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1.
J Microbiol Biotechnol ; 29(8): 1324-1334, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31370117

RESUMO

Fish mycobacteriosis is a common bacterial disease in many species of freshwater and marine fish and has caused severe loss of fish production. Mycobacterium marinum has been the most prevalent pathogen observed in several outbreaks of mycobacteriosis of farmed sturgeons in China. However, the immune responses and pathology of sturgeons in mycobacterial infection are rarely studied. Therefore, we used the Illumina RNA-seq method to analyze the transcriptome profile of Acipenser schrenckii challenged with Mycobacterium marinum. To begin, 168,220 non-redundant contigs were acquired from the infection and control groups, and among these, 33,225 contigs have acquired annotations. A total of 4,043 differently expressed (DE) contigs between the two groups were identified, and among these, 2479 were upregulated and 1564 were down-regulated in the infected fish. A total of 1,340 DE contigs with acquired annotations in KEGG were enriched for 124 pathways including the TNF signaling pathway, and the Toll-like receptor signaling pathway. The roles of DE genes involved in significant pathways and other processes were discussed. The 2,209 DE contigs that have yet to acquire proper annotation may represent candidate genes associated with infection in sturgeons and are expected to serve as immunogenetic resources for further study. To our best knowledge, this is the first transcriptome study on sturgeons under bacterial infection.


Assuntos
Doenças dos Peixes/genética , Doenças dos Peixes/imunologia , Proteínas de Peixes/genética , Peixes/genética , Infecções por Micobactéria não Tuberculosa/genética , Infecções por Micobactéria não Tuberculosa/imunologia , Infecções por Micobactéria não Tuberculosa/veterinária , Transcriptoma , Animais , China , Regulação para Baixo , Doenças dos Peixes/microbiologia , Proteínas de Peixes/metabolismo , Perfilação da Expressão Gênica , Imunidade , Anotação de Sequência Molecular , Mycobacterium marinum/patogenicidade , Regulação para Cima
2.
Int J Med Microbiol ; 309(5): 307-318, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31178418

RESUMO

Mycobacterium abscessus (MAB) is an emerging, rapidly growing non-tuberculous Mycobacterium causing therapy-resistant pulmonary disease especially in patients with cystic fibrosis (CF). Smooth and rough colony type MAB can be isolated from infected patients whereby rough colony type MAB are more often associated with severe disease. Disease severity is also associated with an alternated type I interferon (IFN-I) response of the MAB-infected patients. However the relevance of this response for the outcome of MAB infection is still unknown. In this study, we analyzed the IFNß expression of murine macrophages infected with a MAB rough colony strain (MAB-R) isolated from a patient with progressive CF and compared it to macrophages infected with the MAB smooth colony type reference strain (MAB-S). We found that MAB-R infected macrophages expressed significantly more IFNß mRNA and protein than MAB-S infected macrophages. Higher IFNß induction by MAB-R was associated with higher TNF expression and intracellular killing while low IFNß induction was associated with lower TNF expression and persistence of MAB-S. IFNß induction was independent of the intracellular cGAS-STING recognition pathway. MAB appeared to be recognized extracellularly and induced IFNß expression via TLR2-TLR4-MyD88-TRIF-IRF3 dependent pathways. By using macrophages lacking the IFN-I receptor we demonstrate that MAB induced IFN-I response essentially contributed to restricting MAB-R and MAB-S infections by activating macrophage Nos2 expression and nitric oxide production. Thus IFN-I seem to influence the intrinsic ability of macrophages to control MAB infections. As MAB persists over long time periods in susceptible patients, our findings suggest that virulence of MAB strains is promoted by an insufficient IFN-I response of the host.


Assuntos
Interferon beta/imunologia , Macrófagos/microbiologia , Mycobacterium abscessus/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Óxido Nítrico/imunologia , Receptor 2 Toll-Like/imunologia , Receptor 4 Toll-Like/imunologia , Animais , Interações Hospedeiro-Patógeno/imunologia , Humanos , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Infecções por Micobactéria não Tuberculosa/imunologia , Infecções por Micobactéria não Tuberculosa/microbiologia , Óxido Nítrico Sintase Tipo II/genética , Escarro/microbiologia
3.
Mayo Clin Proc ; 94(8): 1567-1581, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31160063

RESUMO

Nontuberculous mycobacteria (NTM) comprise a heterogeneous group of organisms, with only a small subset known to cause disease in humans. Although NTM infection is not a reportable disease, both the increasing clinical recognition and recent advancements in laboratory diagnostic capabilities of NTM infections in immunocompromised and immunocompetent patients are rapidly evolving. We reviewed antimicrobial agents used to treat the most frequently encountered NTM infections and examined optimized drug dosing strategies, toxicity profiles, drug-drug interactions, and the role of therapeutic drug monitoring. Antimicrobial susceptibility testing and patient monitoring on therapy were also examined. We used PubMed to review the published literature on the management of select NTM pathogens, the common syndromes encountered since 2000, and select pharmacokinetic principles of select antimicrobial agents used since 1990. We included select clinical trials, systematic reviews, published guidelines, and observational studies when applicable. The prolonged duration and the necessity for combination therapy for most forms of NTM disease can be problematic for many patients. A multidisciplinary care team that includes pharmacy engagement may help increase rates of optimal patient tolerability and successful treatment completion.


Assuntos
Antibacterianos/uso terapêutico , Infecções por Micobactéria não Tuberculosa/tratamento farmacológico , Infecções por Micobactéria não Tuberculosa/epidemiologia , Micobactérias não Tuberculosas/efeitos dos fármacos , Monitoramento de Medicamentos , Feminino , Seguimentos , Humanos , Imunocompetência/imunologia , Hospedeiro Imunocomprometido/imunologia , Incidência , Comunicação Interdisciplinar , Masculino , Dose Máxima Tolerável , Testes de Sensibilidade Microbiana , Infecções por Micobactéria não Tuberculosa/diagnóstico , Infecções por Micobactéria não Tuberculosa/imunologia , Micobactérias não Tuberculosas/isolamento & purificação , Segurança do Paciente , Medição de Risco
4.
BMC Infect Dis ; 19(1): 454, 2019 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-31117976

RESUMO

BACKGROUND: Mycolicibacterium phlei (M. phlei) is known to be a non-pathogenic nontuberculous mycobacterium (NTM) which rarely causes diseases in humans. A disseminated NTM infection is mostly caused by the Mycobacterium avium complex (MAC) and is known to develop in immunocompromised hosts, like those with acquired immune deficiency syndrome (AIDS). Here, we report a case of disseminated M. phlei infection in an immunocompetent host carrying anti-interferon gamma (IFN-γ) autoantibodies. CASE PRESENTATION: We detected M. phlei in multiple organs of an elderly woman with no significant medical history except positivity for anti-IFN-γ autoantibodies. She tested negative for human immunodeficiency virus (HIV)-1, 2/ Human T-cell leukemia virus type 1 (HTLV-1) antibody. High-resolution computed tomography (HRCT) of the chest demonstrated a nodule in the left S1 + 2 segment, interlobular septal thickening, multi lymphadenopathy, and osteolysis. A maximum intensity projection image following fluorodeoxyglucose-positron emission tomography (FDG-PET) revealed multifocal hypermetabolic lesions in the nodule and all the swollen lymph nodes seen in HRCT. FDG also accumulated in multiple bones. Advanced primary lung cancer was suspected, and biopsies of each lesion were performed. The pathology revealed caseating granuloma, positive for acid-fast bacteria, and DNA sequencing of the acid-fast bacteria confirmed the organism to be M. phlei. The patient also tested positive for anti-IFN-γ autoantibodies. Based on these findings, she was diagnosed with disseminated M. phlei infection, with anti-IFN-γ autoantibodies. CONCLUSIONS: Though known to be non-pathogenic, we show that M. phlei can be pathogenic like the MAC in immunocompetent individuals carrying anti-IFN-γ autoantibodies.


Assuntos
Interferon gama/imunologia , Infecções por Micobactéria não Tuberculosa/imunologia , Micobactérias não Tuberculosas/patogenicidade , Idoso , Antibacterianos/uso terapêutico , Autoanticorpos , Osso e Ossos/diagnóstico por imagem , Osso e Ossos/patologia , Feminino , Granuloma/diagnóstico por imagem , Granuloma/tratamento farmacológico , Granuloma/microbiologia , Humanos , Imunocompetência , Infecções por Micobactéria não Tuberculosa/diagnóstico por imagem , Infecções por Micobactéria não Tuberculosa/tratamento farmacológico , Infecções por Micobactéria não Tuberculosa/microbiologia , Micobactérias não Tuberculosas/imunologia , Tomografia por Emissão de Pósitrons/métodos , Tomografia Computadorizada por Raios X/métodos , Virulência
6.
Front Immunol ; 10: 125, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30766538

RESUMO

Mycobacterium abscessus complex (MAB) is a rapidly growing mycobacterium(RGM) whose clinical significance as an emerging human pathogen has been increasing worldwide. It has two types of colony morphology, a smooth (S) type, producing high glycopeptidolipid (GPL) content, and a rough (R) type, which produces low levels of GPLs and is associated with increased virulence. However, the mechanism responsible for their difference in virulence is poorly known. By ultrastructural examination of murine macrophages infected, we found that MAB-R strains could replicate more actively in the macrophage phagosome than the S variants and that they could escape into cytosol via phagosomal rupture. The cytosolic access of MAB-R strains via phagosomal rupture led to enhanced Type I interferon (IFN) production and cell death, which resulted in their cell-to-cell spreading. This behavior can provide an additional niche for the survival of MAB-R strains. In addition, we found that their enhancement of cell death mediated cell spreading are dependent on Type I IFN signaling via comparison of wild-type and IFNAR1 knockout mice. In conclusion, our data indicated that a transition of MAB-S strains into MAB-R variants increased their virulence via enhanced Type I IFN production, which led to enhanced survival in infected macrophage via cell death mediated cell-to-cell spreading. This result provides not only a novel insight into the difference in virulence between MAB-R and -S variants but also hints to their treatment strategy.


Assuntos
Interferon Tipo I/metabolismo , Macrófagos/imunologia , Infecções por Micobactéria não Tuberculosa/imunologia , Mycobacterium abscessus/fisiologia , Fagossomos/ultraestrutura , Animais , Morte Celular , Linhagem Celular , Humanos , Evasão da Resposta Imune , Macrófagos/microbiologia , Macrófagos/ultraestrutura , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infecções por Micobactéria não Tuberculosa/transmissão , Mycobacterium abscessus/patogenicidade , Fagossomos/microbiologia , Receptor de Interferon alfa e beta/genética , Transdução de Sinais , Especificidade da Espécie , Virulência
7.
Microb Pathog ; 128: 405-413, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30685363

RESUMO

Mycolyl-arabinogalactan-peptidoglycan (mAGP) is the major content of the mycobacterium cell wall structure and essential for mycobacterial survival. Peptidoglycan (PG) plays an important role in maintenance of cell division, cell wall integrity and pathogenesis. Mycobacterium smegmatis MSMEG_6281, a peptidoglycan amidase, is vital for mycobacterial cell division. However, the effects of MSMEG_6281on cell wall integrity and mycobacterial virulence remain unknown. In the current study, we demonstrate that MSMEG_6281gene knockout in M.smegmatis alters the microbiological characteristics. Our results revealed that MSMEG_6281gene knockout bacteria (M. sm-ΔM_6281) lost their acid-fastness, increased their sensitivity to lipophilic compounds and presented an abnormal morphology. Our results revealed that MSMEG_6281was related to maintaining the cell wall integrity. Furthermore, we investigated the effects of MSMEG_6281 inactivation on mycobacterial virulence using mice models infected by different M.smegmatis strains. MSMEG_6281 inactivation in the M sm-ΔM_6281 infected group caused less mycobacterial colonization, reduced pathological signs, decreased the anti-microbial enzymes production including iNOS and ß-defensins in mouse lungs. Moreover, IL-1ß and TLR2 expression were significantly down-regulated, while the production of IFN-γ and TNF-α was up-regulated. These findings indicated the diversity of host immune responses induced by different strains of M.smegmatis, suggesting that MSMEG_6281 inactivation impact mycobacterial virulence. In conclusion, the MSMEG_6281 protein plays important roles in maintaining cell wall integrity and mycobacterial virulence.


Assuntos
Amidoidrolases/metabolismo , Parede Celular/metabolismo , Mycobacterium smegmatis/enzimologia , Mycobacterium smegmatis/metabolismo , Peptidoglicano/metabolismo , Amidoidrolases/genética , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Divisão Celular , Parede Celular/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Técnicas de Inativação de Genes , Genes Bacterianos/genética , Recombinação Homóloga , Interleucina-1beta/metabolismo , Pulmão/microbiologia , Pulmão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Infecções por Micobactéria não Tuberculosa/imunologia , Infecções por Micobactéria não Tuberculosa/patologia , Mycobacterium smegmatis/genética , Mycobacterium smegmatis/patogenicidade , Óxido Nítrico Sintase Tipo II/metabolismo , Fragmentos de Peptídeos/metabolismo , Receptor 2 Toll-Like/metabolismo , Virulência , beta-Defensinas/metabolismo
9.
Dev Comp Immunol ; 91: 93-100, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30385316

RESUMO

In this study, we investigated the immune responses against Mycobacterium gordonae in ginbuna crucian carp. Cumulative mortality of ginbuna injected with 2.0 × 107 CFU of M. gordonae was 50% at 170 days post-infection. CD4-1, CD8α, T-bet and IFNγ2 gene expression levels were significantly upregulated in ginbuna injected with 1.9 × 108 CFU of M. gordonae at 21 and 28 days post-infection. The CD4-2 level did not change during the experiment. Granulomatous responses consisted of central macrophage accumulation and surrounding lymphocytes, and Ziehl-Neelsen-positive bacteria were observed in the trunk kidney of the challenged fish. Immunohistochemistry using anti-ginbuna IFNγs and anti-ginbuna CD4-1 polyclonal antibody revealed that the marginal lymphocytes were positive for CD4-1, and the IFNγ-producing cells surrounded the mycobacterial cell-laden phagocytes. These results suggest that CD4-1+ cells and IFNγ2 play important roles in the granulomatous inflammation against Mycobacterial infections in teleosts.


Assuntos
Doenças dos Peixes/imunologia , Carpa Dourada/imunologia , Granuloma/imunologia , Inflamação/imunologia , Linfócitos/imunologia , Macrófagos/imunologia , Infecções por Micobactéria não Tuberculosa/imunologia , Micobactérias não Tuberculosas/fisiologia , Animais , Antígenos CD4/metabolismo , Antígenos CD8/metabolismo , Proteínas de Peixes/metabolismo , Imuno-Histoquímica , Interferon gama/metabolismo , Proteínas com Domínio T/metabolismo
11.
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 Micobactéria não Tuberculosa/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
12.
Sci Immunol ; 3(30)2018 12 21.
Artigo em Inglês | MEDLINE | ID: mdl-30578351

RESUMO

Hundreds of patients with autosomal recessive, complete IL-12p40 or IL-12Rß1 deficiency have been diagnosed over the last 20 years. They typically suffer from invasive mycobacteriosis and, occasionally, from mucocutaneous candidiasis. Susceptibility to these infections is thought to be due to impairments of IL-12-dependent IFN-γ immunity and IL-23-dependent IL-17A/IL-17F immunity, respectively. We report here patients with autosomal recessive, complete IL-12Rß2 or IL-23R deficiency, lacking responses to IL-12 or IL-23 only, all of whom, unexpectedly, display mycobacteriosis without candidiasis. We show that αß T, γδ T, B, NK, ILC1, and ILC2 cells from healthy donors preferentially produce IFN-γ in response to IL-12, whereas NKT cells and MAIT cells preferentially produce IFN-γ in response to IL-23. We also show that the development of IFN-γ-producing CD4+ T cells, including, in particular, mycobacterium-specific TH1* cells (CD45RA-CCR6+), is dependent on both IL-12 and IL-23. Last, we show that IL12RB1, IL12RB2, and IL23R have similar frequencies of deleterious variants in the general population. The comparative rarity of symptomatic patients with IL-12Rß2 or IL-23R deficiency, relative to IL-12Rß1 deficiency, is, therefore, due to lower clinical penetrance. There are fewer symptomatic IL-23R- and IL-12Rß2-deficient than IL-12Rß1-deficient patients, not because these genetic disorders are rarer, but because the isolated absence of IL-12 or IL-23 is, in part, compensated by the other cytokine for the production of IFN-γ, thereby providing some protection against mycobacteria. These experiments of nature show that human IL-12 and IL-23 are both required for optimal IFN-γ-dependent immunity to mycobacteria, both individually and much more so cooperatively.


Assuntos
Imunidade Inata/imunologia , Interferon gama/imunologia , Interleucina-12/imunologia , Interleucina-23/imunologia , Infecções por Micobactéria não Tuberculosa/imunologia , Mycobacterium/imunologia , Humanos , Interleucina-12/deficiência , Interleucina-12/genética , Interleucina-23/deficiência , Interleucina-23/genética , Linhagem
13.
J Vis Exp ; (140)2018 10 30.
Artigo em Inglês | MEDLINE | ID: mdl-30451232

RESUMO

The interest in DNA-based vaccination has increased during the past two decades. DNA vaccination is based on the cloning of a sequence of a selected antigen or a combination of antigens into a plasmid, which enables a tailor-made and safe design. The administration of DNA vaccines into host cells leads to the expression of antigens that stimulate both humoral and cell-mediated immune responses. This report describes a protocol for the cloning of antigen sequences into the pCMV-EGFP plasmid, the immunization of adult zebrafish with the vaccine candidates by intramuscular microinjection, and the subsequent electroporation to improve intake. The vaccine antigens are expressed as green fluorescent protein (GFP)-fusion proteins, which allows the confirmation of the antigen expression under UV light from live fish and the quantification of expression levels of the fusion protein with ELISA, as well as their detection with a western blot analysis. The protective effect of the vaccine candidates is tested by infecting the fish with Mycobacterium marinum five weeks postvaccination, followed by the quantification of the bacteria with qPCR four weeks later. Compared to mammalian preclinical screening models, this method provides a cost-effective method for the preliminary screening of novel DNA-based vaccine candidates against a mycobacterial infection. The method can be further applied to screening DNA-based vaccines against various bacterial and viral diseases.


Assuntos
Vacinas Bacterianas/imunologia , Infecções por Micobactéria não Tuberculosa/prevenção & controle , Mycobacterium marinum/imunologia , Vacinas de DNA/imunologia , Peixe-Zebra/imunologia , Animais , Vacinas Bacterianas/administração & dosagem , Modelos Animais de Doenças , Proteínas de Fluorescência Verde/imunologia , Imunidade Celular , Infecções por Micobactéria não Tuberculosa/imunologia , Infecções por Micobactéria não Tuberculosa/microbiologia , Vacinação , Vacinas de DNA/administração & dosagem , Peixe-Zebra/microbiologia
14.
Cell Host Microbe ; 24(4): 514-525.e6, 2018 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-30308157

RESUMO

Mycobacterial infection leads to the formation of characteristic immune aggregates called granulomas, a process accompanied by dramatic remodeling of the host vasculature. As granuloma angiogenesis favors the infecting mycobacteria, it may be actively promoted by bacterial determinants during infection. Using Mycobacterium marinum-infected zebrafish as a model, we identify the enzyme proximal cyclopropane synthase of alpha-mycolates (PcaA) as an important bacterial determinant of granuloma-associated angiogenesis. cis-Cyclopropanation of mycobacterial mycolic acids by pcaA drives the activation of host Vegf signaling within granuloma macrophages. Cyclopropanation of the mycobacterial cell wall glycolipid trehalose dimycolate is both required and sufficient to induce robust host angiogenesis. Inducible genetic inhibition of angiogenesis and Vegf signaling during granuloma formation results in bacterial growth deficits. Together, these data reveal a mechanism by which PcaA-mediated cis-cyclopropanation of mycolic acids promotes bacterial growth and dissemination in vivo by eliciting granuloma vascularization and suggest potential approaches for host-directed therapies.


Assuntos
Proteínas de Bactérias/metabolismo , Metiltransferases/metabolismo , Mycobacterium marinum/enzimologia , Neovascularização Patológica/microbiologia , Receptores de Fatores de Crescimento do Endotélio Vascular/metabolismo , Tuberculoma/microbiologia , Inibidores da Angiogênese/farmacologia , Animais , Proteínas de Bactérias/genética , Fatores Corda/metabolismo , Modelos Animais de Doenças , Humanos , Macrófagos/imunologia , Macrófagos/microbiologia , Metiltransferases/genética , Infecções por Micobactéria não Tuberculosa/imunologia , Infecções por Micobactéria não Tuberculosa/microbiologia , Mycobacterium marinum/genética , Mycobacterium marinum/patogenicidade , Ácidos Micólicos/metabolismo , Neovascularização Patológica/imunologia , Neovascularização Patológica/patologia , Pirimidinas/farmacologia , Receptores de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptores de Fatores de Crescimento do Endotélio Vascular/efeitos dos fármacos , Transdução de Sinais , Sulfonamidas/farmacologia , Tuberculoma/imunologia , Tuberculoma/patologia , Peixe-Zebra
15.
J Vis Exp ; (140)2018 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-30346391

RESUMO

Mycobacterium tuberculosis is currently the deadliest human pathogen causing 1.7 million deaths and 10.4 million infections every year. Exposure to this bacterium causes a wide disease spectrum in humans ranging from a sterilized infection to an actively progressing deadly disease. The most common form is the latent tuberculosis, which is asymptomatic, but has the potential to reactivate into a fulminant disease. Adult zebrafish and its natural pathogen Mycobacterium marinum have recently proven to be an applicable model to study the wide disease spectrum of tuberculosis. Importantly, spontaneous latency and reactivation as well as adaptive immune responses in the context of mycobacterial infection can be studied in this model. In this article, we describe methods for the experimental infection of adult zebrafish, the collection of internal organs for the extraction of nucleic acids for the measurement of mycobacterial loads and host immune responses by quantitative PCR. The in-house-developed, M. marinum-specific qPCR assay is more sensitive than the traditional plating methods as it also detects DNA from non-dividing, dormant or recently dead mycobacteria. As both DNA and RNA are extracted from the same individual, it is possible to study the relationships between the diseased state, and the host and pathogen gene-expression. The adult zebrafish model for tuberculosis thus presents itself as a highly applicable, non-mammalian in vivo system to study host-pathogen interactions.


Assuntos
Modelos Animais de Doenças , Infecções por Micobactéria não Tuberculosa/imunologia , Infecções por Micobactéria não Tuberculosa/microbiologia , Mycobacterium marinum/fisiologia , Peixe-Zebra , Animais , Expressão Gênica , Interações Hospedeiro-Patógeno , Infecções por Micobactéria não Tuberculosa/genética , Mycobacterium marinum/genética , Reação em Cadeia da Polimerase em Tempo Real
16.
Int J Infect Dis ; 76: 35-37, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30201506

RESUMO

A 75-year-old woman with a history of immunosuppressive treatment for rheumatoid arthritis and non-Hodgkin lymphoma, was referred to our reference centre for treatment of tenosynovitis caused by Mycobacterium malmoense, which had disseminated due to immunosuppressive therapy. This rare diagnosis was made after years of treatment for supposed rheumatoid arthritis. The patient presented with relapsing tenosynovitis with wounds on her right middle finger and wounds on her left lower leg, despite 3 months of adequate therapy (rifampicin+ethambutol+clarithromycin). Therapy was intensified with amikacin, clofazimine, moxifloxacin, and interferon-gamma due to the lack of response. Amputation of the right middle finger was necessary due to advanced disease. Treatment was further complicated by a paradoxical reaction, requiring prednisone treatment, which ultimately led to cure.


Assuntos
Hospedeiro Imunocomprometido , Infecções por Micobactéria não Tuberculosa/diagnóstico por imagem , Infecções por Micobactéria não Tuberculosa/imunologia , Idoso , Antibacterianos/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Feminino , Dedos/microbiologia , Dedos/cirurgia , Humanos , Imunossupressores/uso terapêutico , Linfoma não Hodgkin/tratamento farmacológico , Micobactérias não Tuberculosas/isolamento & purificação
17.
J Aquat Anim Health ; 30(4): 312-324, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30120830

RESUMO

No vaccine is yet commercially available against Mycobacterium marinum, the etiological agent of fish mycobacteriosis (also known as "fish tuberculosis"). The mycobacterial gene responsible for invasion and intracellular persistence, iipA, is known to moderate M. marinum pathology in Zebrafish Danio rerio. Two doses of heat-killed, wild-type, virulent M. marinum and two doses of a heat-killed, avirulent M. marinum iipA::kan mutant strain were used in parallel to vaccinate European Seabass Dicentrarchus labrax. The fish were then challenged with live, virulent M. marinum, and the pathogenesis of the infection was monitored. High specific immunoglobulin M (IgM) response and an increase in cytokine tumor necrosis factor alpha (TNF-α) messenger RNA expression levels were observed in all vaccinated fish. At 1 month postchallenge, TNF-α expression levels increased in spleen tissues of fish vaccinated with the virulent type and in those of unvaccinated fish, whereas in the head kidney, expression was up-regulated only in unvaccinated fish. The expression then decreased, and at 2 months postchallenge, expression appeared similar in all vaccination types. The highest survival rate (75%) was recorded in the group of fish that were vaccinated with a high dose of avirulent iipA::kan mutant. The iipA::kan mutant induced a strong immune response accompanied by only modest tissue disruption. Coupled with an effective program of booster treatments, the iipA::kan mutant vaccine may be developed into a powerful preventive measure against fish mycobacteriosis.


Assuntos
Doenças dos Peixes/microbiologia , Infecções por Micobactéria não Tuberculosa/veterinária , Mycobacterium marinum/patogenicidade , Animais , Bass , Doenças dos Peixes/imunologia , Temperatura Alta , Imunidade Celular , Imunidade Humoral , Imunoglobulina M/metabolismo , Infecções por Micobactéria não Tuberculosa/imunologia , Mycobacterium marinum/genética , Mycobacterium marinum/imunologia , RNA Mensageiro , Fator de Necrose Tumoral alfa/metabolismo , Vacinas de Produtos Inativados/imunologia , Vacinas de Produtos Inativados/uso terapêutico
18.
J Immunol ; 201(5): 1478-1490, 2018 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-30061197

RESUMO

In developed countries, pulmonary nontuberculous mycobacteria (NTM) infections are more prevalent than Mycobacterium tuberculosis infections. Given the differences in the pathogenesis of NTM and M. tuberculosis infections, separate studies are needed to investigate the pathological effects of NTM pathogens. Our previous study showed that anti-IFN-γ autoantibodies are detected in NTM-infected patients. However, the role of NK cells and especially NK cell-derived IFN-γ in this context has not been studied in detail. In the current study, we show that NK1.1 cell depletion increases bacterial load and mortality in a mouse model of pulmonary NTM infection. NK1.1 cell depletion exacerbates NTM-induced pathogenesis by reducing macrophage phagocytosis, dendritic cell development, cytokine production, and lung granuloma formation. Similar pathological phenomena are observed in IFN-γ-deficient (IFN-γ-/-) mice following NTM infection, and adoptive transfer of wild-type NK cells into IFN-γ-/- mice considerably reduces NTM pathogenesis. Injection of rIFN-γ also prevents NTM-induced pathogenesis in IFN-γ-/- mice. We observed that NK cells represent the main producers of IFN-γ in the lungs and production starts as soon as 1 d postinfection. Accordingly, injection of rIFN-γ into IFN-γ-/- mice 1 d (but not 2 wk) postinfection significantly improves immunity against NTM infection. NK cells also stimulate mycobacterial killing and IL-12 production by macrophages. Our results therefore indicate that IFN-γ production by NK cells plays an important role in activating and enhancing innate and adaptive immune responses at early stages of pulmonary NTM infection.


Assuntos
Imunidade Inata , Interferon gama/imunologia , Células Matadoras Naturais/imunologia , Pulmão/imunologia , Infecções por Micobactéria não Tuberculosa/imunologia , Mycobacterium/imunologia , Pneumonia Bacteriana/imunologia , Imunidade Adaptativa/genética , Animais , Interferon gama/deficiência , Interleucina-12/genética , Interleucina-12/imunologia , Células Matadoras Naturais/patologia , Pulmão/microbiologia , Pulmão/patologia , Camundongos , Camundongos Knockout , Infecções por Micobactéria não Tuberculosa/genética , Infecções por Micobactéria não Tuberculosa/patologia , Pneumonia Bacteriana/patologia
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