scholarly journals Detection and Quantification of Superoxide Formed within the Periplasm of Escherichia coli

2006 ◽  
Vol 188 (17) ◽  
pp. 6326-6334 ◽  
Author(s):  
Sergei Korshunov ◽  
James A. Imlay
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Phagocytic Cells ◽  
Free Living ◽  
E Coli ◽  
Superoxide Formation ◽  
Genes Encoding ◽  
Copper Zinc ◽  
Primary Substrate

ABSTRACT Many gram-negative bacteria harbor a copper/zinc-containing superoxide dismutase (CuZnSOD) in their periplasms. In pathogenic bacteria, one role of this enzyme may be to protect periplasmic biomolecules from superoxide that is released by host phagocytic cells. However, the enzyme is also present in many nonpathogens and/or free-living bacteria, including Escherichia coli. In this study we were able to detect superoxide being released into the medium from growing cultures of E. coli. Exponential-phase cells do not normally synthesize CuZnSOD, which is specifically induced in stationary phase. However, the engineered expression of CuZnSOD in growing cells eliminated superoxide release, confirming that this superoxide was formed within the periplasm. The rate of periplasmic superoxide production was surprisingly high and approximated the estimated rate of cytoplasmic superoxide formation when both were normalized to the volume of the compartment. The rate increased in proportion to oxygen concentration, suggesting that the superoxide is generated by the adventitious oxidation of an electron carrier. Mutations that eliminated menaquinone synthesis eradicated the superoxide formation, while mutations in genes encoding respiratory complexes affected it only insofar as they are likely to affect the redox state of menaquinone. We infer that the adventitious autoxidation of dihydromenaquinone in the cytoplasmic membrane releases a steady flux of superoxide into the periplasm of E. coli. This endogenous superoxide may create oxidative stress in that compartment and be a primary substrate of CuZnSOD.

Mannose-Binding Activity of Escherichia coli: a Determinant of Attachment and Ingestion of the Bacteria by Macrophages

1980 ◽  
Vol 29 (2) ◽  
pp. 417-424
Author(s):  
Zvi Bar-Shavit ◽  
Rachel Goldman ◽  
Itzhak Ofek ◽  
Nathan Sharon ◽  
David Mirelman
Keyword(s):  
Escherichia Coli ◽  
Stationary Phase ◽  
Pathogenic Bacteria ◽  
Binding Activity ◽  
Exponential Phase ◽  
Restrictive Temperature ◽  
Filamentous Bacteria ◽  
Phagocytic Cells ◽  
E Coli ◽  
Mannose Binding

Recently, it was suggested that a mannose-specific lectin on the bacterial cell surface is responsible for the recognition by phagocytic cells of certain nonopsonized Escherichia coli strains. In this study we assessed the interaction of two strains of E. coli at different phases of growth with a monolayer of mouse peritoneal macrophages and developed a direct method with [ 14 C]mannan to quantitate the bacterial mannose-binding activity. Normal-sized bacteria were obtained from logarithmic and stationary phases of growth. Nonseptated filamentous cells were formed by growing the organisms in the presence of cephalexin or at a restrictive temperature. Attachment to macrophages of all bacterial forms was inhibited by methyl α- d -mannoside and mannan but not by other sugars tested. The attachment of stationary phase and filamentous bacteria to macrophages, as well as their mannose-binding activity, was similar, whereas in the exponential-phase bacteria they were markedly reduced. The results show a linear relation between the two parameters ( R = 0.98, P < 0.001). The internalization of the filamentous cells attached to macrophages during 45 min of incubation was much less efficient (20%) compared to that of exponential-phase, stationary-phase, or antibody-coated filamentous bacteria (90%). The results indicate that the mannose-binding activity of E. coli determines the recognition of the organisms by phagocytes. They further suggest that administration of β-lactam antibiotics may impair elimination of certain pathogenic bacteria by inducing the formation of filaments which are inefficiently internalized by the host's phagocytic cells.

scholarly journals Role of the Escherichia coli glgX Gene in Glycogen Metabolism

2005 ◽  
Vol 187 (4) ◽  
pp. 1465-1473 ◽  
Author(s):  
David Dauvillée ◽  
Isabelle S. Kinderf ◽  
Zhongyi Li ◽  
Behjat Kosar-Hashemi ◽  
Michael S. Samuel ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Glycogen Synthesis ◽  
High Specificity ◽  
Glycogen Metabolism ◽  
Futile Cycle ◽  
E Coli ◽  
Debranching Enzymes ◽  
Genes Encoding ◽  
Definition Of

ABSTRACT A role for the Escherichia coli glgX gene in bacterial glycogen synthesis and/or degradation has been inferred from the sequence homology between the glgX gene and the genes encoding isoamylase-type debranching enzymes; however, experimental evidence or definition of the role of the gene has been lacking. Construction of E. coli strains with defined deletions in the glgX gene is reported here. The results show that the GlgX gene encodes an isoamylase-type debranching enzyme with high specificity for hydrolysis of chains consisting of three or four glucose residues. This specificity ensures that GlgX does not generate an extensive futile cycle during glycogen synthesis in which chains with more than four glucose residues are transferred by the branching enzyme. Disruption of glgX leads to overproduction of glycogen containing short external chains. These results suggest that the GlgX protein is predominantly involved in glycogen catabolism by selectively debranching the polysaccharide outer chains that were previously recessed by glycogen phosphorylase.

scholarly journals Role of Exopolysaccharides in the Survival and Pathogenesis of the Fire Blight Bacterium, Erwinia amylovora

1994 ◽  
Author(s):  
David Gutnick ◽  
David L. Coplin
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Erwinia Amylovora ◽  
Extracellular Polysaccharide ◽  
Gene Clusters ◽  
Site Directed Mutagenesis ◽  
E Coli ◽  
Erwinia Stewartii ◽  
Stewart's Wilt

Fireblight, a disease of apples and pears, is caused by Erwinia amylovora. Mutants of E. amylovora that do not produce the extreacellular polysaccharide (EPS), amylovoran, are avirulent. A similar EPS, stewartan, is produced by E. stewartii, which caused Stewart's wilt of corn, and which has also been implicated in the virulence of this strain. Both stewartan and amylovoran are type 1 capsular polysaccharides, typified by the colanic acid slime produced by Escherichia coli. Extracellular polysaccharide slime and capsules are important for the virulence of bacterial pathogens of plants and animals and to enhance their survival and dissemination outside of the host. The goals of this project were to examine the importance of polysaccharide structure on the pathogenicity and survival properties of three pathogenic bacteria: Erwinia amylovora, Erwinia stewartii and Escherichia coli. The project was a collaboration between the laboratories of Dr. Gutnick (PI, E. coli genetics and biochemistry), Dr. Coplin (co-PI, E. stewartii genetics) and Dr. Geider (unfunded collaborator, E. amylovora genetics and EPS analysis). Structural analysis of the EPSs, sequence analysis of the biosynthetic gene clusters and site-directed mutagenesis of individual cps and ams genes revealed that the three gene clusters shared common features for polysaccharide polymerization, translocation, and precursor synthesis as well as in the modes of transcriptional regulation. Early EPS production resulted in decreased virulence, indicating that EPS, although required for pathogenicity, is anot always advantageous and pathogens must regulate its production carefully.

scholarly journals Potential Role of Diploscapter sp. Strain LKC25, a Bacterivorous Nematode from Soil, as a Vector of Food-Borne Pathogenic Bacteria to Preharvest Fruits and Vegetables

2005 ◽  
Vol 71 (5) ◽  
pp. 2433-2437 ◽  
Author(s):  
Daunte S. Gibbs ◽  
Gary L. Anderson ◽  
Larry R. Beuchat ◽  
Lynn K. Carta ◽  
Phillip L. Williams
Keyword(s):  
Pathogenic Bacteria ◽  
Fruits And Vegetables ◽  
Potential Role ◽  
The United States ◽  
Cow Manure ◽  
Free Living ◽  
E Coli ◽  
Food Borne ◽  
Agar Media

ABSTRACT Diploscapter, a thermotolerant, free-living soil bacterial-feeding nematode commonly found in compost, sewage, and agricultural soil in the United States, was studied to determine its potential role as a vehicle of Salmonella enterica serotype Poona, enterohemorrhagic Escherichia coli O157:H7, and Listeria monocytogenes in contaminating preharvest fruits and vegetables. The ability of Diploscapter sp. strain LKC25 to survive on agar media, in cow manure, and in composted turkey manure and to be attracted to, ingest, and disperse food-borne pathogens inoculated into soil or a mixture of soil and composted turkey manure was investigated. Diploscapter sp. strain LKC25 survived and reproduced in lawns of S. enterica serotype Poona, E. coli O157:H7, and L. monocytogenes on agar media and in cow manure and composted turkey manure. Attraction of Diploscapter sp. strain LKC25 to colonies of pathogenic bacteria on tryptic soy agar within 10, 20, 30, and 60 min and 24 h was determined. At least 85% of the worms initially placed 0.5 to 1 cm away from bacterial colonies migrated to the colonies within 1 h. Within 24 h, ≥90% of the worms were embedded in colonies. The potential of Diploscapter sp. strain LKC25 to shed pathogenic bacteria after exposure to bacteria inoculated into soil or a mixture of soil and composted turkey manure was investigated. Results indicate that Diploscapter sp. strain LKC25 can shed pathogenic bacteria after exposure to pathogens in these milieus. They also demonstrate its potential to serve as a vector of food-borne pathogenic bacteria in soil, with or without amendment with compost, to the surface of preharvest fruits and vegetables in contact with soil.

scholarly journals Association of Escherichia Coli 0157:H7 in Flocs: Role of Extracellular Polymeric Substances and Fate of Pathogenic Organisms

2021 ◽  
Author(s):  
Mitsuko Kanetani
Keyword(s):  
Escherichia Coli ◽  
Extracellular Polymeric Substances ◽  
Natural Environments ◽  
Potential Mechanism ◽  
Free Living ◽  
E Coli ◽  
Adhesion Ability ◽  
Food Borne ◽  
Water Borne

In North America, Escherichia coli (E. coli) O157:H7 has been frequently associated with outbreaks of food-borne and water-borne infection. The association of E. coli O157:H7 to flocs is hypothesized to be a potential mechanism of transport and survival in natural environments. This study examines this hypothesis with a focus on the role of extracellular polymeric substances (EPS) and on E. coli O157:H7 survival/pathogenicity in an aquatic environment. The floc characterization experiment indicted that EPS play a significant role in floc stability. The EPS distribution experiment revealed abundant hydrophobic protein throughout the floc, which contributed to floc stability and microbial adhesion. In examining the survivability of E. coli O157:H7 in a low nutrient water sample for 10-day incubation, more floc-adherent E. coli O157:H7 survived than the free-living form since EPS provided protection, nutrients and stable sites for survival. Surviving E. coli O157:H7 exhibited both decreased host adhesion ability and ª-actinin accumulation; however, their infection ability was not affected. This suggests that low nutrient levels did not affect pathogenicity over 10 days in this study.

scholarly journals Development of a Set of Primers for Drug-Resistance Genes Detection in the Agents of Dangerous Bacterial Infections as Exemplified by Yersinia pestis, Vibrio cholerae, Escherichia coli Strains

2014 ◽  
pp. 57-60 ◽  
Author(s):  
K. A. Nikiforov ◽  
L. V. Anisimova ◽  
G. N. Odinokov ◽  
A. V. Fadeeva ◽  
L. A. Novichkova ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Drug Resistance ◽  
Yersinia Pestis ◽  
Vibrio Cholerae ◽  
Resistance Genes ◽  
Bacterial Infections ◽  
Pathogenic Bacteria ◽  
E Coli ◽  
Genes Encoding ◽  
Drug Resistant Strains

A set of primers for detection of genes encoding resistance to streptomycin ( strA, strB ), tetracyclin ( tetA, tetR ), chloramphenicol ( catА ), kanamycin ( npt , aphA ), vankomycin ( sanA ), polymyxin ( pmrD ) has been developed with the aim of rapid and effective detection of drug-resistant strains of dangerous bacterial infections agents. Efficacy of constructed primers has been confirmed against a panel of 40 Yersinia pestis, 49 Vibrio cholerae, and 2 Escherichia coli strains from the State collection of pathogenic bacteria of the RAPI “Microbe”. Drug-resistance genes ntp and catA have been detected in plague agent strains , strA, strB , npt , aphA , tetA and tetR - in cholera agent; strA , tetR , ntp and aphA - in pathogenic strain E. coli О157:H7. Determined is universal character of the designed primers for drug-resistance genes detection in these pathogenic bacteria species.

scholarly journals Association of Escherichia Coli 0157:H7 in Flocs: Role of Extracellular Polymeric Substances and Fate of Pathogenic Organisms

2021 ◽  
Author(s):  
Mitsuko Kanetani
Keyword(s):  
Escherichia Coli ◽  
Extracellular Polymeric Substances ◽  
Natural Environments ◽  
Potential Mechanism ◽  
Free Living ◽  
E Coli ◽  
Adhesion Ability ◽  
Food Borne ◽  
Water Borne

In North America, Escherichia coli (E. coli) O157:H7 has been frequently associated with outbreaks of food-borne and water-borne infection. The association of E. coli O157:H7 to flocs is hypothesized to be a potential mechanism of transport and survival in natural environments. This study examines this hypothesis with a focus on the role of extracellular polymeric substances (EPS) and on E. coli O157:H7 survival/pathogenicity in an aquatic environment. The floc characterization experiment indicted that EPS play a significant role in floc stability. The EPS distribution experiment revealed abundant hydrophobic protein throughout the floc, which contributed to floc stability and microbial adhesion. In examining the survivability of E. coli O157:H7 in a low nutrient water sample for 10-day incubation, more floc-adherent E. coli O157:H7 survived than the free-living form since EPS provided protection, nutrients and stable sites for survival. Surviving E. coli O157:H7 exhibited both decreased host adhesion ability and ª-actinin accumulation; however, their infection ability was not affected. This suggests that low nutrient levels did not affect pathogenicity over 10 days in this study.

scholarly journals The Escherichia coli Protein YfeX Functions as a Porphyrinogen Oxidase, Not a Heme Dechelatase

mBio ◽  
2011 ◽  
Vol 2 (6) ◽  
Author(s):  
Harry A. Dailey ◽  
Alecia N. Septer ◽  
Lauren Daugherty ◽  
Daniel Thames ◽  
Svetlana Gerdes ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Iron Removal ◽  
Anaerobic Growth ◽  
Limiting Factor ◽  
Alizarin Red ◽  
Content Type ◽  
E Coli ◽  
Genes Encoding

ABSTRACT The protein YfeX from Escherichia coli has been proposed to be essential for the process of iron removal from heme by carrying out a dechelation of heme without cleavage of the porphyrin macrocycle. Since this proposed reaction is unique and would represent the first instance of the biological dechelation of heme, we undertook to characterize YfeX. Our data reveal that YfeX effectively decolorizes the dyes alizarin red and Cibacron blue F3GA and has peroxidase activity with pyrogallal but not guiacol. YfeX oxidizes protoporphyrinogen to protoporphyrin in vitro. However, we were unable to detect any dechelation of heme to free porphyrin with purified YfeX or in cellular extracts of E. coli overexpressing YfeX. Additionally, Vibrio fischeri, an organism that can utilize heme as an iron source when grown under iron limitation, is able to grow with heme as the sole source of iron when its YfeX homolog is absent. Plasmid-driven expression of YfeX in V. fischeri grown with heme did not result in accumulation of protoporphyrin. We propose that YfeX is a typical dye-decolorizing peroxidase (or DyP) and not a dechelatase. The protoporphyrin reported to accumulate when YfeX is overexpressed in E. coli likely arises from the intracellular oxidation of endogenously synthesized protoporphyrinogen and not from dechelation of exogenously supplied heme. Bioinformatic analysis of bacterial YfeX homologs does not identify any connection with iron acquisition but does suggest links to anaerobic-growth-related respiratory pathways. Additionally, some genes encoding homologs of YfeX have tight association with genes encoding a bacterial cytoplasmic encapsulating protein. IMPORTANCE Acquisition of iron from the host during infection is a limiting factor for growth and survival of pathogens. Host heme is the major source of iron in infections, and pathogenic bacteria have evolved complex mechanisms to acquire heme and abstract the iron from heme. Recently Létoffé et al. (Proc. Natl. Acad. Sci. U. S. A. 106:11719–11724, 2009) reported that the protein YfeX from E. coli is able to dechelate heme to remove iron and leave an intact tetrapyrrole. This is totally unlike any other described biological system for iron removal from heme and, thus, would represent a dramatically new feature with potentially profound implications for our understanding of bacterial pathogenesis. Given that this reaction has no precedent in biological systems, we characterized YfeX and a related protein. Our data clearly demonstrate that YfeX is not a dechelatase as reported but is a peroxidase that oxidizes endogenous porphyrinogens to porphyrins.

scholarly journals Plasmids Shape the Diverse Accessory Resistomes of Escherichia coli ST131

2020 ◽  
Author(s):  
Arun G. Decano ◽  
Nghia Tran ◽  
Hawriya Al-Foori ◽  
Buthaina Al-Awadi ◽  
Leigh Campbell ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Gene Diversity ◽  
Gastrointestinal Microbiome ◽  
E Coli ◽  
Type Iv ◽  
Genes Encoding ◽  
Clade C ◽  
Gene Sharing

The human gut microbiome includes beneficial, commensal and pathogenic bacteria that possess antimicrobial resistance (AMR) genes and exchange these predominantly through conjugative plasmids. Escherichia coli is a significant component of the gastrointestinal microbiome and is typically non-pathogenic in this niche. In contrast, extra-intestinal pathogenic E. coli (ExPEC) including ST131 may occupy other environments like the urinary tract or bloodstream where they express genes enabling AMR and host adhesion like type 1 fimbriae. The extent to which non-pathogenic gut E. coli and infectious ST131 share AMR genes and key associated plasmids remains understudied at a genomic level. Here, we examined AMR gene sharing between gut E. coli and ST131 to discover an extensive shared preterm infant resistome. In addition, individual ST131 show extensive AMR gene diversity highlighting that analyses restricted to the core genome may be limiting and could miss AMR gene transfer patterns. We show that pEK499-like segments are ancestral to most ST131 Clade C isolates, contrasting with a minority with substantial pEK204-like regions encoding a type IV fimbriae operon. Moreover, ST131 possess extensive diversity at genes encoding type 1, type IV, P and F17-like fimbriae, particular within subclade C2. The type, structure and composition of AMR genes, plasmids and fimbriae varies widely in ST131 and this may mediate pathogenicity and infection outcomes.

scholarly journals Antimicrobial Resistance Glides in the Sky—Free-Living Birds as a Reservoir of Resistant Escherichia coli With Zoonotic Potential

2021 ◽  
Vol 12 ◽  
Author(s):  
Magdalena Skarżyńska ◽  
Magdalena Zaja̧c ◽  
Arkadiusz Bomba ◽  
Łukasz Bocian ◽  
Wojciech Kozdruń ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antimicrobial Resistance ◽  
Animal Health ◽  
Resistance Mechanisms ◽  
Natural Habitats ◽  
Free Living ◽  
E Coli ◽  
Virulence Potential ◽  
Clonal Spread

Antimicrobial resistance (AMR) is one of the most important global health concerns; therefore, the identification of AMR reservoirs and vectors is essential. Attention should be paid to the recognition of potential hazards associated with wildlife as this field still seems to be incompletely explored. In this context, the role of free-living birds as AMR carriers is noteworthy. Therefore, we applied methods used in AMR monitoring, supplemented by colistin resistance screening, to investigate the AMR status of Escherichia coli from free-living birds coming from natural habitats and rescue centers. Whole-genome sequencing (WGS) of strains enabled to determine resistance mechanisms and investigate their epidemiological relationships and virulence potential. As far as we know, this study is one of the few that applied WGS of that number (n = 71) of strains coming from a wild avian reservoir. The primary concerns arising from our study relate to resistance and its determinants toward antimicrobial classes of the highest priority for the treatment of critical infections in people, e.g., cephalosporins, quinolones, polymyxins, and aminoglycosides, as well as fosfomycin. Among the numerous determinants, blaCTX–M–15, blaCMY–2, blaSHV–12, blaTEM–1B, qnrS1, qnrB19, mcr-1, fosA7, aac(3)-IIa, ant(3”)-Ia, and aph(6)-Id and chromosomal gyrA, parC, and parE mutations were identified. Fifty-two sequence types (STs) noted among 71 E. coli included the global lineages ST131, ST10, and ST224 as well as the three novel STs 11104, 11105, and 11194. Numerous virulence factors were noted with the prevailing terC, gad, ompT, iss, traT, lpfA, and sitA. Single E. coli was Shiga toxin-producing. Our study shows that the clonal spread of E. coli lineages of public and animal health relevance is a serious avian-associated hazard.

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