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Small mammals are known to carryCampylobacterspp.; however, little is known about the genotypes and their role in human infections. We studied intestinal content from small wild mammals collected in their natural habitats in Finland in 2010–2017, and in close proximity to 40 pig or cattle farms in 2017. The animals were trapped using traditional Finnish metal snap traps.Campylobacterspp. were isolated from the intestinal content using direct plating on mCCDA. A total of 19% of the captured wild animals (n= 577) and 41% of the pooled farm samples (n= 227) were positive forC. jejuni, which was the onlyCampylobacterspecies identified. The highest prevalence occurred in yellow-necked mice (Apodemus flavicollis) and bank voles (Myodes glareolus) which carriedCampylobacterspp. in 66.3 and 63.9% of the farm samples and 41.5 and 24.4% of individual animals trapped from natural habitats, respectively. Interestingly, all house mouse (Mus musculus) and shrew (Sorexspp.) samples were negative forCampylobacterspp.C. jejuniisolates (n= 145) were further characterized by whole-genome sequencing. Core genome multilocus sequence typing (cgMLST) clustering showed that mouse and vole strains were separated from the rest of theC. jejunipopulation (636 and 671 allelic differences, 94 and 99% of core loci, respectively). Very little or no alleles were shared withC. jejunigenomes described earlier from livestock or human isolates. FastANI results further indicated thatC. jejunistrains from voles are likely to represent a new previously undescribed species or subspecies ofCampylobacter. Core-genome phylogeny showed that there was no difference between isolates originating from the farm and wild captured animals. Instead, the phylogeny followed the host species-association. There was some evidence (one strain each) of livestock-associatedC. jejunioccurring in a farm-caughtA. flavicollisand a brown rat (Rattus norvegicus), indicating that although small mammals may not be the original reservoir ofCampylobactercolonizing livestock, they may sporadically carryC. jejunistrains occurring mainly in livestock and be associated with disease in humans.

Campylobacter jejuni has caused several campylobacteriosis outbreaks via raw milk consumption. This study reports follow-up of a milk-borne campylobacteriosis outbreak that revealed persistent C. jejuni contamination of bulk tank milk for seven months or longer. Only the outbreak-causing strain, representing sequence type (ST) 883, was isolated from milk, although other C. jejuni STs were also isolated from the farm. We hypothesized that the outbreak strain harbors features that aid its environmental transmission or survival in milk. To identify such phenotypic features, the outbreak strain was characterized for survival in refrigerated raw milk and in aerobic broth culture by plate counting and for biofilm formation on microplates by crystal violet staining and quantification. Furthermore, whole-genome sequences were studied for such genotypic features. For comparison, we characterized isolates representing other STs from the same farm and an ST-883 isolate that persisted on another dairy farm, but was not isolated from bulk tank milk. With high inocula (105 CFU/ml), ST-883 strains survived in refrigerated raw milk longer (4–6 days) than the other strains (≤3 days), but the outbreak strain showed no outperformance among ST-883 strains. This suggests that ST-883 strains may share features that aid their survival in milk, but other mechanisms are required for persistence in milk. No correlation was observed between survival in refrigerated milk and aerotolerance. The outbreak strain formed a biofilm, offering a potential explanation for persistence in milk. Whether biofilm formation was affected by pTet-like genomic element and phase-variable genes encoding capsular methyltransferase and cytochrome C551 peroxidase warrants further study. This study suggests a phenotypic target candidate for interventions and genetic markers for the phenotype, which should be investigated further with the final aim of developing control strategies against C. jejuni infections.

In November 2016, an elderly patient was diagnosed with Listeria monocytogenes bacteremia in Finland. Grocery store loyalty card records and microbiological investigation of foods found in the home fridge and freezer of the patient revealed commercial, modified-atmosphere packaged meatballs as the source of the infection. Investigation of the meatball production plant revealed that the floor drain samples were contaminated with the same L. monocytogenes strain as those isolated from the patient and meatballs. Ready-to-eat meatballs were likely contaminated after heat treatment from the production environment before packaging. Long-term cold storage, modified-atmosphere conditions, and the absence of competing bacteria presumably enhanced the growth of L. monocytogenes. We recommend that collection of shopping details and home fridge and freezer sampling should be part of surveillance of all cases of L. monocytogenes infections to complement information obtained from in-depth interviews.

Shiga toxin-producing Escherichia coli (STEC) and Campylobacter jejuni are notable health hazards associated with the consumption of raw milk. These bacteria may colonize the intestines of asymptomatic cattle and enter bulk tank milk via fecal contamination during milking. We studied the frequency of STEC O157:H7 and C. jejuni contamination in tank milk (n = 785) and the in-line milk filters of milking machines (n = 631) versus the frequency of isolation from cattle feces (n = 257) on three Finnish dairy farms for 1 year. Despite simultaneous isolation of STEC O157:H7 (17%) or C. jejuni (53%) from cattle, these bacteria were rarely isolated from milk filters (2% or

The increased popularity of raw milk consumption has created demand for relaxing legislation, despite the risk of contamination by pathogenic bacteria, notably STEC and C. jejuni. However, the epidemiology of these milk-borne pathogens on the herd level is still poorly understood, and data are lacking on the frequency of milk contamination on farms with cattle shedding these bacteria in their feces. This study suggests (i) that STEC contamination in milk can be reduced, but not prevented, by on-farm hygienic measures while fecal shedding is observable, (ii) that milk filters are more suitable sampling targets for monitoring than milk although pathogen detection from both sample matrices may be challenging, and (iii) that STEC and C. jejuni genotypes may persist in cattle herds for several months. The results can be utilized in developing and targeting pathogen monitoring and risk management on the farm level and contributed to the revision of Finnish legislation in 2017., Shiga toxin-producing Escherichia coli (STEC) and Campylobacter jejuni are notable health hazards associated with the consumption of raw milk. These bacteria may colonize the intestines of asymptomatic cattle and enter bulk tank milk via fecal contamination during milking. We studied the frequency of STEC O157:H7 and C. jejuni contamination in tank milk (n = 785) and the in-line milk filters of milking machines (n = 631) versus the frequency of isolation from cattle feces (n = 257) on three Finnish dairy farms for 1 year. Despite simultaneous isolation of STEC O157:H7 (17%) or C. jejuni (53%) from cattle, these bacteria were rarely isolated from milk filters (2% or

The prevalence of human pathogenic Yersinia enterocolitica was determined in tonsil and intestinal content samples from 388 healthy fattening pigs at the four biggest Finnish slaughterhouses. These slaughterhouses process 73% of pigs in Finland. Tonsil samples were tested by PCR targeted for yadA, and intestinal samples were cultured. All pathogenic Y. enterocolitica isolates represented bioserotype 4/O:3. The prevalence of Y. enterocolitica in tonsil samples was 60% (95% confidence limit, 55.4 to 65.1%), and its prevalence in intestinal samples was 26% (95% confidence limit, 22.1 to 31.2%). The prevalence of Y. enterocolitica in tonsil and intestinal samples varied between the four slaughterhouses. The tonsil prevalence of Y. enterocolitica was higher in slaughterhouse B, and the prevalence in intestinal content was higher in slaughterhouse C. There were more positive results in both tonsil and intestinal samples in pigs coming from fattening farms than in pigs coming from farrowing-and-fattening farms. A seasonal variation was observed in the prevalence of Y. enterocolitica in intestinal samples, with the highest prevalence during July and August, but no seasonal variation was detected in tonsil samples.

A Bayesian statistical temporal-prevalence-concentration model (TPCM) was built to assess the prevalence and concentration of pathogenic campylobacter species in batches of fresh chicken and turkey meat at retail. The data set was collected from Finnish grocery stores in all the seasons of the year. Observations at low concentration levels are often censored due to the limit of determination of the microbiological methods. This model utilized the potential of Bayesian methods to borrow strength from related samples in order to perform under heavy censoring. In this extreme case the majority of the observed batch-specific concentrations was below the limit of determination. The hierarchical structure was included in the model in order to take into account the within-batch and between-batch variability, which may have a significant impact on the sample outcome depending on the sampling plan. Temporal changes in the prevalence of campylobacter were modeled using a Markovian time series. The proposed model is adaptable for other pathogens if the same type of data set is available. The computation of the model was performed using OpenBUGS software.

In response to the EFSA call New approaches in identifying and characterizing microbial and chemical hazards, the project INNUENDO (https://sites.google.com/site/theinnuendoproject/) aimed to design an analytical platform and standard procedures for the use of whole-genome sequencing in surveillance and outbreak investigation of food-borne pathogens. The project firstly attempted to identify existing flaws and needs, and then to provide applicable cross-sectorial solutions. The project focused in developing a platform for small countries with limited economical and personnel resources. To achieve these goals, we applied a user-centered design strategy involving the end-users, such as microbiologists in public health and veterinary authorities, in every step of the design, development and implementation phases. As a result, we delivered the INNUENDO Platform V1.0 (https://innuendo.readthedocs.io/en/latest/), a stand-alone, portable, open-source, end-to-end system for the management, analysis, and sharing of bacterial genomic data. The platform uses Nextflow workflow manager to assemble analytical software modules in species-specific protocols that can be run using a user-friendly interface. The reproducibility of the process is ensured by using Docker containers and throught the annotation of the whole process using an ontology. Several modules, available at https://github.com/TheInnuendoProject, have been developed including: genome assembly and species confirmation; fast genome clustering; in silico typing; standardized species-specific phylogenetic frameworks for Campylobacter jejuni, Yersinia enterocolitica, Salmonella enterica and Escherichia coli based on an innovative gene-by-gene methodology; quality control measures from raw reads to allele calling; reporting system; a built-in communication protocols and a strain classification system enabling smooth communication during outbreak investigation. As proof-of-concepts, the proposed solutions have been thoroughly tested in simulated outbreak conditions by several public health and veterinary agencies across Europe. The results have been widely disseminated through several channels (web-sites, scientific publications, organization of workshops). The INNUENDO Platform V1.0 is effectively one of the models for the usage of open-source software in genomic epidemiology.

Packaged raw milk contaminated with Yersinia pseudotuberculosis mediated a large yersiniosis outbreak in southern Finland in 2014. The outbreak was traced back to a single dairy farm in southern Finland. Here we explore risk factors leading to the outbreak through epidemiologic investigation of the outbreak farm and through genomic and phenotypic characterization of the farm’s outbreak and non-outbreak associated Y. pseudotuberculosis strains. We show that the outbreak strain persisted on the farm throughout the 7-month study, whereas the non-outbreak strains occurred sporadically. Phylogenomic analysis illustrated that the outbreak strain was related to previously published genomes of wild animal isolates from Finland, implying that wild animals were a potential source of the outbreak strain to the farm. We observed allelic differences between the farm’s outbreak and non-outbreak strains in several genes associated with virulence, stress response and biofilm formation, and found that the outbreak strain formed biofilm in vitro and maintained better growth fitness during cold stress than the non-outbreak strains. Finally, we demonstrate the rapid growth of the outbreak strain in packaged raw milk during refrigerated storage. This study provides insight of the risk factors leading to the Y. pseudotuberculosis outbreak, highlights the importance of pest control to avoid the spread of pathogens from wild to domestic animals, and demonstrates that the cold chain is insufficient as the sole risk management strategy to control Y. pseudotuberculosis risk associated with raw drinking milk.

The molecular epidemiology of Listeria monocytogenes was investigated in a longitudinal study of three Finnish dairy farms during 2013 to 2016. A total of 186 bulk tank milk (BTM), 224 milk filter sock (MFS), and 1,702 barn environment samples were analyzed, and isolates of L. monocytogenes were genotyped using pulsed-field gel electrophoresis. L. monocytogenes occurred throughout the year in all sample types, and the prevalence in MFS increased significantly during the indoor season. L. monocytogenes was more prevalent in MFS (29%) than in BTM (13%) samples. However, the prevalence of L. monocytogenes varied more between farms in samples of MFS (13 to 48%) than in BTM (10 to 16%). For each farm, the L. monocytogenes genotypes detected were classified by persistence (defined as persistent if isolated from ≥3 samples during ≥6 months) and predominance (defined as predominant if >5% prevalence on at least one farm visit). The prevalence of sporadic genotypes was 4 to 5% on all three farms. In contrast, the prevalence of persistent predominant genotypes varied between farms by 4% to 16%. The highest prevalence of persistent predominant genotypes was observed on the farm with the poorest production hygiene. Persistent predominant genotypes were most prevalent on feeding surfaces, water troughs, and floors. Genotypes isolated from the milking system or from cow udders had a greater relative risk of occurring in BTM and MFS than genotypes that only occurred elsewhere in the farm, supporting the hypothesis that L. monocytogene s is transmitted to milk from contamination on the udder surface or in the milking equipment. IMPORTANCE Listeria monocytogenes is a ubiquitous environmental bacterium and the causative agent of a serious foodborne illness, listeriosis. Dairy products are common vehicles of listeriosis, and dairy cattle farms harbor L. monocytogenes genotypes associated with human listeriosis outbreaks. Indeed, dairy cattle farms act as a reservoir of L. monocytogenes , and the organism is frequently detected in bulk tank milk (BTM) and in the feces of clinically healthy cows. The ecology of L. monocytogenes in the farm environment is complex and poorly understood. Isolates of the same L. monocytogenes genotype can occur in the farm for years, but the factors contributing to the persistence of genotypes on dairy farms are unknown. Knowledge of the persistence patterns and contamination routes of L. monocytogenes on dairy farms can improve management of the contamination pressure in the farm environment and aid in the development of focused control strategies to reduce BTM contamination.

Summary Shiga toxin-producing, sorbitol-fermenting Escherichia coli O157 (SF O157) has emerged as a cause of severe human illness. Despite frequent human findings, its transmission routes and reservoirs remain largely unknown. Foodborne transmission and reservoir in cattle have been suspected, but with limited supporting evidence. This study describes the outbreak of SF O157 that occurred in Finland in 2012. The outbreak originated from a recreational farm selling unpasteurized milk, as revealed by epidemiologic and microbiological investigations, and involved six hospitalized children and two asymptomatic adults with culture-confirmed infection. An identical strain of SF O157 was isolated from patients, cattle and the farm environment, and epidemiologic analysis suggested unpasteurized milk as the vehicle of transmission. This study reports the first milkborne outbreak of SF O157, provides supporting evidence of cattle as a reservoir and highlights the health risks related to the consumption of unpasteurized milk.

The decreased costs of genome sequencing have increased the capability to apply whole-genome sequencing to epidemiological surveillance of zoonotic Campylobacter jejuni. However, knowledge of the genetic diversity of this bacteria is vital for inferring relatedness between epidemiologically linked isolates and a necessary prerequisite for correct application of this methodology. To address this issue in C. jejuni we investigated the spatial and temporal signals in the genomes of a major clonal complex and generalist lineage, ST-45 CC, by analysing the population structure and genealogy as well as applying genome-wide association analysis of 340 isolates from across Europe collected over a wide time range. The occurrence and strength of the geographical signal varied between sublineages and followed the clonal frame when present, while no evidence of a temporal signal was found. Certain sublineages of ST-45 formed discrete and genetically isolated clades containing isolates with extremely similar genomes regardless of time and location of sampling. Based on a separate data set, these monomorphic genotypes represent successful C. jejuni clones, possibly spread around the globe by rapid animal (migrating birds), food or human movement. In addition, we observed an incongruence between the genealogy of the strains and multilocus sequence typing (MLST), challenging the existing clonal complex definition and the use of whole-genome gene-by-gene hierarchical nomenclature schemes for C. jejuni.

A Bayesian statistical temporal-prevalence-concentration model (TPCM) was built to assess the prevalence and concentration of pathogenic campylobacter species in batches of fresh chicken and turkey meat at retail. The data set was collected from Finnish grocery stores in all the seasons of the year. Observations at low concentration levels are often censored due to the limit of determination of the microbiological methods. This model utilized the potential of Bayesian methods to borrow strength from related samples in order to perform under heavy censoring. In this extreme case the majority of the observed batch-specific concentrations was below the limit of determination. The hierarchical structure was included in the model in order to take into account the within-batch and between-batch variability, which may have a significant impact on the sample outcome depending on the sampling plan. Temporal changes in the prevalence of campylobacter were modeled using a Markovian time series. The proposed model is adaptable for other pathogens if the same type of data set is available. The computation of the model was performed using OpenBUGS software.

Statistical source attribution approaches of food-related zoonoses can generally be based on reported diagnosed human cases and surveillance results from different food sources or reservoirs of bacteria. The attribution model, or probabilistic classifier, can thus be based on the (sub)typing information enabling comparison between human infections and samples derived from source surveillance. Having time series of both data allows analyzing temporal patterns over time providing a repeated natural experiment. A Bayesian approach combining both sources of information over a long time series is presented in the case of Campylobacter in Finland and Norway. The full model is transparently presented and derived from the Bayes theorem. Previous statistical source attribution approaches are here advanced (1) by explicit modeling of the cases not associated with any of the sources under surveillance over time, (2) by modeling uncertain prevalence in a food source by bacteria type over time, and (3) by implementing formal model fit assessment using posterior predictive discrepancy functions. Large proportion of all campylobacteriosis can be attributed to broiler, but considerable uncertainty remains over time. The source attribution is inherently incomplete if only the sources under surveillance are included in the model. All statistical source attribution approaches should include a model fit assessment for judgment of model performance with respect to relevant quantities of interest. It is especially relevant when the model aims at a synthesis of several incomplete information sources under significant uncertainty of explanatory variables.

During a period of 9 months, 194 marinated and non-marinated poultry products were collected from retail shops in a defined area in Western Finland and tested for Campylobacter spp. using a conventional enrichment culture and Polymerase Chain Reaction (PCR) method. For marinated poultry products, the study involved modification of a commercial DNA isolation method. Using either a conventional culture or PCR method, a total of 25 (12.9%) of all investigated samples were Campylobacter positive. In marinated poultry products, Campylobacter was detected at a prevalence of 21.1% and 9.5% in turkey and chicken products, respectively. In August, there was a peak with 28.9% positive Campylobacter samples. Campylobacter inoculation tests were carried out to test the detection limit of both methods. The PCR method used is faster than microbiological analyses. However, enrichment of the samples is necessary due to the low occurrence of Campylobacter spp. in retail Finnish poultry products.

Campylobacter jejuni is the leading cause of bacterial gastroenteritis and chicken is considered a major reservoir and source of human campylobacteriosis. In this study, we investigated temporally related Finnish human (n = 95), chicken (n = 83) and swimming water (n = 20) C. jejuni isolates collected during the seasonal peak in 2012 using multilocus sequence typing (MIST) and whole-genome MIST (wgMLST). Our objective was to trace domestic human C jejuni infections to C jejuni isolates from chicken slaughter batches and swimming water. At MIST level, 79% of the sequence types (STs) of the human isolates overlapped with chicken STs suggesting chicken as an important reservoir. Four STs, the ST-45, ST-230, ST-267 and ST-677, covered 75% of the human and 64% of the chicken isolates. In addition, 50% of the swimming water isolates comprised ST-45, ST-230 and ST-677. Further wgMLST analysis of the isolates within STs, accounting their temporal relationship, revealed that 22 of the human isolates (24%) were traceable back to C jejuni positive chicken slaughter batches. None of the human isolates were traced back to swimming water, which was rather sporadically sampled. The highly discriminatory wgMLST, together with the patient background information and temporal relationship data with possible sources, offers a new, accurate approach to trace back the origin of domestic campylobacteriosis. Our results suggest that potentially a substantial proportion of campylobacteriosis cases during the seasonal peak most probably are due to other sources than chicken meat consumption. These findings warrant further wgMLST-based studies to reassess the role of other reservoirs in the Campylobacter epidemiology both in Finland and elsewhere. (C) 2016 Elsevier B.V. All rights reserved.

Consumption and handling of chicken meat are well-known risk factors for acquiring campylobacteriosis. This study aimed to describe the Campylobacter jejuni population in Finnish chickens and to investigate the distribution of C. jejuni genotypes on Finnish chicken farms over a period of several years. We included 89.8% of the total C. jejuni population recovered in Finnish poultry during 2004, 2006, 2007, 2008, and 2012 and used multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) to characterize the 380 isolates. The typing data was combined with isolate information on collection-time and farm of origin. The C. jejuni prevalence in chicken slaughter batches was low (mean 3.0%, CI95% [1.8%, 4.2%]), and approximately a quarter of Finnish chicken farms delivered at least one positive chicken batch yearly. In general, the C. jejuni population was diverse as represented by a total of 63 sequence types (ST), but certain predominant MLST lineages were identified. ST-45 clonal complex (CC) accounted for 53% of the isolates while ST-21 CC and ST-677 CC covered 11% and 9% of the isolates, respectively. Less than half of the Campylobacter positive farms (40.3%) delivered C. jejuni-contaminated batches in multiple years, but the genotypes (ST and PFGE types) generally varied from year to year. Therefore, no evidence for a persistent C. jejuni source for the colonization of Finnish chickens emerged. Finnish chicken farms are infrequently contaminated with C. jejuni compared to other European Union (EU) countries, making Finland a valuable model for further epidemiological studies of the C. jejuni in poultry flocks.

Results. Of 7392 persons at risk, 111 (1.5%) met the case definition 76 case patients and 172 healthy control subjects were enrolled in the case-control study. Only raw grated carrots were significantl associated with illness in a logistic-regression model (multivariable odds ratio, 5.7 [95% confidenc interval, 1.7–19.5]); a dose response was found for increasing amount of consumption. Y. pseudotuberculosis O:1 isolates from 39 stool specimens and from 5 (42%) of 12 soil samples that contained carrot residue and were obtained from peeling and washing equipment at the production farm were indistinguishable by PFGE. Conclusions. Carrots contaminated early in the production process caused a large point-source outbreak. Our finding enable the development of evidence-based strategies to prevent outbreaks of this emerging foodborne pathogen.

A fatal infection with temporal relation to 2 other febrile infections caused by Vibrio cholerae non-O1, non-O139 (NCV) occurred in Finland in 2003. All infections were associated with contact with seawater. The patient who died had also eaten home-salted whitefish, tested positive for NCV, preceding his symptoms. All patients had compromising factors, and all strains were distinguishable by pulsed-field gel electrophoresis and negative for the ctx gene. These 3 cases illustrate that, despite being uncommon in Finland, NCVs can cause clinically significant and even fatal infections.

In March 2014, a Yersinia pseudotuberculosis (YP) outbreak was detected by a municipal authority in southern Finland. We conducted epidemiological, microbiological and traceback investigations to identify the source. We defined a case as a person with YP infection notified to the National Infectious Disease Registry between February and April 2014, or their household member, with abdominal pain and fever ≥ 38 °C or erythema nodosum. Healthy household members were used as household-matched controls. We identified 43 cases and 50 controls. The illness was strongly associated with the consumption of raw milk from a single producer. The odds ratio of illness increased with the amount of raw milk consumed. Also previously healthy adults became infected by consuming raw milk. Identical YP strains were identified from cases’ stool samples, raw milk sampled from a case’s refrigerator and from the milk filter at the producer’s farm. The producer fulfilled the legal requirements for raw milk production and voluntarily recalled the raw milk and stopped its production. We advised consumers to heat the raw milk to 72 °C for 15 s. Current legislation for raw milk producers should be reviewed and public awareness of health risks linked to raw milk consumption should be increased.

The genomic stability of 12 Campylobacter jejuni strains consisting of two groups of human and chicken isolates was studied by analysis of their PFGE (pulsed-field gel electrophoresis) patterns after passage through newly hatched chicks’ intestines. The patterns of Sma I, Sal I, and Sac II digests remained stable after intestinal passage, except for those of two strains. One originally human strain, FB 6371, changed its genotype from II/A ( Sma I/ Sac II) to I/B. Another strain, BTI, originally isolated from a chicken, changed its genotype from I/B to a new genotype. The genomic instability of the strains was further confirmed by Sal I digestion and ribotyping of the Hae III digests. In addition, heat-stable serotype 57 of strain FB 6371 changed to serotype 27 in all isolates with new genotypes but remained unchanged in an isolate with the original genotype. Serotype 27 of strain BTI remained stable. Our study suggests that during intestinal colonization, genomic rearrangement, as demonstrated by changed PFGE and ribopatterns, may occur.

To the Editor: Illness caused by Yersinia pseudotuberculosis is mainly characterized by fever and acute abdominal pain due to mesenteric lymphadenitis that mimics appendicitis. Secondary manifestations include erythema nodosum and reactive arthritis (1). Outbreaks have been reported in the Northern Hemisphere, including Canada (2,3), Japan (4), and Russia (5). Several community outbreaks have also been reported in Finland since 1982 (1,6–9). Only in a few of the outbreaks has the vector or source of the infection been identified. Recently, fresh produce, such as iceberg lettuce (7) and carrots (9), has been implicated by epidemiologic investigations as a source of infection, but mechanisms of contamination of fresh produce have remained unknown. On April 8, 2004, the National Public Health Institute of Finland was informed of several cases of gastroenteritis in schoolchildren in 1 municipality in northern Finland. On April 13, 2004, stool samples from symptomatic schoolchildren confirmed Y. pseudotuberculosis infections. At the same time, an increase occurred in Y. pseudotuberculosis cases reported to the National Infectious Disease Register (NIDR) from other parts of the country. We conducted epidemiologic, microbiologic, trace-back, and environmental investigations to determine the source of the outbreak and the origin of contamination. In the school outbreak, a survey concerning symptoms of gastrointestinal illness was conducted among all schoolchildren (7–18 years of age) and personnel (N = 900) of the 7 schools in the municipality. A case was defined as a laboratory-confirmed Y. pseudotuberculosis infection in a child or staff member who attended a school that received lunches from the school central kitchen, or as abdominal pain and fever, or erythema nodosum with illness onset from March 8 through March 28, 2004. Of respondents to the survey, 53 met the case definition. Among these respondents, Y. pseudotuberculosis was isolated from stool samples of 5 persons. A case–control study was conducted to identify the source of infection; self-administered questionnaires asked about consumption of items on school menus from March 8 through March 26, 2004. For each of the 53 case-patients identified in the survey, 3 controls were selected from the same class; 39 cases and 107 controls were included the analysis. Univariate analysis showed that a vegetable mixture of carrots and white cabbage served on March 8 and a mixture of cucumber and white cabbage served on March 25 were associated with illness. Multivariate analysis showed that only the carrot–white cabbage mixture was associated with illness. We also conducted a case–control study by mailing questionnaires to 37 persons with microbiologically confirmed Y. pseudotuberculosis infections reported to the NIDR from March 15 through May 7, 2004. These cases were from other parts of the country and were not associated with the school outbreak. For each case, 4 controls matched by age, sex, and municipality were randomly selected from the national population registry. Risk of illness increased with increased frequency of eating fresh carrots. Carrots served in the school were traced back to the farm level. Samples checked included grated carrots, which were available from the school kitchen. The kitchen had received all vegetables from 1 fresh-food processing plant. Samples were taken from the carrot-peeling line, carrot-peeling leftovers, grated carrots, and other vegetable-processing lines at the plant. Carrots originated from only 2 farms, which were inspected, and samples were obtained for bacteriologic examination. Small mammals at the farms were caught in carrot fields and investigated microbiologically to identify the reservoir of Y. pseudotuberculosis. This bacterium was isolated from 1 environmental sample from the carrot-peeling line in the fresh-food processing plant, from spoiled carrots, from fluid draining from spoiled carrots, and from a pooled sample of common shrew (Sorex araneus) intestines from 1 farm. Human and environmental isolates obtained were serotype O:1, subtype O:1b. Pulsed-field gel electrophoresis (PFGE) profiles of isolates from schoolchildren, fluid of spoiled carrots at the infected farm, and shrew intestines were indistinguishable. All 22 isolates from NIDR cases belonged to 2 PFGE genotypes. One genotype had a PFGE profile that was indistinguishable from the profile of the school outbreak isolates and the other genotype differed from these isolates by only 1 fragment. Our study provides microbiologic and epidemiologic evidence that the school outbreak was caused by carrots contaminated at the production farm. We isolated a Y. pseudotuberculosis subtype from human patients that was indistinguishable from isolates from the implicated source and a potential animal reservoir. Although the association between shrews and carrots is uncertain, shrews may have been picked up with carrots by harvesting machinery and ended up dead in wooden storage frames with the carrots. If carrots become contaminated, long storage at cold temperatures favors growth of Y. pseudotuberculosis and may result in human infections. Further studies are needed to determine the mechanism of contamination and other natural reservoirs. After the outbreak, the Finnish Food Safety Authority recommended controlling contamination at the farm level by removing spoiled carrots and paying attention to any subsequent spoilage during handling procedures.

SUMMARYIn 2010, 7/44 (16%) reported foodborne outbreaks in Finland were linked with raw beetroot consumption. We reviewed data from the national outbreak registry in order to hypothesize the aetiology of illness and to prevent further outbreaks. In the seven outbreaks, 124 cases among 623 respondents were identified. Consumption of raw beetroot was strongly associated with gastrointestinal illness (relative risk 8·99, 95% confidence interval 6·06–13·35). The illness was characterized by sudden onset of gastrointestinal symptoms; the median incubation time was 40 min and duration of illness 5 h. No common foodborne pathogens or toxins were found in either clinical or beetroot samples, but all tested beetroot samples were of poor quality according to total bacterial counts. Beta-haemolytic Pseudomonas fluorescens was detected in several beetroot samples but its effect on human health is unknown. No outbreaks were reported after the Finnish Food Safety Authority Evira advised against serving raw beetroot in institutional canteens.

The genetic diversity of Campylobacter jejuni strains isolated from Finnish patients was studied by typing 508 strains by pulsed-field gel electrophoresis: 311 were isolated from domestically acquired infections and 197 from travel-related infections. The strains were systematically selected from a larger collection of previously serotyped strains isolated during two 1-year sampling periods. The four most common SmaI profiles accounted for 45% of the domestic strains but only 3% of the travel-related strains. Of the domestic strains, 69% belonged to SmaI subtypes found during both sampling periods. The predominating SmaI subtypes and strains that were not digested by SmaI were typed by KpnI. Analyzing the temporal diversity of the pulsed-field gel electrophoresis profiles revealed six common persistent SmaI/KpnI subtypes among the domestic strains. Five of them have been identified in cattle, and two in chickens with a temporal association with human infections.

Statistical source attribution approaches of food-related zoonoses can generally be based on reported diagnosed human cases and surveillance results from different food sources or reservoirs of bacteria. The attribution model, or probabilistic classifier, can thus be based on the (sub)typing information enabling comparison between human infections and samples derived from source surveillance. Having time series of both data allows analyzing temporal patterns over time providing a repeated natural experiment. A Bayesian approach combining both sources of information over a long time series is presented in the case of Campylobacter in Finland and Norway. The full model is transparently presented and derived from the Bayes theorem. Previous statistical source attribution approaches are here advanced (1) by explicit modeling of the cases not associated with any of the sources under surveillance over time, (2) by modeling uncertain prevalence in a food source by bacteria type over time, and (3) by implementing formal model fit assessment using posterior predictive discrepancy functions. Large proportion of all campylobacteriosis can be attributed to broiler, but considerable uncertainty remains over time. The source attribution is inherently incomplete if only the sources under surveillance are included in the model. All statistical source attribution approaches should include a model fit assessment for judgment of model performance with respect to relevant quantities of interest. It is especially relevant when the model aims at a synthesis of several incomplete information sources under significant uncertainty of explanatory variables.

SUMMARYA survey of 788 pigs from 120 farms was conducted to determine the within-farm prevalence of pathogenicYersinia enterocoliticaand a questionnaire of management conditions was mailed to the farms afterwards. A univariate statistical analysis with carriage and shedding as outcomes was conducted with random-effects logistic regression with farm as a clustering factor. Variables with aPvalue Y. enterocoliticabetween farms.

To evaluate the effectiveness of bagging of the rectum in mitigating the contamination of carcasses with enteropathogenic Yersinia at the slaughterhouse and to estimate the hidden prevalences of these pathogens in different farm types and capacities, samples from pigs, carcasses, and slaughterhouse environment were collected, and a Bayesian probability model was constructed. In addition, the contamination routes were studied with molecular typing of the isolated strains. According to the model, bagging of the rectum reduced carcass contamination significantly with pathogenic Yersinia enterocolitica, but not with Yersinia pseudotuberculosis, and alone it was insufficient to completely prevent the carcass contamination with enteropathogenic Yersinia. The hidden prevalence of pathogenic Y. enterocolitica was higher at high production capacity than it was in low production capacity, but the 95 % credible intervals overlapped. Slaughterhouse environments can contaminate carcasses with enteropathogenic Yersinia, but the plausible main contamination source is the pig carrying the pathogen.

We describe the long-term multilocus sequence typing (MLST) analysis of the population structure and dynamics of 454 Finnish human Campylobacter jejuni isolates, as well as 208 chicken isolates, collected during the mid-1990s to 2007. The sequence type clonal complexes (ST CC) ST-45 CC, ST-21 CC, and ST-677 CC were the most common ones found among all isolates, and they covered 73.9% of all isolates. The ST-283 CC also was found frequently among chicken isolates (8.2%). The predominant STs among all isolates were ST-45, ST-50, and ST-677. ST-137 and ST-230 were common among human isolates, and ST-267 was found more frequently among chicken isolates than human isolates. The ST-45 CC was significantly associated with chicken isolates ( P < 0.01), whereas the ST-21 CC was associated with human isolates ( P < 0.001). The ST-677 CC was not associated with any host ( P = 0.5), and an opposite temporary trend of this complex was seen among chicken and human isolates, with an increase in the former and a decrease in the latter during the study period. Furthermore, the ST-22 and ST-48 CCs were significantly associated with human isolates ( P < 0.01), but neither of the CCs was found in chicken isolates. The annual overlap between STs from human and chicken isolates decreased from 76% at the beginning of the study to 58% at the end. Our results suggest that the importance of chicken as a reservoir for strains associated with human infections has declined despite the consumption of domestic chicken meat increasing during the follow-up period by 83%.

Background Campylobacter jejuni is the most common bacterial cause of human gastroenteritis worldwide. Due to the sporadic nature of infection, sources often remain unknown. Multilocus sequence typing (MLST) has been successfully applied to population genetics of Campylobacter jejuni and mathematical modelling can be applied to the sequence data. Here, we analysed the population structure of a total of 250 Finnish C. jejuni isolates from bovines, poultry meat and humans collected in 2003 using a combination of Bayesian clustering (BAPS software) and phylogenetic analysis. Results In the first phase we analysed sequence types (STs) of 102 Finnish bovine C. jejuni isolates by MLST and found a high diversity totalling 50 STs of which nearly half were novel. In the second phase we included MLST data from domestic human isolates as well as poultry C. jejuni isolates from the same time period. Between the human and bovine isolates we found an overlap of 72.2%, while 69% of the human isolates were overlapping with the chicken isolates. In the BAPS analysis 44.3% of the human isolates were found in bovine-associated BAPS clusters and 45.4% of the human isolates were found in the poultry-associated BAPS cluster. BAPS reflected the phylogeny of our data very well. Conclusions These findings suggest that bovines and poultry were equally important as reservoirs for human C. jejuni infections in Finland in 2003. Our results differ from those obtained in other countries where poultry has been identified as the most important source for human infections. The low prevalence of C. jejuni in poultry flocks in Finland could explain the lower attribution of human infection to poultry. Of the human isolates 10.3% were found in clusters not associated with any host which warrants further investigation, with particular focus on waterborne transmission routes and companion animals.

Aims: The aim of this study was to evaluate the efficiency of four isolation methods for the detection of pathogenic Yersinia enterocolitica from pig intestinal content. Methods and Results: The four methods comprised of 15 isolation steps using selective enrichments (irgasan–ticarcillin–potassium chlorate and modified Rappaport broth) and mildly selective enrichments at 4 or 25°C. Salmonella–Shigella-desoxycholate–calcium chloride agar, cefsulodin–irgasan–novobiocin agar were used as plating media. The most sensitive method detected 78% (53/68) of the positive samples. Individual isolation steps using cold enrichment as the only enrichment or as a pre-enrichment step with further selective enrichment showed the highest sensitivities (55–66%). All isolation methods resulted in high numbers of suspected colonies not confirmed as pathogenic Y. enterocolitica. Conclusions: Cold enrichment should be used in the detection of pathogenic Y. enterocolitica from pig intestinal contents. In addition, more than one parallel isolation step is needed. Significance and Impact of the Study: The study shows that depending on the isolation method used for Y. enterocolitica, the detected prevalence of Y. enterocolitica in pig intestinal contents varies greatly. More selective and sensitive isolation methods need to be developed for pathogenic Y. enterocolitica.

A substantial sampling among domestic human campylobacter cases, chicken process lots, and cattle at slaughter was performed during the seasonal peak of human infections. Campylobacter jejuni isolates ( n = 419) were subtyped using pulsed-field gel electrophoresis with SmaI, and isolates representing overlapping types ( n = 212) were further subtyped using KpnI for restriction. The SmaI/KpnI profiles of 55.4% (97/175) of the human isolates were indistinguishable from those of the chicken or cattle isolates. The overlapping SmaI/KpnI subtypes accounted for 69.8% (30/43) and 15.9% (32/201) of the chicken and cattle isolates, respectively. The occurrence of identical SmaI/KpnI subtypes with human C. jejuni isolates was significantly associated with animal host species ( P < 0.001). A temporal association of isolates from chickens and patients was possible in 31.4% (55/175) of the human infections. Besides chickens as sources of C. jejuni in the sporadic infections, the role of cattle appears notable. New approaches to restrict the occurrence of campylobacters in other farm animals may be needed in addition to hygienic measures in chicken production. However, only about half of the human infections were attributable to these sources.

The association of four new genetic markers with a chicken, bovine, or human host was studied among 645 Campylobacter jejuni isolates. The γ-glutamate transpeptidase gene and dmsA were common in human and chicken isolates but uncommon among bovine isolates. In the t test, bovine isolates differed significantly ( P < 0.05) from human and chicken isolates.

Background Streptococcus equi subspecies zooepidemicus is a rare infection in humans associated with contact with horses or consumption of unpasteurized milk products. On October 23, 2003, the National Public Health Institute was alerted that within one week three persons had been admitted to Tampere University Central Hospital (TaYS) because of S. equi subsp. zooepidemicus septicaemia. All had consumed fresh goat cheese produced in a small-scale dairy located on a farm. We conducted an investigation to determine the source and the extent of the outbreak. Methods Cases were identified from the National Infectious Disease Register. Cases were persons with S. equi subsp. zooepidemicus isolated from a normally sterile site who had illness onset 15.9-31.10.2003. All cases were telephone interviewed by using a standard questionnaire and clinical information was extracted from patient charts. Environmental and food specimens included throat swabs from two persons working in the dairy, milk from goats and raw milk tank, cheeses made of unpasteurized milk, vaginal samples of goats, and borehole well water. The isolates were characterized by ribotyping and pulsed-field gel electrophoresis (PFGE). Results Seven persons met the case definition; six had septicaemia and one had purulent arthritis. Five were women; the median age was 70 years (range 54–93). None of the cases were immunocompromized and none died. Six cases were identified in TaYS, and one in another university hospital in southern Finland. All had eaten goat cheese produced on the implicated farm. S. equi subsp. zooepidemicus was isolated from throat swabs, fresh goat cheese, milk tank, and vaginal samples of one goat. All human and environmental strains were indistinguishable by ribotyping and PFGE. Conclusion The outbreak was caused by goat cheese produced from unpasteurized milk. Outbreaks caused by S. equi subsp. zooepidemicus may not be detected if streptococcal strains are only typed to the group level. S. equi subsp. zooepidemicus may be a re-emerging disease if unpasteurized milk is increasingly used for food production. Facilities using unpasteurized milk should be carefully monitored to prevent this type of outbreaks.

In March 2004, the number of Yersinia pseudotuberculosis notifications increased suddenly in Finland compared to previous months. Overall, 125 cases were notified to the National Infectious Diseases Register between 1 January and 31 July 2004

The association of Penner heat-stable serotypes and pulsed-field gel electrophoresis genotypes of 208 human and 30 chicken Campylobacter jejuni isolates was studied. Overall, 46% of the human strains had overlapping sero- and genotype combinations with chicken strains. The percentage was reduced to 31% for strains that were considered temporally related. This suggests common environmental sources.

Background Waterborne Campylobacter jejuni outbreaks are common in the Nordic countries, and PFGE (pulsed field gel electrophoresis) remains the genotyping method of choice in outbreak investigations. However, PFGE cannot assess the clonal relationship between isolates, leading to difficulties in molecular epidemiological investigations. Here, we explored the applicability of whole genome sequencing to outbreak investigation by re-analysing three C. jejuni strains (one isolated from water and two from patients) from an earlier resolved Finnish waterborne outbreak from the year 2000. Results One of the patient strains had the same PFGE profile, as well as an identical overall gene synteny and three polymorphisms in comparison with the water strain. However, the other patient isolate, which showed only minor differences in the PFGE pattern relative to the water strain, harboured several polymorphisms as well as rearrangements in the integrated element CJIE2. We reconstructed the genealogy of these strains with ClonalFrame including in the analysis four C. jejuni isolated from chicken in 2012 having the same PFGE profile and sequence type as the outbreak strains. The three outbreak strains exhibited a paraphyletic relationship, implying that the drinking water from 2000 was probably contaminated with at least two different, but related, C. jejuni strains. Conclusions Our results emphasize the capability of whole genome sequencing to unambiguously resolve the clonal relationship between isolates of C. jejuni in an outbreak situation and evaluate the diversity of the C. jejuni population. Electronic supplementary material The online version of this article (doi:10.1186/1471-2164-15-768) contains supplementary material, which is available to authorized users.