Prevalence, characterisation and clinical profiles of Shiga toxin-producing Escherichia coli in The Netherlands.

Detection of Shiga toxin-producing Escherichia coli (STEC) in The Netherlands is traditionally limited to serogroup O157. To assess the relative importance of STEC, including non-O157 serogroups, stool samples submitted nationwide for investigation of enteric pathogens or diarrhoea were screened with real-time PCR for the presence of the Shiga toxin genes. Patients were selected if their stool contained blood upon macroscopic examination, if they had a history of bloody diarrhoea, were diagnosed with haemolytic uraemic syndrome, or were aged <6 years (irrespective of the bloody aspect of the stool). PCR-positive stools were forwarded to a central laboratory for STEC isolation and typing. In total, 4069 stools were examined, with 68 (1.7%) positive PCR results. The highest prevalence was for stools containing macroscopic blood (3.5%), followed by stools from patients with a history of bloody diarrhoea (2.4%). Among young children, the prevalence (1.0%) was not significantly higher than among random, non-bloody, stool samples from diarrhoeal patients (1.4%). STEC strains were isolated from 25 (38%) PCR-positive stools. Eleven O-serogroups were detected, including five STEC O157 strains. As serogroup O157 represented only 20% of the STEC isolates, laboratories should be encouraged to use techniques enabling them to detect non-O157 serogroups, in parallel with culture, for isolation and subsequent characterisation of STEC strains for public health surveillance and detection of outbreaks.

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.

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 americain"), 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) 3-23,211). 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.

A longitudinal study of Escherichia coli O157 in cattle of a Dutch dairy farm and in the farm environment.

From July 1999 till November 2000, a longitudinal study was conducted on a dairy farm in The Netherlands to study within herd prevalence and types of verocytotoxin producing Escherichia coli (VTEC) of serogroup O157 over time, and determine environmental reservoirs and possible transmission routes. Faeces, blood, milk and environmental samples were collected 14 times with intervals varying from 4 to 10 weeks during the study period. Faecal samples were selectively cultured for Escherichia coli O157. Isolates were tested by PCR for the most common virulence genes, VTI, VTII and eae, and typed by pulsed field gel electrophoresis. In total, 71 isolates were obtained, of which 49 from dairy cows, 8 from young stock, 5 from other animals and 9 from the environment. Positive samples were all detected in summer and early fall. VT- and eae-genes were found in all tested isolates, except in one. DNA typing showed that three clusters of O157 isolates could be identified. One of these clusters contained samples of two shedding seasons, indicating persistence on the farm during winter and spring. Repeated measures analysis of variance showed that cows with O157 VTEC infection had higher daily milk production in the period preceding sampling (p = 0.0055). There was no significant association between the results of the LPS-ELISA on serum samples from dairy cows and their O157 status.

Invasive Listeria monocytogenes infections in the Netherlands, 1995-2003.

In order to add to the limited data available about the incidence of invasive Listeria monocytogenes infection in the Netherlands, two studies were conducted. In the first study, data on hospital patients with listeriosis in the period 1995-2003 were obtained from the National Medical Registration (study 1). In the second study, hospital discharge letters for patients whose Listeria isolates were received by the Netherlands Reference Laboratory for Bacterial Meningitis (NRLBM) in the period 1999-2003 were retrieved (study 2). Serotyping and pulsed-field gel electrophoresis (PFGE) were used to subtype the various strains of Listeria. These reviews revealed 283 hospital patients and 159 patients with Listeria isolates. Discharge letters were received for 107 (67%) patients. The mean annual incidence of listeriosis in both studies was 2.0 per million inhabitants. The main clinical manifestations were meningitis (incidence: 0.9 and 1.0 per million in studies 1 and 2, respectively) and septicaemia (incidence: 0.08 and 1.0 per million, respectively). Listeriosis in pregnancy was rare (incidence: 1.3 and 2.4 per 100,000 pregnancies over 24 weeks of gestation, respectively). Predisposing conditions were present in 47 and 71% of the patients in studies 1 and 2, respectively. The mortality due to listeriosis was 18%. Serotypes 4b, 1/2a, and 1/2b were responsible for 96% of the cases of human listeriosis. Listeriosis is rare in the Netherlands, but its clinical course is severe and the resulting mortality is high. Therefore, the current recommendations for pregnant women to avoid high-risk foods should be continued. These dietary recommendations should also be given to individuals with predisposing conditions, since they, too, are at risk of Listeria infection.

Distribution of molecular subtypes within Salmonella enterica serotype Enteritidis phage type 4 and S. Typhimurium definitive phage type 104 in nine European countries, 2000-2004: results of an international multi-centre study.

This study investigates the distribution of pulsed-field gel electrophoresis (PFGE) profiles within Salmonella enterica serotype Enteritidis phage type (PT) 4 and S. Typhimurium definitive phage type (DT) 104, from cases of human infection in nine European countries from 2000 to 2004. Isolates were subtyped using standardized methods and gel images submitted by each participating country to the coordinating centre (Health Protection Agency Centre for Infections, London, UK), where they were entered into a central database, developed within BioNumerics software, and designated using an agreed nomenclature. S. Enteritidis PT4 (n=3637) was differentiated into 38 different profiles. Simpson's index of diversity (D) of profiles ranged from 0.2 to 0.4. Profile SENTXB.0001 represented at least 80% of all profiles in each country. S. Typhimurium DT104 (n=1202) was differentiated into 28 different profile types. Simpson's D was at least 0.6 in all countries except in Austria and Italy. In both these countries over 74% of S. Typhimurium DT104 profiles were STYMXB.0013. Profile STYMXB.0061, was predominant in Denmark, Spain, Finland and England and Wales where it represented between 36% and 45% of profiles. Profile STYMXB.0001 represented nearly half of all profiles in Scotland and 23% in England and Wales. PFGE is proving useful for further discrimination within S. Enteritidis PT4 and S. Typhimurium DT104. Ascertainment of international outbreaks involving common serotypes and phage types may be increased by the timely pooling of PFGE profiles within a central database readily accessible to all participating countries.

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.