scholarly journals DELETION AND AMBER MUTANTS OF fla LOCI IN ESCHERICHIA COLI K-12

Genetics ◽  
1976 ◽  
Vol 84 (3) ◽  
pp. 403-421
Author(s):  
Hisato Kondoh ◽  
Haruo Ozeki
Keyword(s):  
Escherichia Coli ◽  
High Temperature ◽  
Genetic Map ◽  
Screening Method ◽  
The Other ◽  
Rapid Screening ◽  
E Coli ◽  
Flagellar Filament ◽  
Rapid Screening Method ◽  
K 12

ABSTRACT A rapid screening method for amber fla mutants of E. coli was devised and many mutants were obtained. In addition, strains with deletions of the fla genes in the his-uvrC region were isolated from high-temperature survivors of a λcI857 lysogen in which the prophage is located between his and fla. Utilizing these mutants, eleven fla genes (I-XI) and one hag gene were identified in the his-uvrc region, in the following order: his-supD-I-II-(III, IV)-V-(VI, VII)-VIII-IX-hag-(X, XI)-uvrC. The fla genes X and XI and hag are located at about 42.5 min and the other fla genes at about 43.0 min on the E. coli genetic map (Bachmann, Low and Taylor 1976). Mutants of fla gene X showed a slight sensitivity to chi phage, although they lack the flagellar filament.

scholarly journals Quorum-Sensing Escherichia coli Regulator A: a Regulator of the LysR Family Involved in the Regulation of the Locus of Enterocyte Effacement Pathogenicity Island in Enterohemorrhagic E. coli

2002 ◽  
Vol 70 (6) ◽  
pp. 3085-3093 ◽  
Author(s):  
Vanessa Sperandio ◽  
Caiyi C. Li ◽  
James B. Kaper
Keyword(s):  
Escherichia Coli ◽  
Quorum Sensing ◽  
Type Iii Secretion ◽  
Pathogenicity Island ◽  
The Other ◽  
E Coli ◽  
Type Iii ◽  
Enterocyte Effacement ◽  
K 12 ◽  
Lysr Family

ABSTRACT The locus of enterocyte effacement (LEE) is a chromosomal pathogenicity island that encodes the proteins involved in the formation of the attaching and effacing lesions by enterohemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC). The LEE comprises 41 open reading frames organized in five major operons, LEE1, LEE2, LEE3, tir (LEE5), and LEE4, which encode a type III secretion system, the intimin adhesin, the translocated intimin receptor (Tir), and other effector proteins. The first gene of LEE1 encodes the Ler regulator, which activates all the other genes within the LEE. We previously reported that the LEE genes were activated by quorum sensing through Ler (V. Sperandio, J. L. Mellies, W. Nguyen, S. Shin, and J. B. Kaper, Proc. Natl. Acad. Sci. USA 96:15196-15201, 1999). In this study we report that a putative regulator in the E. coli genome is itself activated by quorum sensing. This regulator is encoded by open reading frame b3243; belongs to the LysR family of regulators; is present in EHEC, EPEC, and E. coli K-12; and shares homology with the AphB and PtxR regulators of Vibrio cholerae and Pseudomonas aeruginosa, respectively. We confirmed the activation of b3243 by quorum sensing by using transcriptional fusions and renamed this regulator quorum-sensing E. coli regulator A (QseA). We observed that QseA activated transcription of ler and therefore of the other LEE genes. An EHEC qseA mutant had a striking reduction of type III secretion activity, which was complemented when qseA was provided in trans. Similar results were also observed with a qseA mutant of EPEC. The QseA regulator is part of the regulatory cascade that regulates EHEC and EPEC virulence genes by quorum sensing.

scholarly journals Activation of Prophage eib Genes for Immunoglobulin-Binding Proteins by Genes from the IbrAB Genetic Island of Escherichia coli ECOR-9

2002 ◽  
Vol 184 (13) ◽  
pp. 3640-3648 ◽  
Author(s):  
Carol H. Sandt ◽  
James E. Hopper ◽  
Charles W. Hill
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Phylogenetic Group ◽  
The Other ◽  
Overlapping Genes ◽  
Left Boundary ◽  
E Coli ◽  
Genome Homology ◽  
K 12 ◽  
Immunoglobulin Binding

ABSTRACT Four distinct Escherichia coli immunoglobulin-binding (eib) genes, each of which encodes a surface-exposed protein that binds immunoglobulins in a nonimmune manner, are carried by separate prophages in E. coli reference (ECOR) strain ECOR-9. Each eib gene was transferred to test E. coli strains, both in the form of multicopy recombinant plasmids and as lysogenized prophage. The derived lysogens express little or no Eib protein, in sharp contrast to the parental lysogen, suggesting that ECOR-9 has an expression-enhancing activity that the derived lysogens lack. Supporting this hypothesis, we cloned from ECOR-9 overlapping genes, ibrA and ibrB (designation is derived from “immunoglobulin-binding regulator”), which together activated eib expression in the derived lysogens. The proteins encoded by ibrA and ibrB are very similar to uncharacterized proteins encoded by genes of Salmonella enterica serovar Typhi and E. coli O157:H7 (in a prophage-like element of the Sakai strain and in two O islands of strain EDL933). The genomic segment containing ibrA and ibrB has been designated the IbrAB island. It contains regions of homology to the Shiga toxin-converting prophage, Stx2, as well as genes homologous to phage antirepressor genes. The left boundary between the IbrAB island and the chromosomal framework is located near min 35.8 of the E. coli K-12 genome. Homology to IbrAB was found in certain other ECOR strains, including the other five eib-positive strains and most strains of the phylogenetic group B2. Sequencing of a 1.1-kb portion of ibrAB revealed that the other eib-positive strains diverge by ≤0.1% from ECOR-9, whereas eib-negative ECOR-47 diverges by 16%.

Development of a rapid screening method for the detection of pathogenic Escherichia coli using a combination of Colilert® Quanti-Trays/2000 and PCR

2010 ◽  
Vol 10 (1) ◽  
pp. 7-13 ◽  
Author(s):  
K. B. Omar ◽  
N. Potgieter ◽  
T. G. Barnard
Keyword(s):  
Escherichia Coli ◽  
Internal Control ◽  
Screening Method ◽  
Rapid Screening ◽  
E Coli ◽  
Toilet Seat ◽  
Pathogenic Escherichia Coli ◽  
Wide Range ◽  
Polymerase Chain ◽  
Pcr Targeting

The use of culture based methods for the detection of Escherichia coli (E. coli) only gives information on the occurrence of E. coli and not pathogenicity of the organisms detected. To detect the both indicator and diarrhoeagenic E. coli (DEC) a method was developed using the Colilert® Quanti-Tray/2000 system and polymerase chain reaction (PCR). A total of 619 samples were collected from springs (33), boreholes (24), taps (5), rivers (36), streams (8), domestic storage containers (253), raw sewage (1), final effluents (5), stool (149), soil (5) and toilet seat swab (100) samples from various provinces in South Africa. E. coli was enumerated using the Colilert® Quanti-Tray's/2000 and DNA extracted from E. coli positive wells and used as template for a multiplex-PCR targeting genes specific to entero-pathogenic- (EPEC), entero-haemorrhagic- (EHEC), entero-invasive- (EIEC), entero-toxigenic (ETEC) and entero-aggregative E. coli (EAEC). The internal control was detected in 99% of positive E. coli samples. EAEC was detected in 17%, EIEC in 4%, ETEC in 11%, EHEC in 9% and EPEC in 21% of the E. coli positive samples. It is shown that this method can be used for the detection of DEC from a wide range of samples enriched with Colilert® Quanti-Tray's/2000.

scholarly journals A Novel Putrescine Importer Required for Type 1 Pili-driven Surface Motility Induced by Extracellular Putrescine in Escherichia coli K-12

2011 ◽  
Vol 286 (12) ◽  
pp. 10185-10192 ◽  
Author(s):  
Shin Kurihara ◽  
Hideyuki Suzuki ◽  
Mayu Oshida ◽  
Yoshimi Benno
Keyword(s):  
Escherichia Coli ◽  
Plasmid Vector ◽  
The Other ◽  
Gene Deletions ◽  
E Coli ◽  
Type 1 Pili ◽  
Transport Assay ◽  
Surface Motility ◽  
K 12

Recently, many studies have reported that polyamines play a role in bacterial cell-to-cell signaling processes. The present study describes a novel putrescine importer required for induction of type 1 pili-driven surface motility. The surface motility of the Escherichia coli ΔspeAB ΔspeC ΔpotABCD strain, which cannot produce putrescine and cannot import spermidine from the medium, was induced by extracellular putrescine. Introduction of the gene deletions for known polyamine importers (ΔpotE, ΔpotFGHI, and ΔpuuP) or a putative polyamine importer (ΔydcSTUV) into the ΔspeAB ΔspeC ΔpotABCD strain did not affect putrescine-induced surface motility. The deletion of yeeF, an annotated putative putrescine importer, in the ΔspeAB ΔspeC ΔpotABCD ΔydcSTUV strain abolished surface motility in putrescine-supplemented medium. Complementation of yeeF by a plasmid vector restored surface motility. The surface motility observed in the present study was abolished by the deletion of fimA, suggesting that the surface motility is type 1 pili-driven. A transport assay using the yeeF+ or ΔyeeF strains revealed that YeeF is a novel putrescine importer. The Km of YeeF (155 μm) is 40 to 300 times higher than that of other importers reported previously. On the other hand, the Vmax of YeeF (9.3 nmol/min/mg) is comparable to that of PotABCD, PotFGHI, and PuuP. The low affinity of YeeF for putrescine may allow E. coli to sense the cell density depending on the concentration of extracellular putrescine.

scholarly journals Positive control of sulphate reduction in Escherichia coli. Isolation, characterization and mapping of cysteineless mutants of E. coli K 12

1968 ◽  
Vol 110 (3) ◽  
pp. 589-595 ◽  
Author(s):  
M. C. Jones-Mortimer
Keyword(s):  
Escherichia Coli ◽  
Salmonella Typhimurium ◽  
Genetic Map ◽  
Genetic Data ◽  
Positive Control ◽  
Sulphate Reduction ◽  
Biochemical Data ◽  
E Coli ◽  
K 12 ◽  
Single Enzyme

To determine to what extent the biosynthesis of cysteine in Escherichia coli resembles that in Salmonella typhimurium, the following experiments were performed. (1) Mutants of E. coli K 12 deficient in the biosynthesis of cysteine were isolated. (2) These mutants were classified by nutritional and biochemical criteria; some mutants lacked a single enzyme of sulphate reduction, other mutants appeared to lack two or more enzymes. (3) The genetic map predicted from the biochemical data alone is shown to be incorrect, and an alternative map, consistent with the genetic data, is proposed for the cys mutants of E. coli.

scholarly journals Rapid screening method for enterotoxigenic Escherichia coli

1977 ◽  
Vol 6 (3) ◽  
pp. 314-316
Author(s):  
M Gurwith
Keyword(s):  
Escherichia Coli ◽  
Adrenal Tumor ◽  
Screening Method ◽  
Brain Heart Infusion ◽  
Brain Heart Infusion Broth ◽  
Enterotoxigenic Escherichia Coli ◽  
Rapid Screening ◽  
Broth Culture ◽  
Rapid Screening Method ◽  
Culture Supernatants

A rapid screening method for detection of Escherichia coli producing heat-labile enterotoxin is described. Single colonies are transferred directly from primary culture plates into 96-well microculture plates containing 0.3 ml of brain heart infusion broth in each well. After 24 h at 37 degrees C, each brain heart infusion broth culture is assayed by the miniculture method in the corresponding well of a microculture plate in which Y-1 mouse adrenal tumor cells have been grown. All enterotoxigenic isolates detected by this method were confirmed in the assay but with culture supernatants.

scholarly journals Insertion Element IS3-Based PCR Method for Subtyping Escherichia coli O157:H7

1998 ◽  
Vol 36 (5) ◽  
pp. 1180-1184 ◽  
Author(s):  
Curt J. Thompson ◽  
Claire Daly ◽  
Timothy J. Barrett ◽  
Jane P. Getchell ◽  
Mary J. R. Gilchrist ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Sequences ◽  
Screening Method ◽  
Pcr Amplification ◽  
Rapid Screening ◽  
Insertion Element ◽  
E Coli ◽  
Pcr Method ◽  
Template Dna ◽  
Single Primer

An Escherichia coli O157:H7 subtyping method based on PCR amplification of variable DNA sequences between the repetitive element IS3 was developed. Template DNA was prepared by boiling cells in Chelex. Two separate IS3 PCR amplifications were performed for each isolate: one with a single primer (primer IS3A) and one with two primers (primers IS3A and IS3B). The IS3 PCR subtyping method was applied to 35 epidemiologically related and unrelated E. coli O157:H7 isolates that had been previously characterized by pulsed-field gel electrophoresis (PFGE). PFGE identified 25 different subtypes (difference of one or more bands). PCR with single primer IS3A and primer pair IS3A-IS3B identified 6 and 14 different subtypes, respectively. By combining the results of the two PCR amplifications, 15 different IS3 PCR subtypes were identified. While not as sensitive as PFGE, IS3 PCR subtyping grouped all outbreak-related isolates. IS3 PCR banding patterns were reproducible between amplifications and between subcultures. IS3 PCR could serve as a simple, rapid screening method for the identification of unrelated E. coli O157:H7 isolates.

scholarly journals Role of the rapA Gene in Controlling Antibiotic Resistance of Escherichia coli Biofilms

2007 ◽  
Vol 51 (10) ◽  
pp. 3650-3658 ◽  
Author(s):  
S. V. Lynch ◽  
L. Dixon ◽  
M. R. Benoit ◽  
E. L. Brodie ◽  
M. Keyhan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
High Throughput Screening ◽  
Screening Method ◽  
Resistance Mechanisms ◽  
Planktonic Cell ◽  
Penicillin G ◽  
Wild Type ◽  
E Coli ◽  
K 12 ◽  
Dual Mechanism

ABSTRACT By using a high-throughput screening method, a mutant of a uropathogenic Escherichia coli strain affected in the rapA gene was isolated. The mutant formed normal-architecture biofilms but showed decreased penicillin G resistance, although the mutation did not affect planktonic cell resistance. Transcriptome analysis showed that 22 genes were down-regulated in the mutant biofilm. One of these genes was yhcQ, which encodes a putative multidrug resistance pump. Mutants with mutations in this gene also formed biofilms with decreased resistance, although the effect was less pronounced than that of the rapA mutation. Thus, an additional mechanism(s) controlled by a rapA-regulated gene(s) was involved in wild-type biofilm resistance. The search for this mechanism was guided by the fact that another down-regulated gene in rapA biofilms, yeeZ, is suspected to be involved in extra cell wall-related functions. A comparison of the biofilm matrix of the wild-type and rapA strains revealed decreased polysaccharide quantities and coverage in the mutant biofilms. Furthermore, the (fluorescent) functional penicillin G homologue Bocillin FL penetrated the mutant biofilms more readily. The results strongly suggest a dual mechanism for the wild-type biofilm penicillin G resistance, retarded penetration, and effective efflux. The results of studies with an E. coli K-12 strain pointed to the same conclusion. Since efflux and penetration can be general resistance mechanisms, tests were conducted with other antibiotics. The rapA biofilm was also more sensitive to norfloxacin, chloramphenicol, and gentamicin.

scholarly journals The distribution of plasmids determining citrate utilization in citratè-positive variants ofEscherichia colifrom humans, domestic animals, feral birds and environments

1979 ◽  
Vol 83 (2) ◽  
pp. 331-344 ◽  
Author(s):  
Naotaka Ishiguro ◽  
Gihei Sato
Keyword(s):  
Escherichia Coli ◽  
Domestic Animals ◽  
Domestic Animal ◽  
The Other ◽  
Conjugative Plasmid ◽  
E Coli ◽  
Resistance Determinants ◽  
Sucrose Fermentation ◽  
R Plasmids ◽  
K 12

summarySixty-seven isolates of citrate-positive variants ofEscherichia coliwere isolated from human, domestic animal, feral bird and environmental sources. With the exception of citrate utilization, all isolates were identified as typicalE. coliby their biochemical reactions. The transmission of the ability to utilize citrate on Simmons' citrate agar was demonstrated in 53 (79·1%) out of the 67 citratepositiveE. colivariants obtained from various sources. Drug resistance determinants and citrate utilizing character were co-transmitted intoE. coliK-12 by conjugation among citrate-positiveE. coliisolates carrying R plasmids except for that isolated from horses. The other characters (haemolysin or colicin production, raffinose or sucrose fermentation) were not transmitted together with the citrate utilizing character. These facts suggested that the structural gene responsible for citrate utilizing ability in citrate-positive variants ofE. coliwas located on a conjugative plasmid.

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