Shiga Toxin-producing Escherichia Coli Serotypes Research Articles

Molecular mechanisms that mediate colonization of Shiga toxin-producing Escherichia coli strains.

Shiga toxin-producing Escherichia coli (STEC) is a group of pathogens which cause gastrointestinal disease in humans and have been associated with numerous food-borne outbreaks worldwide. The intimin adhesin has been considered for many years to be the only colonization factor in these strains. However, the rapid progress in whole-genome sequencing of different STEC serotypes has accelerated the discovery of other adhesins (fimbrial and afimbrial), which have emerged as important contributors to the intestinal colonization occurring during STEC infection. This review summarizes recent progress to identify and characterize, at the molecular level, novel adhesion and colonization factors in STEC strains, with an emphasis on their contribution to virulence traits, their host-pathogen interactions, the regulatory mechanisms controlling their expression, and their role as targets eliciting immune responses in the host.

Secondary transmissions during the outbreak of Shiga toxin-producing Escherichia coli O104 in Hesse, Germany, 2011

During the recent outbreak of Shiga toxin-producing Escherichia coli (STEC) O104:H4 in Germany most cases notified in the State of Hesse (6 million inhabitants) were linked to satellite clusters or had travelled to the outbreak area in northern Germany. Intensified surveillance was introduced to rapidly identify cases not linked to known clusters or cases and thus to obtain timely information on possible further contaminated vehicles distributed in Hesse, as well to describe the risk of secondary transmission among known cases. As of 2 August 2011* [corrected], 56 cases of haemolytic uraemic syndrome (HUS) including two fatal cases, and 124 cases of STEC gastroenteritis meeting the national case definitions have been reported in Hesse. Among the 55 HUS and 81 STEC gastroenteritis cases thatmet the outbreak case definition, one HUS case and eight STEC gastroenteritis cases may have acquired their infection through secondary transmission. They include six possible transmissions within the family, two possible nosocomial and one possible laboratory transmission. Our results do not suggest an increased transmissibility of the outbreak strain compared to what is already known about E. coli O157 and other STEC serotypes.

Bacteriophage-Encoding Cytolethal Distending Toxin Type V Gene Induced from Nonclinical Escherichia coli Isolates

Cytolethal distending toxin (Cdt) is produced by a variety of pathogenic bacteria, including pathogenic serotypes of Shiga toxin-producing Escherichia coli (STEC). The Cdt family comprises five variants (Cdt-I to Cdt-V) encoded by three genes located within the chromosome or plasmids or, in the case of Cdt-I, within bacteriophages. In this study, we evaluated the occurrence of the cdt gene in a collection of 140 environmental STEC isolates. cdt was detected in 12.1% of strains, of which five strains carried inducible bacteriophages containing the Cdt-V variant. Two Cdt-V phages of the Siphoviridae morphology lysogenized Shigella sonnei, generating two lysogens: a single Cdt phage lysogen and a double lysogen, containing a Cdt phage and an Stx phage, both from the wild-type strain. The rates of induction of Cdt phages were evaluated by quantitative PCR, and spontaneous induction of Cdt-V phage was observed, whereas induction of Stx phage in the double lysogen was mitomycin C dependent. The Cdt distending effect was observed in HeLa cells inoculated with the supernatant of the Cdt-V phage lysogen. A ClaI fragment containing the cdt-V gene of one phage was cloned, and sequencing confirmed the presence of Cdt-V, as well as a fragment downstream from the cdt homolog to gpA, encoding a replication protein of bacteriophage P2. Evaluation of Cdt-V phages in nonclinical water samples showed densities of 10(2) to 10(9) gene copies in 100 ml, suggesting the high prevalence of Cdt phages in nonclinical environments.

Update on the ongoing outbreak of haemolytic uraemic syndrome due to Shiga toxin-producing Escherichia coli (STEC) serotype O104, Germany, May 2011

Since early May 2011, a large outbreak of haemolytic uraemic syndrome (HUS) and bloody diarrhoea related to infections with Shiga toxin-producing Escherichia coli (STEC) has been observed in Germany. The outbreak is focused in the north, but cases have been reported from all German states and other countries. Since our report last week, the number of HUS cases has increased to 470 and STEC serotype O104 has been confirmed in many cases.

Serological response of Shiga toxin-producing Escherichia coli type III secreted proteins in sera from vaccinated rabbits, naturally infected cattle, and humans.

Escherichia coli O157:H7 is an important zoonotic pathogen, causing hemolytic uremic syndrome (HUS). The colonization of cattle and human hosts is mediated through the action of effectors secreted via a type III secretion system (T3SS). The structural genes for the T3SS and many of the secreted effectors are located on a pathogenicity island called the locus of enterocyte effacement (LEE). We cloned and expressed the genes coding for 66 effectors and purified each to measure the cross-reactivity of type III secreted proteins from Shiga toxin-producing Escherichia coli (STEC) serotypes. These included 37 LEE-encoded proteins and 29 non-LEE effectors. The serological response against each protein was measured by Western blot analysis and enzyme-linked immunosorbent assay (ELISA) using sera from rabbits immunized with type III secreted proteins (T3SPs) from four STEC serotypes, experimentally infected cattle, and human sera from six HUS patients. Twenty proteins were recognized by at least one of the STEC T3SP-vaccinated rabbits by Western blotting. Several structural proteins (EspA, EspB, and EspD) and a number of effectors (Tir, NleA, and TccP) were recognized by O26-, O103-, O111-, and O157-specific sera. Sera from experimentally infected cattle and HUS patients were tested using an ELISA against each of the proteins. Tir, EspB, EspD, EspA, and NleA were recognized by the majority of the samples tested. A number of other proteins also were recognized by individual serum samples. Overall, proteins such as Tir, EspB, EspD, NleA, and EspA were highly immunogenic in vaccinated and naturally infected subjects and could be candidates for a cross-protective STEC vaccine.

The Evolution of Virulence in Non-O157 Shiga Toxin-Producing Escherichia Coli

Shiga toxin-producing Escherichia coli (STEC) are zoonotic foodborne and waterborne pathogens that are a serious public health concern because they cause outbreaks and the potentially fatal hemolytic uremic syndrome (HUS). The most common STEC serotype associated with human disease is O157:H7, but there is a growing recognition of over 100 non-O157 serotypes that also may result in human illness. Some of these non-O157 STEC strains cause outbreaks and severe disease such as HUS and hemorrhagic colitis, whereas others are associated with only mild diarrhea or with no human disease at all. The relative scarceness of whole genome sequence data for non-O157 STEC has limited the scientific discovery into the genetic basis of these differences in virulence. Uncovering the scope of genetic diversity and phylogeny of the non-O157 STEC through targeting sequencing of clinically relevant isolates will offer new biological insight to the pathogenic behavior of these emerging pathogens. These approaches would also enable molecular risk assessment strategies to rapidly identify and respond to emerging non-O157 STEC that pose a serious public health risk to humans.

Characterization of Shiga toxin-producing Escherichia coli isolated from dairy cows in Argentina

To feno-genotypically characterize the Shiga toxin-producing Escherichia coli (STEC) population in Argentinean dairy cows. From 540 STEC positive samples, 170 isolates were analyzed by multiplex PCR and serotyping. Of these, 11% carried stx1, 52% stx2 and 37% stx1/stx2. The ehxA, saa and eae were detected in 77%, 66% and 3%, respectively. Thirty-five per cent of strains harboured the profile stx1, stx2, saa, ehxA and 29% stx2, saa, ehxA. One hundred and fifty-six strains were associated with 29 different O serogroups, and 19 H antigens were distributed among 157 strains. STEC O113:H21, O130:H11 and O178:H19 were the most frequently found serotypes. The STEC O157:H7 were detected in low rate and corresponded to the stx2(+) , eae(+) , ehxA(+) virulence pattern. We detected a diversity of STEC strains in dairy cattle from Argentina, most of them carrying genes linked to human disease. The non-O157 STEC serotypes described in this study are associated worldwide with disease in humans and represent a risk for the public health. For this, any microbiological control in dairy farms should be targeted not only to the search of O157:H7 serotype.

Family Outbreak of Shiga Toxin–producingEscherichia coliO123:H–, France, 2009

To the Editor: Shiga toxin–producing Escherichia coli (STEC) is a major cause of foodborne disease in industrialized countries. We present results of the investigation of a family outbreak in France caused by a rare STEC serotype. Surveillance of STEC infections in France since 1996 has been based on national surveillance of STEC-related pediatric hemolytic uremic syndrome (HUS) (1). On February 11, 2009, two cases of diarrhea were reported to a surveillance coordinator: 1 in a child with HUS and the other in that child’s sibling. The 2 siblings, 2 and 6 years of age, had diarrhea beginning on February 4 and 5, 2009. Bloody diarrhea developed in the younger child, and HUS was diagnosed on February 9. The older child had nonbloody diarrhea for 3 days and abdominal pain. Questioning of the patients’ parents identified no recent history of travel, contact with farm animals, or outdoor bathing. A food history indicated that the 2 patients had shared an undercooked ground beef burger 4–5 days before symptom onset. The patients’ parents also ate burgers from the same package (box); they did not report any gastrointestinal symptoms. Fecal specimens of the patients were tested for STEC by direct PCR for STEC genes (stx); after which culture and identification of stx1, stx2, eae, and ehxA (hlyA) virulence genes; and serotyping with a panel of 22 serum samples were conducted as described (1,2). Molecular serotyping was subsequently conducted on nonagglutinating strains by using the rfb–restriction fragment length polymorphism technique for O antigen (3) and sequencing of the fliC gene for H antigen (4). A trace-back investigation was conducted for the implicated beef burgers, which were obtained from a box of 10, frozen, 100-g ground beef burgers purchased in late January 2009. The remaining beef burger in the box from which the patients had eaten a beef burger was obtained from the family’s freezer for microbiologic testing. Stored production samples from the implicated batch underwent microbiologic testing. After broth enrichment, ground beef samples were tested by PCR for stx and eae virulence genes and O antigens of serotypes O157, O26, O145, O103, and O111 (2,5,6). Subsequently, strains isolated from stx-positive and eae-positive enrichment broths were biochemically tested and underwent serotyping and PCR identification of virulence genes. Genetic relatedness of clinical and ground beef STEC strains was studied by using pulsed-field gel electrophoresis with Xbal as described (7). A nonmotile strain of STEC stx2 eae ehxA, which was not serotypeable by the panel of 22 serum samples, was identified in fecal samples from patients and in the remaining ground beef. Molecular serotyping of clinical isolates and an isolate from the beef identified a strain of STEC O123:H2. Analysis by pulsed-field gel electrophoresis indicated that the clinical and meat isolates were genetically related (Figure). The level of STEC contamination in the meat was 30–40 CFU/g. All stored meat production samples tested were negative for STEC. Figure Representative XbaI pulsed-field gel electrophoresis patterns of Shiga toxin–producing Escherichia coli (STEC) O123:H– strains isolated from patient fecal samples and strains isolated from ground beef obtained from patients’ home, ... A clinical strain and a ground beef STEC strain were sent to the World Health Organization Collaborating Centre for Reference and Research on Escherichia and Klebsiella in Copenhagen, Denmark, in December 2009 for analysis. The clinical strain was confirmed as STEC O123:H–, and the meat strain was confirmed as a nonmotile STEC rough type by serum agglutination. Both strains had virulence genes stx2a, eae, and ehxA (F. Scheutz, pers. comm.). We identified a family outbreak of STEC O123:H– stx2a, eae ehxA infections associated with ingestion of undercooked ground beef. No similar cases of STEC infection were identified by active case finding. This serotype is rarely described as a cause of human clinical infection. No human isolate of serotype O123:H– is recorded in the database of the World Health Organization Collaborating Centre for Reference and Research on Escherichia and Klebsiella (F. Scheutz, pers. comm.). Two strains of STEC O123:H– stx2d were isolated from asymptomatic persons in Germany during 1996–2000 (8). A study in Australia in 2003 reported using a strain of O123:H– stx1 stx2 ehxA from Switzerland that had been isolated from a person with diarrhea (9). We report foodborne transmission of STEC O123:H– that resulted in a cluster of clinical cases of infection. Eating ground beef is a well-established mode of STEC transmission, particularly for serotype O157:H7. STEC serotype O123:H– has been isolated from feces of healthy lambs and sheep in Spain (10) and in southwestern Australia (9) and is considered to be among the predominant ovine STEC serotypes in these countries. This family outbreak shows that STEC serotype O123:H–, albeit rarely described as causing human illness, can cause severe human infection. This serotype can also cause clusters of STEC infections and be transmitted by ingestion of undercooked ground beef.

Non-O157 Shiga toxin-producing Escherichia coli in foods.

Non-O157 Shiga toxin-producing Escherichia coli (STEC) strains have been linked to outbreaks and sporadic cases of illness worldwide. Illnesses linked to STEC serotypes other than O157:H7 appear to be on the rise in the United States and worldwide, indicating that some of these organisms may be emerging pathogens. As more laboratories are testing for these organisms in clinical samples, more cases are uncovered. Some cases of non-O157 STEC illness appear to be as severe as cases associated with O157, although in general cases attributed to non-O157 are less severe. There is much variation in virulence potential within STEC serotypes, and many may not be pathogenic. Of more than 400 serotypes isolated, fewer than 10 serotypes cause the majority of STEC-related human illnesses. Various virulence factors are involved in non-O157 STEC pathogenicity; the combined presence of both eae and stx genes has been associated with enhanced virulence. A scientific definition of a pathogenic STEC has not yet been accepted. Several laboratories have attempted to develop detection and identification methods, and although substantial progress has been made, a practical method of STEC detection has yet to be validated. Worldwide, foods associated with non-O157 STEC illness include sausage, ice cream, milk, and lettuce, among others. Results from several studies suggest that control measures for O157 may be effective for non-O157 STEC. More research is needed to uncover unique characteristics and resistances of non-O157 STEC strains if they exist. The public health significance of non-O157 STEC and the implications for industry practices and regulatory actions are discussed.

Detection, isolation and characterization of Shiga toxin-producing Escherichia coli (STEC) in fresh ground beef from butcher shops in Concepción, Tucumán Province

Shiga toxin-producing Escherichia coli is an emerging foodborne pathogen. There are many STEC serotypes associated with human diseases, being the O157:H7 serotype the most prevalent. Ground beef is the main transmission vehicle. In Concepción city, Tucumán Province, between September and December 2004, two hemolytic uremic syndrome (HUS) cases were diagnosed. The main objective of this work was to detect, isolate and characterize STEC O157 and non-O157 strains in fresh ground beef. Between September and December 2004, 53 fresh ground beef samples were collected from butcher shops in Concepción city. The USDA-FSIS (2002) methodology was used for detection, isolation and characterization of STEC O157:H7. Two PCR techniques for E. coli non-O157 detection and a previous intra-laboratory validated methodology for the isolation and characterization of these strains were used. The stx2 gen was identified in seven samples and the rfbO157 gene also in four of them. However, only one E. coli O157:H7 strain, biotype C, carrying the eae, stx2 and ehxA genes, was isolated. The present study shows the importance of implementing techniques for the detection of this emerging pathogen in meat samples.

Characteristics of O157 versus Non‐O157 Shiga Toxin–ProducingEscherichia coliInfections in Minnesota, 2000–2006

Escherichia coli O157:H7 (O157) is the Shiga toxin-producing E. coli (STEC) serotype most frequently isolated and most often associated with hemolytic uremic syndrome (HUS) in the United States. Non-O157 STEC serotypes can also cause serious illness, but their impact as pathogens remains undefined. We compared characteristics of non-O157 and O157 STEC infections identified through sentinel surveillance. Sentinel sites included a metropolitan health maintenance organization laboratory and a hospital laboratory serving a small city and rural area. We received sorbitol-MacConkey agar plates from every stool culture performed at both sites during 2000-2006. Colony sweeps were screened for stx1 and stx2 by polymerase chain reaction. E. coli identity, serotype, and presence of stx1 and/or stx2 were confirmed on individual isolates. Two hundred six STEC isolates were identified: 108 (52%) were non-O157 serotypes, and 98 (48%) were O157. Of non-O157 cases, 54% involved bloody diarrhea, and 8% involved hospitalization. Non-O157 isolates with at least stx2 were not more likely to cause severe illness (bloody diarrhea, hospitalization, or HUS) than were non-O157 isolates with only stx1. O157 cases were more likely than non-O157 cases to involve bloody diarrhea (78% vs 54%; P < .001), hospitalization (34% vs 8%; P < .001 and HUS (7% vs 0%; P = .005). When including only isolates with at least stx2, O157 cases were still more likely to involve bloody diarrhea (78% vs 56%; P = .02) and hospitalization (33% vs 12%; P = .01) than non-O157 cases. Differences in severity among STEC infections could not be explained by stx2, suggesting that additional factors are important in STEC virulence.

Isolation and detection of Shiga toxin-producing Escherichia coli in clinical stool samples using conventional and molecular methods

The isolation of Shiga toxin-producing Escherichia coli (STEC) other than serogroup O157 from clinical stool samples is problematic due to the lack of differential phenotypic characteristics from non-pathogenic E. coli. The development of molecular reagents capable of identifying both toxin and serogroup-specific genetic determinants holds promise for a more comprehensive characterization of stool samples and isolation of STEC strains. In this study, 876 stool samples from paediatric patients with gastroenteritis were screened for STEC using a cytotoxicity assay, commercial immunoassay and a conventional PCR targeting Shiga-toxin determinants. In addition, routine culture methods for isolating O157 STEC were also performed. The screening assays identified 45 stools presumptively containing STEC, and using non-differential culture techniques a total of 20 O157 and 22 non-O157 strains were isolated. These included STEC serotypes O157 : H7, O26 : H11, O121 : H19, O26 : NM, O103 : H2, O111 : NM, O115 : H18, O121 : NM, O145 : NM, O177 : NM and O5 : NM. Notably, multiple STEC serotypes were isolated from two clinical stool samples (yielding O157 : H7 and O26 : H11, or O157 : H7 and O103 : H2 isolates). These data were compared to molecular serogroup profiles determined directly from the stool enrichment cultures using a LUX real-time PCR assay targeting the O157 fimbrial gene lpfA, a microsphere suspension array targeting allelic variants of espZ and a gnd-based molecular O-antigen serogrouping method. The genetic profile of individual stool cultures indicated that the espZ microsphere array and lpfA real-time PCR assay could accurately predict the presence and provide preliminary typing for the STEC strains present in clinical samples. The gnd-based molecular serogrouping method provided additional corroborative evidence of serogroup identities. This toolbox of molecular methods provided robust detection capabilities for STEC in clinical stool samples, including co-infection of multiple serogroups.

Shiga toxin-producingEscherichia coliand atypical enteropathogenicEscherichia colistrains isolated from healthy sheep of different populations in São Paulo, Brazil

Sheep are important carriers of Shiga toxin-producing Escherichia coli (STEC) in several countries. However, there are a few reports about ovine STEC in American continent. About 86 E. coli strains previously isolated from 172 healthy sheep from different farms were studied. PCR was used for detection of stx(1), stx(2), eae, ehxA and saa genes and for the identification of intimin subtypes. Restriction fragment length polymorphism (RFLP)-PCR was performed to investigate the variants of stx(1) and stx(2), and the flagellar antigen (fliC) genes in nonmotile isolates. Five isolates were eae(+) and stx(-), and belonged to serotypes O128:H2/beta-intimin (2), O145:H2/gamma, O153:H7/beta and O178:H7/epsilon. Eighty-one STEC isolates were recovered, and the stx genotypes identified were stx(1c)stx(2d-O118) (46.9%), stx(1c) (27.2%), stx(2d-O118) (23.4%), and stx(1c)stx(2dOX3a) (2.5%). Pulsed-field gel electrophoresis (PFGE) revealed 27 profiles among 53 STEC and atypical enteropathogenic Escherichia coli (EPEC) isolates. This study demonstrated that healthy sheep in São Paulo, Brazil, can be carriers of potential human pathogenic STEC and atypical EPEC. As some of the STEC serotypes presently found have been involved with haemolytic uraemic syndrome (HUS) in other countries, the important role of sheep as sources of STEC infection in our settings should not be disregarded.

Identification of Shiga Toxin-producing E. coli Isolated from Diarrhea Patients and Cattle in Gwangju Area, Korea

Shiga toxin-producing Escherichia coli (STEC) strains are commensal bacteria in cattle and cause food borne disease in human. We analyzed the isolation rate of STEC in stool specimens of patients with diarrhea and in fecal samples of cattle in Gwangju, Korea. STEC strains were detected from 33 (0.19%) out of 17,148 patients with diarrhea while there has been a progressive increase in the incidence rate from 0.07% in 2004 to 0.33% in 2008. We investigated serotypes, shiga toxin genes, and antimicrobial resistance patterns of the 44 STEC isolates from human and cattle sources. The 33 STEC isolates from human belonged to 14 O serotypes including O157, O26 and O111. The 11 isolates from cattle belonged to 11 O serotypes. PCR detection for stx genes showed that 12 (27.3%) isolates carried stx₁ genes, 20 (45.5%) possessed stx₂ genes, and 12 (27.3%) carried both stx₁ and stx₂. Of the 33 STEC isolates from human, 25 strains (76%) were resistant to one or more antibiotics. High level of resistance to tetracycline (73%) was most common, followed by ticarcillin and ampicillin (64%). But none of the 33 isolates from human were resistant to amikacin, cefazolin, cefepime, cefotetan, cefotaxime, ciprofloxacin, or imipenem. The 5 strains (45%) of the 11 isolates from cattle were resistant to at least one or three antibiotics but most of the isolates were sensitive to the 16 antibiotics employed in this survey. In conclusion, toxin types and serotypes of STEC isolated from human and cattle were diverse, and non-O157 STEC was also observed to be a greater proportion of STEC isolates. According to a specific comparison solely based on the toxin types and serotypes, most of the STEC strains isolated from cattle feces in Gwangju, Korea showed characteristics different from those isolated from patients. Therefore, laboratory surveillance is required to detect and carefully monitor the potentially hypervirulent STEC not only in human and cattle but also in other animals.

Quantitative isolation efficiency of O26, O103, O111, O145 and O157 STEC serotypes from artificially contaminated food and cattle faeces samples using a new isolation protocol

A range of new differential and confirmation plating media for some non-O157 Shiga toxin producing Escherichia coli (STEC) serotypes (O26, O103, O111, O145) and both sorbitol-positive and -negative O157 were evaluated using artificially contaminated samples. Dairy products (raw milk, cheese made from pasteurized milk and raw milk), meat (ground beef, fermented meat) and cattle faeces were artificially contaminated using clinical STEC strains. Isolation efficiency was 100%, 82.3%, 88.5%, 65.9%, 64.3% and 15.8%, respectively, for an inoculum size of </=100 CFU 25 g(-1). The consecutive use of differential and confirmation media limited the incidence of false positive isolates from 0% for raw milk samples, cheese made from pasteurized milk and for fermented meat to 2.1% for cheese made from raw milk, and to 8.9% for ground beef. Data presented in this paper indicated that the efficiency of the applied isolation method was dependent on sample-to-sample variation but not on the inoculum size. Data in this paper indicated that isolation of low levels of non-O157 and sorbitol-positive O157 STEC from food samples is possible.

EhaA is a novel autotransporter protein of enterohemorrhagic Escherichia coli O157:H7 that contributes to adhesion and biofilm formation

Autotransporter (AT) proteins have been identified in many Gram-negative pathogens and are unique in that their primary sequence is sufficient to direct their transport across the bacterial membrane system. Where characterized they are uniformly associated with virulence. Using conserved AT motifs as a search tool, four putative AT proteins were identified in the Enterohemorrhagic Escherichia coli O157:H7 EDL933 genome. The genes encoding these proteins (z0402/ehaA, z0469/ehaB, z3487/ehaC and z3948/ehaD) were PCR amplified, cloned and expressed in an E. coli K-12 MG1655flu background. Preliminary characterization revealed that ehaA, ehaB and ehaD encode proteins associated with increased biofilm formation. One of these genes (ehaA) resides on a genomic island in E. coli O157:H7 strains EDL933 and Sakai. Over-expression of EhaA in E. coli K-12 demonstrated it is located at the cell surface and resulted in the formation of large cell aggregates, promoted significant biofilm formation and mediated adhesion to primary epithelial cells of the bovine terminal rectum. The expression of ehaA was demonstrated in E. coli EDL933 by RT-PCR. An EhaA-specific antibody revealed the EhaA protein was expressed in 24/50 generic Shiga toxin-producing E. coli (STEC) strains of various serotypes including O157:H7. However, the deletion of ehaA from E. coli EDL933 and a STEC strain from serotype O111:H(-) did not affect biofilm growth. Our results suggest that EhaA may contribute to adhesion, colonization and biofilm formation by E. coli O157:H7 and possibly other STEC serotypes.

Isolation and Genetic Characterization of a Coinfection of Non-O157 Shiga Toxin-Producing Escherichia coli

A coinfection of O177:NM and O55:H7 Shiga toxin-producing Escherichia coli (STEC) was identified for a child with acute bloody diarrhea and hemolytic uremic syndrome by using culture and serotype-specific molecular reagents. The profile of O157-related genetic islands revealed that the O55:H7 isolate was highly similar to O157 STEC whereas the O177:NM isolate lacked several fimbrial O islands and non-locus-of-enterocyte-effacement effector determinants. However, both STEC serotypes are known to cause serious disease, and the significant repertoire of virulence determinants in both strains made it impossible to determine their individual contributions to the clinical symptoms.

Subtypes of the Plasmid-Encoded Serine Protease EspP in Shiga Toxin-Producing Escherichia coli : Distribution, Secretion, and Proteolytic Activity

We investigated the prevalence, distribution, and structure of espP in Shiga toxin-producing Escherichia coli (STEC) and assessed the secretion and proteolytic activity of the encoded autotransporter protein EspP (extracellular serine protease, plasmid encoded). espP was identified in 56 of 107 different STEC serotypes. Sequencing of a 3,747-bp region of the 3,900-bp espP gene distinguished four alleles (espPalpha, espPbeta, espPgamma, and espPdelta), with 99.9%, 99.2%, 95.3%, and 95.1% homology, respectively, to espP of E. coli O157:H7 strain EDL933. The espPbeta, espPgamma, and espPdelta genes contained unique insertions and/or clustered point mutations that enabled allele-specific PCRs; these demonstrated the presence of espPalpha, espPbeta, espPgamma, and espPdelta in STEC isolates belonging to 17, 16, 15, and 8 serotypes, respectively. Among four subtypes of EspP encoded by these alleles, EspPalpha (produced by enterohemorrhagic E. coli [EHEC] O157:H7 and the major non-O157 EHEC serotypes) and EspPgamma cleaved pepsin A, human coagulation factor V, and an oligopeptide alanine-alanine-proline-leucine-para-nitroaniline, whereas EspPbeta and EspPdelta either were not secreted or were proteolytically inactive. The lack of proteolysis correlated with point mutations near the active serine protease site. We conclude that espP is widely distributed among STEC strains and displays genetic heterogeneity, which can be used for subtyping and which affects EspP activity. The presence of proteolytically active EspP in EHEC serogroups O157, O26, O111, and O145, which are bona fide human pathogens, suggests that EspP might play a role as an EHEC virulence factor.

Microbiological Characterization of Lamb Carcasses at Commercial Processing Plants in the United States

Although the United States produces 203 million lb (ca. 92.1 kg) of domestic lamb and mutton each year, thorough studies of the microbiological safety during lamb processing are lacking. To address this missing information, a total of 2,548 sponge samples from pelts, preevisceration carcasses, and postintervention carcasses were collected from multiple large commercial lamb processing plants to determine aerobic plate counts, the prevalences of Escherichia coli O157:H7, non-O157 Shiga toxin–producing E. coli (STEC), and Salmonella. The averages of the aerobic plate counts from pelts, the preevisceration carcasses, and the postintervention carcasses were 6.3, 4.4, and 2.4 log CFU/100 cm2, respectively. The prevalences of E. coli O157:H7 from the pelts, the preevisceration carcasses, and the postintervention carcasses were 12.8, 1.6, and 2.9%, respectively. The average Salmonella prevalences were 14.4, 4.3, and 1.8% for pelts, preevisceration carcasses, and postintervention carcasses, respectively. The most frequently identified Salmonella serotype was Heidelberg. The prevalences of non-O157 STEC from pelts, preevisceration carcasses, and postintervention carcasses averaged 86.2, 78.6, and 81.6%, respectively. A total of 488 non-O157 STEC strains were isolated from postintervention carcasses. Sixty-nine different serotypes of non-O157 STEC were identified. The most frequently detected serotypes were O91:H14 (40.8%), followed by O5:H19 (18.4%). A small number of STEC serotypes associated with severe human illness were isolated from postintervention carcasses. These were serotypes O76:H19, O128:H2 (0.8%), O146:H8 (2.1%), O146:H21, O163:H19, and O174:H8 (1.3%). The results of this study establish a baseline for microbiological quality and prevalences of Salmonella, E. coli O157:H7, and STEC in U.S. lamb processing plants.

Metabolic and genetic profiling of clinical O157 and non-O157 Shiga-toxin-producing Escherichia coli

A collection of clinical Shiga-toxin-producing Escherichia coli (STEC) strains, mainly belonging to serotypes O26, O103, O111, O145 and O157, was characterised by a polyphasic approach including molecular serotyping, PCR-based detection of virulence factors ( stx1, stx2, eae, EHEC-hlyA, saa, katP, espP), carbohydrate fermentation profiles using API50 tests and random amplification of polymorphic DNA (RAPD) fingerprinting. An RAPD protocol based on the combination of 2 primers resulted in sufficiently complex patterns enabling discrimination to the serotype level. Moreover, carbohydrate fermentation profiles obtained after evaluating up to 50 different carbohydrates led to separation of different STEC serotypes. Virulence typing results confirm the association of Shiga toxins and intimin subtypes with specific serotypes and clinical diagnosis. Clinical diagnosis of strains did not correlate with either RAPD profiles or carbohydrate fermentation patterns.