scholarly journals Insertion Site Occupancy by stx2 Bacteriophages Depends on the Locus Availability of the Host Strain Chromosome

2007 ◽  
Vol 189 (18) ◽  
pp. 6645-6654 ◽  
Author(s):  
Ruth Serra-Moreno ◽  
Juan Jofre ◽  
Maite Muniesa
Keyword(s):  
Shiga Toxin ◽  
Insertion Site ◽  
Site Occupancy ◽  
Host Strain ◽  
Host Chromosome ◽  
Shiga Toxins ◽  
E Coli ◽  
Emergent Pathogen ◽  
Insertion Sites ◽  
Host Strains

ABSTRACT Shiga toxin-producing Escherichia coli (STEC) is an emergent pathogen characterized by the expression of Shiga toxins, which are encoded in the genomes of lambdoid phages. These phages are infectious for other members of the Enterobacteriaceae and establish lysogeny when they integrate into the host chromosome. Five insertion sites, used mainly by these prophages, have been described to date. In the present study, the insertion of stx 2 prophages in these sites was analyzed in 168 STEC strains isolated from cattle. Additionally, insertion sites were determined for stx 2 phages which (i) converted diverse laboratory host strains, (ii) coexisted with another stx 2 prophage, and (iii) infected a recombinant host strain lacking the most commonly used insertion site. Results show that depending on the host strain, phages preferentially use one insertion site. For the most part, yehV is occupied in STEC strains while wrbA is preferentially selected by the same stx phages in E. coli laboratory strains. If this primary insertion site is unavailable, then a secondary insertion site is selected. It can be concluded that insertion site occupancy by stx phages depends on the host strain and on the availability of the preferred locus in the host strain.

scholarly journals Greater Diversity of Shiga Toxin-Encoding Bacteriophage Insertion Sites among Escherichia coli O157:H7 Isolates from Cattle than in Those from Humans

2006 ◽  
Vol 73 (3) ◽  
pp. 671-679 ◽  
Author(s):  
Thomas E. Besser ◽  
Nurmohammad Shaikh ◽  
Nicholas J. Holt ◽  
Phillip I. Tarr ◽  
Michael E. Konkel ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Genetic Diversity ◽  
Shiga Toxin ◽  
Insertion Site ◽  
Reservoir Host ◽  
Clinical Isolates ◽  
Escherichia Coli O157 ◽  
Human Pathogen ◽  
E Coli ◽  
Insertion Sites

ABSTRACT Escherichia coli O157:H7, a zoonotic human pathogen for which domestic cattle are a reservoir host, produces a Shiga toxin(s) (Stx) encoded by bacteriophages. Chromosomal insertion sites of these bacteriophages define three principal genotypes (clusters 1 to 3) among clinical isolates of E. coli O157:H7. Stx-encoding bacteriophage insertion site genotypes of 282 clinical and 80 bovine isolates were evaluated. A total of 268 (95.0%) of the clinical isolates, but only 41 (51.3%) of the bovine isolates, belonged to cluster 1, 2, or 3 (P < 0.001). Thirteen additional genotypes were identified in isolates from both cattle and humans (four genotypes), from only cattle (seven genotypes), or from only humans (two genotypes). Two other markers previously associated with isolates from cattle or with clinical isolates showed similar associations with genotype groups within bovine isolates; the tir allele sp-1 and the Q 933W allele were under- and overrepresented, respectively, among cluster 1 to 3 genotypes. Stx-encoding bacteriophage insertion site typing demonstrated that there is broad genetic diversity of E. coli O157:H7 in the bovine reservoir and that numerous genotypes are significantly underrepresented among clinical isolates, consistent with the possibility that there is reduced virulence or transmissibility to humans of some bovine E. coli O157:H7 genotypes.

scholarly journals Diversity of Shiga Toxin-Producing Escherichia coli (STEC) O26:H11 Strains Examined viastxSubtypes and Insertion Sites of Stx and EspK Bacteriophages

2015 ◽  
Vol 81 (11) ◽  
pp. 3712-3721 ◽  
Author(s):  
Ludivine Bonanno ◽  
Estelle Loukiadis ◽  
Patricia Mariani-Kurkdjian ◽  
Eric Oswald ◽  
Lucille Garnier ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Sequence Type ◽  
Phylogenetic Groups ◽  
Content Type ◽  
Shiga Toxins ◽  
E Coli ◽  
Food Borne ◽  
Insertion Sites ◽  
Human Infections

ABSTRACTShiga toxin-producingEscherichia coli(STEC) is a food-borne pathogen that may be responsible for severe human infections. Only a limited number of serotypes, including O26:H11, are involved in the majority of serious cases and outbreaks. The main virulence factors, Shiga toxins (Stx), are encoded by bacteriophages. Seventy-four STEC O26:H11 strains of various origins (including human, dairy, and cattle) were characterized for theirstxsubtypes and Stx phage chromosomal insertion sites. The majority of food and cattle strains possessed thestx1asubtype, while human strains carried mainlystx1aorstx2a. ThewrbAandyehVgenes were the main Stx phage insertion sites in STEC O26:H11, followed distantly byyecEandsbcB. Interestingly, the occurrence of Stx phages inserted in theyecEgene was low in dairy strains. In most of the 29stx-negativeE. coliO26:H11 strains also studied here, these bacterial insertion sites were vacant. Multilocus sequence typing of 20stx-positive orstx-negativeE. coliO26:H11 strains showed that they were distributed into two phylogenetic groups defined by sequence type 21 (ST21) and ST29. Finally, an EspK-carrying phage was found inserted in thessrAgene in the majority of the STEC O26:H11 strains but in only a minority of thestx-negativeE. coliO26:H11 strains. The differences in thestxsubtypes and Stx phage insertion sites observed in STEC O26:H11 according to their origin might reflect that strains circulating in cattle and foods are clonally distinct from those isolated from human patients.

Detection, enumeration and isolation of strains carrying the stx2 gene from urban sewage

2003 ◽  
Vol 47 (3) ◽  
pp. 109-116 ◽  
Author(s):  
A.R. Blanch ◽  
C. García-Aljaro ◽  
M. Muniesa ◽  
J. Jofre
Keyword(s):  
Shiga Toxin ◽  
Colony Growth ◽  
Coliform Bacteria ◽  
Shiga Toxins ◽  
E Coli ◽  
Urban Sewage ◽  
Shiga Toxin 2 ◽  
Shiga Toxin Genes ◽  
Lateral Transmission ◽  
Stx2 Gene

Verotoxigenic Escherichia coli strains have been related with waterborne outbreaks. Besides 0157:H7, several serotypes of E. coli and other enterobacteria have been implicated in outbreaks and reported to carry the shiga toxin genes. Shiga toxins, stx1 and stx2, are important virulence factors of these strains. These genes have been linked to bacteriophages and consequently are susceptible to lateral transmission. To better understand the ecology of these genes a study of the presence of the shiga toxin 2 gene (stx2) among coliform bacteria present in sewage samples was carried out. A procedure based on colony hybridisation was developed for the isolation of enterobacteria carrying this gene. Colony growth on Chromocult® agar was transferred to a membrane and hybridised with a gene specific probe. The procedure allowed detection of about one colony carrying the gene among around 1,000 faecal coliform colonies. The numbers of bacteria carrying the gene in sewage were also estimated by PCR indicating that the numbers of bacteria carrying the stx2 gene were about 1/1,000 faecal coliforms. The detected numbers by both methods were similar. Positive colony hybridisation was detected in four sewage origins. Fifty-two colonies showing positive signal were isolated from the Chromocult® agar plates, confirmed to be stx2 positive by PCR and phenotypically characterised. Results of the characterisation showed certain diversity among the isolates even in isolates from the same sample. Most of these isolates would not have been isolated with the methods regularly used for the isolation of E. coli 0157:H7 strains. The method will allow study of the numbers and characteristics of bacteria carrying the stx2 gene in different water environments and isolate them in order to determine their role in the spread of the gene.

scholarly journals Virulence Characteristics and Antibiotic Resistance Profiles of Shiga Toxin-Producing Escherichia coli Isolates from Diverse Sources

Antibiotics ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 587
Author(s):  
Momna Rubab ◽  
Deog-Hwan Oh
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Shiga Toxin ◽  
Multidrug Resistant ◽  
Control Measures ◽  
Diffusion Method ◽  
Shiga Toxins ◽  
E Coli ◽  
And Control ◽  
High Degree

Shiga toxin-producing Escherichia coli (STEC) is an enteric pathogen that causes several gastrointestinal ailments in humans across the world. STEC’s ability to cause ailment is attributed to the presence of a broad range of known and putative virulence factors (VFs) including those that encode Shiga toxins. A total of 51 E. coli strains belonging to serogroups O26, O45, O103, O104, O113, O121, O145, and O157 were tested for the presence of nine VFs via PCR and for their susceptibility to 17 frequently used antibiotics using the disc diffusion method. The isolates belonged to eight different serotypes, including eight O serogroups and 12 H types. The frequency of the presence of key VFs were stx1 (76.47%), stx2 (86.27%), eae (100%), ehxA (98.03%), nleA (100%), ureC (94.11%), iha (96.07%), subA (9.80%), and saa (94.11%) in the E. coli strains. All E. coli strains carried seven or more distinct VFs and, among these, four isolates harbored all tested VFs. In addition, all E. coli strains had a high degree of antibiotic resistance and were multidrug resistant (MDR). These results show a high incidence frequency of VFs and heterogeneity of VFs and MDR profiles of E. coli strains. Moreover, half of the E. coli isolates (74.5%) were resistant to > 9 classes of antibiotics (more than 50% of the tested antibiotics). Thus, our findings highlight the importance of appropriate epidemiological and microbiological surveillance and control measures to prevent STEC disease in humans worldwide.

scholarly journals Shiga Toxin Gene Loss and Transfer In Vitro and In Vivo during Enterohemorrhagic Escherichia coli O26 Infection in Humans

2007 ◽  
Vol 73 (10) ◽  
pp. 3144-3150 ◽  
Author(s):  
Martina Bielaszewska ◽  
Rita Prager ◽  
Robin Köck ◽  
Alexander Mellmann ◽  
Wenlan Zhang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Gene Loss ◽  
Pulsed Field Gel Electrophoresis ◽  
Enterohemorrhagic Escherichia Coli ◽  
Toxin Gene ◽  
Shiga Toxins ◽  
E Coli

ABSTRACT Escherichia coli serogroup O26 consists of enterohemorrhagic E. coli (EHEC) and atypical enteropathogenic E. coli (aEPEC). The former produces Shiga toxins (Stx), major determinants of EHEC pathogenicity, encoded by bacteriophages; the latter is Stx negative. We have isolated EHEC O26 from patient stools early in illness and aEPEC O26 from stools later in illness, and vice versa. Intrapatient EHEC and aEPEC isolates had quite similar pulsed-field gel electrophoresis (PFGE) patterns, suggesting that they might have arisen by conversion between the EHEC and aEPEC pathotypes during infection. To test this hypothesis, we asked whether EHEC O26 can lose stx genes and whether aEPEC O26 can be lysogenized with Stx-encoding phages from EHEC O26 in vitro. The stx 2 loss associated with the loss of Stx2-encoding phages occurred in 10% to 14% of colonies tested. Conversely, Stx2- and, to a lesser extent, Stx1-encoding bacteriophages from EHEC O26 lysogenized aEPEC O26 isolates, converting them to EHEC strains. In the lysogens and EHEC O26 donors, Stx2-converting bacteriophages integrated in yecE or wrbA. The loss and gain of Stx-converting bacteriophages diversifies PFGE patterns; this parallels findings of similar but not identical PFGE patterns in the intrapatient EHEC and aEPEC O26 isolates. EHEC O26 and aEPEC O26 thus exist as a dynamic system whose members undergo ephemeral interconversions via loss and gain of Stx-encoding phages to yield different pathotypes. The suggested occurrence of this process in the human intestine has diagnostic, clinical, epidemiological, and evolutionary implications.

scholarly journals A New Immunoassay for Detecting All Subtypes of Shiga Toxins Produced by Shiga Toxin-Producing E. coli in Ground Beef

PLoS ONE ◽  
2016 ◽  
Vol 11 (1) ◽  
pp. e0148092 ◽  
Author(s):  
Xiaohua He ◽  
Qiulian Kong ◽  
Stephanie Patfield ◽  
Craig Skinner ◽  
Reuven Rasooly
Keyword(s):  
Shiga Toxin ◽  
Ground Beef ◽  
Shiga Toxins ◽  
E Coli

scholarly journals Emergence of Escherichia coli encoding Shiga toxin 2f in human Shiga toxin-producing E. coli (STEC) infections in the Netherlands, January 2008 to December 2011

2014 ◽  
Vol 19 (17) ◽  
Author(s):  
I Friesema ◽  
K van der Zwaluw ◽  
T Schuurman ◽  
M Kooistra-Smid ◽  
E Franz ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
The Netherlands ◽  
Shiga Toxin ◽  
Distinct Group ◽  
Shiga Toxins ◽  
E Coli ◽  
Mild Disease ◽  
Shiga Toxin 2 ◽  
Almost All ◽  
Human Infections

The Shiga toxins of Shiga toxin-producing Escherichia coli (STEC) can be divided into Shiga toxin 1 (Stx1) and Shiga toxin 2 (Stx2) with several sub-variants. Variant Stx2f is one of the latest described, but has been rarely associated with symptomatic human infections. In the enhanced STEC surveillance in the Netherlands, 198 STEC O157 cases and 351 STEC non-O157 cases, including 87 stx2f STEC isolates, were reported between 2008 and 2011. Most stx2f strains belonged to the serogroups O63:H6 (n=47, 54%), O113:H6 (n=12, 14%) and O125:H6 (n=12, 14%). Of the 87 stx2f isolates, 84 (97%) harboured the E. coli attaching and effacing (eae) gene, but not the enterohaemorrhagic E. coli haemolysin (hly) gene. Stx2f STEC infections show milder symptoms and a less severe clinical course than STEC O157 infections. Almost all infections with stx2f (n=83, 95%) occurred between June and December, compared to 170/198 (86%) of STEC O157 and 173/264 (66%) of other STEC non-O157. Stx2f STEC infections in the Netherlands are more common than anticipated, and form a distinct group within STEC with regard to virulence genes and the relatively mild disease.

scholarly journals Contribution and Interaction of Shiga Toxin Genes to Escherichia coli O157:H7 Virulence

Toxins ◽  
2019 ◽  
Vol 11 (10) ◽  
pp. 607 ◽  
Author(s):  
Gillian A.M. Tarr ◽  
Taryn Stokowski ◽  
Smriti Shringi ◽  
Phillip I. Tarr ◽  
Stephen B. Freedman ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Population Level ◽  
Escherichia Coli O157 ◽  
Additive Interaction ◽  
Shiga Toxins ◽  
E Coli ◽  
Uremic Syndrome ◽  
Shiga Toxin Genes

Escherichia coli O157:H7 is the predominant cause of diarrhea-associated hemolytic uremic syndrome (HUS) worldwide. Its cardinal virulence traits are Shiga toxins, which are encoded by stx genes, the most common of which are stx1a, stx2a, and stx2c. The toxins these genes encode differ in their in vitro and experimental phenotypes, but the human population-level impact of these differences is poorly understood. Using Shiga toxin-encoding bacteriophage insertion typing and real-time polymerase chain reaction, we genotyped isolates from 936 E. coli O157:H7 cases and verified HUS status via chart review. We compared the HUS risk between isolates with stx2a and those with stx2a and another gene and estimated additive interaction of the stx genes. Adjusted for age and symptoms, the HUS incidence of E. coli O157:H7 containing stx2a alone was 4.4% greater (95% confidence interval (CI) −0.3%, 9.1%) than when it occurred with stx1a. When stx1a and stx2a occur together, the risk of HUS was 27.1% lower (95% CI −87.8%, −2.3%) than would be expected if interaction were not present. At the population level, temporal or geographic shifts toward these genotypes should be monitored, and stx genotype may be an important consideration in clinically predicting HUS among E. coli O157:H7 cases.

scholarly journals Identification of Human-Pathogenic Strains of Shiga Toxin-Producing Escherichia coli from Food by a Combination of Serotyping and Molecular Typing of Shiga Toxin Genes

2007 ◽  
Vol 73 (15) ◽  
pp. 4769-4775 ◽  
Author(s):  
Lothar Beutin ◽  
Angelika Miko ◽  
Gladys Krause ◽  
Karin Pries ◽  
Sabine Haby ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Severe Illness ◽  
Shiga Toxins ◽  
E Coli ◽  
Food Borne ◽  
Length Polymorphisms ◽  
Beef Products ◽  
Pcr Products ◽  
Shiga Toxin Genes

ABSTRACT We examined 219 Shiga toxin-producing Escherichia coli (STEC) strains from meat, milk, and cheese samples collected in Germany between 2005 and 2006. All strains were investigated for their serotypes and for genetic variants of Shiga toxins 1 and 2 (Stx1 and Stx2). stx 1 or variant genes were detected in 88 (40.2%) strains and stx 2 and variants in 177 (80.8%) strains. Typing of stx genes was performed by stx-specific PCRs and by analysis of restriction fragment length polymorphisms (RFLP) of PCR products. Major genotypes of the Stx1 (stx 1, stx 1c, and stx 1d) and the Stx2 (stx 2, stx 2d, stx 2-O118, stx 2e, and stx 2g) families were detected, and multiple types of stx genes coexisted frequently in STEC strains. Only 1.8% of the STEC strains from food belonged to the classical enterohemorrhagic E. coli (EHEC) types O26:H11, O103:H2, and O157:H7, and only 5.0% of the STEC strains from food were positive for the eae gene, which is a virulence trait of classical EHEC. In contrast, 95 (43.4%) of the food-borne STEC strains carried stx 2 and/or mucus-activatable stx 2d genes, an indicator for potential high virulence of STEC for humans. Most of these strains belonged to serotypes associated with severe illness in humans, such as O22:H8, O91:H21, O113:H21, O174:H2, and O174:H21. stx 2 and stx 2d STEC strains were found frequently in milk and beef products. Other stx types were associated more frequently with pork (stx 2e), lamb, and wildlife meat (stx 1c). The combination of serotyping and stx genotyping was found useful for identification and for assignment of food-borne STEC to groups with potential lower and higher levels of virulence for humans.

scholarly journals Construction of high copy number and highly stable Escherichia coli-Bacillus subtilis shuttle plasmids based on pWB980

2019 ◽  
Author(s):  
XingYa Zhao ◽  
JianYong Xu ◽  
Ming Tan ◽  
Jie Zhen ◽  
WenJu Shu ◽  
...  
Keyword(s):  
Copy Number ◽  
Insertion Site ◽  
Pectate Lyase ◽  
High Copy Number ◽  
Expression Vectors ◽  
Upstream Region ◽  
Transformation Rate ◽  
E Coli ◽  
Shuttle Plasmid ◽  
Insertion Sites

Abstract Background: pWB980 derived from pUB110 is a promising expression vector in Bacillus for its high copy number and high stability. However, the low transformation rate of recombinant plasmids to the wild cells limited the application of it. On the basis of pWB980, constructing an E. coli - B. subtilis shuttle plasmid could facilitate the transformation rate to Bacillus cells. Because the insertion site for E. coli replication origin sequence ( ori ) is not unique in pWB980, in order to investigate the best insertion site, eight shuttle plasmids (pUC980-1~pUC980-8) containing all possible insertion sites and directions were constructed.Results: The results showed that all the selected insertion sites could be used to construct shuttle plasmid but some sites required a specific direction. And different insertion sites led to different properties of the shuttle plasmids. The best shuttle plasmids pUC980-1 and pUC980-2, which showed copies more than 450 per cell and segregational stabilities up to 98%, were selected for heterologous expressions of an alkaline pectate lyase gene pelN , an alkaline protease spro1 and a pullulanase gene pulA11 , respectively. The highest extracellular activities of PelN, Spro1 and PulA11 were upto 5200 U/mL, 21537 U/mL and 504 U/mL correspondingly after 54 h, 60 h and 48 h fermentation in a 10 L fermentor. Notably, PelN and Spro1 showed remarkably higher yields in Bacillus than previous reports.Conclusion: In this work we can conclude that the optimum ori insertion site was the upstream region of BA3-1 in pWB980 which resulted in shuttle plasmids with higher copy numbers and higher stabilities. The novel shuttle plasmids pUC980-1 and pUC980-2 will be promising expression vectors in B. subtilis . And the ori insertion mechanism revealed in this work could provide theoretical guidance for further studies of pWB980 and constructions of other shuttle plasmids.

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