß was released by macrophages in response to S. aureus exoproteins through activation of the NOD-like receptors NLRP3 and NLRP4 and enhanced phagocytosis and superoxide-dependent killing, stimulated proinflammatory cytokine production, and promoted calcium-dependent neutrophil recruitment. TrkA knockdown in zebrafish increased susceptibility to S. aureus infection, confirming an evolutionarily conserved role for NGFß-TRKA signaling in pathogen-specific host immunity.


Assuntos
Fator de Crescimento Neural/imunologia , Receptor trkA/imunologia , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/imunologia , Animais , Drosophila melanogaster/genética , Drosophila melanogaster/imunologia , Evolução Molecular , Técnicas de Silenciamento de Genes , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Macrófagos/imunologia , Fator de Crescimento Neural/genética , Fagocitose/genética , Fagocitose/imunologia , Receptor trkA/genética , Infecções Estafilocócicas/genética , Peixe-Zebra/genética , Peixe-Zebra/imunologia
11.
Dev Comp Immunol ; 46(1): 53-62, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24690566

RESUMO

Inflammatory diseases are a significant burden on global healthcare systems, and tackling these diseases is a major focus of modern medicine. Key to many inflammatory diseases is the cytokine, Interleukin-1 (IL-1). Due to its apical role in initiating the inflammatory response, dysregulated IL-1 signalling results in a number of pathologies. Treatment of inflammatory diseases with anti-IL-1 therapies has offered many therapeutic benefits, however current therapies are protein based, with all the accompanying limitations. The non-conventional pathways involved in IL-1 signalling provide a number of potential therapeutic targets for clinical intervention and this has led to the exploitation of a number of model organisms for the study of IL-1 biology. Murine models have long been used to study IL-1 processing and release, but do not allow direct visualisation in vivo. Recently, fish models have emerged as genetically tractable and optically transparent inflammatory disease models. These models have raised questions on the evolutionary origins of the IL-1 family and the conservation in its processing and activation. Here we review the current understanding of IL-1 evolution in fish and discuss the study of IL-1 processing in these models.


Assuntos
Evolução Biológica , Proteínas de Peixes/imunologia , Peixes/genética , Peixes/imunologia , Inflamassomos/genética , Interleucina-1/imunologia , Animais , Caspase 1/metabolismo , Modelos Animais de Doenças , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Inflamassomos/imunologia , Interleucina-1/química , Interleucina-1/genética , Interleucina-1/metabolismo , Filogenia
12.
Dis Model Mech ; 7(2): 259-64, 2014 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-24203886

RESUMO

Interleukin-1 (IL-1), the 'gatekeeper' of inflammation, is the apical cytokine in a signalling cascade that drives the early response to injury or infection. Expression, processing and secretion of IL-1 are tightly controlled, and dysregulated IL-1 signalling has been implicated in a number of pathologies ranging from atherosclerosis to complications of infection. Our understanding of these processes comes from in vitro monocytic cell culture models as lines or primary isolates, in which a range and spectra of IL-1 secretion mechanisms have been described. We therefore investigated whether zebrafish embryos provide a suitable in vivo model for studying IL-1-mediated inflammation. Structurally, zebrafish IL-1ß shares a ß-sheet-rich trefoil structure with its human counterpart. Functionally, leukocyte expression of IL-1ß was detectable only following injury, which activated leukocytes throughout zebrafish embryos. Migration of macrophages and neutrophils was attenuated by inhibitors of either caspase-1 or P2X7, which similarly inhibited the activation of NF-κB at the site of injury. Zebrafish offer a new and versatile model to study the IL-1ß pathway in inflammatory disease and should offer unique insights into IL-1 biology in vivo.


Assuntos
Caspase 1/metabolismo , Inflamação/patologia , Interleucina-1beta/metabolismo , Regulação para Cima , Peixe-Zebra/metabolismo , 1-(5-Isoquinolinasulfonil)-2-Metilpiperazina/análogos & derivados , 1-(5-Isoquinolinasulfonil)-2-Metilpiperazina/farmacologia , Nadadeiras de Animais/efeitos dos fármacos , Nadadeiras de Animais/patologia , Animais , Inibidores de Caspase/farmacologia , Sequência Conservada , Regulação para Baixo/efeitos dos fármacos , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Embrião não Mamífero/patologia , Humanos , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , NF-kappa B/metabolismo , Corantes de Rosanilina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos , Peixe-Zebra/embriologia
13.
Dis Model Mech ; 7(1): 163-9, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24291762

RESUMO

Cell migration is fundamental to the inflammatory response, but uncontrolled cell migration and excess recruitment of neutrophils and other leukocytes can cause damage to the tissue. Here we describe the use of an in vivo model - the Tg(mpx:GFP)(i114) zebrafish line, in which neutrophils are labelled by green fluorescent protein (GFP) - to screen a natural product library for compounds that can affect neutrophil migratory behaviour. Among 1040 fungal extracts screened, two were found to inhibit neutrophil migration completely. Subfractionation of these extracts identified sterigmatocystin and antibiotic PF1052 as the active components. Using the EZ-TAXIScan chemotaxis assay, both compounds were also found to have a dosage-dependent inhibitory effect on murine neutrophil migration. Furthermore, neutrophils treated with PF1052 failed to form pseudopods and appeared round in shape, suggesting a defect in PI3-kinase (PI3K) signalling. We generated a transgenic neutrophil-specific PtdIns(3,4,5)P3 (PIP3) reporter zebrafish line, which revealed that PF1052 does not affect the activation of PI3K at the plasma membrane. In human neutrophils, PF1052 neither induced apoptosis nor blocked AKT phosphorylation. In conclusion, we have identified an antibiotic from a natural product library with potent anti-inflammatory properties, and have established the utility of the mpx:GFP transgenic zebrafish for high-throughput in vivo screens for novel inhibitors of neutrophil migration.


Assuntos
Antibacterianos/farmacologia , Quimiotaxia/efeitos dos fármacos , Naftalenos/farmacologia , Neutrófilos/citologia , Pirrolidinonas/farmacologia , Esterigmatocistina/farmacologia , Animais , Animais Geneticamente Modificados , Apoptose , Membrana Celular/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Humanos , Inflamação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Infiltração de Neutrófilos , Neutrófilos/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Frações Subcelulares , Transgenes , Peixe-Zebra
14.
Infect Immun ; 81(11): 4271-9, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24002065

RESUMO

Enterococcus faecalis is an opportunistic pathogen responsible for a wide range of life-threatening nosocomial infections, such as septicemia, peritonitis, and endocarditis. E. faecalis infections are associated with a high mortality and substantial health care costs and cause therapeutic problems due to the intrinsic resistance of this bacterium to antibiotics. Several factors contributing to E. faecalis virulence have been identified. Due to the variety of infections caused by this organism, numerous animal models have been used to mimic E. faecalis infections, but none of them is considered ideal for monitoring pathogenesis. Here, we studied for the first time E. faecalis pathogenesis in zebrafish larvae. Using model strains, chosen isogenic mutants, and fluorescent derivatives expressing green fluorescent protein (GFP), we analyzed both lethality and bacterial dissemination in infected larvae. Genetically engineered immunocompromised zebrafish allowed the identification of two critical steps for successful establishment of disease: (i) host phagocytosis evasion mediated by the Epa rhamnopolysaccharide and (ii) tissue damage mediated by the quorum-sensing Fsr regulon. Our results reveal that the zebrafish is a novel, powerful model for studying E. faecalis pathogenesis, enabling us to dissect the mechanism of enterococcal virulence.


Assuntos
Modelos Animais de Doenças , Enterococcus faecalis/patogenicidade , Infecções por Bactérias Gram-Positivas/microbiologia , Infecções por Bactérias Gram-Positivas/patologia , Peixe-Zebra/microbiologia , Animais , Animais Geneticamente Modificados , Evasão da Resposta Imune , Hospedeiro Imunocomprometido , Infecções Oportunistas/microbiologia , Infecções Oportunistas/patologia , Análise de Sobrevida , Virulência , Imagem Corporal Total
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