scholarly journals CS5 Pilus Biosynthesis Genes from Enterotoxigenic Escherichia coli O115:H40

1999 ◽  
Vol 181 (18) ◽  
pp. 5847-5851 ◽  
Author(s):  
Thomas G. Duthy ◽  
Lothar H. Staendner ◽  
Paul A. Manning ◽  
Michael W. Heuzenroeder
Keyword(s):  
Escherichia Coli ◽  
Dna Sequences ◽  
Surface Expression ◽  
Open Reading Frames ◽  
Enterotoxigenic Escherichia Coli ◽  
Cell Surface Expression ◽  
E Coli ◽  
Pilus Biogenesis ◽  
K 12 ◽  
And Function

ABSTRACT We have sequenced the entire region of DNA required for the biosynthesis of CS5 pili from enterotoxigenic Escherichia coli O115:H40 downstream of the major subunit gene, designatedcsfA (for coli surface factor five A). Five more open reading frames (ORFs) (csfB, csfC,csfE, csfF, and csfD) which are transcribed in the same direction as the major subunit and are flanked by a number of insertion sequence regions have been identified. T7 polymerase-mediated overexpression of the cloned csf ORFs confirmed protein sizes based on the DNA sequences that encode them. The expression of only the csf region in E. coli K-12 resulted in the hemagglutination of human erythrocytes and the cell surface expression of CS5 pili, suggesting that the cluster contains all necessary information for CS5 pilus biogenesis and function.

scholarly journals afa-8 Gene Cluster Is Carried by a Pathogenicity Island Inserted into the tRNAPhe of Human and Bovine Pathogenic Escherichia coli Isolates

2001 ◽  
Vol 69 (2) ◽  
pp. 937-948 ◽  
Author(s):  
Lila Lalioui ◽  
Chantal Le Bouguénec
Keyword(s):  
Escherichia Coli ◽  
Direct Repeat ◽  
Pathogenicity Island ◽  
Genomic Region ◽  
Open Reading Frames ◽  
Blood Isolate ◽  
Distinctive Features ◽  
E Coli ◽  
Pathogenic Escherichia Coli ◽  
K 12

ABSTRACT We recently described a new afimbrial adhesin, AfaE-VIII, produced by animal strains associated with diarrhea and septicemia and by human isolates associated with extraintestinal infections. Here, we report that the afa-8 operon, encoding AfaE-VIII adhesin, from the human blood isolate Escherichia coli AL862 is carried by a 61-kb genomic region with characteristics typical of a pathogenicity island (PAI), including a size larger than 10 kb, the presence of an integrase-encoding gene, the insertion into a tRNA locus (pheR), and the presence of a small direct repeat at each extremity. Moreover, the G+C content of the afa-8 operon (46.4%) is lower than that of the E. coli K-12/MG1655 chromosome (50.8%). Within this PAI, designated PAI IAL862, we identified open reading frames able to code for products similar to proteins involved in sugar utilization. Four probes spanning these sequences hybridized with 74.3% of pathogenicafa-8-positive E. coli strains isolated from humans and animals, 25% of human pathogenic afa-8-negativeE. coli strains, and only 8% of fecal strains (P = 0.05), indicating that these sequences are strongly associated with the afa-8 operon and that this genetic association may define a PAI widely distributed among human and animal afa-8-positive strains. One of the distinctive features of this study is that E. coli AL862 also carries another afa-8-containing PAI (PAI IIAL862), which appeared to be similar in size and genetic organization to PAI IAL862 and was inserted into the pheV gene. We investigated the insertion sites of afa-8-containing PAI in human and bovine pathogenic E. coli strains and found that this PAI preferentially inserted into the pheV gene.

scholarly journals Identification of Candidates for a Subunit Vaccine against Extraintestinal Pathogenic Escherichia coli

2006 ◽  
Vol 75 (4) ◽  
pp. 1916-1925 ◽  
Author(s):  
Lionel Durant ◽  
Arnaud Metais ◽  
Coralie Soulama-Mouze ◽  
Jean-Marie Genevard ◽  
Xavier Nassif ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Subunit Vaccine ◽  
Iron Acquisition ◽  
Passive Immunization ◽  
Open Reading Frames ◽  
Lethal Challenge ◽  
E Coli ◽  
Protective Antigens ◽  
K 12 ◽  
Genomic Regions

ABSTRACT Extraintestinal pathogenic Escherichia coli (ExPEC) strains cause a large spectrum of infections. The majority of ExPEC strains are closely related to the B2 or the D phylogenetic group. The aim of our study was to develop a protein-based vaccine against these ExPEC strains. To this end, we identified ExPEC-specific genomic regions, using a comparative genome analysis, between the nonpathogenic E. coli strain K-12 MG1655 and ExPEC strains C5 (meningitis isolate) and CFT073 (urinary tract infection isolate). The analysis of these genomic regions allowed the selection of 40 open reading frames, which are conserved among B2/D clinical isolates and encode proteins with putative outer membrane localization. These genes were cloned, and recombinant proteins were purified and assessed as vaccine candidates. After immunization of BALB/c mice, five proteins induced a significant protective immunity against a lethal challenge with a clinical E. coli strain of the B2 group. In passive immunization assays, antigen-specific antibodies afforded protection to naive mice against a lethal challenge. Three of these antigens were related to iron acquisition metabolism, an important virulence factor of the ExPEC, and two corresponded to new, uncharacterized proteins. Due to the large number of genetic differences that exists between commensal and pathogenic strains of E. coli, our results demonstrate that it is possible to identify targets that elicit protective immune responses specific to those strains. The five protective antigens could constitute the basis for a preventive subunit vaccine against diseases caused by ExPEC strains.

Genes encoding osmoregulatory proline/glycine betaine transporters and the proline catabolic system are present and expressed in diverse clinical Escherichia coli isolates

1994 ◽  
Vol 40 (5) ◽  
pp. 397-402 ◽  
Author(s):  
Doreen E. Culham ◽  
Katherine S. Emmerson ◽  
Bonnie Lasby ◽  
Daniel Mamelak ◽  
Brian A. Steer ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Dna Sequences ◽  
Glycine Betaine ◽  
Clinical Isolates ◽  
E Coli ◽  
High Osmolality ◽  
Proline Catabolism ◽  
K 12 ◽  
Physiological Tests

Sixty-three clinical isolates identified as Escherichia coli, 30 from the human urinary tract and 33 derived from other human origins, were screened for proline/glycine betaine transporters similar to those that support proline catabolism and proline- or glycine betaine-based osmoregulation in E. coli K-12. Both molecular (DNA- and protein-based) analyses and physiological tests were performed. All tests were calibrated with E. coli K-12 derivatives from which genetic loci putP (encoding a proline transporter required for proline catabolism), proP, and (or) proU (loci encoding osmoregulatory proline/glycine betaine transporters) had been deleted. All clinical isolates showed both enhanced sensitivity to the toxic proline analogue azetidine-2-carboxylate on media of high osmolality and growth stimulation by glycine betaine in an artificial urine preparation of high osmolality. DNA sequences similar to the putP, proP, and proU loci of E. coli K-12 were detected by DNA amplification and (or) hybridization and protein specifically reactive with antibodies raised against the ProX protein of E. coli K-12 (a ProU constituent) was detected by western blotting in over 95% of the isolates. Two anomalous isolates were reclassified as non-E. coli on the basis of the API 20E series of tests. A protein immunochemically cross-reactive with the ProP protein of E. coli K-12 was also expressed by the clinical isolates. Since all three transporters were ubiquitous, no particular correlation between clinical origin and PutP, ProP, or ProU activity was observed. These data suggest that the transporters encoded in loci putP, proP, and proU perform housekeeping functions essential for the survival of E. coli cells in diverse habitats.Key words: osmoregulation, betaine transport, urinary tract infection, Escherichia coli.

scholarly journals Gene Cluster for Assembly of Pilus Colonization Factor Antigen III of Enterotoxigenic Escherichia coli

2001 ◽  
Vol 69 (9) ◽  
pp. 5864-5873 ◽  
Author(s):  
Tooru Taniguchi ◽  
Yukihiro Akeda ◽  
Ayako Haba ◽  
Yoko Yasuda ◽  
Koichiro Yamamoto ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Nucleotide Sequence ◽  
Gene Cluster ◽  
Enterotoxigenic Escherichia Coli ◽  
Human Colon Carcinoma Cell ◽  
E Coli ◽  
Colonization Factor ◽  
Type Iv ◽  
K 12 ◽  
Factor Antigen

ABSTRACT The assembly of pilus colonization factor antigen III (CFA/III) of enterotoxigenic Escherichia coli (ETEC) requires the processing of CFA/III major pilin (CofA) by a prepilin peptidase (CofP), similar to other type IV pilus formation systems. CofA is produced initially as a 26.5-kDa preform pilin (prepilin) and then processed to a 20.5-kDa mature pilin by CofP which is predicted to be localized in the inner membrane. In the present experiment, we determined the nucleotide sequence of the whole region for CFA/III formation and identified a cluster of 14 genes, includingcofA and cofP. Several proteins encoded bycof genes were similar to previously described proteins, such as the toxin-coregulated pili of Vibrio cholerae and the bundle-forming pili of enteropathogenic E. coli. The G+C content of the cof gene cluster was 37%, which was significantly lower than the average for the E. coli genome (50%). The introduction of a recombinant plasmid containing thecof gene cluster into the E. coli K-12 strain conferred CFA/III biogenesis and the ability of adhesion to the human colon carcinoma cell line Caco-2. This is the first report of a complete nucleotide sequence of the type IV pili found in human ETEC, and our results provide a useful model for studying the molecular mechanism of CFA/III biogenesis and the role of CFA/III in ETEC infection.

Some Characteristics of the Outer Membrane Material Released by Growing Enterotoxigenic Escherichia coli

1980 ◽  
Vol 29 (2) ◽  
pp. 704-713 ◽  
Author(s):  
Henk Gankema ◽  
Jan Wensink ◽  
Pieter A. M. Guinée ◽  
Wim H. Jansen ◽  
Bernard Witholt
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Freeze Fracture ◽  
Sucrose Density Gradient ◽  
Enterotoxigenic Escherichia Coli ◽  
E Coli ◽  
Freeze Fracture Electron Microscopy ◽  
K 12

The high-molecular-weight material released into the medium by Escherichia coli AP1, an enterotoxigenic strain of porcine origin, has been isolated and resolved into two clearly distinct fractions, based on sucrose density gradient and differential centrifugation, chemical analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and freeze-fracture electron microscopy. These two fractions, referred to as “medium vesicles” and “medium lipopolysaccharides”, were compared with the cellular outer and cytoplasmic membranes, the periplasmic fraction, and the cytoplasmic fraction. The medium vesicles closely resembled outer membrane and accounted for 3 to 5% of the total cellular outer membrane. They contained most of the heat-labile enterotoxin (LT) activity released into the medium by E. coli AP1. The medium lipopolysaccharide consisted mostly of lipopolysaccharide and a small amount of outer membrane and contained relatively little LT activity. Based on experiments with E. coli K-12 strains, in which about 5% of the newly synthesized outer membrane is lost from areas of outer membrane synthesis, it is proposed that enterotoxigenic E. coli strains release LT as part of such newly synthesized outer membrane fragments and that released outer membrane fragments may function as physiologically significant LT carriers.

scholarly journals Representational Difference Analysis between Afa/Dr Diffusely Adhering Escherichia coli and Nonpathogenic E. coli K-12

2002 ◽  
Vol 70 (10) ◽  
pp. 5503-5511 ◽  
Author(s):  
Anne-Beatrice Blanc-Potard ◽  
Colin Tinsley ◽  
Isabel Scaletsky ◽  
Chantal Le Bouguenec ◽  
Julie Guignot ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Sequences ◽  
Effector Proteins ◽  
Representational Difference Analysis ◽  
Transport Systems ◽  
Difference Analysis ◽  
E Coli ◽  
Tract Infections ◽  
K 12 ◽  
Specific Sequences

ABSTRACT Diffusely adhering Escherichia coli strains harboring Afa/Dr adhesins (Afa/Dr DAEC) have been associated with diarrhea and urinary tract infections (UTIs). The present work is the first extensive molecular study of a Afa/Dr DAEC strain using the representational difference analysis technique. We have searched for DNA sequences present in strain C1845, recovered from a diarrheagenic child, but absent from a nonpathogenic K-12 strain. Strain C1845 harbors part of a pathogenicity island (PAICFT073) and several iron transport systems found in other E. coli pathovars. We did not find genes encoding factors known to subvert host cell proteins, such as type III secretion system or effector proteins. Several C1845-specific sequences are homologous to putative virulence genes or show no homology with known sequences, and we have analyzed their distribution among Afa/Dr and non-Afa/Dr clinical isolates and among strains from the E. coli Reference Collection. Three C1845-specific sequences (MO30, S109, and S111) have a high prevalence (77 to 80%) among Afa/Dr strains and a low prevalence (12 to 23%) among non-Afa/Dr strains. In addition, our results indicate that strain IH11128, an Afa/Dr DAEC strain recovered from a patient with a UTI, is genetically closely related to strain C1845.

scholarly journals Genetic Structure and Distribution of Four Pathogenicity Islands (PAI I536 to PAI IV536) of Uropathogenic Escherichia coli Strain 536

2002 ◽  
Vol 70 (11) ◽  
pp. 6365-6372 ◽  
Author(s):  
Ulrich Dobrindt ◽  
Gabriele Blum-Oehler ◽  
Gabor Nagy ◽  
György Schneider ◽  
André Johann ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Genetic Structure ◽  
Dna Sequences ◽  
Coli Strain ◽  
Open Reading Frames ◽  
Pathogenicity Islands ◽  
Virulence Determinants ◽  
Core Element ◽  
E Coli ◽  
Virulence Potential

ABSTRACT For the uropathogenic Escherichia coli strain 536 (O6:K15:H31), the DNA sequences of three pathogenicity islands (PAIs) (PAI I536 to PAI III536) and their flanking regions (about 270 kb) were determined to further characterize the virulence potential of this strain. PAI I536 to PAI III536 exhibit features typical of PAIs, such as (i) association with tRNA-encoding genes; (ii) G+C content differing from that of the host genome; (iii) flanking repeat structures; (iv) a mosaic-like structure comprising a multitude of functional, truncated, and nonfunctional putative open reading frames (ORFs) with known or unknown functions; and (v) the presence of many fragments of mobile genetic elements. PAI I536 to PAI III536 range between 68 and 102 kb in size. Although these islands contain several ORFs and known virulence determinants described for PAIs of other extraintestinal pathogenic E. coli (ExPEC) isolates, they also consist of as-yet-unidentified ORFs encoding putative virulence factors. The genetic structure of PAI IV536, which represents the core element of the so-called high-pathogenicity island encoding a siderophore system initially identified in pathogenic yersiniae, was further characterized by sample sequencing. For the first time, multiple PAI sequences (PAI I536 to PAI IV536) in uropathogenic E. coli were studied and their presence in several wild-type E. coli isolates was extensively investigated. The results obtained suggest that these PAIs or at least large fragments thereof are detectable in other pathogenic E. coli isolates. These results support our view that the acquisition of large DNA regions, such as PAIs, by horizontal gene transfer is an important factor for the evolution of bacterial pathogens.

scholarly journals The Iron- and Temperature-RegulatedcjrBC Genes of Shigella and Enteroinvasive Escherichia coli Strains Code for Colicin Js Uptake

2001 ◽  
Vol 183 (13) ◽  
pp. 3958-3966 ◽  
Author(s):  
David Šmajs ◽  
George M. Weinstock
Keyword(s):  
Escherichia Coli ◽  
Open Reading Frames ◽  
Cosmid Library ◽  
E Coli ◽  
Order Of Magnitude ◽  
Fur Protein ◽  
The Difference ◽  
K 12 ◽  
Reading Frames ◽  
Made In

ABSTRACT A cosmid library of DNA from colicin Js-sensitive enteroinvasiveEscherichia coli (EIEC) strain O164 was made in colicin Js-resistant strain E. coli VCS257, and colicin Js-sensitive clones were identified. Sensitivity to colicin Js was associated with the carriage of a three-gene operon upstream of and partially overlapping senB. The open reading frames were designated cjrABC (for colicin Js receptor), coding for proteins of 291, 258, and 753 amino acids, respectively. Tn7 insertions in any of them led to complete resistance to colicin Js. A near-consensus Fur box was found upstream ofcjrA, suggesting regulation of the cjroperon by iron levels. CjrA protein was homologous to iron-regulatedPseudomonas aeruginosa protein PhuW, whose function is unknown; CjrB was homologous to the TonB protein fromPseudomonas putida; and CjrC was homologous to a putative outer membrane siderophore receptor from Campylobacter jejuni. Cloning experiments showed that the cjrBand cjrC genes are sufficient for colicin Js sensitivity. Uptake of colicin Js into sensitive bacteria was dependent on the ExbB protein but not on the E. coli K-12 TonB and TolA, -B, and -Q proteins. Sensitivity to colicin Js is positively regulated by temperature via the VirB protein and negatively controlled by the iron source through the Fur protein. Among EIEC strains, two types of colicin Js-sensitive phenotypes were identified that differed in sensitivity to colicin Js by 1 order of magnitude. The difference in sensitivity to colicin Js is not due to differences between the sequences of the CjrB and CjrC proteins.

A surface-exposed DraD protein of uropathogenic Escherichia coli bearing Dr fimbriae may be expressed and secreted independently from DraC usher and DraE adhesin

Microbiology ◽  
2005 ◽  
Vol 151 (7) ◽  
pp. 2477-2486 ◽  
Author(s):  
Beata Zalewska ◽  
Rafał Piątek ◽  
Katarzyna Bury ◽  
Alfred Samet ◽  
Bogdan Nowicki ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Bacterial Cell ◽  
Surface Expression ◽  
Uropathogenic Escherichia Coli ◽  
Bacterial Attachment ◽  
Open Reading Frames ◽  
E Coli ◽  
Fimbrial Subunit ◽  
Reading Frames

The dra gene cluster, expressed by uropathogenic Escherichia coli strains, determines bacterial attachment and invasion. The Dr fimbrial structures formed at the bacterial cell surface are composed of DraE subunits. The Dr fimbriae-coding cluster contains six open reading frames – draA, draB, draC, draD, draP and draE – among which the draE gene encodes the structural fimbrial subunit DraE. Very little is known about E. coli surface expression of the draD gene product. The expression of DraD and its role in the biogenesis of Dr fimbriae were determined by constructing mutants in the dra operon and by immunoblot and immunofluorescence experiments. In this study, DraD was found to be a surface-exposed protein. The expression of DraD was independent of the DraC usher and DraE fimbrial subunits. Polymerization of DraE fimbrial subunits into fimbrial structures did not require expression of the DraD protein.

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