scholarly journals O-Antigen-Dependent Colicin Insensitivity of UropathogenicEscherichia coli

2018 ◽  
Vol 201 (4) ◽  
Author(s):  
Connor Sharp ◽  
Christine Boinett ◽  
Amy Cain ◽  
Nicholas G. Housden ◽  
Sandip Kumar ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Growth Conditions ◽  
Membrane Receptors ◽  
Cell Surface Receptor ◽  
Dominant Factor ◽  
Content Type ◽  
E Coli ◽  
O Antigen ◽  
K 12

ABSTRACTThe outer membrane of Gram-negative bacteria presents a significant barrier for molecules entering the cell. Nevertheless, colicins, which are antimicrobial proteins secreted byEscherichia coli, can target otherE. colicells by binding to cell surface receptor proteins and activating their import, resulting in cell death. Previous studies have documented high rates of nonspecific resistance (insensitivity) of variousE. colistrains toward colicins that is independent of colicin-specific immunity and is instead associated with lipopolysaccharide (LPS) in the outer membrane. This observation poses a contradiction: why doE. colistrains have colicin-expressing plasmids, which are energetically costly to retain, if cells around them are likely to be naturally insensitive to the colicin they produce? Here, using a combination of transposon sequencing and phenotypic microarrays, we show that colicin insensitivity of uropathogenicE. colisequence type 131 (ST131) is dependent on the production of its O-antigen but that minor changes in growth conditions render the organism sensitive toward colicins. The reintroduction of O-antigen intoE. coliK-12 demonstrated that it is the density of O-antigen that is the dominant factor governing colicin insensitivity. We also show, by microscopy of fluorescently labelled colicins, that growth conditions affect the degree of occlusion by O-antigen of outer membrane receptors but not the clustered organization of receptors. The result of our study demonstrate that environmental conditions play a critical role in sensitizingE. colitoward colicins and that O-antigen in LPS is central to this role.IMPORTANCEEscherichia coliinfections can be a major health burden, especially with the organism becoming increasingly resistant to “last-resort” antibiotics such as carbapenems. Although colicins are potent narrow-spectrum antimicrobials with potential as future antibiotics, high levels of naturally occurring colicin insensitivity have been documented which could limit their efficacy. We identify O-antigen-dependent colicin insensitivity in a clinically relevant uropathogenicE. colistrain and show that this insensitivity can be circumvented by minor changes to growth conditions. The results of our study suggest that colicin insensitivity amongE. coliorganisms has been greatly overestimated, and as a consequence, colicins could in fact be effective species-specific antimicrobials targeting pathogenicE. colisuch as uropathogenicE. coli(UPEC).

scholarly journals Flocculation of Escherichia coli Cells in Association with Enhanced Production of Outer Membrane Vesicles

2015 ◽  
Vol 81 (17) ◽  
pp. 5900-5906 ◽  
Author(s):  
Yoshihiro Ojima ◽  
Minh Hong Nguyen ◽  
Reiki Yajima ◽  
Masahito Taya
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Sodium Dodecyl ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Laser Scanning Microscopy ◽  
Content Type ◽  
E Coli ◽  
Enhanced Production ◽  
K 12

ABSTRACTMicrobial flocculation is a phenomenon of aggregation of dispersed bacterial cells in the form of flocs or flakes. In this study, the mechanism of spontaneous flocculation ofEscherichia colicells by overexpression of thebcsBgene was investigated. The flocculation induced by overexpression ofbcsBwas consistent among the variousE. colistrains examined, including the K-12, B, and O strains, with flocs that resembled paper scraps in structure being about 1 to 2 mm. The distribution of green fluorescent protein-labeledE. colicells within the floc structure was investigated by three-dimensional confocal laser scanning microscopy. Flocs were sensitive to proteinase K, indicating that the main component of the flocs was proteinous. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and nano-liquid chromatography tandem mass spectrometry analyses of the flocs strongly suggested the involvement of outer membrane vesicles (OMVs) inE. coliflocculation. The involvement of OMVs in flocculation was supported by transmission electron microscopy observation of flocs. Furthermore,bcsB-inducedE. coliflocculation was greatly suppressed in strains with hypovesiculation phenotypes (ΔdsbAand ΔdsbBstrains). Thus, our results demonstrate the strong correlation between spontaneous flocculation and enhanced OMV production ofE. colicells.

scholarly journals A Rhodobacter sphaeroides Protein Mechanistically Similar to Escherichia coli DksA Regulates Photosynthetic Growth

mBio ◽  
2014 ◽  
Vol 5 (3) ◽  
Author(s):  
Christopher W. Lennon ◽  
Kimberly C. Lemmer ◽  
Jessica L. Irons ◽  
Max I. Sellman ◽  
Timothy J. Donohue ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Structure Function ◽  
Rhodobacter Sphaeroides ◽  
Fatty Acid Content ◽  
Regulatory Protein ◽  
Growth Conditions ◽  
Globular Domain ◽  
Content Type ◽  
E Coli

ABSTRACTDksA is a global regulatory protein that, together with the alarmone ppGpp, is required for the “stringent response” to nutrient starvation in the gammaproteobacteriumEscherichia coliand for more moderate shifts between growth conditions. DksA modulates the expression of hundreds of genes, directly or indirectly. Mutants lacking a DksA homolog exhibit pleiotropic phenotypes in other gammaproteobacteria as well. Here we analyzed the DksA homolog RSP2654 in the more distantly relatedRhodobacter sphaeroides, an alphaproteobacterium. RSP2654 is 42% identical and similar in length toE. coliDksA but lacks the Zn finger motif of theE. coliDksA globular domain. Deletion of the RSP2654 gene results in defects in photosynthetic growth, impaired utilization of amino acids, and an increase in fatty acid content. RSP2654 complements the growth and regulatory defects of anE. colistrain lacking thedksAgene and modulates transcriptionin vitrowithE. coliRNA polymerase (RNAP) similarly toE. coliDksA. RSP2654 reduces RNAP-promoter complex stabilityin vitrowith RNAPs fromE. coliorR. sphaeroides, alone and synergistically with ppGpp, suggesting that even though it has limited sequence identity toE. coliDksA (DksAEc), it functions in a mechanistically similar manner. We therefore designate the RSP2654 protein DksARsp. Our work suggests that DksARsphas distinct and important physiological roles in alphaproteobacteria and will be useful for understanding structure-function relationships in DksA and the mechanism of synergy between DksA and ppGpp.IMPORTANCEThe role of DksA has been analyzed primarily in the gammaproteobacteria, in which it is best understood for its role in control of the synthesis of the translation apparatus and amino acid biosynthesis. Our work suggests that DksA plays distinct and important physiological roles in alphaproteobacteria, including the control of photosynthesis inRhodobacter sphaeroides. The study of DksARsp, should be useful for understanding structure-function relationships in the protein, including those that play a role in the little-understood synergy between DksA and ppGpp.

scholarly journals Antibiotic-Mediated Modulations of Outer Membrane Vesicles in Enterohemorrhagic Escherichia coli O104:H4 and O157:H7

2017 ◽  
Vol 61 (9) ◽  
Author(s):  
Andreas Bauwens ◽  
Lisa Kunsmann ◽  
Helge Karch ◽  
Alexander Mellmann ◽  
Martina Bielaszewska
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Shiga Toxin ◽  
Membrane Vesicles ◽  
Polymyxin B ◽  
Outer Membrane Vesicles ◽  
Enterohemorrhagic Escherichia Coli ◽  
Content Type ◽  
E Coli ◽  
Major Virulence Factor

ABSTRACT Ciprofloxacin, meropenem, fosfomycin, and polymyxin B strongly increase production of outer membrane vesicles (OMVs) in Escherichia coli O104:H4 and O157:H7. Ciprofloxacin also upregulates OMV-associated Shiga toxin 2a, the major virulence factor of these pathogens, whereas the other antibiotics increase OMV production without the toxin. These two effects might worsen the clinical outcome of infections caused by Shiga toxin-producing E. coli. Our data support the existing recommendations to avoid antibiotics for treatment of these infections.

scholarly journals Molecular Control of Sucrose Utilization in Escherichia coli W, an Efficient Sucrose-Utilizing Strain

2012 ◽  
Vol 79 (2) ◽  
pp. 478-487 ◽  
Author(s):  
Suriana Sabri ◽  
Lars K. Nielsen ◽  
Claudia E. Vickers
Keyword(s):  
Escherichia Coli ◽  
Growth Rate ◽  
Good Growth ◽  
Knockout Mutant ◽  
Sucrose Transport ◽  
Molecular Control ◽  
Content Type ◽  
E Coli ◽  
K 12 ◽  
Sucrose Utilization

ABSTRACTSucrose is an industrially important carbon source for microbial fermentation. Sucrose utilization inEscherichia coli, however, is poorly understood, and most industrial strains cannot utilize sucrose. The roles of the chromosomally encoded sucrose catabolism (csc) genes inE. coliW were examined by knockout and overexpression experiments. At low sucrose concentrations, thecscgenes are repressed and cells cannot grow. Removal of either the repressor protein (cscR) or the fructokinase (cscK) gene facilitated derepression. Furthermore, combinatorial knockout ofcscRandcscKconferred an improved growth rate on low sucrose. The invertase (cscA) and sucrose transporter (cscB) genes are essential for sucrose catabolism inE. coliW, demonstrating that no other genes can provide sucrose transport or inversion activities. However,cscKis not essential for sucrose utilization. Fructose is excreted into the medium by thecscK-knockout strain in the presence of high sucrose, whereas at low sucrose (when carbon availability is limiting), fructose is utilized by the cell. Overexpression ofcscA,cscAK, orcscABcould complement the WΔcscRKABknockout mutant or confer growth on a K-12 strain which could not naturally utilize sucrose. However, phenotypic stability and relatively good growth rates were observed in the K-12 strain only when overexpressingcscAB, and full growth rate complementation in WΔcscRKABalso requiredcscAB. Our understanding of sucrose utilization can be used to improveE. coliW and engineer sucrose utilization in strains which do not naturally utilize sucrose, allowing substitution of sucrose for other, less desirable carbon sources in industrial fermentations.

scholarly journals Increased Survival of Antibiotic-Resistant Escherichia coli inside Macrophages

2012 ◽  
Vol 57 (1) ◽  
pp. 189-195 ◽  
Author(s):  
Migla Miskinyte ◽  
Isabel Gordo
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Bacterial Infections ◽  
Resistance Mutation ◽  
Fitness Cost ◽  
Resistance Mutations ◽  
Content Type ◽  
E Coli ◽  
Fitness Effects ◽  
K 12

ABSTRACTMutations causing antibiotic resistance usually incur a fitness cost in the absence of antibiotics. The magnitude of such costs is known to vary with the environment. Little is known about the fitness effects of antibiotic resistance mutations when bacteria confront the host's immune system. Here, we study the fitness effects of mutations in therpoB,rpsL, andgyrAgenes, which confer resistance to rifampin, streptomycin, and nalidixic acid, respectively. These antibiotics are frequently used in the treatment of bacterial infections. We measured two important fitness traits—growth rate and survival ability—of 12Escherichia coliK-12 strains, each carrying a single resistance mutation, in the presence of macrophages. Strikingly, we found that 67% of the mutants survived better than the susceptible bacteria in the intracellular niche of the phagocytic cells. In particular, allE. colistreptomycin-resistant mutants exhibited an intracellular advantage. On the other hand, 42% of the mutants incurred a high fitness cost when the bacteria were allowed to divide outside of macrophages. This study shows that single nonsynonymous changes affecting fundamental processes in the cell can contribute to prolonged survival ofE. coliin the context of an infection.

scholarly journals Ib-M6 Antimicrobial Peptide: Antibacterial Activity against Clinical Isolates of Escherichia coli and Molecular Docking

Antibiotics ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 79 ◽  
Author(s):  
J. M. Flórez-Castillo ◽  
P. Rondón-Villareal ◽  
J. L. Ropero-Vega ◽  
S. Y. Mendoza-Espinel ◽  
J. A. Moreno-Amézquita ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antibacterial Activity ◽  
Molecular Docking ◽  
Outer Membrane ◽  
Docking Simulations ◽  
E Coli ◽  
Pathogenic Escherichia Coli ◽  
Membrane Components ◽  
K 12 ◽  
Susceptibility Assay

The Ib-M6 peptide has antibacterial activity against non-pathogenic Escherichia coli K-12 strain. The first part of this study determines the antibacterial activity of Ib-M6 against fourteen pathogenic strains of E. coli O157:H7. Susceptibility assay showed that Ib-M6 had values of Minimum Inhibitory Concentration (MIC) lower than streptomycin, used as a reference antibiotic. Moreover, to predict the possible interaction between Ib-M6 and outer membrane components of E. coli, we used molecular docking simulations where FhuA protein and its complex with Lipopolysaccharide (LPS–FhuA) were used as targets of the peptide. FhuA/Ib-M6 complexes had energy values between −39.5 and −40.5 Rosetta Energy Units (REU) and only one hydrogen bond. In contrast, complexes between LPS–FhuA and Ib-M6 displayed energy values between −25.6 and −40.6 REU, and the presence of five possible hydrogen bonds. Hence, the antimicrobial activity of Ib-M6 peptide shown in the experimental assays could be caused by its interaction with the outer membrane of E. coli.

scholarly journals Bioinformatic and Molecular Analysis of Inverse Autotransporters from Escherichia coli

mSphere ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Kelvin G. K. Goh ◽  
Danilo G. Moriel ◽  
Steven J. Hancock ◽  
Minh-Duy Phan ◽  
Mark A. Schembri
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Biofilm Formation ◽  
Secretion System ◽  
Content Type ◽  
E Coli ◽  
Wide Range ◽  
Type V Secretion ◽  
K 12 ◽  
Type V

ABSTRACT Proteins secreted by the type V secretion system possess multiple functions, including the capacity to mediate adhesion, aggregation, and biolfilm formation. The type V secretion system can be divided into five subclasses, one of which is the type Ve system. Proteins of the type Ve secretion system are also referred to as inverse autotransporters (IATs). In this study, we performed an in silico analysis of 126 completely sequenced Escherichia coli genomes available in the NCBI database and identified several distinct IAT-encoding gene families whose distribution varied throughout the E. coli phylogeny. The genes included three characterized IATs (intimin, fdeC, and yeeJ) and four uncharacterized IATs (here named iatA, iatB, iatC, and iatD). The four iat genes were cloned from the completely sequenced environmental E. coli strain SMS-3-5 and characterized. Three of these IAT proteins (IatB, IatC, and IatD) were expressed at the cell surface and possessed the capacity to mediate biofilm formation in a recombinant E. coli K-12 strain. Further analysis of the iatB gene, which showed a unique association with extraintestinal E. coli strains, suggested that its regulation is controlled by the LeuO global regulator. Overall, this study provides new data describing the prevalence, sequence variation, domain structure, function, and regulation of IATs found in E. coli. IMPORTANCE Escherichia coli is one of the most prevalent facultative anaerobes of the human gut. E. coli normally exists as a harmless commensal but can also cause disease following the acquisition of genes that enhance its pathogenicity. Adhesion is an important first step in colonization of the host and is mediated by an array of cell surface components. In E. coli, these include a family of adhesins secreted by the type V secretion system. Here, we identified and characterized new proteins from an emerging subclass of the type V secretion system known as the inverse autotransporters (IATs). We found that IAT-encoding genes are present in a wide range of strains and showed that three novel IATs were localized on the E. coli cell surface and mediated biofilm formation. Overall, this study provides new insight into the prevalence, function, and regulation of IATs in E. coli.

scholarly journals Additional Og-Typing PCR Techniques Targeting Escherichia coli-Novel and Shigella-Unique O-Antigen Biosynthesis Gene Clusters

2020 ◽  
Vol 58 (11) ◽  
Author(s):  
Atsushi Iguchi ◽  
Hironobu Nishii ◽  
Kazuko Seto ◽  
Jiro Mitobe ◽  
Kenichi Lee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Strong Association ◽  
Gene Clusters ◽  
Epidemiological Studies ◽  
Content Type ◽  
E Coli ◽  
O Antigen ◽  
Wide Range ◽  
Biosynthesis Gene ◽  
Almost All

ABSTRACT The O-serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and controls. O-serogroup diversification shows a strong association with the genetic diversity in some O-antigen biosynthesis gene clusters. Through genomic studies, in addition to the types of O-antigen biosynthesis gene clusters (Og-types) from conventional O-serogroup strains, a number of novel Og-types have been found in E. coli isolates. To assist outbreak investigations and surveillance of pathogenic E. coli at inspection institutes, in previous studies, we developed PCR methods that could determine almost all conventional O-serogroups and some novel Og-types. However, there are still many Og-types that may not be determined by simple genetic methods such as PCR. Thus, in the present study, we aimed to develop an additional Og-typing PCR system. Based on the novel Og-types, including OgN32, OgN33, and OgN34, presented in this study, we designed an additional 24 PCR primer pairs targeting 14 novel and 2 diversified E. coli Og-types and 8 Shigella-unique Og-types. Subsequently, we developed 5 new multiplex PCR sets consisting of 33 primers, including the aforementioned 24 primers and 9 primers reported in previous studies. The accuracy and specificity of the PCR system was validated using approximately 260 E. coli and Shigella O-serogroup and Og-type reference strains. The Og-typing PCR system reported here can determine a wide range of Og-types of E. coli and may help epidemiological studies, in addition to the surveillance of pathogenic E. coli.

scholarly journals The c-Type Cytochrome OmcA Localizes to the Outer Membrane upon Heterologous Expression in Escherichia coli

2008 ◽  
Vol 190 (14) ◽  
pp. 5127-5131 ◽  
Author(s):  
James W. Donald ◽  
Matthew G. Hicks ◽  
David J. Richardson ◽  
Tracy Palmer
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Shewanella Oneidensis ◽  
Type Ii Secretion ◽  
Type Ii ◽  
E Coli ◽  
Expression In Escherichia Coli ◽  
Type Ii Secretion System ◽  
Surface Localization ◽  
K 12

ABSTRACT We have functionally produced the outer membrane cytochrome OmcA from Shewanella oneidensis in Escherichia coli. Substrate accessibility experiments indicate that OmcA is surface exposed in an E. coli B strain but not in a K-12 strain. We show that a functional type II secretion system is required for surface localization.

scholarly journals Global Lysine Acetylation in Escherichia coli Results from Growth Conditions That Favor Acetate Fermentation

2019 ◽  
Vol 201 (9) ◽  
Author(s):  
Birgit Schilling ◽  
Nathan Basisty ◽  
David G. Christensen ◽  
Dylan Sorensen ◽  
James S. Orr ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Enzyme Activity ◽  
Regulatory Mechanism ◽  
Growth Conditions ◽  
Lysine Acetylation ◽  
Label Free ◽  
Content Type ◽  
E Coli ◽  
Regulatory Mode ◽  
Specific Sugar

ABSTRACT Lysine acetylation is thought to provide a mechanism for regulating metabolism in diverse bacteria. Indeed, many studies have shown that the majority of enzymes involved in central metabolism are acetylated and that acetylation can alter enzyme activity. However, the details regarding this regulatory mechanism are still unclear, specifically with regard to the signals that induce lysine acetylation. To better understand this global regulatory mechanism, we profiled changes in lysine acetylation during growth of Escherichia coli on the hexose glucose or the pentose xylose at both high and low sugar concentrations using label-free mass spectrometry. The goal was to see whether lysine acetylation differed during growth on these two different sugars. No significant differences, however, were observed. Rather, the initial sugar concentration was the principal factor governing changes in lysine acetylation, with higher sugar concentrations causing more acetylation. These results suggest that acetylation does not target specific metabolic pathways but rather simply targets accessible lysines, which may or may not alter enzyme activity. They further suggest that lysine acetylation principally results from conditions that favor accumulation of acetyl phosphate, the principal acetate donor in E. coli. IMPORTANCE Bacteria alter their metabolism in response to nutrient availability, growth conditions, and environmental stresses using a number of different mechanisms. One is lysine acetylation, a posttranslational modification known to target many metabolic enzymes. However, little is known about this regulatory mode. We investigated the factors inducing changes in lysine acetylation by comparing growth on glucose and xylose. We found that the specific sugar used for growth did not alter the pattern of acetylation; rather, the amount of sugar did, with more sugar causing more acetylation. These results imply that lysine acetylation is a global regulatory mechanism that is responsive not to the specific carbon source per se but rather to the accumulation of downstream metabolites.

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