scholarly journals Human Diffusely Adhering Escherichia coli Expressing Afa/Dr Adhesins That Use Human CD55 (Decay-Accelerating Factor) as a Receptor Does Not Bind the Rodent and Pig Analogues of CD55

2004 ◽  
Vol 72 (8) ◽  
pp. 4859-4863 ◽  
Author(s):  
Sylvie Hudault ◽  
O. Brad Spiller ◽  
B. Paul Morgan ◽  
Alain L. Servin
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Cho Cells ◽  
Chinese Hamster ◽  
Intestinal Cells ◽  
Gastrointestinal Infections ◽  
Decay Accelerating Factor ◽  
E Coli ◽  
Empty Vector ◽  
Human Intestinal Cells

ABSTRACT Afa/Dr diffusely adhering Escherichia coli (DAEC) bacteria that are responsible for recurrent urinary tract and gastrointestinal infections recognized as a receptor the glycosylphosphatidylinositol (GPI)-anchored protein decay-accelerating factor (DAF; CD55) at the brush border of cultured human intestinal cells. Results show that Afa/Dr DAEC C1845 bacteria were poorly associated with the mucosa of the gastrointestinal tract of infected mice. We conducted experiments with Chinese hamster ovary (CHO) cells stably transfected with mouse (GPI or transmembrane forms), pig, or human CD55 or mouse Crry cDNAs or transfected with empty vector pDR2EF1α. Recombinant E. coli AAEC185 bacteria expressing Dr or F1845 adhesins bound strongly to CHO cells expressing human CD55 but not to the CHO cells expressing mouse (transmembrane and GPI anchored), rat, or pig CD55 or mouse Crry. Positive clustering of CD55 around Dr-positive bacteria was observed in human CD55-expressing CHO cells but not around the rarely adhering Dr-positive bacteria randomly distributed at the cell surface of CHO cells expressing mouse, rat, or pig CD55.

scholarly journals Role of Decay-Accelerating Factor Domains and Anchorage in Internalization of Dr-Fimbriated Escherichia coli

2000 ◽  
Vol 68 (3) ◽  
pp. 1391-1399 ◽  
Author(s):  
R. Selvarangan ◽  
P. Goluszko ◽  
V. Popov ◽  
J. Singhal ◽  
T. Pham ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Interaction Model ◽  
Electron Microscopic ◽  
Decay Accelerating Factor ◽  
Glycosylphosphatidylinositol Anchor ◽  
E Coli ◽  
Short Consensus Repeat

ABSTRACT Dr-fimbriated Escherichia coli capable of invading epithelial cells recognizes human decay-accelerating factor (DAF) as its cellular receptor. The role of extracellular domains and the glycosylphosphatidylinositol anchor of DAF in the process of internalization of Dr+ E. coli was characterized in a cell-cell interaction model. Binding of Dr+ E. coli to the short consensus repeat 3 domain of DAF expressed by Chinese hamster ovary cells was critical for internalization to occur. Deletion of short consensus repeat 3 domain or replacement of Ser165 by Leu in this domain, or the use of a monoclonal antibody to this region abolished internalization. Replacing the glycosylphosphatidylinositol anchor of DAF with the transmembrane anchor of membrane cofactor protein or HLA-B44 resulted in abolition or reduction of internalization respectively. Cells expressing glycosylphosphatidylinositol-anchored DAF but not the transmembrane-anchored DAF internalized Dr+ E. coli through a glycolipid pathway, since the former cells were more sensitive to inhibition by methyl-β-cyclodextrin, a sterol-chelating agent. Electron microscopic studies revealed that the intracellular vacuoles containing the internalized Dr+ E. coli were morphologically distinct between the anchor variants of DAF. The cells expressing glycosylphosphatidylinositol-anchored DAF contained a single bacterium in tight-fitting vacuoles, while the cells expressing transmembrane-anchored DAF contained multiple (two or three) bacteria in spacious phagosomes. This finding suggests that distinct postendocytic events operate in the cells expressing anchor variants of DAF. We provide direct evidence for the DAF-mediated internalization of Dr+ E. coli and demonstrate the significance of the glycosylphosphatidylinositol anchor, which determines the ability and efficiency of the internalization event.

scholarly journals Assay method for Vibrio cholerae and Escherichia coli enterotoxins by automated counting of floating chinese hamster ovary cells in culture medium

1978 ◽  
Vol 7 (5) ◽  
pp. 479-485
Author(s):  
R T Nozawa ◽  
T Yokota ◽  
S Kuwahara
Keyword(s):  
Escherichia Coli ◽  
Chinese Hamster Ovary ◽  
Cho Cells ◽  
Culture Medium ◽  
Culture Media ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Assay Method ◽  
Ovary Cells ◽  
E Coli

As Chinese hamster ovary (CHO) cells on plastic proliferate, many cells float off into the medium instead of piling up after they form a monolayer. Fewer cells were floating in the medium when CHO cells were incubated with cholera toxin at a concentration as low as 10 pg/ml. The toxin increased the adhesiveness of the cells forming confluent monolayers so that the floating cells accumulated on the adherent monolayers. On the basis of this finding, a simple, quantitative assay method for cholera and Escherichia coli enterotoxins was devised by cultivating CHO cells in a Linbro multidish and counting the cells in the medium with a Coulter Counter. The method was sensitive enough to detect toxins in 100- to 200-fold-diluted culture media of toxigenic E. coli strains. Little or no activity was detected by this method in the culture medium of nontoxigenic E. coli.

scholarly journals Escherichia coli B2 Phylogenetic Subgroups in the Infant Gut Microbiota: Predominance of Uropathogenic Lineages in Swedish Infants and Enteropathogenic Lineages in Pakistani Infants

2019 ◽  
Vol 85 (24) ◽  
Author(s):  
Forough L. Nowrouzian ◽  
Olivier Clermont ◽  
Mona Edin ◽  
Anna Östblom ◽  
Erick Denamur ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Gut Microbiota ◽  
Urinary Tract Infections ◽  
Intestinal Microbiota ◽  
Gastrointestinal Infections ◽  
Content Type ◽  
E Coli ◽  
Virulence Markers ◽  
Tract Infections

ABSTRACT Escherichia coli segregates into phylogenetic groups, with group B2 containing both extraintestinal pathogenic E. coli (ExPEC) and enteropathogenic E. coli (EPEC) strains. Ten main B2 subgroups (subgroups I to X)/sequence type complexes (STcs), as well as EPEC lineages, have been identified. In the current study, we characterized ExPEC and EPEC strains of E. coli B2 phylogenetic subgroups/STcs that colonize Swedish and Pakistani infants. Gut commensal E. coli B2 strains, 120 from Swedish infants (n = 87) and 19 from Pakistani infants (n = 12), were assigned to B2 subgroups. Carriage of the bundle-forming pili and intimin adhesin was examined in the EPEC lineages. The ExPEC virulence markers and the time of persistence of the strains in the microbiota were previously determined. In total, 84% of the Swedish strains and 47% of the Pakistani strains belonged to 1 of the 10 main B2 subgroups (P = 0.001). Among the Swedish strains, the most common B2 subgroups were IX/STc95 (19%), II/STc73 (17%), VI/STc12 (13%), and III/STc127 (11%), with each subgroup carrying distinctive sets of ExPEC virulence markers. EPEC lineages with few ExPEC features constituted 47% of the Pakistani B2 strains but only 7% of the Swedish B2 strains (P = 0.0001). The subgroup distribution within phylogenetic group B2 strains colonizing the gut differed between Swedish and Pakistani infants. B2 subgroups with uropathogenic characteristics dominated the gut microbiota of Swedish infants, while EPEC lineage 1 strains frequently colonized the intestines of Pakistani infants. Moreover, within the B2 subgroups, ExPEC virulence genes were more prevalent in Swedish strains than in Pakistani strains. Thus, ExPEC traits exemplify the intestinal B2 strains from Western populations. IMPORTANCE The intestinal microbiota is an important reservoir for bacteria that cause extraintestinal infections. Escherichia coli is found ubiquitously in the gut microbiota, and it also causes urinary tract infections, infantile septicemia, and meningitis. Urinary tract infections are usually caused by E. coli strains that originate in the intestinal microbiota. E. coli also causes gastrointestinal infections and is a major cause of diarrhea in infants worldwide. The abilities of certain E. coli strains to cause infections are attributed to their virulence factors, i.e., bacterial components that contribute to the development of different diseases. Our study shows that different subtypes of potentially pathogenic E. coli strains dominate in the gut microbiota of infants in different geographical areas and expands our knowledge of the interplay between bacterial commensalism and pathogenicity.

scholarly journals Iron Acquisition of Urinary Tract Infection Escherichia coli Involves Pathogenicity in Caenorhabditis elegans

2021 ◽  
Vol 9 (2) ◽  
pp. 310
Author(s):  
Masayuki Hashimoto ◽  
Yi-Fen Ma ◽  
Sin-Tian Wang ◽  
Chang-Shi Chen ◽  
Ching-Hao Teng
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Caenorhabditis Elegans ◽  
Large Scale ◽  
Iron Acquisition ◽  
Infection Model ◽  
High Throughput Analysis ◽  
E Coli ◽  
C Elegans ◽  
Tract Infections

Uropathogenic Escherichia coli (UPEC) is a major bacterial pathogen that causes urinary tract infections (UTIs). The mouse is an available UTI model for studying the pathogenicity; however, Caenorhabditis elegans represents as an alternative surrogate host with the capacity for high-throughput analysis. Then, we established a simple assay for a UPEC infection model with C. elegans for large-scale screening. A total of 133 clinically isolated E. coli strains, which included UTI-associated and fecal isolates, were applied to demonstrate the simple pathogenicity assay. From the screening, several virulence factors (VFs) involved with iron acquisition (chuA, fyuA, and irp2) were significantly associated with high pathogenicity. We then evaluated whether the VFs in UPEC were involved in the pathogenicity. Mutants of E. coli UTI89 with defective iron acquisition systems were applied to a solid killing assay with C. elegans. As a result, the survival rate of C. elegans fed with the mutants significantly increased compared to when fed with the parent strain. The results demonstrated, the simple assay with C. elegans was useful as a UPEC infectious model. To our knowledge, this is the first report of the involvement of iron acquisition in the pathogenicity of UPEC in a C. elegans model.

scholarly journals Characterization of Anti-Bacterial Effect of the Two New Phages against Uropathogenic Escherichia coli

Viruses ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1348
Author(s):  
Lívia Slobodníková ◽  
Barbora Markusková ◽  
Michal Kajsík ◽  
Michal Andrezál ◽  
Marek Straka ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Urinary Tract Infections ◽  
Therapeutic Potential ◽  
Medical Intervention ◽  
Uropathogenic Escherichia Coli ◽  
E Coli ◽  
Phage Cocktail ◽  
Causative Agents ◽  
Tract Infections

Urinary tract infections (UTIs) are among the events that most frequently need medical intervention. Uropathogenic Escherichia coli are frequently their causative agents and the infections are sometimes complicated by the presence of polyresistant nosocomial strains. Phage therapy is a tool that has good prospects for the treatment of these infections. In the present study, we isolated and characterized two bacteriophages with broad host specificity against a panel of local uropathogenic E. coli strains and combined them into a phage cocktail. According to genome sequencing, these phages were closely related and belonged to the Tequatrovirus genus. The newly isolated phages showed very good activity on a panel of local clinical E. coli strains from urinary tract infections. In the form of a two-phage cocktail, they were active on E. coli strains belonging to phylogroups B2 and D, with relatively lower activity in B1 and no response in phylogroup A. Our study is a preliminary step toward the establishment of a national phage bank containing local, well-characterized phages with therapeutic potential for patients in Slovakia.

Viability and survival capability of quinolone-resistant uropathogenic Escherichia coli

2010 ◽  
Vol 5 (6) ◽  
pp. 827-830
Author(s):  
Georgi Slavchev ◽  
Nadya Markova
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Expression System ◽  
Antibiotic Sensitivity ◽  
Uropathogenic Escherichia Coli ◽  
Resistant Bacteria ◽  
E Coli ◽  
Serum Bactericidal Assay ◽  
Tract Infections ◽  
Macrophage Killing

AbstractUropathogenic strains of E. coli isolated from urine of patients with urinary tract infections were tested for antibiotic sensitivity using bio-Merieux kits and ATB-UR 5 expression system. The virulence of strains was evaluated by serum bactericidal assay, macrophage “killing” and bacterial adhesive tests. Survival capability of strains was assessed under starvation in saline. The results showed that quinolone-resistant uropathogenic strains of E. coli exhibit significantly reduced adhesive potential but relatively high resistance to serum and macrophage bactericidity. In contrast to laboratory strains, the quinolone-resistant uropathogenic clinical isolate demonstrated increased viability during starvation in saline. Our study suggests that quinolone-resistant uropathogenic strains are highly adaptable clones of E. coli, which can exhibit compensatory viability potential under unfavorable conditions. The clinical occurrence of such phenotypes is likely to contribute to the survival, persistence and spread strategy of resistant bacteria.

scholarly journals Single Clinical Isolates from Acute Uncomplicated Urinary Tract Infections Are Representative of Dominant In Situ Populations

mBio ◽  
2014 ◽  
Vol 5 (2) ◽  
Author(s):  
Dana Willner ◽  
Serene Low ◽  
Jason A. Steen ◽  
Narelle George ◽  
Graeme R. Nimmo ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Urinary Tract Infections ◽  
Clinical Isolates ◽  
Content Type ◽  
E Coli ◽  
Strain Diversity ◽  
Amplicon Pyrosequencing ◽  
Culture Independent ◽  
Tract Infections

ABSTRACTUrinary tract infections (UTIs) are one of the most commonly acquired bacterial infections in humans, and uropathogenicEscherichia colistrains are responsible for over 80% of all cases. The standard method for identification of uropathogens in clinical laboratories is cultivation, primarily using solid growth media under aerobic conditions, coupled with morphological and biochemical tests of typically a single isolate colony. However, these methods detect only culturable microorganisms, and characterization is phenotypic in nature. Here, we explored the genotypic identity of communities in acute uncomplicated UTIs from 50 individuals by using culture-independent amplicon pyrosequencing and whole-genome and metagenomic shotgun sequencing. Genus-level characterization of the UTI communities was achieved using the 16S rRNA gene (V8 region). Overall UTI community richness was very low in comparison to other human microbiomes. We strain-typedEscherichia-dominated UTIs using amplicon pyrosequencing of the fimbrial adhesin gene,fimH. There were nine highly abundantfimHtypes, and each UTI sample was dominated by a single type. Molecular analysis of the corresponding clinical isolates revealed that in the majority of cases the isolate was representative of the dominant taxon in the community at both the genus and the strain level. Shotgun sequencing was performed on a subset of eightE. coliurine UTI and isolate pairs. The majority of UTI microbial metagenomic sequences mapped to isolate genomes, confirming the results obtained using phylogenetic markers. We conclude that for the majority of acute uncomplicatedE. coli-mediated UTIs, single cultured isolates are diagnostic of the infection.IMPORTANCEIn clinical practice, the diagnosis and treatment of acute uncomplicated urinary tract infection (UTI) are based on analysis of a single bacterial isolate cultured from urine, and it is assumed that this isolate represents the dominant UTI pathogen. However, these methods detect only culturable bacteria, and the existence of multiple pathogens as well as strain diversity within a single infection is not examined. Here, we explored bacteria present in acute uncomplicated UTIs using culture-independent sequence-based methods.Escherichia coliwas the most common organism identified, and analysis ofE. colidominant UTI samples and their paired clinical isolates revealed that in the majority of infections the cultured isolate was representative of the dominant taxon at both the genus and the strain level. Our data demonstrate that in most cases single cultured isolates are diagnostic of UTI and are consistent with the notion of bottlenecks that limit strain diversity during UTI pathogenesis.

scholarly journals degS Is Necessary for Virulence and Is among Extraintestinal Escherichia coli Genes Induced in Murine Peritonitis

2003 ◽  
Vol 71 (6) ◽  
pp. 3088-3096 ◽  
Author(s):  
Peter Redford ◽  
Paula L. Roesch ◽  
Rodney A. Welch
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract Infection ◽  
Urinary Tract ◽  
Tract Infection ◽  
Luria Bertani ◽  
Broth Culture ◽  
Wild Type ◽  
E Coli ◽  
Virulence Attenuation

ABSTRACT Extraintestinal Escherichia coli strains cause meningitis, sepsis, urinary tract infection, and other infections outside the bowel. We examined here extraintestinal E. coli strain CFT073 by differential fluorescence induction. Pools of CFT073 clones carrying a CFT073 genomic fragment library in a promoterless gfp vector were inoculated intraperitoneally into mice; bacteria were recovered by lavage 6 h later and then subjected to fluorescence-activated cell sorting. Eleven promoters were found to be active in the mouse but not in Luria-Bertani (LB) broth culture. Three are linked to genes for enterobactin, aerobactin, and yersiniabactin. Three others are linked to the metabolic genes metA, gltB, and sucA, and another was linked to iha, a possible adhesin. Three lie before open reading frames of unknown function. One promoter is associated with degS, an inner membrane protease. Mutants of the in vivo-induced loci were tested in competition with the wild type in mouse peritonitis. Of the mutants tested, only CFT073 degS was found to be attenuated in peritoneal and in urinary tract infection, with virulence restored by complementation. CFT073 degS shows growth similar to that of the wild type at 37°C but is impaired at 43°C or in 3% ethanol LB broth at 37°C. Compared to the wild type, the mutant shows similar serum survival, motility, hemolysis, erythrocyte agglutination, and tolerance to oxidative stress. It also has the same lipopolysaccharide appearance on a silver-stained gel. The basis for the virulence attenuation is unclear, but because DegS is needed for σE activity, our findings implicate σE and its regulon in E. coli extraintestinal pathogenesis.

scholarly journals Human milk glycosaminoglycans inhibit in vitro the adhesion of Escherichia coli and Salmonella fyris to human intestinal cells

2015 ◽  
Vol 79 (4) ◽  
pp. 603-607 ◽  
Author(s):  
Giovanni V. Coppa ◽  
Bruna Facinelli ◽  
Gloria Magi ◽  
Emanuela Marini ◽  
Lucia Zampini ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Human Milk ◽  
Intestinal Cells ◽  
Human Intestinal Cells

scholarly journals Biological Cost of Single and Multiple Norfloxacin Resistance Mutations in Escherichia coli Implicated in Urinary Tract Infections

2005 ◽  
Vol 49 (6) ◽  
pp. 2343-2351 ◽  
Author(s):  
Patricia Komp Lindgren ◽  
Linda L. Marcusson ◽  
Dorthe Sandvang ◽  
Niels Frimodt-Møller ◽  
Diarmaid Hughes
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Infection Model ◽  
Resistance Mutations ◽  
Topoisomerase Iv ◽  
E Coli ◽  
Tract Infections ◽  
Subsequent Selection

ABSTRACT Resistance to fluoroquinolones in urinary tract infection (UTIs) caused by Escherichia coli is associated with multiple mutations, typically those that alter DNA gyrase and DNA topoisomerase IV and those that regulate AcrAB-TolC-mediated efflux. We asked whether a fitness cost is associated with the accumulation of these multiple mutations. Mutants of the susceptible E. coli UTI isolate Nu14 were selected through three to five successive steps with norfloxacin. Each selection was performed with the MIC of the selected strain. After each selection the MIC was measured; and the regions of gyrA, gyrB, parC, and parE, previously associated with resistance mutations, and all of marOR and acrR were sequenced. The first selection step yielded mutations in gyrA, gyrB, and marOR. Subsequent selection steps yielded mutations in gyrA, parE, and marOR but not in gyrB, parC, or acrR. Resistance-associated mutations were identified in almost all isolates after selection steps 1 and 2 but in less than 50% of isolates after subsequent selection steps. Selected strains were competed in vitro, in urine, and in a mouse UTI infection model against the starting strain, Nu14. First-step mutations were not associated with significant fitness costs. However, the accumulation of three or more resistance-associated mutations was usually associated with a large reduction in biological fitness, both in vitro and in vivo. Interestingly, in some lineages a partial restoration of fitness was associated with the accumulation of additional mutations in late selection steps. We suggest that the relative biological costs of multiple mutations may influence the evolution of E. coli strains that develop resistance to fluoroquinolones.

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