E. M. Nielsen

Multilocus sequence typing performed on Campylobacter coli isolates from humans, broilers, pigs and cattle originating in Denmark

Aims:  To assess whether Campylobacter coli isolated from different sources in Denmark constitute separate populations.

Outbreak with multi-resistant Salmonella Typhimurium DT104 linked to carpaccio, Denmark, 2005

We report an outbreak with Salmonella Typhimurium DT104 resistant to six different antibiotic classes. The outbreak occurred in Denmark in July/August 2005 and was traced to a single restaurant. In addition to patient interviews, an important tool in the investigation of this outbreak was comparison by multi-locus variable number of tandem repeat analysis (MLVA) typing of patient strains with strains from the food surveillance system. This showed that the source of the outbreak was imported beef served as carpaccio. Carpaccio, thinly sliced raw fillet of beef, has not previously been associated with outbreaks, but should be considered a high-risk food item. This outbreak was one of two in different European Union (EU) countries traced back to beef from one company in a third EU country. This underscores the importance of efficient international Salmonella surveillance and food-safety control systems enabling timely interventions within the EU.

Development of real-time PCR and hybridization methods for detection and identification of thermophilic Campylobacter spp. in pig faecal samples

Aims:  To develop a real‐time (rt) PCR for species differentiation of thermophilic Campylobacter and to develop a method for assessing co‐colonization of pigs by Campylobacter spp.

Outbreaks of Shigella sonnei infections in Denmark and Australia linked to consumption of imported raw baby corn

We investigated an outbreak of Shigella sonnei infections in Denmark and Australia associated with imported baby corn from one packing shed in Thailand. We reviewed nationwide surveillance and undertook case finding, food trace-back and microbiological investigation of human, food and environmental samples. A recall of baby corn and sugar snaps was based on descriptive epidemiological evidence. In Denmark, we undertook a retrospective cohort study in one workplace. In total, 215 cases were laboratory-confirmed in Denmark, and 12 in Australia. In a multivariable analysis, baby corn was the only independent risk factor. Antibiotic resistance and PFGE outbreak profiles in Denmark and Australia were indistinguishable, linking the outbreaks. Although we did not detect S. sonnei in baby corn, we isolated high levels of other enteric pathogens. We identified a packing shed in Thailand that supplied baby corn to Denmark and Australia, and uncovered unhygienic practices in the supply chain. This outbreak highlights the importance of international communication in linking outbreaks and pinpointing the source.

Source attribution of human campylobacteriosis in Denmark

SUMMARY This study assesses the contribution of different sources of human campylobacteriosis in Denmark using two different source-attribution approaches. In total, 794 non-human isolates and 406 isolates from human cases (domestic, travel related, and cases with unknown travel history) were collected. Isolates were characterized by multilocus sequence typing, flaA typing and susceptibility to antibiotics. Both models used indicate that the major burden of human campylobacteriosis in Denmark originates from the domestic broiler chicken reservoir. The second most important reservoir was found to be cattle. The Asymmetric Island model attributed 52% [95% credibility interval (CrI) 37-67] to Danish chicken, 17% (95% CrI 3-33) to imported chicken, and 17% (95% CrI 7-28) to cattle. Similarly, the Campylobacter source-attribution model apportioned 38% (95% CrI 28-47) to Danish chicken, 14% (95% CrI 10-18) to imported chicken, and 16% (95% CrI 7-25) to cattle. The addition of flaA type as an extra discriminatory typing parameter did not change the attribution of cases markedly.

First general outbreak of verocytotoxin-producing Escherichia Coli O157 in Denmark

This report describes the first general outbreak of verocytotoxin-producing E. coli (VTEC) in Denmark. Twenty five patients, 18 children and seven adults, with culture-confirmed VTEC O157:H- infection and indistinguishable pulsed-field gel electrophoresis DNA profiles, were identified during a six month period from September 2003 to March 2004. The outbreak strain possessed the virulence genes: eae, vtx1 and vtx2c. All patients but one presented with diarrhoea; none developed haemolytic uraemic syndrome. The outbreak was restricted to Copenhagen and surrounding areas. A case-control study including 11 cases and 55 matched controls revealed an association between VTEC O157:H- infection and shopping in a specific supermarket chain in Copenhagen and surrounding area, matched odds ratio (OR): 8.7 (95% confidence interval (CI): 1.1-71). After exclusion of three assumed secondary cases, only consumption of a particular kind of organic milk from a small dairy was associated with disease OR: 8.7 (95% CI 1.6-48). Environmental and microbiological investigations at the suspected dairy did not confirm the presence of the outbreak strain, but the outbreak stopped once the dairy was closed and thoroughly cleaned.

Epidemiology of campylobacteriosis in Denmark 2000–2015

Campylobacter is the most frequently occurring cause of bacterial gastroenteritis in Europe. Unlike other zoonotic diseases, European‐wide incidences of Campylobacter infections have increased during the past decade, resulting in a significant disease burden. In Denmark, campylobacteriosis is notifiable by laboratory and a unique registration system of electronic transfer and storage of notified Campylobacter cases linked to the national person register of age, gender and geographical location allows collection of comprehensive case data. Using national surveillance data, we describe Campylobacter infections in Denmark from 2000 to 2015, focusing on age‐specific incidences, geography, seasonality and outbreaks. During the observed period, a total of 60,725 Campylobacter infections were registered with a mean annual incidence of 69.3 cases/100,000 population. From 2000 to 2014, the incidence of campylobacteriosis decreased by 20%, followed by an apparent increase of 20% from 2014 to 2015. Approximately one‐third of cases were travel‐related. Incidences were highest in males, young adults aged 20–29 years and children under 5 years of age. Generally, children under 10 years of age living in rural areas were at higher risk of infection. Infection patterns were seasonal with an increase from May to October, peaking in August. Outbreaks were identified each year, including four large waterborne outbreaks which all occurred following heavy rainfall events. For the most part, patterns of Campylobacter infection in Denmark during 2000 to 2015 remained remarkably constant and followed what is known about the disease with respect to demographic, temporal and spatial characteristics. To establish better targeted prevention and control measures, the current knowledge gaps regarding both Campylobacter microbiology (degree of clonal diversity and clustering) and the importance of different risk factors (food versus environment/climate) need to be filled.

Use of virulence determinants and seropathotypes to distinguish high- and low-risk Escherichia coli O157 and non-O157 isolates from Europe

The presence of 10 virulence genes was examined using polymerase chain reaction (PCR) in 365 European O157 and non-O157 Escherichia coli isolates associated with verotoxin production. Strain-specific PCR data were analysed using hierarchical clustering. The resulting dendrogram clearly separated O157 from non-O157 strains. The former clustered typical high-risk seropathotype (SPT) A strains from all regions, including Sweden and Spain, which were homogenous by Cramers V statistic, and strains with less typical O157 features mostly from Hungary. The non-O157 strains divided into a high-risk SPTB harbouring O26, O111 and O103 strains, a group pathogenic to pigs, and a group with few virulence genes other than for verotoxin. The data demonstrate SPT designation and selected PCR separated verotoxigenic E. coli of high and low risk to humans; although more virulence genes or pulsed-field gel electrophoresis will need to be included to separate high-risk strains further for epidemiological tracing.

External quality assessment for molecular typing of Salmonella 2013–2015: performance of the European national public health reference laboratories

We report the results of three consecutive External Quality Assessments (EQAs) for molecular subtyping of Salmonella to assess the performance of the European national public health reference laboratories (NPHRLs). The EQA included the molecular typing methods used for European enhanced surveillance of human Salmonella infections: pulsed field gel electrophoresis (PFGE), including gel analysis by the use of the software BioNumerics, and 5-locus multiple locus variable number of tandem repeat analysis (MLVA) for serovar Typhimurium. The participation in the PFGE laboratory part was higher (27/35) than in the gel analysis (19/35) and MLVA (15/35), suggestive of the need for capacity building in methods requiring specialized equipment (MLVA) or software (gel analysis). The majority (25/27) of the participating NPHRLs produced inter-laboratory comparable PFGE gel(s). Two laboratories continued to produce low-quality gels and should have additional technical assistance in the future. In particular, two gel quality evaluation parameters, measuring “image acquisition and running conditions” and “bands”, were identified to cause gel quality problems throughout the EQAs. Despite the high number of laboratories participating in the PFGE laboratory part, the participation in gel analysis was low, although increasing. In the MLVA part, the NPHRLs correctly assigned 96% (405/420) allelic profiles according to the nomenclature. In conclusion, the EQAs identified critical parameters for unsuccessful performance and helped to offer assistance to those laboratories that needed it most. The assessments supported the development of quality in molecular typing and promoted the harmonization of subtyping methods used for EU/EEA-wide surveillance of human Salmonella infections.