Shiga Toxin-producing Escherichia Coli O157 Infections Research Articles

Linking active rectal mucosa-attached microbiota to host immunity reveals its role in host-pathogenic STEC O157 interactions.

The rectal anal junction (RAJ) is the major colonization site of Shiga toxin-producing Escherichia coli (STEC) O157 in beef cattle, leading to transmission of this foodborne pathogen from farms to food chains. To date, there is limited understanding on whether mucosa-attached microbiome has a profound impact on host-STEC interactions. In this study, the active RAJ mucosa-attached microbiota and its potential role in host immunity-STEC-commensal interactions were investigated using RAJ mucosal biopsies collected from calves orally challenged with two STEC O157 strains with or without functional stx2a (stx2a + or stx2a-). The results revealed that shifts of microbial diverstives, topological, and assembly patterns were subjected to stx2a production post-challenge and Paeniclostridium and Gallibacterium being the keystone taxa for both microbial interactions and assembly. Additional mucosal transcriptome profiling showed stx2a-dependent host immune responses (i.e. B and T cell signaling, antigen processing and presentation) post-challenge. Further integrated analysis revealed that mucosa-attached beneficial microbes (i.e. Provotella, Faecalibacterium, and Dorea) interacted with host immune genes pre-challenge to maintain host homeostasis, however, opportunistic pathogenic microbes (i.e. Paeniclostridium) could interact with host immune genes after the STEC O157 colonization and interactions were stx2a-dependent. Furthermore, bacterial predicted functions involved in pathogen (O157 and Paeniclostridium) colonization and metabolism were related to host immunities. These findings suggest that host-microbe interactions could shift from beneficial to opportunistic pathogenic bacteria-driven during the pathogen colonization and be dependent on the production of particular virulence factors, highlighting the potential regulatory role of mucosa-attached microbiota in affecting pathogen-commensal-host interactions under STEC O157 infection in calves.

Linking active rectal mucosa-attached microbiota to host immunity reveals its role in host-pathogenic STEC O157 interactions.

The rectal anal junction (RAJ) is the major colonization site of Shiga toxin-producing Escherichia coli (STEC) O157 in beef cattle, leading to transmission of this foodborne pathogen from farms to food chains. To date, there is limited understanding on whether mucosa-attached microbiome has a profound impact on host-STEC interactions. In this study, the active RAJ mucosa-attached microbiota and its potential role in host immunity-STEC-commensal interactions were investigated using RAJ mucosal biopsies collected from calves orally challenged with two STEC O157 strains with or without functional stx2a (stx2a + or stx2a-). The results revealed that shifts of microbial diverstives, topological, and assembly patterns were subjected to stx2a production post-challenge and Paeniclostridium and Gallibacterium being the keystone taxa for both microbial interactions and assembly. Additional mucosal transcriptome profiling showed stx2a-dependent host immune responses (i.e. B and T cell signaling, antigen processing and presentation) post-challenge. Further integrated analysis revealed that mucosa-attached beneficial microbes (i.e. Provotella, Faecalibacterium, and Dorea) interacted with host immune genes pre-challenge to maintain host homeostasis, however, opportunistic pathogenic microbes (i.e. Paeniclostridium) could interact with host immune genes after the STEC O157 colonization and interactions were stx2a-dependent. Furthermore, bacterial predicted functions involved in pathogen (O157 and Paeniclostridium) colonization and metabolism were related to host immunities. These findings suggest that host-microbe interactions could shift from beneficial to opportunistic pathogenic bacteria-driven during the pathogen colonization and be dependent on the production of particular virulence factors, highlighting the potential regulatory role of mucosa-attached microbiota in affecting pathogen-commensal-host interactions under STEC O157 infection in calves.

The epidemiology of Shiga toxin-producing Escherichia coli serogroup O157 in England, 2009-2019.

Shiga toxin-producing Escherichia coli (STEC) serogroup O157 is a zoonotic, foodborne gastrointestinal pathogen of major public health concern. We describe the epidemiology of STEC O157 infection in England by exploring the microbiological and clinical characteristics, the demographic and geographical distribution of cases, and examining changes in environmental exposures over 11 years of enhanced surveillance. Enhanced surveillance data including microbiological subtyping, clinical presentations and exposures were extracted for all cases resident in England with evidence of STEC O157 infection, either due to faecal culture or serology detection. Incidence rates were calculated based on mid-year population estimates from the Office of National Statistics (ONS). Demographics, geography, severity and environmental exposures were compared across the time periods 2009-2014 and 2015-2019. The number of cases reported to national surveillance decreased, with the mean cases per year dropping from 887 for the period 2009-2014 to 595 for the period 2015-2019. The decline in STEC O157 infections appears to be mirrored by the decrease in cases infected with phage type 21/28. Although the percentage of cases that developed HUS decreased, the percentage of cases reporting bloody diarrhoea and hospitalisation remained stable. The number of outbreaks declined over time, although more refined typing methods linked more cases to each outbreak. Integration of epidemiological data with microbiological typing data is essential to understanding the changes in the burden of STEC infection, assessment of the risks to public health, and the prediction and mitigation of emerging threats.

Spatial Effects of Livestock Farming on Human Infections With Shiga Toxin-Producing Escherichia coli O157 in Small but Densely Populated Regions: The Case of the Netherlands.

The role of environmental transmission of typically foodborne pathogens like Shiga toxin‐producing Escherichia coli (STEC) O157 is increasingly recognized. To gain more insights into spatially restricted risk factors that play a role in this transmission, we assessed the spatial association between sporadic STEC O157 human infections and the exposure to livestock (i.e. small ruminants, cattle, poultry, and pigs) in a densely populated country: the Netherlands. This was done for the years 2007–2016, using a state‐of‐the‐art spatial analysis method in which hexagonal areas with different sizes (90, 50, 25 and 10 km2) were used in combination with a novel probability of exposure metric: the population‐weighted number of animals per hexagon. To identify risk factors for STEC O157 infections and their population attributable fraction (PAF), a spatial regression model was fitted using integrated nested Laplace approximation (INLA). Living in hexagonal areas of 25, 50 and 90 km2 with twice as much population‐weighted small ruminants was associated with an increase of the incidence rate of human STEC O157 infections in summer (RR of 1.09 [95%CI;1.01–1.17], RR of 1.17 [95%CI;1.07–1.28] and RR of 1.13 [95%CI;1.01–1.26]), with a PAF of 49% (95%CI;8–72%). Results suggest exposure to small ruminants to be a risk factor, although no evidence on the mode of transmission is provided. Therefore, the underlying mechanisms warrant further investigation and could offer new targets for control. The newly proposed exposure metric has potential to improve existing spatial modeling studies on infectious diseases related to livestock exposure, especially in densely populated countries like the Netherlands.

Shiga Toxin-Producing Escherichia coli Infections Associated With Romaine Lettuce-United States, 2018.

Produce-associated outbreaks of Shiga toxin-producing Escherichia coli (STEC) were first identified in 1991. In April 2018, New Jersey and Pennsylvania officials reported a cluster of STEC O157 infections associated with multiple locations of a restaurant chain. The Centers for Disease Control and Prevention (CDC) queried PulseNet, the national laboratory network for foodborne disease surveillance, for additional cases and began a national investigation. A case was defined as an infection between 13 March and 22 August 2018 with 1 of the 22 identified outbreak-associated E. coli O157:H7 or E. coli O61 pulsed-field gel electrophoresis pattern combinations, or with a strain STEC O157 that was closely related to the main outbreak strain by whole-genome sequencing. We conducted epidemiologic and traceback investigations to identify illness subclusters and common sources. A US Food and Drug Administration-led environmental assessment, which tested water, soil, manure, compost, and scat samples, was conducted to evaluate potential sources of STEC contamination. We identified 240 case-patients from 37 states; 104 were hospitalized, 28 developed hemolytic uremic syndrome, and 5 died. Of 179 people who were interviewed, 152 (85%) reported consuming romaine lettuce in the week before illness onset. Twenty subclusters were identified. Product traceback from subcluster restaurants identified numerous romaine lettuce distributors and growers; all lettuce originated from the Yuma growing region. Water samples collected from an irrigation canal in the region yielded the outbreak strain of STEC O157. We report on the largest multistate leafy greens-linked STEC O157 outbreak in several decades. The investigation highlights the complexities associated with investigating outbreaks involving widespread environmental contamination.

Epidemiology of Shiga Toxin-Producing Escherichia coli O157 in the Province of Alberta, Canada, 2009-2016.

Shiga toxin-producing Escherichia coli (STEC) infections are the product of the interaction between bacteria, phages, animals, humans, and the environment. In the late 1980s, Alberta had one of the highest incidences of STEC infections in North America. Herein, we revisit and contextualize the epidemiology of STEC O157 human infections in Alberta for the period 2009–2016. STEC O157 infections were concentrated in large urban centers, but also in rural areas with high cattle density. Hospitalization was often required when the Shiga toxin genotype stx2a stx2c was involved, however, only those aged 60 years or older and infection during spring months (April to June) independently predicted that need. Since the late 1980s, the rate of STEC O157-associated hemolytic uremic syndrome (HUS) in Alberta has remained unchanged at 5.1%, despite a marked drop in the overall incidence of the infection. While Shiga toxin genotypes stx1a stx2c and stx2a stx2c seemed associated with HUS, only those aged under 10 years and infection during spring months were independently predictive of that complication. The complexity of the current epidemiology of STEC O157 in Alberta highlights the need for a One Health approach for further progress to be made in mitigating STEC morbidity.

An outbreak of Shiga toxin-producing Escherichia coli serogroup O157 linked to a lamb-feeding event.

Fifteen confirmed cases and 15 possible cases of Shiga toxin-producing Escherichia coli (STEC) O157 phage type 21/28 were linked to direct contact with lambs at a 'Lambing Live' event in the North West of England between 29 March and 21 April 2014. Twenty-one (70%) of the cases were female, 23 (77%) were children aged <16 years, of whom 14 (46%) were in the 0-5 years age group. Five children developed haemolytic uraemic syndrome. Multilocus variable number tandem repeat analysis (MLVA) profiles on 14 human cases were indistinguishable, and 6/10 animal isolates had a MLVA profile identical to the outbreak profile. Whole-genome sequencing analysis revealed that all isolates, both human and animal, fell within a 5-single nucleotide polymorphism cluster indicating the isolates belonged to the same point source. On inspection of the premises, extensive and uncontrolled physical contact between visitors and animals was occuring within the animal pens and during bottle-feeding. Public areas were visibly contaminated with animal faeces. Information to visitors, and the infection control awareness demonstrated by staff, was inadequate. Managing the risk to visitors of STEC O157 infection at animal petting events and open farms requires implementation of stringent control measures by the operator, as outlined in the industry code of practice. Enforcement action is sometimes required to prevent high-risk activities taking place at both permanent and temporary attractions.

Characterization of the Shiga toxin-producing Escherichia coli O26 isolated from human in Poland between 1996 and 2014.

Shiga toxin-producing Escherichia coli (STEC) O26 infections can be comparable with STEC O157 infections in severity of the acute haemolytic-uremic syndrome HUS and long-term sequelae. Among O26 STEC isolates, highly virulent clone O26:H11/H- Sequence Type 29 (ST 29) emerged in Germany in mid-1990s and spread to European countries. However, up to date, no STEC O26:H11/H- belonging to ST29 has been documented in Poland. In this study, we determined the relationship and clonal structure, stx genotypes, plasmid gene profiles and antimicrobial resistance of nine human STEC O26:H11/H- strains from human patients in Poland between 1996 and 2014. Of the 9 human STEC O26:H11/H- strains, two belonged to ST29 and were isolated from two children with HUS and renal failure with sepsis respectively. These strains showed the molecular characteristics of the emerging human-pathogenic ST29 clone (stx1-, stx2a+, eae+, ehxA+, etpD+, katP-, espP-). The remaining STEC O26:H11/H- strains examined in this study, belonged to ST21, with plasmid genes profiles frequently reported in ST21 strains in Europe. STEC O26 infections with serious human health consequences highlight the need of continuous surveillance of non-O157 STEC and implementation of the diagnostic approaches focused on their detection. Significance and impact of the study: These study provides the first data on the occurrence of emerging Shiga toxin-producing Escherichia coli O26:H11 ST 29 clone in human patients in Poland. Those strains show the molecular characteristics of highly virulent new ST29 pathotype (stx1-, stx2a+, eae+ ehxA+, etpD+, katP-, espP-). These results demonstrated prompt efforts to implement diagnostic approaches detection of those pathogen in the European countries.

Genotypic characterization of Shiga toxin-producing Escherichia coli O157:H7 isolates in food products from china between 2005 and 2010

Shiga toxin-producing Escherichia coli (STEC) O157 is an important foodborne pathogen. The aims of this study were to determine genetic relatedness of STEC O157 isolated from foods in China. STEC O157 isolates from food were characterized by virulence gene typing, antibiotyping, Multi-locus sequence typing (MLST), pulsed-field gel electrophoresis analysis (PFGE), Multi-locus variable number tandem repeat analysis (MLVA), and the single nucleotide polymorphism (SNP) clade typing. Of the 30 STEC O157 isolates analyzed, all isolates harbored eae, exhA, stx1 and/or stx2 genes with stx2c subtype predominating. By MLST, they were relatively homogenous with only 4 STs. PFGE and MLVA generated 22 pulsotyples and 23 patterns, respectively, which showed considerable diversity. Only one clade 8 isolate was detected. These results indicate that STEC O157 isolates from foods in China were heterogeneous. There was no correlation between genotypic characteristics and sources of isolates. Since different subtyping methods often give different discriminatory powers, the use of more than one subtyping approach is necessary in providing a more accurate picture of the genetic diversity of STEC O157. Four isolates were from ready-to-eat meat or salads, underscores the risk of infections. There is a need for surveillance of STEC O157 in foods and clinically and implementation of prevention strategies to prevent outbreaks of STEC O157 infections in China.

The Dilemma of Antimicrobial Treatment of Shiga Toxin-producing Escherichia coli

The Dilemma of Antimicrobial Treatment of Shiga Toxin-producing Escherichia coli

Comparison between ImmunoCard STAT!® and real-time PCR as screening tools for both O157:H7 and non-O157 Shiga toxin-producing Escherichia coli in Southern Alberta, Canada

An increasing number of non-O157 Shiga toxin-producing Escherichia coli (STEC) infections and outbreaks have been reported. In this study, we evaluated the performance of ImmunoCard STAT!® (Meridian Bioscience, Inc., Cincinnati, OH, USA) as a method to screen stool specimens for STEC (O157 and non-O157). An in-house real-time PCR method was used as the “gold standard”. We also evaluated the prevalence and clinical characteristics of STEC infections in the Alberta South West Zone. From July to November 2011, 819 stool specimens submitted for routine stool culture were tested. With our in-house real-time PCR, 7 O157:H7 and 10 non-O157 STEC isolates were identified for a total of 17 STECs. In comparison, ImmunoCard STAT!® identified a total of 6, resulting in a sensitivity and specificity of 35% and 99%, respectively (P < 0.05). Because of the low sensitivity, ImmunoCard STAT!® cannot be recommended as a routine screening test for STEC from enriched stool specimens. The rate of STEC positivity as detected by PCR was 2.08%, of which 0.86% was O157:H7 and 1.22% non-O157 STEC. Five of the 7 cases of STEC O157 infection experienced bloody diarrhea, and 1 developed hemolytic uremic syndrome.

Attributing sporadic and outbreak-associated infections to sources: blending epidemiological data

Common sources of shiga toxin-producing Escherichia coli (STEC) O157 infection have been identified by investigating outbreaks and by case-control studies of sporadic infections. We conducted an analysis to attribute STEC O157 infections ascertained in 1996 and 1999 by the Foodborne Diseases Active Surveillance Network (FoodNet) to sources. Multivariable models from two case-control studies conducted in FoodNet and outbreak investigations that occurred during the study years were used to calculate the annual number of infections attributable to six sources. Using the results of the outbreak investigations alone, 27% and 15% of infections were attributed to a source in 1996 and 1999, respectively. Combining information from both data sources, 65% of infections in 1996 and 34% of infections in 1999 were attributed. The results suggest that methods to incorporate data from multiple surveillance systems and over several years are needed to improve estimation of the number of illnesses attributable to exposure sources.

Pork Implicated in a Shiga Toxin-producing Escherichia coli O157:H7 Outbreak in Ontario, Canada

To describe an outbreak of Shiga toxin-producing Escherichia coli (STEC) O157:H7 infection following a four-day family gathering in Ontario. This is the first published account of a STEC O157 outbreak in Canada linked to consumption of pork. The outbreak investigation included interviews with food handlers and other key associated persons, inspection of food preparation premises, traceback investigations, case finding, analysis of data from an outbreak questionnaire, and laboratory analysis of samples collected from various sources associated with the outbreak. Several meals, including pork from a pig roast, were served to the 59 attendees, 29 of whom developed gastrointestinal illness following the event. Six cases developed bloody diarrhoea and seven were hospitalized. Leftover pork served the day after the pig roast was the item most significantly associated with an increased risk of illness (p<0.001). STEC O157:H7 was isolated from 11 of the 29 ill attendees, and also from the pork. By pulse-field gel electrophoresis (PFGE), all STEC O157:H7 pork isolates were either identical or closely related to the 11 clinical isolates. No STEC was detected in any other samples. Three Clostridium perfringens isolates, unrelated by PFGE, were obtained from two STEC-positive cases and the pork. This outbreak highlights the need for increased awareness of pork as a potential source of STEC O157 infection, and for enhanced education regarding the safe handling, cooking and storage of food, specifically where large cuts of meat are cooked outdoors at events such as pig roasts, a cultural norm in some communities.

Epidemiology of Shiga toxin producing Escherichia coli in Australia, 2000-2010.

BackgroundShiga toxin-producing Escherichia coli (STEC) are an important cause of gastroenteritis in Australia and worldwide and can also result in serious sequelae such as haemolytic uraemic syndrome (HUS). In this paper we describe the epidemiology of STEC in Australia using the latest available data.MethodsNational and state notifications data, as well as data on serotypes, hospitalizations, mortality and outbreaks were examined.ResultsFor the 11 year period 2000 to 2010, the overall annual Australian rate of all notified STEC illness was 0.4 cases per 100,000 per year. In total, there were 822 STEC infections notified in Australia over this period, with a low of 1 notification in the Australian Capital Territory (corresponding to a rate of 0.03 cases per 100,000/year) and a high of 413 notifications in South Australia (corresponding to a rate of 2.4 cases per 100,000/year), the state with the most comprehensive surveillance for STEC infection in the country. Nationally, 71.2% (504/708) of STEC infections underwent serotype testing between 2001 and 2009, and of these, 58.0% (225/388) were found to be O157 strains, with O111 (13.7%) and O26 (11.1%) strains also commonly associated with STEC infections. The notification rate for STEC O157 infections Australia wide between 2001-2009 was 0.12 cases per 100,000 per year. Over the same 9 year period there were 11 outbreaks caused by STEC, with these outbreaks generally being small in size and caused by a variety of serogroups. The overall annual rate of notified HUS in Australia between 2000 and 2010 was 0.07 cases per 100,000 per year. Both STEC infections and HUS cases showed a similar seasonal distribution, with a larger proportion of reported cases occurring in the summer months of December to February.ConclusionsSTEC infections in Australia have remained fairly steady over the past 11 years. Overall, the incidence and burden of disease due to STEC and HUS in Australia appears comparable or lower than similar developed countries.

Nationwide outbreak of STEC O157 infection in the Netherlands, December 2008-January 2009: continuous risk of consuming raw beef products

The Netherlands experienced a nationwide outbreak of Shiga toxin-producing Escherichia coli (STEC) O157 with onset of symptoms from the end of December 2008 until the end of January 2009. A total of 20 laboratory-confirmed cases were linked to the outbreak strain, serotype O157: H-, stx1, stx2, eae and e-hly positive. The investigation into the source of this outbreak is still ongoing, but evidence so far suggests that infection occurred as a result of consuming contaminated raw meat (steak tartare).

An international outbreak of Shiga toxin-producing Escherichia coli O157 infection due to lettuce, September-October 2007.

Between 14 September and 20 October 2007, an outbreak of Shiga toxin-producing Escherichia coli (STEC) O157 simultaneously occurred in the Netherlands and Iceland. A total of 50 laboratory-confirmed cases were reported with a STEC O157 infection caused by the same clone. The strain was of type O157:H-, PT8, positive for stx1, stx2, eae and e-hly, and sorbitol negative. The most probable cause of this international outbreak was contaminated lettuce, shredded and pre-packed in a Dutch food processing plant. Samples of the environment, raw produce and end products, taken at several vegetable growers and processing plants all tested negative for STEC O157. However, the only epidemiological link between the cases in the Netherlands and in Iceland was the implicated Dutch processing plant. In Europe, food products are often widely distributed posing the risk of potential spread of food borne pathogens simultaneously to several countries. This international outbreak emphasises the importance of common alert and surveillance systems in earlier detection of international outbreaks and better assessment of their spread.

Risk Factors of Sporadic Human Infection of Shiga Toxin Producing Escherichia coli O157 and Non-O157 in Australia

Background: Shiga toxin producing Escherichia coli (STEC) causes acute gastroenteritis characterised by abdominal cramps and bloody diarrhoea. In Australia and elsewhere, serotype O157 is the predominant strain of STEC. To investigate risk factors for STEC O157 and non-O157 infection we conducted an Australia-wide case control study between 2003 and 2007. Methods: Patients infected with STEC that were notified to health departments were eligible for the study and three controls per case were selected from a bank of controls based on sex and five year age groups. Cases were recruited in South Australia from July 2003, with all other Australian states participating from 2005. A standardised questionnaire was used to collect information regarding demographic data, food, behavioural and environmental exposures. All factors associated with O157 and non-O157 infection at the univariate level (p < 0.1) were included in separate logistic regression models. Results: The O157 analysis included 43 cases and 117 controls; the non-O157 analysis included 71 cases and 187 controls. In multivariate analysis, STEC O157 infection was associated with eating hamburgers (OR = 6.6, 95% CI = 2.2–20.0), eating out of the home (OR = 4.8, 95% CI = 1.7–13.6) and living on or visiting a farm (OR = 4.6, 95% CI = 1.2–17.6). Non-O157 STEC infection was associated with consuming chicken deli meat (OR = 5.3, 95% CI = 1.5–18.5), working with animals (OR = 3.2, 95% CI = 1.2-8.3) and asthma (OR = 2.7, 95% CI = 1.1–6.3). Bush camping in Australia had an elevated odds ratio, although this was not significant (OR = 3.3, 95% CI = 0.9–11.9). Conclusions: This national case control study is the first to provide risk factors for both O157 and non-O157 STEC cases. That O157 was associated with farm exposure and eating hamburgers is consistent with that reported internationally. However, some of the risk factors for non-0157 cases have not been reported previously. It is important to consider these risk factors in developing prevention programmes for this serious illness.

Shiga toxin-producing Escherichia coli (STEC) O157 outbreak, The Netherlands, September - October 2005.

In September 2005, the first national food-related outbreak of Shiga toxin (Stx)-producing Escherichia coli (STEC) O157 was investigated in the Netherlands. A total of 21 laboratory-confirmed cases (including one secondary case), and another 11 probable cases (two primary and nine secondary cases) were reported in patients who became ill between 11 September and 10 October 2005. Preliminary investigation suggested consumption of a raw beef product, steak tartare (in the Netherlands also known as 'filet américain'), and contact with other symptomatic persons as possible risk factors. A subsequent case-control study supported the hypothesis that steak tartare was the source of the outbreak (matched odds ratio (OR) 272, 95% confidence interval (CI) 3 - 23211). Consumption of ready-to-eat vegetables was also associated with STEC O157 infection (matched OR 24, 95% CI 1.1 - 528), but was considered a less likely source, as only 40% of the cases were exposed. Samples of steak tartare collected from one chain of supermarkets where it is likely that most patients (67%) bought steak tartare, all tested negative for STEC O157. However, sampling was done three days after the date of symptom onset of the last reported case. Since 88% of the cases became ill within a two week period, point source contamination may explain these negative results. It is concluded that steak tartare was the most likely cause of the first national food-related outbreak of STEC O157 in the Netherlands.

Enhanced laboratory-based surveillance of Shiga-toxin-producing Escherichia coli O157 in The Netherlands.

The aim of this study was to analyse the results of a programme in the Netherlands for enhanced surveillance of Shiga-toxin-producing Escherichia coli (STEC) O157. In this programme, implemented in January 1999, all laboratories report positive cases to the public health services and submit isolates for typing to the reference laboratory. Public health services collect clinical and risk factor information of patients, using a standardised questionnaire. Results were analysed for the first two and a half years of the programme. In February 2000, a questionnaire was sent to all laboratories to assess (i) the criteria for testing faecal samples for STEC O157, (ii) the diagnostic tools used, and (iii) the level of participation in the surveillance programme. Between January 1999 and June 2001, 93 cases of symptomatic STEC O157 infection were reported, 25% of which occurred in children aged 0-4 years. Serotyping for O, H and stx types showed that two types dominated, O157:H7, s tx2 positive (48%) and O157:H-, stx1 and stx2 positive (24%). Analysis of the 93 isolates by pulsed-field gel electrophoresis showed 17 clusters of isolates with at least 95% fragments in common, including isolates with unknown epidemiological links. Of the patients for whom questionnaire information was reported, 38% were hospitalised, 15% developed haemolytic uraemic syndrome, and 52% reported a known risk factor, such as contact with farm animals or manure, consumption of raw or undercooked beef, consumption of raw milk or cheese made from raw milk, or contact with a symptomatic individual. Response to the laboratory survey was high (97%). Only 6% of the laboratories carried out testing for non-O157 STEC, although 95% performed testing for STEC O157. The majority (88%) used culture on sorbitol MacConkey agar or sorbitol MacConkey agar with cefixime and tellurite as the method of detection of STEC O157. The identity of the strains was confirmed primarily with commercially available latex agglutination assays (95% of laboratories) and biochemical characterisation with the API 20E test (bioMérieux, France) (42% of laboratories). Most laboratories (92%) used selection criteria for testing, especially bloody diarrhoea and other clinical information (81% of laboratories) and young age (10%). It is concluded that STEC O157 is a limited public health problem in the Netherlands, although the selective testing policy and the low sensitivity of the culture techniques used probably caused the incidence of STEC O157 infection to be underestimated.