scholarly journals Structural Analysis of Phage-Borne stx Genes and Their Flanking Sequences in Shiga Toxin-Producing Escherichia coli and Shigella dysenteriae Type 1 Strains

2000 ◽  
Vol 68 (9) ◽  
pp. 4856-4864 ◽  
Author(s):  
Alexandra Unkmeir ◽  
Herbert Schmidt
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Shigella Dysenteriae ◽  
Nucleotide Sequencing ◽  
Restriction Patterns ◽  
E Coli ◽  
Pcr Products ◽  
Flanking Region ◽  
Flanking Regions

ABSTRACT The stx-flanking regions of 49 Shiga toxin-producingEscherichia coli strains and nine Shigella dysenteriae serotype 1 strains containing either stx,stx 1, stx 2, orstx 2 variant genes, were examined. We analyzed these regions by PCR using a set of primers with one primer specific for the respective stx gene and a second primer complementary to sequences of Stx phages H-19B and 933W. We further characterized the amplification products by restriction endonuclease digestion and nucleotide sequencing. PCR products ofstx 1-containing E. coli strains of serogroups O157, O26, and 0103 showed the same lengths and similar restriction patterns. However, we failed to amplify the 3′stx-flanking region instx 1-harboring E. coliO111:H− strains. Stx2-producing E. colistrains revealed amplification products of different lengths and restriction patterns, suggesting greater heterogeneity than instx 1-positive strains. We also obtained specific PCR products for two Stx2c-producing and seven Stx2f-producingE. coli strains when they were subjected to PCR analysis. In nine S. dysenteriae type 1 strains, H-19B- and 933W-specific primers amplified only the 3′ stx-flanking region. The results of our study demonstrate that the stxgenes of all strains investigated are continuous with phage sequences. Whereas almost all strains except E. coliO111:H− strains were associated with a S-like gene, association with Q could not be demonstrated in nine S. dysenteriae type 1 strains and three E. coli strains. Furthermore, we showed that the organization of thestx-flanking regions is similar in all strains investigated, whereas fine-structure analysis showed subtle differences among the sequences examined. Our results support the hypothesis thatstx genes in E. coli and S. dysenteriae are generally phage-borne.

scholarly journals Characterization of a Shiga Toxin-Encoding Temperate Bacteriophage of Shigella sonnei

2001 ◽  
Vol 69 (12) ◽  
pp. 7588-7595 ◽  
Author(s):  
Eckhard Strauch ◽  
Rudi Lurz ◽  
Lothar Beutin
Keyword(s):  
Shiga Toxin ◽  
Regulatory Region ◽  
Shigella Sonnei ◽  
Shigella Dysenteriae ◽  
Nucleotide Sequencing ◽  
Temperate Bacteriophage ◽  
Cross Hybridization ◽  
E Coli ◽  
K 12

ABSTRACT A Shiga toxin (Stx)-encoding temperate bacteriophage ofShigella sonnei strain CB7888 was investigated for its morphology, DNA similarity, host range, and lysogenization inShigella and Escherichia coli strains. Phage 7888 formed plaques on a broad spectrum of Shigella strains belonging to different species and serotypes, including Stx-producingShigella dysenteriae type 1. With E. coli, only strains with rough lipopolysaccharide were sensitive to this phage. The phage integrated into the genome of nontoxigenic S. sonneiand laboratory E. coli K-12 strains, which became Stx positive upon lysogenization. Moreover, phage 7888 is capable of transducing chromosomal genes in E. coli K-12. The relationships of phage 7888 with the E. coli Stx1-producing phage H-19B and the E. coli Stx2-producing phage 933W were investigated by DNA cross-hybridization of phage genomes and by nucleotide sequencing of an 8,053-bp DNA region of the phage 7888 genome flanking the stx genes. By these methods, a high similarity was found between phages 7888 and 933W. Much less similarity was found between phages H-19B and 7888. As in the other Stx phages, a regulatory region involved in Q-dependent expression is found upstream of stxA and stxB (stx gene) in phage 7888. The morphology of phage 7888 was similar to that of phage 933W, which shows a hexagonal head and a short tail. Our findings demonstrate that stx genes are naturally transferable and are expressed in strains of S. sonnei, which points to the continuous evolution of human-pathogenic Shigella by horizontal gene transfer.

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.

Haemolytic uraemic syndrome associated with Pseudomonas aeruginosa sepsis

2013 ◽  
Vol 62 (11) ◽  
pp. 1760-1762 ◽  
Author(s):  
Parameswaran Narayanan ◽  
Rashi S. Rustagi ◽  
Prabha Sivaprakasam ◽  
Mahadevan Subramanian ◽  
Sreejith Parameswaran ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Renal Failure ◽  
Influenza Virus ◽  
Shiga Toxin ◽  
Haemolytic Uraemic Syndrome ◽  
Shigella Dysenteriae ◽  
Micro Organisms

Haemolytic uraemic syndrome (HUS) is a recognized complication of infection with Shiga toxin-producing Escherichia coli (STEC) and Shigella dysenteriae type 1. Infections with other micro-organisms, especially Streptococcus pneumoniae, have been cited as causes of HUS. In addition, influenza virus and other viruses may rarely be associated with this syndrome. A 2-year-old girl presented with severe Pseudomonas aeruginosa sepsis with renal failure and ecthyma gangrenosum. Further investigations revealed features of HUS. She was managed with antibiotics and other supportive measures including peritoneal dialysis, and subsequently made a full recovery. A possible role of neuraminidase in the pathogenesis of P. aeruginosa-associated HUS was proposed. This is the first reported case of P. aeruginosa sepsis leading to HUS.

scholarly journals Promiscuous Origin of a Chimeric Sequence in the Escherichia coli O157:H7 Genome

1999 ◽  
Vol 181 (24) ◽  
pp. 7614-7617 ◽  
Author(s):  
J. Eugene LeClerc ◽  
Baoguang Li ◽  
William L. Payne ◽  
Thomas A. Cebula
Keyword(s):  
Escherichia Coli ◽  
Insertion Sequence ◽  
Chromosomal Region ◽  
Shigella Dysenteriae ◽  
Escherichia Coli O157 ◽  
Sequence Element ◽  
E Coli ◽  
K 12 ◽  
Insertion Sequence Element

ABSTRACT A novel sequence of 2.9 kb in the intergenic region between the mutS and rpoS genes of Escherichia coli O157:H7 and closely related strains replaces a sequence of 6.1 kb in E. coli K-12 strains. At the same locus in Shigella dysenteriae type 1, a sequence identical to that in O157:H7 is bounded by the IS1 insertion sequence element. Extensive polymorphism in the mutS-rpoSchromosomal region is indicative of horizontal transfer events.

scholarly journals Lineage and Host Source Are Both Correlated with Levels of Shiga Toxin 2 Production by Escherichia coli O157:H7 Strains

2009 ◽  
Vol 76 (2) ◽  
pp. 474-482 ◽  
Author(s):  
Yongxiang Zhang ◽  
Chad Laing ◽  
Zhengzhong Zhang ◽  
Jennyka Hallewell ◽  
Chunping You ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Enzyme Linked Immunosorbent Assay ◽  
Escherichia Coli O157 ◽  
Rt Pcr ◽  
Genetic Lineages ◽  
E Coli ◽  
Shiga Toxin 2 ◽  
Flanking Region ◽  
Q Gene

ABSTRACT Escherichia coli O157:H7 strains fall into three major genetic lineages that differ in their distribution among humans and cattle. Several recent studies have reported differences in the expression of virulence factors between E. coli O157:H7 strains from these two host species. In this study, we wished to determine if important virulence-associated “mobile genetic elements” such as Shiga toxin 2 (Stx2)-encoding prophage are lineage restricted or are host source related and acquired independently of the pathogen genotype. DNA sequencing of the stx 2 flanking region from a lineage II (LII) strain, EC970520, revealed that the transcriptional activator gene Q in LI strain EDL933 (upstream of stx 2) is replaced by a pphA (serine/threonine phosphatase) homologue and an altered Q gene in this and all other LII strains tested. In addition, nearly all LI strains carried stx 2, whereas all LII strains carried variant stx 2c and 4 of 14 LI/II strains had copies of both stx 2 and variant stx 2c. Real-time PCR (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) demonstrated that LI and LI/II strains produce significantly more stx 2 mRNA and Stx2 than LII strains. However, among LI strains significantly more Stx2 is also produced by strains from humans than from cattle. Therefore, lineage-associated differences among E. coli O157:H7 strains such as prophage content, toxin type, and toxin expression may contribute to host isolation bias. However, the level of Stx2 production alone may also play an important role in the within-lineage association of E. coli O157:H7 strains with human clinical disease.

scholarly journals Detection of Hemolysin Variants of Shiga Toxin-Producing Escherichia coli by PCR and Culture on VancomycinCefixime-Cefsulodin Blood Agar

1998 ◽  
Vol 64 (7) ◽  
pp. 2449-2453 ◽  
Author(s):  
Anselm Lehmacher ◽  
Heidi Meier ◽  
Stojanka Aleksic ◽  
Jochen Bockemühl
Keyword(s):  
Escherichia Coli ◽  
Nucleotide Sequence ◽  
Shiga Toxin ◽  
Raw Milk ◽  
Food Samples ◽  
Blood Agar ◽  
Simple Method ◽  
E Coli ◽  
Alpha Hemolysin ◽  
Pcr Products

ABSTRACT The presence of a hemolysin-encoding gene, elyA orhlyA, from Shiga toxin-producing Escherichia coli (STEC) was detected by PCR in each of 95 strains tested. PCR products of elyA from human STEC isolates of serovars frequently detected in Germany, such as O157:H−, O103:H2, O103:H−, O26:H11, and O26:H−, showed nucleotide sequences identical to previously reported ones for O157:H7 and O111:H− strains. Compared to them, four elyA amplicons derived from human isolates of rare STEC serovars showed identity of about 98% but lacked anAluI restriction site. However, the nucleotide sequence of an amplicon derived from a porcine O138:K81:H− STEC strain was identical to the corresponding region of hlyA, encoding alpha-hemolysin, from E. coli. This hlyAamplicon showed 68% identity with the nucleotide sequence of the corresponding elyA fragment. It differed from theelyA PCR product in restriction fragments generated byAluI, EcoRI, and MluI. Of the 95 representative STEC strains, 88 produced hemolysin on blood agar supplemented with vancomycin (30 mg/liter), cefixime (20 μg/liter), and cefsulodin (3 mg/liter) (BVCC). The lowest added numbers of two to six STEC CFU per g of stool or per ml of raw milk were detectable on BVCC plates after seeding of the preenrichment broth, modified tryptic soy broth (mTSB) supplemented with novobiocin (10 mg/liter), with 16 STEC strains. These strains represented the seven prevailing serovars diagnosed from German patients. However, with ground-beef samples, PCR was essential to identify the lowest added numbers of two to six STEC CFU among colonies of hemolyzing Enterobacteriaceae, such as Serratia spp. and alpha-hemolysin-producing E. coli. We conclude that preenrichment of stool and food samples in mTSB for 6 h followed by overnight culturing on BVCC is a simple method for the isolation and presumptive identification of STEC.

Polyester Cloth–Based Hybridization Array System for Identification of Enterohemorrhagic Escherichia coli Serogroups O26, O45, O103, O111, O121, O145, and O157

2012 ◽  
Vol 75 (9) ◽  
pp. 1691-1697 ◽  
Author(s):  
BURTON W. BLAIS ◽  
MARTINE GAUTHIER ◽  
MYLÈNE DESCHÊNES ◽  
GEORGE HUSZCZYNSKI
Keyword(s):  
Escherichia Coli ◽  
Marker Gene ◽  
Enterohemorrhagic Escherichia Coli ◽  
Oligonucleotide Probes ◽  
E Coli ◽  
Polyester Cloth ◽  
Gene Profiles ◽  
Pcr Products ◽  
Different Strains ◽  
Immunoenzymatic Assay

A cloth-based hybridization array system (CHAS) was developed for the identification of foodborne colony isolates of seven priority enterohemorrhagic Escherichia coli (EHEC-7) serogroups targeted by U.S. food inspection programs. Gene sequences associated with intimin; Shiga-like toxins 1 and 2; and the antigenic markers O26, O45, O103, O111, O121, O145, and O157 were amplified in a multiplex PCR incorporating a digoxigenin label, and detected by hybridization of the PCR products with an array of specific oligonucleotide probes immobilized on a polyester cloth support, with subsequent immunoenzymatic assay of the captured amplicons. The EHEC-7 CHAS exhibited 100% inclusivity and 100% exclusivity characteristics with respect to detection of the various markers among 89 different E. coli strains, with various marker gene profiles and 15 different strains of non–E. coli bacteria.

Comparison of colony and stool blots for detection of enteropathogens by DNA probes

1991 ◽  
Vol 37 (5) ◽  
pp. 407-410
Author(s):  
Mônica A. M. Vieira ◽  
Beatriz E. C. Guth ◽  
Tânia A. T. Gomes
Keyword(s):  
Escherichia Coli ◽  
Key Words ◽  
Sensitivity And Specificity ◽  
Dna Probes ◽  
Type I ◽  
Enteropathogenic Escherichia Coli ◽  
Key Words Dna ◽  
E Coli ◽  
Heat Stable

DNA probes that identify genes coding for heat-labile type I (LT-I) and heat-stable type 1 (ST-I) enterotoxins, enteropathogenic Escherichia coli adherence factor (EAF), and Shigella-like, invasiveness (INV) are used to evaluate the sensitivity and specificity of stool blots in comparison with the sensitivity and specificity of colony blots in detecting enteropathoghens. The sensitivities of the probes in stool blots are 91.7% for the LT-I probe, 76.9% for the ST-I probes, 78.9% for the EAF probe, and 45.5% for the INV probe. The specificity of all probes is higher than 95%. In general, the stool blot method identifies as many if not more LT-I-, ST-I-, and EAF-producing E. coli infections than the colony blots. Key words: DNA probes, stool blots, enteropathogens, diagnosis.

Sign in / Sign up

Export Citation Format

Share Document