Production of cytolethal distending toxin and other virulence characteristics of Escherichia coli strains of serogroup O86

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.

Treatment of severe infectious purpura in children with human plasma from donors immunized with Escherichia coli J5: a prospective double-blind study. J5 study Group.

To evaluate the efficacy of anti-J5 serum in the treatment of severe infectious purpura, 73 children were randomized to receive either anti-J5 (40) or control (33) plasma. Age, blood pressure, and biologic risk factors were similar in both groups. At admission, however, tumor necrosis factor serum concentrations were 974 +/- 173 pg/ml compared with 473 +/- 85 pg/ml (P = .023) and interleukin-6 serum concentrations were 129 +/- 45 compared with 19 +/- 5 ng/ml (P = .005) in the control and treated groups, respectively. The duration of shock and the occurrence of complications were similar in both groups. The mortality rate was 36% in the control group and 25% in the treated group (P = .317; odds ratio, 0.76; 95% confidence interval, 0.46-1.26). This trend disappeared after correction for unbalances in risk factors at randomization using a logistic regression model. These results suggest that anti-j5 plasma did not affect the course or mortality of severe infectious purpura in children.

Phenotypic and genotypic characteristics of shiga toxin-producing Escherichia coli strains isolated from children in São Paulo, Brazil

The biochemical and serological characteristics, virulence properties, and genetic relatedness of Shiga toxin-producing Escherichia coli (STEC) strains isolated in São Paulo, from April 1989 through March 1990, were determined. This is also the first report on clinic findings of human STEC infections in Brazil. The only three STEC strains identified in that period were lysine decarboxylase negative, belonged to serotype O111ac: non-motile, were Stx1 producers, carried the eae and astA genes, and 2 of them also presented the EHEC-hly sequence. The children carrying STEC were all boys, with less than two years old, and had no previous history of hospitalization. None of them presented blood in stools. Vomiting, cough and coryza were the most common clinical manifestations observed. Although the STEC strains were isolated during summer months, and presented similar phenotypic and genotypic characteristics, carbohydrate fermentation patterns and PFGE analysis suggested that these diarrheal episodes were not caused by a single clone.

Diarrheagenic Escherichia coli categories among the traditional enteropathogenic E. coli O serogroups: a review

The socalled enteropathogenic Escherichia coli (EPEC) O serogroups include typical and atypical EPEC, enterohaemorrragic E. coli, enterotoxigenic E. coli, and enteroaggregative E. coli. The aim of this article is to review the composition of each O serogroup and the major serotypes, clones, and additional virulence characteristics of each of these diarrheageniccategories. Their adherence patterns and genetic relationships are also presented. The review is based on the study of 805 strains of serogroups O26, O55, O86, O111, O114, O119, O125, O126, O1127, O128, and O142 most of which isolated in São Paulo from children with diarrhea between 1970 and 1990. Since some O serogroups include more than one diarrheageniccategory O serogrouping only should be abandoned as a diagnostic method. However serotyping is a reliable method for those serotypes that correspond to clones.

Engineering, conjugation, and immunogenicity assessment of Escherichia coli O121 O antigen for its potential use as a typhoid vaccine component.

State-of-the-art production technologies for conjugate vaccines are complex, multi-step processes. An alternative approach to produce glycoconjugates is based on the bacterial N-linked protein glycosylation system first described in Campylobacter jejuni. The C. jejuni N-glycosylation system has been successfully transferred into Escherichia coli, enabling in vivo production of customized recombinant glycoproteins. However, some antigenic bacterial cell surface polysaccharides, like the Vi antigen of Salmonella enterica serovar Typhi, have not been reported to be accessible to the bacterial oligosaccharyltransferase PglB, hence hamper development of novel conjugate vaccines against typhoid fever. In this report, Vi-like polysaccharide structures that can be transferred by PglB were evaluated as typhoid vaccine components. A polysaccharide fulfilling these requirements was found in Escherichia coli serovar O121. Inactivation of the E. coli O121 O antigen cluster encoded gene wbqG resulted in expression of O polysaccharides reactive with antibodies raised against the Vi antigen. The structure of the recombinantly expressed mutant O polysaccharide was elucidated using a novel HPLC and mass spectrometry based method for purified undecaprenyl pyrophosphate (Und-PP) linked glycans, and the presence of epitopes also found in the Vi antigen was confirmed. The mutant O antigen structure was transferred to acceptor proteins using the bacterial N-glycosylation system, and immunogenicity of the resulting conjugates was evaluated in mice. The conjugate-induced antibodies reacted in an enzyme-linked immunosorbent assay with E. coli O121 LPS. One animal developed a significant rise in serum immunoglobulin anti-Vi titer upon immunization.

Prevalence of diarrheagenic Escherichia coli in children with diarrhea in Salvador, Bahia, Brazil

We report the frequency of the different diarrheagenic Escherichia coli (DEC) categories isolated from children with acute endemic diarrhea in Salvador, Bahia. The E. coli isolates were investigated by colony blot hibridization whit the following genes probes: eae, EAF, bfpA, Stx1, Stx2, ST-Ih, ST-Ip, LT-I, LT-II, INV, and EAEC, as virulence markers to distinguish typical and atypical EPEC, EHEC/STEC, ETEC, EIEC, and EAEC. Seven of the eight categories of DEC were detected. The most frequently isolated was atypical EPEC (10.1%) followed by ETEC (7.5%), and EAEC (4.2%). EHEC, STEC, EIEC, and typical EPEC were each detected once. The strains of ETEC, EAEC, and atypical EPEC belonged to a wide variety of serotypes. The serotypes of the others categories were O26:H11 (EHEC), O21:H21 (STEC), O142:H34 (typical EPEC), and O?H55 (EIEC). We also present the clinical manifestations and other pathogenic species observed in children with DEC. This is the first report of EHEC and STEC in Salvador, and one of the first in Brazil.

Membrane fluidization triggers membrane remodeling which affects the thermotolerance in Escherichia coli.

Treatment of Escherichia coli with non-lethal doses of heat or benzyl alcohol (BA) causes transient membrane fluidization and permeabilization, and induces the rapid transcription of heat-shock genes in a sigma32-dependent manner. This early response is followed by a rapid adaptation (priming) of the cells to otherwise lethal elevated temperature, in strong correlation with an observed remodeling of the composition and alkyl chain unsaturation of membrane lipids. The acquisition of cellular thermotolerance in BA-primed cells is unrelated to protein denaturation and is not accompanied by the formation of major heat-shock proteins, such as GroEL and DnaK. This suggests that the rapid remodeling of membrane composition is sufficient for the short-term bacterial thermotolerance.

Genetic variability of hepatitis A virus strain HAF-203 isolated in Brazil and expression of the VP1 gene in Escherichia coli

The hepatitis A virus (HAV) HAF-203 strain was isolated from an acute case of HAV infection. The primary isolation of HAF-203 in Brazil and its adaptation to the FRhK-4 cell lineage allowed the production of large amounts of viral particles enabling molecular characterization of the first HAV isolate in Brazil. The aim of our study was to determine the nucleotide sequence of the HAF-203 strain genome, compare it to other HAV genomes and highlight its genetic variability. The complete nucleotide sequence of the HAF-203 strain (7472 nucleotides) was compared to those obtained earlier by others for other HAV isolates. These analyses revealed 19 HAF-specific nucleotide sequence differences with 10 amino acid substitutions. Most of the non-conservative changes were located at VP1, 2C, and 3D genes, but the 3B region was the most variable. The availability of HAF-203 complementary DNA was useful for the production of the recombinant VP1 protein, which is a major determinant of viral infectivity. This recombinant protein was shown by enzyme-linked immunoassay and blotting, to be immunogenic and resemble the native protein, therefore suggesting its value as a reagent for incorporation into diagnostic tests.

Production, characterization, and application of antibodies against heat-labile type-I toxin for detection of enterotoxigenic Escherichia coli

Strains of enterotoxigenic Escherichia coli (ETEC) are responsible for significant rates of morbidity and mortality among children, particularly in developing countries. The majority of clinical and public health laboratories are capable of isolating and identifying Salmonella, Shigella, Campylobacter, and Escherichia coli O157:H7 from stool samples, but ETEC cannot be identified by routine methods. The method most often used to identify ETEC is polymerase chain reaction for heat-stable and heat-labile enterotoxin genes, and subsequent serotyping, but most clinical and public health laboratories do not have the capacity or resources to perform these tests. In this study, polyclonal rabbit and monoclonal mouse IgG2b antibodies against ETEC heat-labile toxin-I (LT) were characterized and the potential applicability of a capture assay was analyzed. IgG-enriched fractions from rabbit polyclonal and the IgG2b monoclonal antibodies recognized LT in a conformational shape and they were excellent tools for detection of LT-producing strains. These findings indicate that the capture immunoassay could be used as a diagnostic assay of ETEC LT-producing strains in routine diagnosis and in epidemiological studies of diarrhea in developing countries as enzyme linked immunosorbent assay techniques remain as effective and economical choice for the detection of specific pathogen antigens in cultures.

Escherichia coli (E.coli) Bacteraemia Surveillance

Charts showing E.coli and ESBL Bacteraemias in Northern Ireland, by quarter, from 2005 to 2014.

Genetic relationship of diarrheagenic Escherichia coli pathotypes among the enteropathogenic Escherichia coli O serogroup

The genetic relationship among the Escherichia coli pathotypes was investigated. We used random amplified polymorphic DNA (RAPD) data for constructing a dendrogram of 73 strains of diarrheagenic E. coli. A phylogenetic tree encompassing 15 serotypes from different pathotypes was constructed using multilocus sequence typing data. Phylogram clusters were used for validating RAPD data on the clonality of enteropathogenic E. coli (EPEC) O serogroup strains. Both analyses showed very similar topologies, characterized by the presence of two major groups: group A includes EPEC H6 and H34 strains and group B contains the other EPEC strains plus all serotypes belonging to atypical EPEC, enteroaggregative E. coli (EAEC) and enterohemorrhagic E. coli (EHEC). These results confirm the existence of two evolutionary divergent groups in EPEC: one is genetically and serologically very homogeneous whereas the other harbors EPEC and non-EPEC serotypes. The same situation was found for EAEC and EHEC.

Detection of diarrheagenic Escherichia coli from children with and without diarrhea in Salvador, Bahia, Brazil

We identified different diarrheagenic (DEC) Escherichia coli pathotypes isolated from 1,207 children with and without acute endemic diarrhea in Salvador, Bahia, Brazil collected as part of a case-control study. Since the identification of DEC cannot be based on only biochemical and culture criteria, we used a multiplex polymerase chain reaction developed by combining five specific primer pairs for Enteropathogenic Escherichia coli (EPEC), Shiga toxin-producing E. coli/ Enterohaemorrhagic E. coli (STEC/EHEC), Enterotoxigenic E. coli (ETEC) and Enteroaggregative E. coli (EAEC) to detect these pathotypes simultaneously in a single-step reaction. In order to distinguish typical and atypical EPEC strains, these were tested for the presence of EAF plasmid. The prevalence of diarrheagenic E. coli in this sample of a global case-control study was 25.4% (259 patients) and 18.7% (35 patients) in the diarrhea group (1,020 patients) and the control group (187 patients), respectively. The most frequently isolated pathotype was EAEC (10.7%), followed by atypical EPEC (9.4%), ETEC (3.7%), and STEC (0.6%). Typical EPEC was detected only in one sample. The prevalence of the pathotypes studied in children with diarrhea was not significantly different from that in children without diarrhea.

In vitro activity of fosfomycin against extended-spectrum-beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae: comparison of susceptibility testing procedures

The agar dilution, broth microdilution, and disk diffusion methods were compared to determine the in vitro susceptibility of 428 extended-spectrum-beta-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae to fosfomycin. Fosfomycin showed very high activity against all ESBL-producing strains. Excellent agreement between the three susceptibility methods was found for E. coli, whereas marked discrepancies were observed for K. pneumoniae.

In Vivo Selection of Fluoroquinolone-Resistant Escherichia coli Isolates Expressing Plasmid-Mediated Quinolone Resistance and Expanded-Spectrum β-Lactamase

A ciprofloxacin-resistant Escherichia coli isolate, isolate 1B, was obtained from a urinary specimen of a Canadian patient treated with norfloxacin for infection due to a ciprofloxacin-susceptible isolate, isolate 1A. Both isolates harbored a plasmid-encoded sul1-type integron with qnrA1 and blaVEB-1 genes. Isolate 1B had amino acid substitutions in gyrase and topoisomerase.

Identification of uropathogenic Escherichia coli clonal group A (CgA) in hospitalised patients

This study provides the first description of healthcare-associated infections with Escherichia coli clonal group A (CgA) isolates in Latin America. Isolates were typed by enterobacterial repetitive intergenic consensus-PCR, pulsed-field gel electrophoresis, E. coli phylogenetic grouping, multilocus sequence typing and fimH single nucleotide polymorphism analysis. Out of 42 E. coli hospital isolates studied, three belonged to E. coli phylogenetic group D and ST69 and had fimH sequences identical to that of the CgA reference strain ATCC BAA-457. E. coli CgA is another potential source of resistant infections in hospitals.