Characterization of the flexible genome complement of the commensal Escherichia coli strain A0 34/86 (O83 : K24 : H31)

Microbiology ◽  
2005 ◽  
Vol 151 (2) ◽  
pp. 385-398 ◽  
Author(s):  
Jana Hejnova ◽  
Ulrich Dobrindt ◽  
Radka Nemcova ◽  
Christophe Rusniok ◽  
Alojz Bomba ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Gene Clusters ◽  
Coli Strain ◽  
Bac Clone ◽  
Sequence Comparisons ◽  
Multiple Sequence ◽  
E Coli ◽  
Gut Colonization ◽  
K 12

Colonization by the commensal Escherichia coli strain A0 34/86 (O83 : K24 : H31) has proved to be safe and efficient in the prophylaxis and treatment of nosocomial infections and diarrhoea of preterm and newborn infants in Czech paediatric clinics over the past three decades. In searching for traits contributing to this beneficial effect related to the gut colonization capacity of the strain, the authors have analysed its genome by DNA–DNA hybridization to E. coli K-12 (MG1655) genomic DNA arrays and to ‘Pathoarrays’, as well as by multiplex PCR, bacterial artificial chromosome (BAC) library cloning and shotgun sequencing. Four hundred and ten E. coli K-12 ORFs were absent from A0 34/86, while 72 out of 456 genes associated with pathogenicity islands of E. coli and Shigella were also detected in E. coli A0 34/86. Furthermore, extraintestinal pathogenic E. coli-related genes involved in iron uptake and adhesion were detected by multiplex PCR, and genes encoding the HlyA and cytotoxic necrotizing factor toxins, together with 21 genes of the uropathogenic E. coli 536 pathogenicity island II, were identified by analysis of 2304 shotgun and 1344 BAC clone sequences of A0 34/86 DNA. Multiple sequence comparisons identified 31 kb of DNA specific for E. coli A0 34/86; some of the genes carried by this DNA may prove to be implicated in the colonization capacity of the strain, enabling it to outcompete pathogens. Among 100 examined BAC clones roughly covering the A0 34/86 genome, one reproducibly conferred on the laboratory strain DH10B an enhanced capacity to persist in the intestine of newborn piglets. Sequencing revealed that this BAC clone carried gene clusters encoding gluconate and mannonate metabolism, adhesion (fim), invasion (ibe) and restriction/modification functions. Hence, the genome of this clinically safe and highly efficient colonizer strain appears to harbour many ‘virulence-associated’ genes. These results highlight the thin line between bacterial ‘virulence’ and ‘fitness' or ‘colonization’ factors, and question the definition of enterobacterial virulence factors.

scholarly journals The lipopolysaccharide of the mastitis isolate Escherichia coli strain 1303 comprises a novel O-antigen and the rare K-12 core type

Microbiology ◽  
2011 ◽  
Vol 157 (6) ◽  
pp. 1750-1760 ◽  
Author(s):  
Katarzyna A. Duda ◽  
Buko Lindner ◽  
Helmut Brade ◽  
Andreas Leimbach ◽  
Elżbieta Brzuszkiewicz ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Inner Core ◽  
Bovine Mastitis ◽  
Outer Core ◽  
Coli Strain ◽  
Core Oligosaccharide ◽  
2D Nmr Spectroscopy ◽  
E Coli ◽  
O Antigen ◽  
K 12

Mastitis represents one of the most significant health problems of dairy herds. The two major causative agents of this disease are Escherichia coli and Staphylococcus aureus. Of the first, its lipopolysaccharide (LPS) is thought to play a prominent role during infection. Here, we report the O-antigen (OPS, O-specific polysaccharide) structure of the LPS from bovine mastitis isolate E. coli 1303. The structure was determined utilizing chemical analyses, mass spectrometry, and 1D and 2D NMR spectroscopy methods. The O-repeating unit was characterized as -[→4)-β-d-Quip3NAc-(1→3)-α-l-Fucp2OAc-(1→4)-β-d-Galp-(1→3)-α-d-GalpNAc-(1→]- in which the O-acetyl substitution was non-stoichiometric. The nucleotide sequence of the O-antigen gene cluster of E. coli 1303 was also determined. This cluster, located between the gnd and galF genes, contains 13 putative open reading frames, most of which represent unknown nucleotide sequences that have not been described before. The O-antigen of E. coli 1303 was shown to substitute O-7 of the terminal ld-heptose of the K-12 core oligosaccharide. Interestingly, the non-OPS-substituted core oligosaccharide represented a truncated version of the K-12 outer core – namely terminal ld-heptose and glucose were missing; however, it possessed a third Kdo residue in the inner core. On the basis of structural and genetic data we show that the mastitis isolate E. coli 1303 represents a new serotype and possesses the K-12 core type, which is rather uncommon among human and bovine isolates.

A SitABCD homologue from an avian pathogenic Escherichia coli strain mediates transport of iron and manganese and resistance to hydrogen peroxide

Microbiology ◽  
2006 ◽  
Vol 152 (3) ◽  
pp. 745-758 ◽  
Author(s):  
Mourad Sabri ◽  
Simon Léveillé ◽  
Charles M. Dozois
Keyword(s):  
Escherichia Coli ◽  
Hydrogen Peroxide ◽  
Metal Ion ◽  
Growth Promotion ◽  
Coli Strain ◽  
Transport Systems ◽  
Virulence Plasmid ◽  
E Coli ◽  
K 12 ◽  
Iron And Manganese

An operon encoding a member of the family of ATP-binding cassette (ABC) divalent metal ion transporters, homologous to Salmonella enterica SitABCD, has been identified in the avian pathogenic Escherichia coli (APEC) strain χ7122. The sitABCD genes were located on the virulence plasmid pAPEC-1, and were highly similar at the nucleotide level to the chromosomally encoded sitABCD genes present in Shigella spp. A cloned copy of sitABCD conferred increased growth upon a siderophore-deficient E. coli strain grown in nutrient broth supplemented with the chelator 2,2′-dipyridyl. Ion rescue demonstrated that Sit-mediated growth promotion of this strain was due to the transport of iron. SitABCD mediated increased transport of both iron and manganese as demonstrated by uptake of 55Fe, 59Fe or 54Mn in E. coli K-12 strains deficient for the transport of iron (aroB feoB) and manganese (mntH) respectively. Isotope uptake and transport inhibition studies showed that in the iron transport deficient strain, SitABCD demonstrated a greater affinity for iron than for manganese, and SitABCD-mediated transport was higher for ferrous iron, whereas in the manganese transport deficient strain, SitABCD demonstrated greater affinity for manganese than for iron. Introduction of the APEC sitABCD genes into an E. coli K-12 mntH mutant also conferred increased resistance to the bactericidal effects of hydrogen peroxide. APEC strain χ7122 derivatives lacking either a functional SitABCD or a functional MntH transport system were as resistant to hydrogen peroxide as the wild-type strain, whereas a Δsit ΔmntH double mutant was more sensitive to hydrogen peroxide. Overall, the results demonstrate that in E. coli SitABCD represents a manganese and iron transporter that, in combination with other ion transport systems, may contribute to acquisition of iron and manganese, and resistance to oxidative stress.

scholarly journals Cluster II che Genes from Pseudomonas aeruginosa Are Required for an Optimal Chemotactic Response

2002 ◽  
Vol 184 (16) ◽  
pp. 4374-4383 ◽  
Author(s):  
Abel Ferrández ◽  
Andrew C. Hawkins ◽  
Douglas T. Summerfield ◽  
Caroline S. Harwood
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Gene Clusters ◽  
Soft Agar ◽  
Chemotactic Response ◽  
Wild Type ◽  
E Coli ◽  
Chemotaxis Proteins ◽  
Complete Set ◽  
K 12

ABSTRACT Pseudomonas aeruginosa, a γ-proteobacterium, is motile by means of a single polar flagellum and is chemotactic to a variety of organic compounds and phosphate. P. aeruginosa has multiple homologues of Escherichia coli chemotaxis genes that are organized into five gene clusters. Previously, it was demonstrated that genes in cluster I and cluster V are essential for chemotaxis. A third cluster (cluster II) contains a complete set of che genes, as well as two genes, mcpA and mcpB, encoding methyl-accepting chemotaxis proteins. Mutations were constructed in several of the cluster II che genes and in the mcp genes to examine their possible contributions to P. aeruginosa chemotaxis. A cheB2 mutant was partially impaired in chemotaxis in soft-agar swarm plate assays. Providing cheB2 in trans complemented this defect. Further, overexpression of CheB2 restored chemotaxis to a completely nonchemotactic, cluster I, cheB-deficient strain to near wild-type levels. An mcpA mutant was defective in chemotaxis in media that were low in magnesium. The defect could be relieved by the addition of magnesium to the swarm plate medium. An mcpB mutant was defective in chemotaxis when assayed in dilute rich soft-agar swarm medium or in minimal-medium swarm plates containing any 1 of 60 chemoattractants. The mutant phenotype could be complemented by the addition of mcpB in trans. Overexpression of either McpA or McpB in P. aeruginosa or Escherichia coli resulted in impairment of chemotaxis, and these cells had smooth-swimming phenotypes when observed under the microscope. Expression of P. aeruginosa cheA2, cheB2, or cheW2 in E. coli K-12 completely disrupted wild-type chemotaxis, while expression of cheY2 had no effect. These results indicate that che cluster II genes are expressed in P. aeruginosa and are required for an optimal chemotactic response.

Cloning and expression of the beta-D-phosphogalactoside galactohydrolase gene of Lactobacillus casei in Escherichia coli K-12

1982 ◽  
Vol 152 (3) ◽  
pp. 1138-1146
Author(s):  
L J Lee ◽  
J B Hansen ◽  
E K Jagusztyn-Krynicka ◽  
B M Chassy
Keyword(s):  
Escherichia Coli ◽  
Lactobacillus Casei ◽  
Specific Activity ◽  
Protein Product ◽  
Coli Strain ◽  
Cloning And Expression ◽  
E Coli ◽  
Lactose Metabolism ◽  
Gene Coding ◽  
K 12

Lactose metabolism in Lactobacillus casei 64H is associated with the presence of plasmid pLZ64. This plasmid determines both phosphoenolpyruvate-dependent phosphotransferase uptake of lactose and beta-D-phosphogalactoside galactohydrolase. A shotgun clone bank of chimeric plasmids containing restriction enzyme digest fragments of pLZ64 DNA was constructed in Escherichia coli K-12. One clone contained the gene coding for beta-D-phosphogalactoside galactohydrolase on a 7.9-kilobase PstI fragment cloned into the vector pBR322 in E. coli strain chi 1849. The beta-D-phosphogalactoside galactohydrolase enzyme isolated from E. coli showed no difference from that isolated from L. casei, and specific activity of beta-D-phosphogalactoside galactohydrolase was stimulated 1.8-fold in E. coli by growth in media containing beta-galactosides. A restriction map of the recombinant plasmid was compiled, and with that information, a series of subclones was constructed. From an analysis of the proteins produced by minicells prepared from transformant E. coli cells containing each of the recombinant subclone plasmids, it was found that the gene for the 56-kilodalton beta-D-phosphogalactoside galactohydrolase was transcribed from an L. casei-derived promoter. The gene for a second protein product (43 kilodaltons) was transcribed in the opposite direction, presumably under the control of a promoter in pBR322. The relationship of this second product to the lactose metabolism genes of L. casei is at present unknown.

The contribution of small cryptic plasmids to the antibiotic resistance of enteropathogenic Escherichia coli E2348/69

2009 ◽  
Vol 55 (11) ◽  
pp. 1229-1239 ◽  
Author(s):  
Cynthia L. Handford ◽  
Charma T. Stang ◽  
Tracy L. Raivio ◽  
Jonathan J. Dennis
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Dna Sequence ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Gene Clusters ◽  
Coli Strain ◽  
Enteropathogenic Escherichia Coli ◽  
E Coli ◽  
Cryptic Plasmids

Two uncharacterized small cryptic plasmids (SCPs) were isolated from enteropathogenic Escherichia coli strain E2348/69. Genomic DNA sequence analysis of both SCPs indicated that the smaller plasmid, p5217, encoded several mobilization genes, whereas the larger plasmid, p6148, encoded several putative antibiotic resistance determinants. Complementation analysis showed that p6148 encodes functional streptomycin resistance genes but, owing to the presence of several frameshift mutations, a nonfunctional sulfonamide resistance determinant. A plasmid similar to p6148 has previously been shown to confer a slight growth advantage on E. coli. However, we were unable to observe any significant growth advantage in different E. coli strains transformed with p6148. The p6148 DNA sequence is homologous in sequence and arrangement to DNA from other plasmid families, including large conjugative plasmids and SXT integrative and conjugative elements. This study suggests that gene clusters of the sul2–strAB antibiotic resistance genes are widespread and highly transferable, owing to their presence in a wide variety of mobile genetic elements.

scholarly journals A Commensal Gone Bad: Complete Genome Sequence of the Prototypical Enterotoxigenic Escherichia coli Strain H10407

2010 ◽  
Vol 192 (21) ◽  
pp. 5822-5831 ◽  
Author(s):  
Lisa C. Crossman ◽  
Roy R. Chaudhuri ◽  
Scott A. Beatson ◽  
Timothy J. Wells ◽  
Mickael Desvaux ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Genomic Sequence ◽  
Epidemiological Data ◽  
Coli Strain ◽  
Enterotoxigenic Escherichia Coli ◽  
E Coli ◽  
Extraintestinal Disease ◽  
K 12 ◽  
Genetic Context ◽  
Complete Genomic

ABSTRACT In most cases, Escherichia coli exists as a harmless commensal organism, but it may on occasion cause intestinal and/or extraintestinal disease. Enterotoxigenic E. coli (ETEC) is the predominant cause of E. coli-mediated diarrhea in the developing world and is responsible for a significant portion of pediatric deaths. In this study, we determined the complete genomic sequence of E. coli H10407, a prototypical strain of enterotoxigenic E. coli, which reproducibly elicits diarrhea in human volunteer studies. We performed genomic and phylogenetic comparisons with other E. coli strains, revealing that the chromosome is closely related to that of the nonpathogenic commensal strain E. coli HS and to those of the laboratory strains E. coli K-12 and C. Furthermore, these analyses demonstrated that there were no chromosomally encoded factors unique to any sequenced ETEC strains. Comparison of the E. coli H10407 plasmids with those from several ETEC strains revealed that the plasmids had a mosaic structure but that several loci were conserved among ETEC strains. This study provides a genetic context for the vast amount of experimental and epidemiological data that have been published.

Escherichia coliserogroup O2 and O28ac O-antigen gene cluster sequences and detection of pathogenicE. coliO2 and O28ac by PCR

2010 ◽  
Vol 56 (4) ◽  
pp. 308-316 ◽  
Author(s):  
Pina M. Fratamico ◽  
Xianghe Yan ◽  
Yanhong Liu ◽  
Chitrita DebRoy ◽  
Brian Byrne ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Gene Cluster ◽  
Gene Clusters ◽  
Rapid Identification ◽  
Open Reading Frames ◽  
E Coli ◽  
Antigen Gene ◽  
O Antigen ◽  
Pcr Assays

The O-antigen gene clusters of Escherichia coli serogroups O2 and O28ac were sequenced, and PCR assays were developed to identify strains belonging to these 2 serogroups. Sixteen and 8 open reading frames were mapped to these loci in E. coli O2:H4 U 9-41 and E. coli O28ac:H25 96-3286, respectively. The wzx (O-antigen flippase) and wzy (O-antigen polymerase) genes in the E. coli O2 and O28ac O-antigen gene clusters were selected as targets for PCR assays for their identification. PCR assays targeting the wzx and wzy genes were specific for these serogroups, with one exception. Escherichia coli serogroup O42 strains gave positive results with wzx and wzy PCR assays targeting E. coli O28ac, and antiserum raised against O42 cross-reacted with serogroup O28ac strains. The O-antigen gene cluster of a strain of E. coli serogroup O42 was sequenced, and there were only 3 nt differences between the O-antigen gene clusters of the O28ac and O42 strains. Multiplex PCR assays targeting the O2 wzx gene, the stx1, stx2, hly, eae, and saa genes, and the O28ac wzx, ial, ipaC, and ipaH genes were developed for detecting Shiga toxin-producing E. coli O2 strains and enteroinvasive E. coli O28ac strains, respectively. The O2 and O28ac wzx and wzy genes can be used as diagnostic markers in PCR assays for rapid identification of these serogroups as an alternative to serotyping, and the multiplex PCR assays targeting serogroup-specific genes in combination with virulence genes can be used to identify and to detect pathogenic serogroup O2 and O28ac strains.

scholarly journals Bacterial Interspersed Mosaic Elements (BIMEs) Are a Major Source of Sequence Polymorphism in Escherichia coli Intergenic Regions Including Specific Associations With a New Insertion Sequence

Genetics ◽  
1997 ◽  
Vol 145 (3) ◽  
pp. 551-562 ◽  
Author(s):  
Sophie Bachellier ◽  
Jean-Marie Clément ◽  
Maurice Hofnung ◽  
Eric Gilson
Keyword(s):  
Escherichia Coli ◽  
Dna Sequences ◽  
Multiple Sequence ◽  
E Coli ◽  
Sequence Organization ◽  
Is Element ◽  
Integration Sites ◽  
Intergenic Regions ◽  
Natural Isolates ◽  
K 12

A significant fraction of Escherichia coli intergenic DNA sequences is composed of two families of repeated bacterial interspersed mosaic elements (BIME-1 and BIME-2). In this study, we determined the sequence organization of six intergenic regions in 51 E. coli and Shigella natural isolates. Each region contains a BIME in E. coli K-12. We found that multiple sequence variations are located within or near these BIMEs in the different bacteria. Events included excisions of a whole BIME-1, expansion/deletion within a BIME-2 and insertions of non-BIME sequences like the boxC repeat or a new IS element, named IS 1397. Remarkably, 14 out of 14 IS 1397 integration sites correspond to a BIME sequence, strongly suggesting that this IS element is specifically associated with BIMEs, and thus inserts only in extragenic regions. Unlike BIMEs, IS 1397 is not detected in all E. coli isolates. Possible relationships between the presence of this IS element and the evolution of BIMEs are discussed.

scholarly journals Development of PCR Assays Targeting Genes in O-Antigen Gene Clusters for Detection and Identification of Escherichia coli O45 and O55 Serogroups

2005 ◽  
Vol 71 (8) ◽  
pp. 4919-4924 ◽  
Author(s):  
Chitrita DebRoy ◽  
Pina M. Fratamico ◽  
Elisabeth Roberts ◽  
Michael A. Davis ◽  
Yanhong Liu
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Gene Cluster ◽  
Real Time Pcr ◽  
Gene Clusters ◽  
E Coli ◽  
Antigen Gene ◽  
O Antigen ◽  
Detection And Identification ◽  
Pcr Assays

ABSTRACT The Escherichia coli O45 O-antigen gene cluster of strain O45:H2 96-3285 was sequenced, and conventional (singleplex), multiplex, and real-time PCR assays were designed to amplify regions in the wzx (O-antigen flippase) and wzy (O-antigen polymerase) genes. In addition, PCR assays targeting the E. coli O55 wzx and wzy genes were designed based on previously published sequences. PCR assays targeting E. coli O45 showed 100% specificity for this serogroup, whereas by PCR assays specific for E. coli O55, 97/102 strains serotyped as E. coli O55 were positive for wzx and 98/102 for wzy. Multiplex PCR assays targeting the E. coli O45 and the E. coli O55 wzx and wzy genes were used to detect the organisms in fecal samples spiked at levels of 106 and 108 CFU/0.2 g feces. Thus, the PCR assays can be used to detect and identify E. coli serogroups O45 and O55.

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