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1.
Mem Inst Oswaldo Cruz ; 115: e190398, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32187326

RESUMO

BACKGROUND: Streptococcus agalactiae capsular type III strains are a leading cause of invasive neonatal infections. Many pathogens have developed mechanisms to escape from host defense response using the host membrane microdomain machinery. Lipid rafts play an important role in a variety of cellular functions and the benefit provided by interaction with lipid rafts can vary from one pathogen to another. OBJECTIVES: This study aims to evaluate the involvement of membrane microdomains during infection of human endothelial cell by S. agalactiae. METHODS: The effects of cholesterol depletion and PI3K/AKT signaling pathway activation during S. agalactiae-human umbilical vein endothelial cells (HUVEC) interaction were analysed by pre-treatment with methyl-ß-cyclodextrin (MßCD) or LY294002 inhibitors, immunofluorescence and immunoblot analysis. The involvement of lipid rafts was analysed by colocalisation of bacteria with flotillin-1 and caveolin-1 using fluorescence confocal microscopy. FINDINGS: In this work, we demonstrated the importance of the integrity of lipid rafts microdomains and activation of PI3K/Akt pathway during invasion of S. agalactiae strain to HUVEC cells. Our results suggest the involvement of flotillin-1 and caveolin-1 during the invasion of S. agalactiae strain in HUVEC cells. CONCLUSIONS: The collection of our results suggests that lipid microdomain affects the interaction of S. agalactiae type III belonging to the hypervirulent ST-17 with HUVEC cells through PI3K/Akt signaling pathway.

2.
Infect Genet Evol ; 80: 104195, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31954181

RESUMO

Streptococcus agalactiae are important pathogenic bacteria that cause severe infections in humans, especially neonates. The mechanism by which ST-17 causes invasive infections than other STs is not well understood. In this study, we sequenced the first genome of a S. agalactiae ST-17 strain isolated in Brazil using the Illumina HiSeq 2500 technology. S. agalactiae GBS90356 ST-17 belongs to the capsular type III and was isolated from a neonatal with a fatal case of meningitis. The genome presented a size of 2.03 Mbp and a G + C content of 35.2%. S. agalactiae has 706 genes in its core genome and an open pan-genome with a size of 5.020 genes, suggesting a high genomic plasticity. GIPSy software was used to identify 10 Pathogenicity islands (PAIs) which corresponded to 15% of the genome size. IslandViewer4 corroborated the prediction of six PAIs. The pathogenicity islands showed important virulence factors genes for S. agalactiae e.g. neu, cps, dlt, fbs, cfb, lmb. SignalP detected 20 proteins with signal peptides among the 352 proteins found in PAIs, which 60% were located in the SagPAI_5. SagPAI_2 and 5 were mainly detected in ST-17 strains studied. Moreover, we identified 51 unique genes, 9 recombination regions and a large number of SNPs with an average of 760.3 polymorphisms, which can be related with high genomic plasticity and virulence during host-pathogen interactions. Our results showed implications for pathogenesis, evolution, concept of species and in silico analysis value to understand the epidemiology and genome plasticity of S. agalactiae.

3.
Mem Inst Oswaldo Cruz ; 113(6): e140421, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29641644

RESUMO

BACKGROUND Streptococcus agalactiae can causes sepsis, pneumonia, and meningitis in neonates, the elderly, and immunocompromised patients. Although the virulence properties of S. agalactiae have been partially elucidated, the molecular mechanisms related to reactive oxygen species (ROS) generation in infected human endothelial cells need further investigation. OBJECTIVES This study aimed to evaluate the influence of oxidative stress in human umbilical vein endothelial cells (HUVECs) during S. agalactiae infection. METHODS ROS production during S. agalactiae-HUVEC infection was detected using the probe CM-H2DCFDA. Microfilaments labelled with phalloidin-FITC and p47phox-Alexa 546 conjugated were analysed by immunofluorescence. mRNA levels of p47phox (NADPH oxidase subunit) were assessed using Real Time qRT-PCR. The adherence and intracellular viability of S. agalactiae in HUVECs with or without pre-treatment of DPI, apocynin (NADPH oxidase inhibitors), and LY294002 (PI3K inhibitor) were evaluated by penicillin/gentamicin exclusion. Phosphorylation of p47phox and Akt activation by S. agalactiae were evaluated by immunoblotting analysis. FINDINGS Data showed increased ROS production 15 min after HUVEC infection. Real-Time qRT-PCR and western blotting performed in HUVEC infected with S. agalactiae detected alterations in mRNA levels and activation of p47phox. Pre-treatment of endothelial cells with NADPH oxidase (DPI and apocynin) and PI3K/Akt pathway (LY294002) inhibitors reduced ROS production, bacterial intracellular viability, and generation of actin stress fibres in HUVECs infected with S. agalactiae. CONCLUSIONS ROS generation via the NADPH oxidase pathway contributes to invasion of S. agalactiae in human endothelial cells accompanied by cytoskeletal reorganisation through the PI3K/Akt pathway, which provides novel evidence for the involvement of oxidative stress in S. agalactiae pathogenesis.


Assuntos
Células Endoteliais da Veia Umbilical Humana/metabolismo , NADPH Oxidases/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Infecções Estreptocócicas/metabolismo , Streptococcus agalactiae/metabolismo , Análise de Variância , Aderência Bacteriana , Western Blotting , Células Endoteliais da Veia Umbilical Humana/microbiologia , Humanos , NADPH Oxidases/análise , Estresse Oxidativo/fisiologia , Espécies Reativas de Oxigênio/análise , Reação em Cadeia da Polimerase em Tempo Real , Valores de Referência , Transdução de Sinais/fisiologia , Fatores de Tempo
4.
Mem. Inst. Oswaldo Cruz ; 113(6): e140421, 2018. graf
Artigo em Inglês | LILACS | ID: biblio-894933

RESUMO

BACKGROUND Streptococcus agalactiae can causes sepsis, pneumonia, and meningitis in neonates, the elderly, and immunocompromised patients. Although the virulence properties of S. agalactiae have been partially elucidated, the molecular mechanisms related to reactive oxygen species (ROS) generation in infected human endothelial cells need further investigation. OBJECTIVES This study aimed to evaluate the influence of oxidative stress in human umbilical vein endothelial cells (HUVECs) during S. agalactiae infection. METHODS ROS production during S. agalactiae-HUVEC infection was detected using the probe CM-H2DCFDA. Microfilaments labelled with phalloidin-FITC and p47phox-Alexa 546 conjugated were analysed by immunofluorescence. mRNA levels of p47phox (NADPH oxidase subunit) were assessed using Real Time qRT-PCR. The adherence and intracellular viability of S. agalactiae in HUVECs with or without pre-treatment of DPI, apocynin (NADPH oxidase inhibitors), and LY294002 (PI3K inhibitor) were evaluated by penicillin/gentamicin exclusion. Phosphorylation of p47phox and Akt activation by S. agalactiae were evaluated by immunoblotting analysis. FINDINGS Data showed increased ROS production 15 min after HUVEC infection. Real-Time qRT-PCR and western blotting performed in HUVEC infected with S. agalactiae detected alterations in mRNA levels and activation of p47phox. Pre-treatment of endothelial cells with NADPH oxidase (DPI and apocynin) and PI3K/Akt pathway (LY294002) inhibitors reduced ROS production, bacterial intracellular viability, and generation of actin stress fibres in HUVECs infected with S. agalactiae. CONCLUSIONS ROS generation via the NADPH oxidase pathway contributes to invasion of S. agalactiae in human endothelial cells accompanied by cytoskeletal reorganisation through the PI3K/Akt pathway, which provides novel evidence for the involvement of oxidative stress in S. agalactiae pathogenesis.


Assuntos
Humanos , Espécies Reativas de Oxigênio/análise , NADPH Oxidases/análise , NADPH Oxidases/metabolismo , Células Endoteliais da Veia Umbilical Humana/microbiologia , Transdução de Sinais/fisiologia , Reação em Cadeia da Polimerase em Tempo Real
5.
Stand Genomic Sci ; 11: 39, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27274785

RESUMO

Streptococcus agalactiae, also referred to as Group B Streptococcus, is a frequent resident of the rectovaginal tract in humans, and a major cause of neonatal infection. The pathogen can also infect adults with underlying disease, particularly the elderly and immunocompromised ones. In addition, S. agalactiae is a known fish pathogen, which compromises food safety and represents a zoonotic hazard. This study provides valuable structural, functional and evolutionary genomic information of a human S. agalactiae serotype Ia (ST-103) GBS85147 strain isolated from the oropharynx of an adult patient from Rio de Janeiro, thereby representing the first human isolate in Brazil. We used the Ion Torrent PGM platform with the 200 bp fragment library sequencing kit. The sequencing generated 578,082,183 bp, distributed among 2,973,022 reads, resulting in an approximately 246-fold mean coverage depth and was assembled using the Mira Assembler v3.9.18. The S. agalactiae strain GBS85147 comprises of a circular chromosome with a final genome length of 1,996,151 bp containing 1,915 protein-coding genes, 18 rRNA, 63 tRNA, 2 pseudogenes and a G + C content of 35.48 %.

6.
Antonie Van Leeuwenhoek ; 107(6): 1387-99, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25828766

RESUMO

Biofilm-related infections are considered a major cause of morbidity and mortality in hospital environments. Biofilms allow microorganisms to exchange genetic material and to become persistent colonizers and/or multiresistant to antibiotics. Corynebacterium pseudodiphtheriticum (CPS), a commensal bacterium that colonizes skin and mucosal sites has become progressively multiresistant and responsible for severe nosocomial infections. However, virulence factors of this emergent pathogen remain unclear. Herein, we report the adhesive properties and biofilm formation on hydrophilic (glass) and hydrophobic (plastic) abiotic surfaces by CPS strains isolated from patients with localized (ATCC10700/Pharyngitis) and systemic (HHC1507/Bacteremia) infections. Adherence to polystyrene attributed to hydrophobic interactions between bacterial cells and this negatively charged surface indicated the involvement of cell surface hydrophobicity in the initial stage of biofilm formation. Attached microorganisms multiplied and formed microcolonies that accumulated as multilayered cell clusters, a step that involved intercellular adhesion and synthesis of extracellular matrix molecules. Further growth led to the formation of dense bacterial aggregates embedded in the exopolymeric matrix surrounded by voids, typical of mature biofilms. Data also showed CPS recognizing human fibrinogen (Fbg) and fibronectin (Fn) and involvement of these sera components in formation of "conditioning films". These findings suggested that biofilm formation may be associated with the expression of different adhesins. CPS may form biofilms in vivo possibly by an adherent biofilm mode of growth in vitro currently demonstrated on hydrophilic and hydrophobic abiotic surfaces. The affinity to Fbg and Fn and the biofilm-forming ability may contribute to the establishment and dissemination of infection caused by CPS.


Assuntos
Aderência Bacteriana , Biofilmes/crescimento & desenvolvimento , Corynebacterium/fisiologia , Microbiologia Ambiental , Fibrinogênio/metabolismo , Fibronectinas/metabolismo , Adesinas Bacterianas/metabolismo , Bacteriemia/microbiologia , Técnicas Bacteriológicas , Corynebacterium/crescimento & desenvolvimento , Corynebacterium/isolamento & purificação , Infecções por Corynebacterium/microbiologia , Vidro , Humanos , Microscopia , Faringite/microbiologia , Plásticos
7.
Mem Inst Oswaldo Cruz ; 109(7): 940-3, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25410999

RESUMO

Endothelial dysfunction is a major component of the pathophysiology of septicaemic group B Streptococcus (GBS) infections. Although cytokines have been shown to activate human umbilical vein endothelial cells (HUVECs), the capacity of interferon (IFN)-γ to enhance the microbicidal activity of HUVECs against GBS has not been studied. We report that the viability of intracellular bacteria was reduced in HUVECs activated by IFN-γ. Enhanced fusion of lysosomes with bacteria-containing vacuoles was observed by acid phosphatase and the colocalisation of Rab-5, Rab-7 and lysosomal-associated membrane protein-1 with GBS in IFN-γ-activated HUVECs. IFN-γ resulted in an enhancement of the phagosome maturation process in HUVECs, improving the capacity to control the intracellular survival of GBS.


Assuntos
Anti-Infecciosos/farmacologia , Células Endoteliais da Veia Umbilical Humana/microbiologia , Interferon gama/farmacologia , Viabilidade Microbiana/efeitos dos fármacos , Infecções Estreptocócicas/tratamento farmacológico , Streptococcus agalactiae/efeitos dos fármacos , Fosfatase Ácida/metabolismo , Aderência Bacteriana/efeitos dos fármacos , Endocitose , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Lisossomos/efeitos dos fármacos , Fagossomos/efeitos dos fármacos , Cultura Primária de Células , Infecções Estreptocócicas/prevenção & controle , Análise de Sobrevida
8.
Mem. Inst. Oswaldo Cruz ; 109(7): 940-943, 11/2014. graf
Artigo em Inglês | LILACS | ID: lil-728811

RESUMO

Endothelial dysfunction is a major component of the pathophysiology of septicaemic group B Streptococcus (GBS) infections. Although cytokines have been shown to activate human umbilical vein endothelial cells (HUVECs), the capacity of interferon (IFN)-γ to enhance the microbicidal activity of HUVECs against GBS has not been studied. We report that the viability of intracellular bacteria was reduced in HUVECs activated by IFN-γ. Enhanced fusion of lysosomes with bacteria-containing vacuoles was observed by acid phosphatase and the colocalisation of Rab-5, Rab-7 and lysosomal-associated membrane protein-1 with GBS in IFN-γ-activated HUVECs. IFN-γ resulted in an enhancement of the phagosome maturation process in HUVECs, improving the capacity to control the intracellular survival of GBS.


Assuntos
Humanos , Anti-Infecciosos/farmacologia , Células Endoteliais da Veia Umbilical Humana/microbiologia , Interferon gama/farmacologia , Viabilidade Microbiana/efeitos dos fármacos , Infecções Estreptocócicas/tratamento farmacológico , Streptococcus agalactiae/efeitos dos fármacos , Fosfatase Ácida/metabolismo , Aderência Bacteriana/efeitos dos fármacos , Endocitose , Células Endoteliais da Veia Umbilical Humana/metabolismo , Lisossomos/efeitos dos fármacos , Cultura Primária de Células , Fagossomos/efeitos dos fármacos , Análise de Sobrevida , Infecções Estreptocócicas/prevenção & controle
9.
Microbiology ; 160(Pt 3): 537-546, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24344208

RESUMO

Although infection by Corynebacterium diphtheriae is a model of extracellular mucosal pathogenesis, different clones have been also associated with invasive infections such as sepsis, endocarditis, septic arthritis and osteomyelitis. The mechanisms that promote C. diphtheriae infection and haematogenic dissemination need further investigation. In this study we evaluated the association and invasion mechanisms with human umbilical vein endothelial cells (HUVECs) and experimental arthritis in mice of endocarditis-associated strains and control non-invasive strains. C. diphtheriae strains were able to adhere to and invade HUVECs at different levels. The endocarditis-associated strains displayed an aggregative adherence pattern and a higher number of internalized viable cells in HUVECs. Transmission electron microscopy (TEM) analysis revealed intracellular bacteria free in the cytoplasm and/or contained in a host-membrane-confined compartment as single micro-organisms. Data showed bacterial internalization dependent on microfilament and microtubule stability and involvement of protein phosphorylation in the HUVEC signalling pathway. A high number of affected joints and high arthritis index in addition to the histopathological features indicated a strain-dependent ability of C. diphtheriae to cause severe polyarthritis. A correlation between the arthritis index and increased systemic levels of IL-6 and TNF-α was observed for endocarditis-associated strains. In conclusion, higher incidence of potential mechanisms by which C. diphtheriae may access the bloodstream through the endothelial barrier and stimulate the production of pro-inflammatory cytokines such as IL-6 and TNF-α, in addition to the ability to affect the joints and induce arthritis through haematogenic spread are thought to be related to the pathogenesis of endocarditis-associated strains.


Assuntos
Corynebacterium diphtheriae/fisiologia , Endocardite/microbiologia , Células Endoteliais/microbiologia , Animais , Artrite/microbiologia , Aderência Bacteriana , Linhagem Celular , Citocinas/biossíntese , Endocardite/metabolismo , Células Endoteliais/metabolismo , Feminino , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/microbiologia , Humanos , Camundongos
10.
Antonie Van Leeuwenhoek ; 104(6): 1125-33, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24052367

RESUMO

We explored Group B Streptococcus (GBS)-induced apoptosis in human umbilical vein endothelial cells (HUVEC) and the role of phosphoramidon, a zinc metalloprotease inhibitor, in this process. GBS 90186 strain (serotype V, a blood isolate) and concentrated supernatant (CS) were used to investigate the viability and morphological alterations in HUVEC by Trypan blue uptake, electrophoresis in 2 % agarose gel and scanning electron microscopy assays. Apoptosis before and after phosphoramidon-treatment were verified by flow cytometry using annexin V-FITC labeling. Differences were considered significant when P < 0.05 using unpaired Student's t test. GBS and CS induced HUVEC death by apoptosis (76.5 and 32 %, respectively) with an increasing pro-apoptotic Bax expression and decreasing anti-apoptotic Bcl-2 expression. Caspase-3 was activated during GBS-induced endothelial apoptosis. Phosphoramidon reduced 89.3 and 100 % of GBS and CS cell death by apoptosis, respectively. Some GBS strains may induce cell death by apoptosis with involvement of metalloproteases and signaling through the intrinsic pathway of apoptosis, which may contribute to GBS survival during sepsis of adults and neonates.


Assuntos
Apoptose , Células Endoteliais/microbiologia , Células Endoteliais/fisiologia , Glicopeptídeos/metabolismo , Metaloproteases/metabolismo , Inibidores de Proteases/metabolismo , Streptococcus agalactiae/enzimologia , Anexina A5/análise , Sobrevivência Celular , Células Cultivadas , Eletroforese , Humanos , Metaloproteases/antagonistas & inibidores , Microscopia Eletrônica , Coloração e Rotulagem/métodos , Azul Tripano/metabolismo
11.
Braz. j. microbiol ; 44(3): 869-872, July-Sept. 2013. tab
Artigo em Inglês | LILACS | ID: lil-699819

RESUMO

GBS serotypes III and V were the most prevalent in pregnant women and exhibited resistance to tetracycline, clindamycin and sulfamethoxazole/trimethoprim. Serotype III showed high sialic acid content and PFGE analysis discerned 33 heterogeneous profiles. Phenotypic and genotypic characterization could be relevant to control GBS infections unaffected by intra-partum chemoprophylaxis.


Assuntos
Feminino , Humanos , Recém-Nascido , Gravidez , Complicações Infecciosas na Gravidez/epidemiologia , Sorogrupo , Infecções Estreptocócicas/epidemiologia , Streptococcus agalactiae/isolamento & purificação , Antibacterianos/farmacologia , Brasil/epidemiologia , Análise por Conglomerados , Farmacorresistência Bacteriana , Eletroforese em Gel de Campo Pulsado , Genótipo , Tipagem Molecular , Prevalência , Complicações Infecciosas na Gravidez/microbiologia , Infecções Estreptocócicas/microbiologia , Streptococcus agalactiae/classificação , Streptococcus agalactiae/efeitos dos fármacos
12.
13.
Mem Inst Oswaldo Cruz ; 107(4): 486-93, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22666859

RESUMO

Corynebacterium pseudodiphtheriticum is a well-known human pathogen that mainly causes respiratory disease and is associated with high mortality in compromised hosts. Little is known about the virulence factors and pathogenesis of C. pseudodiphtheriticum. In this study, cultured human epithelial (HEp-2) cells were used to analyse the adherence pattern, internalisation and intracellular survival of the ATCC 10700 type strain and two additional clinical isolates. These microorganisms exhibited an aggregative adherence-like pattern to HEp-2 cells characterised by clumps of bacteria with a "stacked-brick" appearance. The differences in the ability of these microorganisms to invade and survive within HEp-2 cells and replicate in the extracellular environment up to 24 h post infection were evaluated. The fluorescent actin staining test demonstrated that actin polymerisation is involved in the internalisation of the C. pseudodiphtheriticum strains. The depolymerisation of microfilaments by cytochalasin E significantly reduced the internalisation of C. pseudodiphtheriticum by HEp-2 cells. Bacterial internalisation and cytoskeletal rearrangement seemed to be partially triggered by the activation of tyrosine kinase activity. Although C. pseudodiphtheriticum strains did not demonstrate an ability to replicate intracellularly, HEp-2 cells were unable to fully clear the pathogen within 24 h. These characteristics may explain how some C. pseudodiphtheriticum strains cause severe infection in human patients.


Assuntos
Aderência Bacteriana/fisiologia , Corynebacterium/patogenicidade , Células Epiteliais/microbiologia , Corynebacterium/fisiologia , Células Hep G2 , Humanos , Virulência
14.
Mem. Inst. Oswaldo Cruz ; 107(4): 486-493, June 2012. ilus, graf
Artigo em Inglês | LILACS | ID: lil-626442

RESUMO

Corynebacterium pseudodiphtheriticum is a well-known human pathogen that mainly causes respiratory disease and is associated with high mortality in compromised hosts. Little is known about the virulence factors and pathogenesis of C. pseudodiphtheriticum. In this study, cultured human epithelial (HEp-2) cells were used to analyse the adherence pattern, internalisation and intracellular survival of the ATCC 10700 type strain and two additional clinical isolates. These microorganisms exhibited an aggregative adherence-like pattern to HEp-2 cells characterised by clumps of bacteria with a "stacked-brick" appearance. The differences in the ability of these microorganisms to invade and survive within HEp-2 cells and replicate in the extracellular environment up to 24 h post infection were evaluated. The fluorescent actin staining test demonstrated that actin polymerisation is involved in the internalisation of the C. pseudodiphtheriticum strains. The depolymerisation of microfilaments by cytochalasin E significantly reduced the internalisation of C. pseudodiphtheriticum by HEp-2 cells. Bacterial internalisation and cytoskeletal rearrangement seemed to be partially triggered by the activation of tyrosine kinase activity. Although C. pseudodiphtheriticum strains did not demonstrate an ability to replicate intracellularly, HEp-2 cells were unable to fully clear the pathogen within 24 h. These characteristics may explain how some C. pseudodiphtheriticum strains cause severe infection in human patients.


Assuntos
Humanos , Aderência Bacteriana/fisiologia , Corynebacterium/patogenicidade , Células Epiteliais/microbiologia , Corynebacterium/fisiologia , Virulência
15.
Microb Pathog ; 52(3): 165-76, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22239957

RESUMO

Although Corynebacterium diphtheriae has been classically described as an exclusively extracellular pathogen, there is growing evidence that it may be internalized by epithelial cells. The aim of the present report was to investigate the nature and involvement of the surface-exposed non-fimbrial 67-72 kDa proteins (67-72p), previously characterized as adhesin/hemagglutinin, in C. diphtheriae internalization by HEp-2 cells. Transmission electron microscopy and bacterial internalization inhibition assays indicated the role of 67-72p as invasin for strains of varied sources. Cytoskeletal changes with accumulation of polymerized actin in HEp-2 cells beneath adherent 67-72p-adsorbed microspheres were observed by the Fluorescent actin staining test. Trypan blue staining method and Methylthiazole tetrazolium reduction assay showed a significant decrease in viability of HEp-2 cells treated with 67-72p. Morphological changes in HEp-2 cells observed after treatment with 67-72p included vacuolization, nuclear fragmentation and the formation of apoptotic bodies. Flow cytometry revealed an apoptotic volume decrease in HEp-2 cells treated with 67-72p. Moreover, a double-staining assay using Propidium Iodide/Annexin V gave information about the numbers of vital vs. early apoptotic cells and late apoptotic or secondary necrotic cells. The comparative analysis of MALDI-TOF MS experiments with the probes provided for 67-72p CDC-E8392 with an in silico proteome deduced from the complete genome sequence of C. diphtheriae identified with significant scores 67-72p as the protein DIP0733. In conclusion, DIP0733 (67-72p) may be directly implicated in bacterial invasion and apoptosis of epithelial cells in the early stages of diphtheria and C. diphtheriae invasive infection.


Assuntos
Apoptose , Corynebacterium diphtheriae/patogenicidade , Endocitose , Hemaglutininas/metabolismo , Hepatócitos/microbiologia , Hepatócitos/fisiologia , Fatores de Virulência/metabolismo , Actinas/metabolismo , Linhagem Celular , Sobrevivência Celular , Corynebacterium diphtheriae/genética , Hemaglutininas/genética , Humanos , Multimerização Proteica , Fatores de Virulência/genética
16.
Rev. saúde pública ; 45(6): 1176-1191, dez. 2011. tab
Artigo em Português | LILACS | ID: lil-606869

RESUMO

O artigo revisa a literatura sobre a emergência de infecções humanas causadas por Corynebacterium ulcerans em diversos países, incluindo o Brasil. Foi realizada análise de artigos publicados entre 1926 e 2011 nas bases Medline/PubMed e SciELO, bem como artigos e informes do Ministério da Saúde. Apresenta-se um esquema de triagem, rápido, econômico e de fácil execução, capaz de permitir a realização do diagnóstico presuntivo de C. ulcerans e C. diphtheriae na maioria dos laboratórios brasileiros públicos e privados. A circulação de C. ulcerans em vários países, aliada aos recentes casos de isolamento do patógeno no Rio de Janeiro, é um alerta a clínicos, veterinários e microbiologistas sobre a ocorrência de difteria zoonótica e a circulação do C. ulcerans em regiões urbanas e rurais do território nacional e/ou da América Latina.


The article is a literature review on the emergence of human infections caused by Corynebacterium ulcerans in many countries including Brazil. Articles in Medline/PubMed and SciELO databases published between 1926 and 2011 were reviewed, as well as articles and reports of the Brazilian Ministry of Health. It is presented a fast, cost-effective and easy to perform screening test for the presumptive diagnosis of C. ulcerans and C. diphtheriae infections in most Brazilian public and private laboratories. C. ulcerans spread in many countries and recent isolation of this pathogen in Rio de Janeiro, southeastern Brazil, is a warning to clinicians, veterinarians, and microbiologists on the occurrence of zoonotic diphtheria and C. ulcerans dissemination in urban and rural areas of Brazil and/or Latin America.


El articulo revisa la literatura sobre la emergencia de infecciones humanas causadas por Corynebacterium ulcerans en diversos países, incluyendo Brasil. Se realizó análisis de artículos publicados entre 1926 y 2011 en las bases Medline/Pubmed y SciELO, así como artículos e informes del Ministerio Brasileño de la Salud. Se presenta un esquema de selección, rápido, económico y de fácil ejecución, capaz de permitir la realización del diagnóstico presuntivo de C. ulcerans y C. diphtheriae en la mayoría de los laboratorios brasileños públicos y privados. La circulación de C. ulcerans en varios países, aliada a los recientes casos de aislamiento del patógeno en Rio de Janeiro (Sureste de Brasil), es un alerta a clínicos, veterinarios y microbiólogos sobre la ocurrencia de difteria zoológica y la circulación de C. ulcerans en regiones urbanas y rurales del territorio nacional y/o de América Latina.


Assuntos
Animais , Humanos , Infecções por Corynebacterium/epidemiologia , Corynebacterium/classificação , Difteria/epidemiologia , Epidemias , Zoonoses/epidemiologia , Brasil/epidemiologia , Técnicas de Laboratório Clínico , Doenças Transmissíveis Emergentes/diagnóstico , Doenças Transmissíveis Emergentes/epidemiologia , Doenças Transmissíveis Emergentes/microbiologia , Infecções por Corynebacterium/diagnóstico , Corynebacterium diphtheriae/isolamento & purificação , Notificação de Doenças , Difteria/diagnóstico , Difteria/microbiologia , Saúde Global
17.
Rev Saude Publica ; 45(6): 1176-91, 2011 Dec.
Artigo em Inglês, Português | MEDLINE | ID: mdl-22124745

RESUMO

The article is a literature review on the emergence of human infections caused by Corynebacterium ulcerans in many countries including Brazil. Articles in Medline/PubMed and SciELO databases published between 1926 and 2011 were reviewed, as well as articles and reports of the Brazilian Ministry of Health. It is presented a fast, cost-effective and easy to perform screening test for the presumptive diagnosis of C. ulcerans and C. diphtheriae infections in most Brazilian public and private laboratories. C. ulcerans spread in many countries and recent isolation of this pathogen in Rio de Janeiro, southeastern Brazil, is a warning to clinicians, veterinarians, and microbiologists on the occurrence of zoonotic diphtheria and C. ulcerans dissemination in urban and rural areas of Brazil and/or Latin America.


Assuntos
Infecções por Corynebacterium/epidemiologia , Corynebacterium/classificação , Difteria/epidemiologia , Epidemias , Zoonoses/epidemiologia , Animais , Brasil/epidemiologia , Técnicas de Laboratório Clínico , Doenças Transmissíveis Emergentes/diagnóstico , Doenças Transmissíveis Emergentes/epidemiologia , Doenças Transmissíveis Emergentes/microbiologia , Infecções por Corynebacterium/diagnóstico , Corynebacterium diphtheriae/isolamento & purificação , Difteria/diagnóstico , Difteria/microbiologia , Notificação de Doenças , Saúde Global , Humanos
18.
Int J Mol Med ; 27(5): 739-44, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21369696

RESUMO

Although group B Streptococcus (GBS) has been classically described as an exclusively extracellular pathogen, growing evidence suggests that it may be internalized by epithelial cells. However, the fates of intracellular GBS and of infected respiratory epithelial cells remain unclear. Little is known about the bacterial components involved in these processes. The present study investigated the bacterial internalization by A549 cells and the apoptosis/necrosis of the infected human epithelial cells. The morphological changes in A549 cells observed from 2 h post-infection with GBS included vacuolization and the formation of apoptotic bodies. Flow cytometry revealed that 81.2% of apoptotic A549 cells were infected with GBS serotype III 90356-liquor. Moreover, a double-staining assay using propidium iodide (PI)/Annexin V (AV) gave information about the numbers of viable (PI-/AV-) (18.27%) vs. early apoptotic (PI-/AV+) (73.83%) and late apoptotic cells (PI+/AV+) (7.37%) during infection of A549 cells with GBS III 90356-liquor. In addition, 37% necrotic cells were observed in A549 cells infected with GBS serotype V 90186-blood. In conclusion, GBS serotypes III and V induce apoptosis of epithelial cells in the early stages of GBS infection, resulting in tissue destruction, bacterial spreading and, in consequence, invasive disease or systemic infection.


Assuntos
Apoptose , Células Epiteliais/microbiologia , Streptococcus agalactiae/fisiologia , Aderência Bacteriana , Linhagem Celular Tumoral , Sobrevivência Celular , Contagem de Colônia Microbiana , Citofagocitose , Células Epiteliais/patologia , Células Epiteliais/ultraestrutura , Humanos , Necrose , Sorotipagem
19.
Int J Mol Med ; 26(4): 511-6, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20818490

RESUMO

Group B streptococci (GBS) are the most common cause of pneumonia and sepsis during the neonatal period. However, the pathogenesis of invasive infection is poorly understood. We investigated the ability of GBS grown at 37 degrees C and 40 degrees C to adhere and invade human umbilical vein endothelial cells (HUVECs) at different periods of incubation (0, 0.5, 1, 2, 18 and 24 h). All strains tested, except strain 88641-vagina survived for 24 h in the intracellular environment at 40 degrees C. For serotype III grown at 40 degrees C, both strains (80340-vagina and 90356-liquor) showed increased adherence and intracellular survival when compared to bacteria grown at 37 degrees C (P<0.01). GBS serotype V strains (88641-vagina and 90186-blood) showed ability to survive inside HUVECs until 2 and 24 h post-infection at 40 degrees C and 37 degrees C, respectively (P<0.01). Influence of growth temperature in bacterial interaction with endothelial cells was partially dependent of serotypes and the clinical origin of strains. Serotypes III and V strains grown at both temperatures remained viable within acidic endothelial vacuoles which acquired Rab7 and LAMP-1 endosomal markers. The data emphasize the influence of temperature on cellular events of phagocytosis and pathogenesis of GBS diseases.


Assuntos
Células Endoteliais/microbiologia , Febre/microbiologia , Infecções Estreptocócicas/microbiologia , Streptococcus agalactiae/fisiologia , Aderência Bacteriana , Temperatura Corporal , Linhagem Celular , Interações Hospedeiro-Patógeno , Humanos
20.
Mem Inst Oswaldo Cruz ; 105(5): 706-11, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20835621

RESUMO

The production of fibrinous exudates may play an important role in determining the outcome of bacterial infection. Although pseudomembrane formation is a characteristic feature of diphtheria, little is known about the fibrinogen (Fbn)-binding properties of Corynebacterium diphtheriae strains and the influence of the gene that codes for diphtheria toxin (tox gene) in this process. In this study we demonstrated the ability of C. diphtheriae strains to bind to Fbn and to convert Fbn to fibrin. Bacterial interaction with rabbit plasma was evaluated by both slide and tube tests. Interaction of microorganisms with human Fbn was evaluated by both enzyme linked immunosorbent assay (ELISA) and fluorescein isothiocyanate-conjugated (FITC) Fbn binding assays. Nontoxigenic and toxigenic strains formed bacterial aggregates in the presence of plasma in the slide tests. The ability to convert Fbn to a loose web of fibrin in the plasma solution in the tube tests appeared to be a common characteristic of the species, including strains that do not carry the tox gene. Fbn binding to C. diphtheriae strains occurred at varying intensities, as demonstrated by the FITC-Fbn and ELISA binding assays. Our data suggest that the capacity to bind to Fbn and to convert Fbn to fibrin may play a role in pseudomembrane formation and act as virulence determinants of both nontoxigenic and toxigenic strains.


Assuntos
Corynebacterium diphtheriae/metabolismo , Toxina Diftérica/metabolismo , Fibrinogênio/metabolismo , Animais , Corynebacterium diphtheriae/genética , Toxina Diftérica/genética , Ensaio de Imunoadsorção Enzimática , Fibrinogênio/genética , Humanos , Coelhos , Virulência/genética
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