scholarly journals Bactericidal Monoclonal Antibodies Specific to the Lipopolysaccharide O Antigen from Multidrug-Resistant Escherichia coli Clone ST131-O25b:H4 Elicit Protection in Mice

2015 ◽  
Vol 59 (6) ◽  
pp. 3109-3116 ◽  
Author(s):  
Valéria Szijártó ◽  
Luis M. Guachalla ◽  
Zehra C. Visram ◽  
Katharina Hartl ◽  
Cecília Varga ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Monoclonal Antibodies ◽  
Human Serum ◽  
Significant Proportion ◽  
Multidrug Resistant ◽  
Bacterial Killing ◽  
Content Type ◽  
E Coli

ABSTRACTTheEscherichia colisequence type 131 (ST131)-O25b:H4 clone has spread worldwide and become responsible for a significant proportion of multidrug-resistant extraintestinal infections. We generated humanized monoclonal antibodies (MAbs) that target the lipopolysaccharide O25b antigen conserved within this lineage. These MAbs bound to the surface of live bacterial cells irrespective of the capsular type expressed. In a serum bactericidal assayin vitro, MAbs induced >95% bacterial killing in the presence of human serum as the complement source. Protective efficacy at low antibody doses was observed in a murine model of bacteremia. The mode of actionin vivowas investigated by using aglycosylated derivatives of the protective MAbs. The significant binding to liveE. colicells and thein vitroandin vivoefficacy were corroborated in assays using bacteria grown in human serum to mimic relevant clinical conditions. Given the dry pipeline of novel antibiotics against multidrug-resistant Gram-negative pathogens, passive immunization with bactericidal antibodies offers a therapeutic alternative to control infections caused byE. coliST131-O25b:H4.

scholarly journals Genomic and Functional Analysis of Emerging Virulent and Multidrug-Resistant Escherichia coli Lineage Sequence Type 648

2019 ◽  
Vol 63 (6) ◽  
Author(s):  
Katharina Schaufler ◽  
Torsten Semmler ◽  
Lothar H. Wieler ◽  
Darren J. Trott ◽  
Johann Pitout ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
High Risk ◽  
Multidrug Resistant ◽  
Phenotypic Analysis ◽  
Content Type ◽  
Extended Spectrum Beta Lactamase ◽  
Niche Adaptation ◽  
High Level

ABSTRACT The pathogenic extended-spectrum-beta-lactamase (ESBL)-producing Escherichia coli lineage ST648 is increasingly reported from multiple origins. Our study of a large and global ST648 collection from various hosts (87 whole-genome sequences) combining core and accessory genomics with functional analyses and in vivo experiments suggests that ST648 is a nascent and generalist lineage, lacking clear phylogeographic and host association signals. By including large numbers of ST131 (n = 107) and ST10 (n = 96) strains for comparative genomics and phenotypic analysis, we demonstrate that the combination of multidrug resistance and high-level virulence are the hallmarks of ST648, similar to international high-risk clonal lineage ST131. Specifically, our in silico, in vitro, and in vivo results demonstrate that ST648 is well equipped with biofilm-associated features, while ST131 shows sophisticated signatures indicative of adaption to urinary tract infection, potentially conveying individual ecological niche adaptation. In addition, we used a recently developed NFDS (negative frequency-dependent selection) population model suggesting that ST648 will increase significantly in frequency as a cause of bacteremia within the next few years. Also, ESBL plasmids impacting biofilm formation aided in shaping and maintaining ST648 strains to successfully emerge worldwide across different ecologies. Our study contributes to understanding what factors drive the evolution and spread of emerging international high-risk clonal lineages.

scholarly journals A Toxic Environment: a Growing Understanding of How Microbial Communities Affect Escherichia coli O157:H7 Shiga Toxin Expression

2020 ◽  
Vol 86 (24) ◽  
Author(s):  
Erin M. Nawrocki ◽  
Hillary M. Mosso ◽  
Edward G. Dudley
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Microbial Interactions ◽  
Severe Illness ◽  
Content Type ◽  
E Coli ◽  
Wide Range ◽  
Lambdoid Prophage

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) strains, including E. coli O157:H7, cause severe illness in humans due to the production of Shiga toxin (Stx) and other virulence factors. Because Stx is coregulated with lambdoid prophage induction, its expression is especially susceptible to environmental cues. Infections with Stx-producing E. coli can be difficult to model due to the wide range of disease outcomes: some infections are relatively mild, while others have serious complications. Probiotic organisms, members of the gut microbiome, and organic acids can depress Stx production, in many cases by inhibiting the growth of EHEC strains. On the other hand, the factors currently known to amplify Stx act via their effect on the stx-converting phage. Here, we characterize two interactive mechanisms that increase Stx production by O157:H7 strains: first, direct interactions with phage-susceptible E. coli, and second, indirect amplification by secreted factors. Infection of susceptible strains by the stx-converting phage can expand the Stx-producing population in a human or animal host, and phage infection has been shown to modulate virulence in vitro and in vivo. Acellular factors, particularly colicins and microcins, can kill O157:H7 cells but may also trigger Stx expression in the process. Colicins, microcins, and other bacteriocins have diverse cellular targets, and many such molecules remain uncharacterized. The identification of additional Stx-amplifying microbial interactions will improve our understanding of E. coli O157:H7 infections and help elucidate the intricate regulation of pathogenicity in EHEC strains.

scholarly journals Role of F1C Fimbriae, Flagella, and Secreted Bacterial Components in the Inhibitory Effect of Probiotic Escherichia coli Nissle 1917 on Atypical Enteropathogenic E. coli Infection

2014 ◽  
Vol 82 (5) ◽  
pp. 1801-1812 ◽  
Author(s):  
Sylvia Kleta ◽  
Marcel Nordhoff ◽  
Karsten Tedin ◽  
Lothar H. Wieler ◽  
Rafal Kolenda ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Molecular Mechanisms ◽  
Inhibitory Effect ◽  
Host Cells ◽  
Single Infection ◽  
Content Type ◽  
E Coli ◽  
Initial Adhesion

ABSTRACTEnteropathogenicEscherichia coli(EPEC) is recognized as an important intestinal pathogen that frequently causes acute and persistent diarrhea in humans and animals. The use of probiotic bacteria to prevent diarrhea is gaining increasing interest. The probioticE. colistrain Nissle 1917 (EcN) is known to be effective in the treatment of several gastrointestinal disorders. While bothin vitroandin vivostudies have described strong inhibitory effects of EcN on enteropathogenic bacteria, including pathogenicE. coli, the underlying molecular mechanisms remain largely unknown. In this study, we examined the inhibitory effect of EcN on infections of porcine intestinal epithelial cells with atypical enteropathogenicE. coli(aEPEC) with respect to single infection steps, including adhesion, microcolony formation, and the attaching and effacing phenotype. We show that EcN drastically reduced the infection efficiencies of aEPEC by inhibiting bacterial adhesion and growth of microcolonies, but not the attaching and effacing of adherent bacteria. The inhibitory effect correlated with EcN adhesion capacities and was predominantly mediated by F1C fimbriae, but also by H1 flagella, which served as bridges between EcN cells. Furthermore, EcN seemed to interfere with the initial adhesion of aEPEC to host cells by secretion of inhibitory components. These components do not appear to be specific to EcN, but we propose that the strong adhesion capacities enable EcN to secrete sufficient local concentrations of the inhibitory factors. The results of this study are consistent with a mode of action whereby EcN inhibits secretion of virulence-associated proteins of EPEC, but not their expression.

scholarly journals Activity of Tigecycline or Colistin in Combination with Zidovudine against Escherichia coli Harboring tet(X) and mcr-1

2020 ◽  
Vol 65 (1) ◽  
pp. e01172-20 ◽  
Author(s):  
Yu-Feng Zhou ◽  
Ping Liu ◽  
Shu-He Dai ◽  
Jian Sun ◽  
Ya-Hong Liu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Multidrug Resistant ◽  
Therapeutic Options ◽  
Infection Model ◽  
Synergistic Activity ◽  
Colistin Resistance ◽  
Content Type ◽  
E Coli ◽  
Antiretroviral Drug

ABSTRACTAlternative therapeutic options are urgently needed against multidrug-resistant Escherichia coli infections, especially in situations of preexisting tigecycline and colistin resistance. Here, we investigated synergistic activity of the antiretroviral drug zidovudine in combination with tigecycline or colistin against E. coli harboring tet(X) and mcr-1 in vitro and in a murine thigh infection model. Zidovudine and tigecycline/colistin combinations achieved synergistic killing and significantly decreased bacterial burdens by >2.5-log10 CFU/g in thigh tissues compared to each monotherapy.

scholarly journals Escherichia coli CFT073 Fitness Factors during Urinary Tract Infection: Identification Using an Ordered Transposon Library

2020 ◽  
Vol 86 (13) ◽  
Author(s):  
Allyson E. Shea ◽  
Juan Marzoa ◽  
Stephanie D. Himpsl ◽  
Sara N. Smith ◽  
Lili Zhao ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Urinary Tract Infections ◽  
Human Urine ◽  
Transposon Mutagenesis ◽  
Content Type ◽  
E Coli ◽  
Tract Infections

ABSTRACT Urinary tract infections (UTI), the second most diagnosed infectious disease worldwide, are caused primarily by uropathogenic Escherichia coli (UPEC), placing a significant financial burden on the health care system. High-throughput transposon mutagenesis combined with genome-targeted sequencing is a powerful technique to interrogate genomes for fitness genes. Genome-wide analysis of E. coli requires random libraries of at least 50,000 mutants to achieve 99.99% saturation; however, the traditional murine model of ascending UTI does not permit testing of large mutant pools due to a bottleneck during infection. To address this, an E. coli CFT073 transposon mutant ordered library of 9,216 mutants was created and insertion sites were identified. A single transposon mutant was selected for each gene to assemble a condensed library consisting of 2,913 unique nonessential mutants. Using a modified UTI model in BALB/c mice, we identified 36 genes important for colonizing the bladder, including purB, yihE, and carB. Screening of the condensed library in vitro identified yigP and ubiG to be essential for growth in human urine. Additionally, we developed a novel quantitative PCR (qPCR) technique to identify genes with fitness defects within defined subgroups of related genes (e.g., genes encoding fimbriae, toxins, etc.) following UTI. The number of mutants within these subgroups circumvents bottleneck restriction and facilitates validation of multiple mutants to generate individual competitive indices. Collectively, this study investigates the bottleneck effects during UTI, provides two techniques for evading those effects that can be applied to other disease models, and contributes a genetic tool in prototype strain CFT073 to the field. IMPORTANCE Uropathogenic Escherichia coli strains cause most uncomplicated urinary tract infections (UTI), one of the most common infectious diseases worldwide. Random transposon mutagenesis techniques have been utilized to identify essential bacterial genes during infection; however, this has been met with limitations when applied to the murine UTI model. Conventional high-throughput transposon mutagenesis screens are not feasible because of inoculum size restrictions due to a bottleneck during infection. Our study utilizes a condensed ordered transposon library, limiting the number of mutants while maintaining the largest possible genome coverage. Screening of this library in vivo, and in human urine in vitro, identified numerous candidate fitness factors. Additionally, we have developed a novel technique using qPCR to quantify bacterial outputs following infection with small subgroups of transposon mutants. Molecular approaches developed in this study will serve as useful tools to probe in vivo models that are restricted by anatomical, physiological, or genetic bottleneck limitations.

scholarly journals Restrictive Streptomycin Resistance Mutations Decrease the Formation of Attaching and Effacing Lesions in Escherichia coli O157:H7 Strains

2013 ◽  
Vol 57 (9) ◽  
pp. 4260-4266 ◽  
Author(s):  
Chun Chen ◽  
Carla A. Blumentritt ◽  
Meredith M. Curtis ◽  
Vanessa Sperandio ◽  
Alfredo G. Torres ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Ribosomal Subunit ◽  
Streptomycin Resistance ◽  
Resistance Mutations ◽  
Content Type ◽  
E Coli ◽  
Enterocyte Effacement ◽  
Translocated Proteins

ABSTRACTStreptomycin binds to the bacterial ribosome and disrupts protein synthesis by promoting misreading of mRNA. Restrictive mutations on the ribosomal subunit protein S12 confer a streptomycin resistance (Strr) phenotype and concomitantly increase the accuracy of the decoding process and decrease the rate of translation. Spontaneous Strrmutants ofEscherichia coliO157:H7 have been generated forin vivostudies to promote colonization and to provide a selective marker for this pathogen. The locus of enterocyte effacement (LEE) ofE. coliO157:H7 encodes a type III secretion system (T3SS), which is required for attaching and effacing to the intestinal epithelium. In this study, we observed decreases in both the expression and secretion levels of the T3SS translocated proteins EspA and EspB inE. coliO157:H7 Strrrestrictive mutants, which have K42T or K42I mutations in S12. However, mildly restrictive (K87R) and nonrestrictive (K42R) mutants showed slight or indistinguishable changes in EspA and EspB secretion. Adherence and actin staining assays indicated that restrictive mutations compromised the formation of attaching and effacing lesions inE. coliO157:H7. Therefore, we suggest thatE. coliO157:H7 strains selected for Strrshould be thoroughly characterized beforein vivoandin vitroexperiments that assay for LEE-directed phenotypes and that strains carrying nonrestrictive mutations such as K42R make better surrogates of wild-type strains than those carrying restrictive mutations.

scholarly journals Antibiotic Trapping by Plasmid-Encoded CMY-2 β-Lactamase Combined with Reduced Outer Membrane Permeability as a Mechanism of Carbapenem Resistance in Escherichia coli

2013 ◽  
Vol 57 (8) ◽  
pp. 3941-3949 ◽  
Author(s):  
Wil H. F. Goessens ◽  
Akke K. van der Bij ◽  
Ria van Boxtel ◽  
Johann D. D. Pitout ◽  
Peter van Ulsen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Membrane Permeability ◽  
Outer Membrane ◽  
Carbapenem Resistance ◽  
Covalent Modification ◽  
Content Type ◽  
E Coli ◽  
Outer Membrane Permeability

ABSTRACTA liver transplant patient was admitted with cholangitis, for which meropenem therapy was started. Initial cultures showed a carbapenem-susceptible (CS)Escherichia colistrain, but during admission, a carbapenem-resistant (CR)E. colistrain was isolated. Analysis of the outer membrane protein profiles showed that both CS and CRE. colilacked the porins OmpF and OmpC. Furthermore, PCR and sequence analysis revealed that both CS and CRE. colipossessedblaCTX-M-15andblaOXA-1. The CRE. colistrain additionally harboredblaCMY-2and demonstrated a >15-fold increase in β-lactamase activity against nitrocefin, but no hydrolysis of meropenem was detected. However, nitrocefin hydrolysis appeared strongly inhibited by meropenem. Furthermore, the CMY-2 enzyme demonstrated lower electrophoretic mobility after its incubation eitherin vitroorin vivowith meropenem, indicative of its covalent modification with meropenem. The presence of the acyl-enzyme complex was confirmed by mass spectrometry. By transformation of the CMY-2-encoding plasmid into variousE. colistrains, it was established that both porin deficiency and high-level expression of the enzyme were needed to confer meropenem resistance. In conclusion, carbapenem resistance emerged by a combination of elevated β-lactamase production and lack of porin expression. Due to the reduced outer membrane permeability, only small amounts of meropenem can enter the periplasm, where they are trapped but not degraded by the large amount of the β-lactamase. This study, therefore, provides evidence that the mechanism of “trapping” by CMY-2 β-lactamase plays a role in carbapenem resistance.

scholarly journals Complete Genome Sequence of Escherichia coli 81009, a Representative of the Sequence Type 131 C1-M27 Clade with a Multidrug-Resistant Phenotype

2018 ◽  
Vol 6 (8) ◽  
Author(s):  
Michele Mutti ◽  
Ágnes Sonnevend ◽  
Tibor Pál ◽  
Sini Junttila ◽  
Heinz Ekker ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Monoclonal Antibodies ◽  
Genome Sequence ◽  
Complete Genome Sequence ◽  
Complete Genome ◽  
Significant Proportion ◽  
Multidrug Resistant ◽  
Sequence Type ◽  
E Coli ◽  
Resistant Phenotype

ABSTRACT The sequence type 131 (ST131)- H 30 clone is responsible for a significant proportion of multidrug-resistant extraintestinal Escherichia coli infections. Recently, the C1-M27 clade of ST131- H 30, associated with bla CTX-M-27 , has emerged. The complete genome sequence of E. coli isolate 81009 belonging to this clone, previously used during the development of ST131-specific monoclonal antibodies, is reported here.

scholarly journals Peptidoglycan sensing prevents quiescence and promotes quorum-independent growth of uropathogenic Escherichia coli

2019 ◽  
Author(s):  
Eric C. DiBiasio ◽  
Hilary J. Ranson ◽  
James R. Johnson ◽  
David C. Rowley ◽  
Paul S. Cohen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Structural Integrity ◽  
Recurrent Infection ◽  
Multidrug Resistant ◽  
Quiescent State ◽  
E Coli ◽  
Tract Infections ◽  
Patients Experience

AbstractThe layer of peptidoglycan surrounding bacteria provides structural integrity for the bacterial cell wall. Many organisms, including human cells and diverse bacteria, detect peptidoglycan fragments that are released as bacteria grow. Uropathogenic Escherichia coli (UPEC) strains are the leading cause of human urinary tract infections (UTIs) and many patients experience recurrent infection after successful antibiotic treatment. The source of recurrent infections may be persistent bacterial reservoirs in vivo that are in a quiescent state and, thus, are not susceptible to antibiotics. Here, we show that multiple UPEC strains require a quorum to proliferate in vitro with glucose as the sole carbon source; at low density, the bacteria remain viable but enter a quiescent, non-proliferative state. Of all clinical UPEC isolates tested to date, 35% (51/145) enter this quiescent state, including archetypal strains CFT073 (from classic endemic lineage ST73) and JJ1886 (from recently emerged, multidrug-resistant pandemic lineage ST131). We further show that quorum-dependent UPEC quiescence is prevented and reversed by small molecules, called proliferants, that stimulate growth, such as L-lysine, L-methionine, and peptidoglycan (PG) stem peptides, including an isolated PG pentapeptide from Staphylococcus aureus. Together, our results indicate that (i) uptake of L-lysine and (ii) PG peptide sensing by UPEC modulate the quorum-regulated decision to proliferate and further demonstrate that PG fragments are important for intra- and interspecies signaling in pathogenic E. coli.

scholarly journals A SimpleIn VitroGut Model for Studying the Interaction betweenEscherichia coliand the Intestinal Commensal Microbiota in Cecal Mucus

2018 ◽  
Vol 84 (24) ◽  
Author(s):  
Matthew E. Mokszycki ◽  
Mary Leatham-Jensen ◽  
Jon L. Steffensen ◽  
Ying Zhang ◽  
Karen A. Krogfelt ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Gene Profiling ◽  
Rrna Gene ◽  
Mucus Layer ◽  
Content Type ◽  
E Coli ◽  
Commensal Microbiota ◽  
Intestinal Mucus

ABSTRACTA novelin vitrogut model was developed to better understand the interactions betweenEscherichia coliand the mouse cecal mucus commensal microbiota. The gut model is simple and inexpensive while providing an environment that largely replicates the nonadherent mucus layer of the mouse cecum. 16S rRNA gene profiling of the cecal microbial communities of streptomycin-treated mice colonized withE. coliMG1655 orE. coliNissle 1917 and the gut model confirmed that the gut model properly reflected the community structure of the mouse intestine. Furthermore, the results from thein vitrogut model mimic the results of publishedin vivocompetitive colonization experiments. The gut model is initiated by the colonization of streptomycin-treated mice, and then the community is serially transferred in microcentrifuge tubes in an anaerobic environment generated in anaerobe jars. The nutritional makeup of the cecum is simulated in the gut model by using a medium consisting of porcine mucin, mouse cecal mucus, HEPES-Hanks buffer (pH 7.2), Cleland’s reagent, and agarose. Agarose was found to be essential for maintaining the stability of the microbial community in the gut model. The outcome of competitions betweenE. colistrains in thein vitrogut model is readily explained by the “restaurant hypothesis” of intestinal colonization. This simple model system potentially can be used to more fully understand how different members of the microbiota interact physically and metabolically during the colonization of the intestinal mucus layer.IMPORTANCEBoth commensal and pathogenic strains ofEscherichia coliappear to colonize the mammalian intestine by interacting physically and metabolically with other members of the microbiota in the mucus layer that overlays the cecal and colonic epithelium. However, the use of animal models and the complexity of the mammalian gut make it difficult to isolate experimental variables that might dictate the interactions betweenE. coliand other members of the microbiota, such as those that are critical for successful colonization. Here, we describe a simple and relatively inexpensivein vitrogut model that largely mimicsin vivoconditions and therefore can facilitate the manipulation of experimental variables for studying the interactions ofE. coliwith the intestinal microbiota.

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