RESUMO
Enteropathogenic Escherichia coli (EPEC) are frequently isolated as a cause of infantile diarrhea in developing countries. Its pathogenicity is distinguished by histopathological alterations at the site of infection, known as attaching and effacing (A/E) lesions, in which bacterial virulence factors and host proteins participate. Intimin, a bacterial adhesin expressed by all EPEC described to date, is responsible for the intimate adherence of the bacteria to host cells and is essential for the formation of A/E lesions. Mucosal vaccination may represent an efficacious intervention to prevent EPEC infection and lower morbidity and mortality rates. Strategies for mucosal vaccinations that use lactic acid bacteria for the delivery of heterologous antigens rely on their safety profile and ability to stimulate the immune system. In the present work, we have constructed Lactobacillus casei strains expressing different fragments of intimin â, a subtype that is frequently expressed by EPEC strains. Mucosal immunization of mice with L. casei expressing intimin fragments induced specific systemic and mucosal antibodies. These antibodies were able to recognize native intimin on the surface of EPEC and to inhibit in vitro EPEC binding to epithelial cells.
Assuntos
Animais , Camundongos , Diarreia Infantil/terapia , Infecções por Escherichia coli/terapia , Lacticaseibacillus casei , ImunizaçãoRESUMO
Escherichia coli isolates recovered from 182 fecal specimens from dogs up to five months old from the cities of São Paulo and Campinas, SP, Brazil, were examined by polymerase chain reaction (PCR) for several virulence factors and properties. The eae gene was found in 23 isolates of E. coli from 22 dogs, 19 of 146 (13%) from dogs with diarrhea and 3 of 36 (8.3%) from dogs with no diarrhea. Two different eae+ isolates were recovered from one dog with diarrhea. Isolates from two dogs with diarrhea harbored the bfpA gene, and none of the isolates possessed genes for enterotoxins, the EAF plasmid or Shiga toxins. PCR showed that, among the 23 isolates, eight were positive for beta intimin, six for gamma, two for, one for alpha, one for kappa, and five showed no amplification with any of the nine pairs of specific intimin primers used. PCR also showed that the LEE (locus of enterocyte effacement) was inserted in selC in four isolates, likely in pheU in seven isolates, and in undetermined sites in twelve isolates. Fifteen isolates adhered to HEp-2 cells and were fluorescence actin staining (FAS) positive. The predominant adherence pattern was the localized adherence-like (LAL) pattern. The eae-positive isolates belonged to a wide diversity of serotypes, including O111:H25, O119:H2 and O142:H6, which are serotypes that are common among human EPEC. These results confirmed the presence of EPEC in dogs (DEPEC) with and without diarrhea. The virulence factors found in these strains were similar to those in human EPEC, leading to the possibility that EPEC may move back and forth among human and canine populations.
Assuntos
Aderência Bacteriana , Doenças do Cão/microbiologia , Infecções por Escherichia coli/veterinária , Escherichia coli/patogenicidade , Adesinas Bacterianas/genética , Animais , Sequência de Bases , Brasil , Diarreia/microbiologia , Diarreia/veterinária , Cães , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli/genética , Fezes/microbiologia , Proteínas de Fímbrias/genética , Genes Bacterianos , Humanos , Reação em Cadeia da Polimerase/veterinária , Sorotipagem/veterinária , Virulência/genéticaRESUMO
A total of 102 Escherichia coli strains belonging to serogroups O127 and O142 were examined for genotypic and phenotypic characteristics. The most frequent serotypes found were O127:H21, O127:H40 and O142:H34. The virulence properties were evaluated by adhesion to HeLa cells and hybridization with gene probes for diarrhoeagenic E. coli. Most strains in the two serogroups were categorized as enteropathogenic E. coli, but enteroaggregative E. coli was also detected in both serogroups. All strains that carried the eae sequence presented the LEE region inserted in selC. Five ribotypes were detected in serogroup O127 and four in serogroup O142 and a correlation between serotypes and ribotypes was observed mainly in serogroup O142.
Assuntos
Diarreia Infantil/microbiologia , Escherichia coli/classificação , Escherichia coli/patogenicidade , Virulência/genética , Brasil , Pré-Escolar , Genótipo , Humanos , Lactente , FenótipoRESUMO
Enteroaggregative Escherichia coli (EAEC) is defined by the ability to produce aggregative adherence (AA) to cultured cells. We analysed 128 EAEC strains, isolated from children with and without diarrhoea, regarding the presence of 11 EAEC virulence genes. Seventy strains carried and 58 lacked the EAEC probe sequence; 17 probe positive and 31 probe negative strains showed variations in the AA pattern. All EAEC probe positive strains carried at least one EAEC marker; aspU (94.3%), irp2 (91.4%), and aggR (74.3%) were the most prevalent. Conversely, among the EAEC probe negative strains, 41.4% were devoid of any marker and astA predominated (44.8%). No significant statistical difference in the prevalence of any marker between cases and controls in both EAEC probe groups or AA variants was found. We suggest that the EAEC probe positive strains may have a higher pathogenic potential or alternatively, EAEC probe negative strains may harbour virulence factors as yet undescribed.
Assuntos
Diarreia/microbiologia , Escherichia coli/patogenicidade , Biomarcadores , Criança , Humanos , Sondas Moleculares , Virulência/genéticaRESUMO
Genetic and phenotypic virulence markers of different categories of diarrhoeagenic Escherichia coli were investigated in 106 strains of enteropathogenic E. coli (EPEC) serogroup O86. The most frequent serotype found was O86:H34 (86%). Strains of this serotype and the non motile ones behaved as EPEC i.e., carried eae, bfpA and EAF DNA sequences and presented localised adherence to HeLa cells. Serotypes O86:H2, O86:H6, O86:H10, O86:H18, O86:H27 and O86:H non determined, belonged to other categories. The majority of the strains of serotype O86:H34 and non motile strains produced cytolethal-distending toxin (CDT). The ribotyping analysis showed a correlation among ribotypes, virulence markers and serotypes, thus suggesting that CDT production might be a property associated with a universal clone represented by the O86:H34 serotype.
Assuntos
Toxinas Bacterianas/biossíntese , Diarreia/microbiologia , Escherichia coli/metabolismo , Escherichia coli/patogenicidade , Brasil , Criança , Diarreia/metabolismo , Escherichia coli/classificação , Marcadores Genéticos , Humanos , Fenótipo , Ribotipagem , Sorotipagem , VirulênciaRESUMO
BACKGROUND: Enteroaggregative Escherichia coli (EAEC) is an important agent of the persistent diarrhea among low socioeconomic level children in developing countries that may be associated with chronic undernourishment. Breast-feeding is effective in protecting infants against diarrhea and other infectious diseases. The aim of the study is to verify the ability of human colostrum to inhibit aggregative adhesion of EAEC to HEp-2 cells and the presence of antibodies reactive to antigenic fractions of EAEC in colostrum samples. METHODS: Enzyme-linked immunosorbent assay, immunoblotting and adhesion assays of EAEC to HEp-2 cells were done with pooled or individual colostrum samples (n = 35). Assays were performed with a well-known EAEC strain, 044:H18 E. coli (strain 042). Colostral IgA was isolated by affinity chromatography in Sepharose anti-human alpha chain column. RESULTS: Total colostrum and isolated IgA inhibited EAEC adhesion, and this ability was associated with the presence of IgA antibodies against a 15-kDa band, compatible with the subunits of aggregative adherence fimbrial adhesin II, characteristic of the 042 strain, absent in its plasmid-cured isogenic strain, that was used as control. Individual colostrum samples also inhibited adhesion, showed variable antibody titles against EAEC antigens in enzyme-linked immunosorbent assay and recognized many antigenic fractions in immunoblotting assays, including the 15-kDa band. CONCLUSIONS: These results confirm that IgA from human colostrum inhibits adhesion of EAEC to HEp-2 cells and suggest that colostrum IgA antibodies reactive to EAEC antigens may play a role in protection of infants against diarrhea caused by these bacteria.
Assuntos
Colostro/imunologia , Infecções por Escherichia coli/fisiopatologia , Escherichia coli/patogenicidade , Imunoglobulina A Secretora/metabolismo , Adulto , Aderência Bacteriana , Brasil , Ensaio de Imunoadsorção Enzimática , Infecções por Escherichia coli/imunologia , Feminino , Células HeLa , Humanos , Imunoglobulina A Secretora/imunologia , GravidezRESUMO
Genomic diversity among 34 strains of Escherichia coli belonging to different serotypes of the O26 serogroup -- encompassing strains from different geographical origins and Shiga toxin-negative Brazilian strains -- was evaluated through random amplified polymorphic DNA (RAPD) analysis. Our results indicate that Brazilian and non-Brazilian O26 strains fall under distinct but closely related differentiation clusters. RFLP-PCR analysis of the fliC gene sequence was done in order to identify the H(-) serotypes and served to confirm the clustering pattern obtained in the dendrogram generated from RAPD data. The epidemiological significance of these data is discussed.
Assuntos
Adesinas Bacterianas , Proteínas de Transporte , Proteínas de Escherichia coli , Escherichia coli/classificação , Escherichia coli/genética , Proteínas da Membrana Bacteriana Externa/genética , Técnicas de Tipagem Bacteriana , Brasil , Escherichia coli/isolamento & purificação , Escherichia coli/patogenicidade , Infecções por Escherichia coli/microbiologia , Flagelina/genética , Variação Genética , Humanos , Filogenia , Reação em Cadeia da Polimerase/métodos , Polimorfismo de Fragmento de Restrição , Técnica de Amplificação ao Acaso de DNA Polimórfico/métodos , Sorotipagem , Toxina Shiga I/genética , Toxina Shiga II/genética , Toxinas ShigaRESUMO
BACKGROUND: enteropathogenic Escherichia coli (EPEC) is the main etiological agent of infantile diarrhea in Brazil and other developing countries. Human milk IgA protects newborn intestinal mucosa by inhibiting bacterial adhesion to epithelial cells and this effect is shown by in vitro assays of EPEC adhesion to HEp-2 cultured cells. Bovine milk, if effective in promoting this protection, could be an useful tool in the absence of the natural breastfeeding, in high-risk nurseries or in hospital infections. METHODS: the effect of colostrum, milk, and serum from dairy cows on the adherence to EPEC to HEp-2 cells was investigated. Colostrum from immunized and control animals and industrialized milk formulas were fractionated through a membrane device with a molecular weight cut off 10 kDa. The high molecular weight fraction (HMWF) of bovine colostrum was depleted of IgG through an affinity column and absorbed with an EPEC adherent strain. Antibodies were searched by ELISA and immunoblotting (IB). RESULTS: colostrum and milk from EPEC-immunized animals showed and inhibitory activity on adherence similar to that of control non-immunized animals. The inhibitory effect on adhesion was related to the HMWF. IgG-depleted colostrum partially retained the inhibitory effect, whereas IgG-rich eluate lost this property. The EPEC-absorbed fraction retained the inhibitory property. Industrialized milk formulas and respective HMWF also inhibited bacterial adherence. In IB assays, colostrum and milk samples from immunized animals recognized proteins of 30-40 kDa and 94 kDa, a molecular weight consistent with the adhesin intimin, in EPEC extracts. CONCLUSIONS: the inhibitory effect of EPEC adherence may be mediated by HMWF components, and IgG was not the only component responsible for this phenomenon.
Assuntos
Aderência Bacteriana/efeitos dos fármacos , Colostro , Proteínas de Escherichia coli , Escherichia coli/efeitos dos fármacos , Leite , Adesinas Bacterianas , Animais , Anticorpos Antibacterianos/análise , Vacinas Bacterianas , Proteínas de Transporte/antagonistas & inibidores , Bovinos , Linhagem Celular , Fracionamento Químico , Colostro/química , Depressão Química , Diarreia Infantil/microbiologia , Escherichia coli/fisiologia , Infecções por Escherichia coli/microbiologia , Feminino , Humanos , Imunização , Immunoblotting , Imunoglobulina G/análise , Lactente , Alimentos Infantis/análise , Leite/química , Peso Molecular , GravidezRESUMO
A total of 919 Escherichia coli isolates from 125 children with diarrhoea (cases) and 98 controls were assayed for adherence to HEp-2 cells. Localised adherence was found only in isolates from cases. Diffuse, aggregative (AA), chain-like adherence (CLA) and variants of the AA pattern were found in both cases and controls. The AA isolates were tested for gene sequences associated with enteroaggregative E. coli (EAEC). Only 25% of the isolates hybridised with the EAEC probe, and the aafA, astA and pet gene sequences were found in 7.9%, 44.7% and 7.9% of the isolates, respectively. The aggA gene was not found, although 7.9% were positive for aggC. The CLA isolates reacted with the EAEC probe (55.6%), and the aggC, astA and pet gene sequences were found in 66.7%, 33.3% and 11.1%, respectively. The aggR (55.6%), aspU (55.6%), shf (33.3%) and she (22.2%) genes were also found in CLA isolates.
Assuntos
Aderência Bacteriana/fisiologia , Diarreia/microbiologia , Infecções por Escherichia coli/microbiologia , Escherichia coli/fisiologia , Escherichia coli/patogenicidade , Adesinas de Escherichia coli/genética , Adesinas de Escherichia coli/metabolismo , Brasil/epidemiologia , Criança , Pré-Escolar , Diarreia/epidemiologia , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Infecções por Escherichia coli/epidemiologia , Humanos , Prevalência , Células Tumorais CultivadasRESUMO
One hundred and five strains of Escherichia coli that were isolated from calves with diarrhea in the state of São Paulo, Brazil, and were negative for enterotoxins and cytotoxins, were examined for the eae gene. Four (3.8%) strains were positive by polymerase chain reaction (PCR) and were shown to produce intimin by using Western blot with specific antiserum against the conserved N-terminal region of intimin. Subtyping of the intimins was done by PCR with specific primers and by Western blot with specific antisera against the C-terminal variable region of the protein. Three of these isolates (O?:H11, O26:H-, O123:H1) produced the beta subtype of intimin, and the 4th (0103:H2) produced intimin that was not typable. The 0103:H2 and the O26:H-isolates adhered to HEp-2 cells with diffuse adherence and localized-like adherence patterns, respectively. The other strains did not adhere to HEp-2 cells. To our knowledge, this is the first report of the occurrence of a subtype of intimin described for human enteropathogenic E. coli among bovine diarrheogenic E. coli. It is also the first report from Brazil demonstrating the presence of bovine E. coli harboring the eae gene.
Assuntos
Adesinas Bacterianas , Proteínas da Membrana Bacteriana Externa/genética , Proteínas de Transporte , Doenças dos Bovinos/microbiologia , Diarreia/veterinária , Infecções por Escherichia coli/veterinária , Proteínas de Escherichia coli , Escherichia coli/genética , Animais , Brasil , Bovinos , Doenças dos Bovinos/genética , Escherichia coli/patogenicidade , Reação em Cadeia da PolimeraseRESUMO
BACKGROUND: In Brazil, enteropathogenic Escherichia coli (EPEC) diarrhoea is endemic in young infants. A characteristic feature of EPEC adhesion to host cells is intimate attachment leading to the formation of distinctive "attaching and effacing" (A/E) lesions on mammalian cells. Two genes directly involved in intimate adhesion, eae and tir, encode the adhesion molecule intimin and its translocated receptor Tir, respectively. The intimin-binding domain of Tir was recently mapped to the middle part of the polypeptide (Tir-M), and the amino (Tir-N) and carboxy (Tir-C) termini were found to be located within infected host cells. Recently, it was shown that colostrum samples from mothers living in Sao Paulo contain IgA-class antibodies reactive with a number of proteins associated with EPEC virulence. It has also been shown that patients infected with verocytotoxin-producing E. coli O157 can produce antibodies to Tir. In the current study antibody responses to the different Tir domains were analyzed in sera and colostrum samples collected in an EPEC-endemic area of Brazil. METHODS: Recombinant Tir, Tir-N, Tir-M, and Tir-C were expressed as His-tagged protein in E. coli BL21a and purified on nickel columns. Western blot analysis was used to investigate colostrum IgA- and serum IgG-class antibodies reactive with the Tir fragments. RESULTS: Anti-Tir IgG antibodies were detected in the serum of children, with (63%) or without (50%) diarrhoea. Anti-Tir IgA-class antibodies were detected in all the colostrum pools tested. With the use of both serum IgG- and colostrum IgA-class antibodies, an immunodominant domain of the Tir-polypeptide, Tir M, was identified. CONCLUSION: The intimin-binding region of Tir (Tir-M) is the immunodominant region of the polypeptide in humans. Both serum IgG-class and colostrum IgA-class antibodies reacted predominantly with the Tir-M domain.
Assuntos
Adesinas Bacterianas , Anticorpos Antibacterianos/análise , Anticorpos Antibacterianos/sangue , Proteínas de Transporte , Colostro/imunologia , Proteínas de Escherichia coli , Escherichia coli/imunologia , Receptores de Superfície Celular/imunologia , Proteínas da Membrana Bacteriana Externa/química , Proteínas da Membrana Bacteriana Externa/imunologia , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Bactérias/imunologia , Sítios de Ligação , Brasil , Mapeamento de Epitopos , Escherichia coli O157/imunologia , Feminino , Humanos , Imunoglobulina A/análise , Imunoglobulina G/sangue , Fragmentos de Peptídeos/imunologia , Proteínas Recombinantes/imunologiaRESUMO
Enteropathogenic Escherichia coli (EPEC) produces a plasmid-encoded type IV pilus, called the bundle-forming pilus (BFP), involved in the formation of the localized adhesion onto epithelial cells. In this study, we demonstrate that clinical isolates of serotypes O128ab:H2 and O119:H2 contain a ca. 13-kb deletion in the bfp operon, resulting in a lack of expression of these pili. An IS sequence with homology to the IS66 of Agrobacterium tumefaciens replaced the deleted bfp genes. These results suggest that the bfp operon was deleted through a transpositional event and that other adherence factors may mediate attachment of these bacteria to the host cells.
Assuntos
Escherichia coli/genética , Fímbrias Bacterianas/genética , Deleção de Genes , Óperon , Agrobacterium tumefaciens/genética , Criança , Mapeamento Cromossômico , Elementos de DNA Transponíveis , Diarreia/microbiologia , Escherichia coli/isolamento & purificação , Escherichia coli/patogenicidade , Escherichia coli/ultraestrutura , Infecções por Escherichia coli/microbiologia , Fímbrias Bacterianas/ultraestrutura , Humanos , Plasmídeos , Homologia de Sequência do Ácido NucleicoRESUMO
A total of 398 diffusely adhering Escherichia coli (DAEC) strains of fecal origin were analyzed for the presence of sequences homologous to the structural subunit gene (daaE) of the F1845 fimbria. For that purpose, a DNA fragment homologous to daaE, obtained by PCR, was used as a probe in colony hybridization assays. Only two strains carried daaE and expressed F1845, suggesting that this fimbria is rare among DAEC strains.
Assuntos
Adesinas de Escherichia coli/genética , Antígenos de Bactérias , Proteínas de Bactérias/genética , Diarreia/microbiologia , Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli , Escherichia coli/genética , Proteínas de Fímbrias , Adesinas de Escherichia coli/biossíntese , Proteínas de Bactérias/biossíntese , Pré-Escolar , Fezes/microbiologia , Expressão Gênica , Células HeLa , Humanos , LactenteRESUMO
The genetic relatedness among 96 invasive Escherichia coli belonging to several serogroups and 13 non-invasive of several serotypes that share the same O antigen was investigated by multilocus enzyme electrophoresis analysis. The invasive strains were isolated in different parts of the world and most of them recovered from dysentery. Twenty-nine electrophoretic types were distinguished and the most invasive strains were found to belong to two major lineages. These results suggested that the invasive ability in these strains has evolved in divergent chromosomal backgrounds, presumably through the horizontal spread of plasmid-borne invasion genes. The maintenance of invasive phenotypes in separate lineages suggests that this ability confers a selective advantage to invasive strains.
Assuntos
Escherichia coli/genética , Disenteria/microbiologia , Eletroforese em Gel de Amido/métodos , Escherichia coli/classificação , Escherichia coli/enzimologia , Humanos , Fenótipo , Polimorfismo Genético/genética , Análise de Sequência de DNA , SorotipagemRESUMO
Enteropathogenic Escherichia coli (EPEC) express a plasmid-encoded type IV pilus termed bundle-forming pilus, which is associated with the formation of bacterial microcolonies on cultured epithelial cells. Bacterial attachment and effacement of the enterocyte brush border membrane is attributed to a surface outer membrane protein adhesin termed intimin and EPEC-secreted proteins EspA, EspB, and EspD. Except for intimin, production in vivo or antibody response against these virulence determinants during natural EPEC infections in young children has not been demonstrated. Antibody responses against BfpA, intimin, EspA, and EspB were investigated in Brazilian children naturally infected with EPEC. Generally, IgG antibodies against BfpA and EspB were the most commonly found, followed by anti-EspA and intimin antibodies. Thus, bundle-forming pilus and locus of enterocyte attachment-encoded products are produced in vivo during natural EPEC infections and elicit an immune response against heterologous EPEC virulence determinants. These findings have important implications in the immunoprophylaxis against EPEC infections.
Assuntos
Adesinas Bacterianas , Anticorpos Antibacterianos/biossíntese , Proteínas de Transporte , Infecções por Escherichia coli/imunologia , Proteínas de Escherichia coli , Escherichia coli/imunologia , Escherichia coli/patogenicidade , Proteínas de Fímbrias , Fímbrias Bacterianas/imunologia , Antígenos de Bactérias/genética , Aderência Bacteriana/genética , Aderência Bacteriana/imunologia , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/imunologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Estudos de Casos e Controles , Pré-Escolar , Diarreia/imunologia , Diarreia/microbiologia , Diarreia/prevenção & controle , Células Epiteliais/microbiologia , Escherichia coli/genética , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/prevenção & controle , Fímbrias Bacterianas/genética , Humanos , Imunoglobulina G/biossíntese , Lactente , Intestinos/microbiologia , Virulência/genética , Virulência/imunologiaRESUMO
The penicillin-binding proteins of 11 pathogenic Escherichia coli strains, including enteropathogenic, enterotoxigenic, enteroinvasive, enteroaggregative, and enterohemorrhagic E. coli, were detected in gels following the labeling of isolated cell envelopes with [3H]benzylpenicillin. The electrophoretic profiles, sensitivities to and morphological changes induced by beta-lactam antibiotics showed that the penicillin-binding proteins of most pathogenic E. coli possess structural and physiological functions similar to those of E. coli K12.
Assuntos
Proteínas de Bactérias , Proteínas de Transporte/química , Proteínas de Transporte/metabolismo , Diarreia/microbiologia , Infecções por Escherichia coli/microbiologia , Escherichia coli/química , Hexosiltransferases , Muramilpentapeptídeo Carboxipeptidase/química , Muramilpentapeptídeo Carboxipeptidase/metabolismo , Peptidil Transferases , Antibacterianos/metabolismo , Antibacterianos/farmacologia , Proteínas de Transporte/isolamento & purificação , Eletroforese em Gel de Poliacrilamida , Escherichia coli/classificação , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Humanos , Lactamas , Testes de Sensibilidade Microbiana , Muramilpentapeptídeo Carboxipeptidase/isolamento & purificação , Proteínas de Ligação às Penicilinas , SorotipagemRESUMO
BACKGROUND: In Brazil, enteropathogenic Escherichia coli diarrhoea is endemic among infants born into low economic levels, and it is one of the main causes of morbidity and mortality in this group. Binding of enteropathogenic E. coli to the brush border mucosa triggers a cascade of transmembrane and intracellular signals, causing cytoskeletal reorganization and formation of a specific lesion, termed the attaching and effacing lesion. Several enteropathogenic E. coli gene products have been implicated in formation of attaching and effacing lesions. Evaluation of pathogen-specific protective factors shows that breast feeding is effective against enteropathogenic E. coli infection. To investigate the nature of the protection, defatted colostrum and secretory immunoglobulin A obtained from mothers living in Sao Paulo were investigated for the ability to recognise selected enteropathogenic E. coli-associated virulence factors. METHODS: Western blot analysis was used to investigate the IgA repertoire in pooled colostrum that is reactive with specific enteropathogenic E. coli proteins. Whole enteropathogenic E. coli bacterial cell extracts, nonpathogenic E. coli strains overexpressing specific virulence factors, and purified polypeptides were used as antigen sources in this study. RESULTS: Reaction of the colostrum samples in Western blots of whole bacterial cell extracts and selected purified enteropathogenic E. coli proteins showed that they contained a secretory immunoglobulin A reactive with all the virulence-associated proteins studied. CONCLUSION: These results suggest that maternal antibodies may protect infants from enteropathogenic E. coli infection by interfering with adherence processes (anti-intimin and anti-bundle-forming pili antibodies) and cell signaling (anti-enteropathogenic Escherichia coli-secreted protein A and B antibodies.
Assuntos
Adesinas Bacterianas , Anticorpos Antibacterianos/análise , Proteínas de Bactérias/imunologia , Proteínas de Transporte , Colostro/imunologia , Proteínas de Escherichia coli , Escherichia coli/imunologia , Proteínas de Fímbrias , Imunoglobulina A/análise , Adolescente , Adulto , Anticorpos Antibacterianos/imunologia , Proteínas da Membrana Bacteriana Externa/imunologia , Western Blotting , Brasil , Feminino , Humanos , Imunoglobulina A/imunologia , Transdução de SinaisRESUMO
Genetic variation of 33 enteroinvasive Escherichia coli (EIEC), 12 non-EIEC and 39 Shigella strains (representing the 4 species of this genus) was analyzed using the random amplified polymorphic DNA (RAPD) technique. Reproducible polymorphisms were generated and the combined data allowed us to construct a dendrogram using Jaccard's distance. Two main groups were obtained: one for Shigella and the other for EIEC and non-EIEC strains. The first group contained four clusters, one for each Shigella species. The second group contained one cluster for EIEC and another for non-EIEC strains. The main clusters encompassed many small clusters corresponding to different serotypes. It was possible to characterize each one of the 84 strains under study as well as the boundaries among Shigella species and between this genus and EIEC strains.
Assuntos
Escherichia coli/genética , Polimorfismo Genético/genética , Shigella/genética , Técnicas de Tipagem Bacteriana , Impressões Digitais de DNA , DNA Bacteriano/genética , Eletroforese em Gel de Ágar , Escherichia coli/classificação , Variação Genética , Humanos , Técnica de Amplificação ao Acaso de DNA Polimórfico , Shigella/classificaçãoRESUMO
All proteins involved in the attachment and effacement lesion produced by enteropathogenic Escherichia coli (EPEC) and Shiga-toxin producing E. coli (STEC) are encoded by the locus of enterocyte effacement (LEE). We studied the presence and insertion site of the LEE in different EPEC and STEC strains. In serotypes O119:H6/H-, O55:H6, O55:H7, O142:H6, O111ac:H9/H-, O111ab:H9/H- LEE is inserted downstream of selC as previously described for EPEC O127:H6 and STEC O157:H7. In serotypes O111ac:H8/H- and O26:H11/H- the LEE is inserted in pheU as previously described for STEC O26:H-. However in EPEC from serotype O111ab:H25 the LEE is not inserted in either site suggesting a third insertion site in the K12 chromosome. We also cloned fragments of 2.3 kb and 1.0 kb from the right and left hand sides of the LEE of a O111ac:H- strain and identified additional insertion sequences on these LEE fragments, suggesting that the LEE may be larger and may have undergone more recombination events in these serotypes.
Assuntos
Aderência Bacteriana/genética , Proteínas de Bactérias/genética , DNA Bacteriano/genética , Escherichia coli/genética , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/metabolismo , Escherichia coli/química , Humanos , Reação em Cadeia da Polimerase , Toxinas ShigaRESUMO
The ability of ribotyping and enterobacterial repetitive intergenic concensus-polymerase chain reaction (ERIC-PCR) to discriminate diarrhoeagenic Escherichia coli clones of 122 strains belonging to 26 distinct serotypes was evaluated. The 26 serotypes corresponded to 24 ribotypes and 25 ERIC-types. Correlation between multilocus enzyme electrophoresis, ERIC-PCR and ribotyping was c. 90% for the dominant ribotypes. Related clones such as O55:H7 and O157:H7 presented similar ribotypes and clustered together in a dendrogram, and the two divergent clonal groups of enteropathogenic E. coli (EPEC) and enterohaemorrhagic E. coli (EHEC) were included in distinct branches. The results suggest the possibility of applying these two simpler techniques as tools to identify clones of diarrhoeagenic E. coli.