Congenital Tooth Agenesis and Risk of Early-Onset Cancer.

Importance: There is some evidence that tooth agenesis (congenital absence of 1 or more teeth) is associated with cancer risk, especially carcinomas of the colon and ovaries, but results of previous studies are conflicting, and associations have not yet been evaluated in a population-based setting. Objective: To examine the association between tooth agenesis and specific cancer types before 40 years of age. Design, Setting, and Participants: This population-based cohort study used linking data from nationwide registries in Denmark to assess all Danish live-born singletons born from January 1, 1977, to December 31, 2018, and followed up for up to 40 years. Data were analyzed from January through June 2023. Exposure: Tooth agenesis as documented by the Danish Central Registry of Odontology (Danish municipal pediatric dental care) from January 1, 1988, to December 31, 2018, and from hospital encounters in the Danish National Patient Registry within the entire study period. Main Outcome and Measures: The primary outcome was first cancer diagnosis before 40 years of age obtained from the Danish Cancer Registry. Associations between tooth agenesis and specific cancers were estimated by Cox proportional hazards regression as hazard ratios (HRs) with 95% CIs. Analyses were split into age groups: younger than 1 year, 1 to younger than 3 years, 3 to younger than 10 years, 10 to younger than 20 years, 20 to younger than 30 years, and 30 to younger than 40 years. Associations with nonsyndromic tooth agenesis were evaluated after exclusion of individuals with known syndromes. Results: Among 2 501 715 included individuals (1 284 292 [51.3%] male), 70 288 (2.8%) had a diagnosis of tooth agenesis (mean [SD] age at diagnosis, 13.2 [4.1] years) and 26 308 (1.1%) had a diagnosis of early-onset cancer within the study period; 778 individuals had co-occurrence of tooth agenesis and cancer. Overall, tooth agenesis was positively associated with several cancer types, including neuroblastoma (age 1 to <3 years; HR, 4.20; 95% CI, 2.24-7.88), nephroblastoma (age 1 to <3 years; HR, 4.59; 95% CI, 2.37-8.91), hepatoblastoma (age 1 to <3 years; HR, 7.10; 95% CI, 2.70-18.68), osteosarcoma (age 10 to <20 years; HR, 2.19; 95% CI, 1.11-4.32), colorectal carcinomas (age 30 to <40 years; HR, 2.81; 95% CI, 1.38-5.71), and carcinomas of bladder (age 20 to <30 years; HR, 3.35; 95% CI, 1.35-8.30). Conclusions and Relevance: This cohort study found associations between congenital tooth agenesis and several cancer types, from childhood to early adulthood. Further evaluation of these associations is needed to assess possible clinical implications.

Early mortality in children with cancer in Denmark and Sweden: The role of social background in a setting with universal healthcare.

Socioeconomic differences in overall survival from childhood cancer have been shown previously, but the underlying mechanisms remain unclear. We aimed to investigate if social inequalities were seen already for early mortality in settings with universal healthcare. From national registers, all children diagnosed with cancer at ages 0-19 years, during 1991-2014, in Sweden and Denmark, were identified, and information on parental social characteristics was collected. We estimated odds ratios (OR) and 95% confidence intervals (CI) of early mortality (death within 90 days after cancer diagnosis) by parental education, income, employment, cohabitation, and country of birth using logistic regression. For children with acute lymphoblastic leukaemia (ALL), clinical characteristics were obtained. Among 13,926 included children, 355 (2.5%) died within 90 days after diagnosis. Indications of higher early mortality were seen among the disadvantaged groups, with the most pronounced associations observed for maternal education (ORadj_Low_vs_High 1.65 [95% CI 1.22-2.23]) and income (ORadj_Q1(lowest)_vs_Q4(highest) 1.77 [1.25-2.49]). We found attenuated or null associations between social characteristics and later mortality (deaths occurring 1-5 years after cancer diagnosis). In children with ALL, the associations between social factors and early mortality remained unchanged when adjusting for potential mediation by clinical characteristics. In conclusion, this population-based cohort study indicated differences in early mortality after childhood cancer by social background, also in countries with universal healthcare. Social differences occurring this early in the disease course requires further investigation, also regarding the timing of diagnosis.

The effect of changing diagnostic method from culture to PCR on the number of episodes of human campylobacteriosis in Denmark: a retrospective study (2015-2022).

IMPORTANCE: This study is important because it shows the potential epidemiological silence associated with the use of culture as the primary diagnostic method for the laboratory identification of human campylobacteriosis. Also, we show how polymerase chain reaction methods are associated with a systematic increase in the number of human campylobacteriosis episodes as reported by routine disease surveillance. These findings are operationally relevant and have public health implications because they tell how crucial it is to consider changes in diagnostic methods, e.g., in the epidemiological analysis of historical data and in the interpretation of future data in light of the past. We also believe that this study highlights how the synergy between microbiology and epidemiology is essential for disease surveillance.

Coping strategies and considerations regarding low anterior resection syndrome and quality of life among patients with rectal cancer; a qualitative interview study.

Introduction: Low anterior resection syndrome (LARS) is defined as disordered bowel function following rectal resection, which is detrimental to quality of life (QoL). A recent international consensus definition of LARS stresses the importance of focusing on both the symptoms and the consequences that the symptoms have for the individual patient as studies indicate that LARS has a negative impact on patients' QoL. However, an ongoing PROM study investigating late sequelae after rectal cancer finds that a minor proportion of patients scoring major LARS experience none or only little impact on quality of life. Aim: The aim of this study was to identify patients' considerations and coping strategies to establish why the burden caused by major LARS had little or no influence on their QoL. Materials and methods: This was a qualitative interview study based on 21 semi-structured individual telephone interviews with patients treated for rectal cancer. Data were analysed using a hermeneutic inspired thematic analysis. Results and conclusion: Three themes emerged from the analysis; Adapting new life situation, Altering life perception and the Importance of relationships. Major LARS and its consequences following rectal cancer may be managed or altered by adopting problem-focused and emotion-focused coping strategies. Maintaining a positive attitude and having a good network of family and friends constitute a surplus, allowing patients to cope with the need for changed behaviour and appreciate the life that they have been given. Accepting that major LARS and its consequences cause limitations in life allowed patients to change their normality threshold over time.

Long-Term Risk of Hospitalization for Somatic Diseases Among Survivors of Childhood Acute Lymphoblastic Leukemia.

BACKGROUND: Survivors of childhood acute lymphoblastic leukemia (ALL) may be at increased long-term risk of hospitalization for somatic diseases. However, large population-based cohort studies with risk estimates for survivors successfully cured without experiencing a relapse or requiring hematopoietic stem cell transplantation (HSCT) are lacking. METHODS: Danish and Swedish patients diagnosed with ALL before age 20 years in 1982-2008 were identified in the national cancer registries. Five-year survivors and matched population comparisons without childhood cancer were followed for hospitalization for 120 somatic disease categories in the national hospital registries from 5 years postdiagnosis until 2017, and disease-specific hospitalization rate ratios (RR) were calculated. The mean cumulative count method was used to estimate the mean number of multiple and recurrent disease-specific hospitalizations per individual. RESULTS: A total of 2024 5-year survivors and 9797 population comparisons were included. The overall hospitalization rate was more than twice as high compared with comparisons (RR = 2.30, 95% confidence interval [CI] = 2.09 to 2.52). At 30 years postdiagnosis, the mean cumulative hospitalization count was 1.69 (95% CI = 1.47 to 1.90) per survivor and 0.80 (95% CI = 0.73 to 0.86) per comparison. In the subcohort without relapse or HSCT (n = 1709), the RR was 1.41 (95% CI = 1.27 to 1.58). CONCLUSIONS: Survivors of childhood ALL were at increased long-term risk for disease-specific hospitalizations; however, in survivors without relapse or HSCT, the rate was only modestly higher than in population comparisons without a childhood cancer. The absolute mean numbers of multiple and recurrent hospitalizations were generally low.

Late mortality among survivors of childhood acute lymphoblastic leukemia diagnosed during 1971-2008 in Denmark, Finland, and Sweden: A population-based cohort study.

OBJECTIVE: Investigate all-cause and cause-specific late mortality after childhood acute lymphoblastic leukemia (ALL) in a population-based Nordic cohort. METHODS: From the cancer registries of Denmark, Finland, and Sweden, we identified 3765 five-year survivors of ALL, diagnosed before age 20 during 1971-2008. For each survivor, up to five matched comparison subjects were randomly selected from the general population (n = 18,323). Causes of death were classified as relapse related, health related, and external. Late mortality was evaluated by cumulative incidences of death from 5-year survival date. Mortality hazard ratios (HR) were evaluated with Cox proportional models. RESULTS: Among the survivors, 315 deaths occurred during a median follow-up of 16 years from 5-year survival date (range 0-42). The majority were attributable to relapse (n = 224), followed by second neoplasm (n = 45). Cumulative incidence of all-cause late mortality at 15 years from diagnosis decreased gradually over treatment decades, from 14.4% (95% confidence interval [CI]: 11.6-17.2) for survivors diagnosed during 1971-1981, to 2.5% (95% CI: 1.3-3.7) for those diagnosed during 2002-2008. This was mainly attributable to a reduction in relapse-related deaths decreasing from 13.4% (95% CI: 10.7-16.1) for survivors diagnosed during 1971-1981 to 1.9% (95% CI: 0.9-2.8) for those diagnosed during 2002-2008. Health-related late mortality was low and did not change substantially across treatment decades. Compared to comparison subjects, all-cause mortality HR was 40 (95% CI: 26-61) 5-9 years from diagnosis, and 4.4 (95% CI: 3.4-5.6) ≥10 years from diagnosis. CONCLUSIONS: Survivors of ALL have higher late mortality than population comparison subjects. Among the survivors, there was a temporal reduction in risk of death from relapse, without increments in health-related death.

Sentinel surveillance and epidemiology of Clostridioides difficile in Denmark, 2016 to 2019.

BackgroundSince 2008, Danish national surveillance of Clostridioides difficile has focused on binary toxin-positive strains in order to monitor epidemic types such as PCR ribotype (RT) 027 and 078. Additional surveillance is needed to provide a more unbiased representation of all strains from the clinical reservoir.AimSetting up a new sentinel surveillance scheme for an improved understanding of type distribution relative to time, geography and epidemiology, here presenting data from 2016 to 2019.MethodsFor 2─4 weeks in spring and autumn each year between 2016 and 2019, all 10 Danish Departments of Clinical Microbiology collected faecal samples containing toxigenic C. difficile. Isolates were typed at the national reference laboratory at Statens Serum Institut. The typing method in 2016-17 used tandem-repeat-sequence typing, while the typing method in 2018-19 was whole genome sequencing.ResultsDuring the study period, the sentinel surveillance scheme included ca 14-15% of all Danish cases of C. difficile infections. Binary toxin-negative strains accounted for 75% and 16 of the 20 most prevalent types. The most common sequence types (ST) were ST2/13 (RT014/020) (19.5%), ST1 (RT027) (10.8%), ST11 (RT078) (6.7%), ST8 (RT002) (6.6%) and ST6 (RT005/117) (5.1%). The data also highlighted geographical differences, mostly related to ST1 and temporal decline of ST1 (p = 0.0008) and the increase of ST103 (p = 0.002), ST17 (p = 0.004) and ST37 (p = 0.003), the latter three binary toxin-negative.ConclusionSentinel surveillance allowed nationwide monitoring of geographical differences and temporal changes in C. difficile infections in Denmark, including emerging types, regardless of binary toxin status.

Combining Salmonella Dublin genome information and contact-tracing to substantiate a new approach for improved detection of infectious transmission routes in cattle populations.

This study presents a new method for detection of between-herd livestock movements to facilitate disease tracing and more accurately describe network behaviour of relevance for spread of infectious diseases, including within-livestock business risk-carrying contacts that are not necessarily recorded anywhere. The study introduces and substantiates the concept of grouping livestock herds into business-units based on ownership and location in the tracing analysis of animal movement-based contact networks. To test the utility of this approach, whole core genome sequencing of 196 Salmonella Dublin isolates stored from previous surveillance and project activities was combined with information on cattle movements recorded in the Danish Cattle Database between 1997 and 2017. The aim was to investigate alternative explanations for S. Dublin circulation in groups of herds connected by ownership, but without complete records of livestock movements. The EpiContactTrace R-package was used to trace the contact networks between businesses and compare the network characteristics of businesses sharing strains of S. Dublin with different levels of genetic relatedness. The ownership-only definition proved to be an unreliable grouping approach for large businesses, which could have internal distances larger than 250 km and therefore do not represent useful epidemiological units. Therefore, the grouping was refined using spatial analysis. More than 90% of final business units formed were composed of one single cattle property, whereas multi-property businesses could reach up to eight properties in a given year, with up to 15 cattle herds having been part of the same business through the study period. Results showed markedly higher probabilities of introduction of infectious animals between proposed businesses from which the same clone of S. Dublin had been isolated, when compared to businesses with non-related strains, thus substantiating the business-unit as an important epidemiological feature to consider in contact network analysis and tracing of infection routes. However, this approach may overestimate real-life contacts between cattle properties and putatively overestimate the degree of risk-contacts within each business, since it is based solely on information about property ownership and location. This does not consider administrative and individual farmers behaviours that essentially keep two properties separated. Despite this, we conclude that defining epidemiological units based on businesses is a promising approach for future disease tracing tasks.

Epidemiology of Salmonella enterica Serovar Dublin in Cattle and Humans in Denmark, 1996 to 2016: a Retrospective Whole-Genome-Based Study.

Salmonella enterica serovar Dublin is a cattle-adapted S. enterica serovar causing both intestinal and systemic infection in its bovine host, and it is also a serious threat to human health. The present study aimed to determine the population structure of S Dublin isolates obtained from Danish cattle herds and to investigate how cattle isolates relate to Danish human isolates, as well as to non-Danish human and bovine isolates. Phylogenetic analysis of 197 Danish cattle isolates from 1996 to 2016 identified three major clades corresponding to distinct geographical regions of cattle herds. Persistence of closely related isolates within the same herd and their circulation between epidemiologically linked herds for a period of more than 20 years were demonstrated. These findings suggest that a lack of internal biosecurity and, to some extent, also a lack of external biosecurity in the herds have played an important role in the long-term persistence of S Dublin in Danish cattle herds in the period investigated. Global population analysis revealed that Danish cattle isolates clustered separately from bovine isolates from other countries, whereas human isolates were geographically spread. Resistance genes were not commonly demonstrated in Danish bovine isolates; only the isolates within one Danish clade were found to often harbor two plasmids of IncFII/IncFIB and IncN types, the latter plasmid carrying blaTEM-1, tetA, strA, and strB antibiotic resistance genes.IMPORTANCES Dublin causes economic losses in cattle production, and the bacterium is a public health concern. A surveillance and control program has been in place in Denmark since 2002 with the ultimate goal to eradicate S Dublin from Danish cattle herds; however, a small proportion of herds have remained positive for many years. In this study, we demonstrate that herds with persistent infection often were infected with the same strain for many years, indicating that internal biosecurity has to be improved to curb the infection. Further, domestic cases of S Dublin infection in humans were found to be caused both by Danish cattle isolates and by isolates acquired abroad. This study shows the strength of whole-genome sequencing to obtain detailed information on epidemiology of S Dublin and allows us to suggest internal biosecurity as a main way to control this bacterium in Danish cattle herds.

WGS based study of the population structure of Salmonella enterica serovar Infantis.

BACKGROUND: Salmonella Infantis (S. Infantis) is one of the most frequent Salmonella serovars isolated from human cases of salmonellosis and the most detected serovar from animal and food sources in Europe. The serovar is commonly associated with poultry and there is increasing concern over multidrug resistant clones spreading worldwide, as the dominating clones are characterized by presence of large plasmids carrying multiple resistance genes. Increasing the knowledge of the S. Infantis population and evolution is important for understanding and preventing further spread. In this study, we analysed a collection of strains representing different decades, sources and geographic locations. We analysed the population structure and the accessory genome, in particular we identified prophages with a view to understand the role of prophages in relation to the evolution of this serovar. RESULTS: We sequenced a global collection of 100 S. Infantis strains. A core-genome SNP analysis separated five strains in e-Burst Group (eBG) 297 with a long branch. The remaining strains, all in eBG31, were divided into three lineages that were estimated to have separated approximately 150 years ago. One lineage contained the vast majority of strains. In five of six clusters, no obvious correlation with source or geographical locations was seen. However, one cluster contained mostly strains from human and avian sources, indicating a clone with preference for these sources. The majority of strains within this cluster harboured a pESI-like plasmid with multiple resistance genes. Another lineage contained three genetic clusters with more rarely isolated strains of mainly animal origin, possibly less sampled or less infectious clones. Conserved prophages were identified in all strains, likely representing bacteriophages which integrated into the chromosome of a common ancestor to S. Infantis. We also saw that some prophages were specific to clusters and were probably introduced when the clusters were formed. CONCLUSIONS: This study analysed a global S. Infantis population and described its genetic structure. We hypothesize that the population has evolved in three separate lineages, with one more successfully emerging lineage. We furthermore detected conserved prophages present in the entire population and cluster specific prophages, which probably shaped the population structure.

Cross-Border Transmission of Salmonella Choleraesuis var. Kunzendorf in European Pigs and Wild Boar: Infection, Genetics, and Evolution.

Salmonella enterica subspecies enterica serotype Choleraesuis is a swine adapted serovar. S. Choleraesuis variant Kunzendorf is responsible for the majority of outbreaks among pigs. S. Choleraesuis is rare in Europe, although there have been serious outbreaks in pigs including two outbreaks in Denmark in 1999-2000 and 2012-2013. Here, we elucidate the epidemiology, possible transmission routes and sources, and clonality of European S. Choleraesuis isolates including the Danish outbreak isolates. A total of 102 S. Choleraesuis isolates from different European countries and the United States, covering available isolates from the last two decades were selected for whole genome sequencing. We applied a temporally structured sequence analysis within a Bayesian framework to reconstruct a temporal and spatial phylogenetic tree. MLST type, resistance genes, plasmid replicons, and accessory genes were identified using bioinformatics tools. Fifty-eight isolates including 11 out of 12 strains from wild boars were pan-susceptible. The remaining isolates carried multiple resistance genes. Eleven different plasmid replicons in eight plasmids were determined among the isolates. Accessory genes were associated to the identified resistance genes and plasmids. The European S. Choleraesuis was estimated to have emerged in ∼1837 (95% credible interval, 1733-1983) with the mutation rate of 1.02 SNPs/genome/year. The isolates were clustered according to countries and neighbor countries. There were transmission events between strains from the United States and European countries. Wild boar and pig isolates were genetically linked suggesting cross-border transmission and transmission due to a wildlife reservoir. The phylogenetic tree shows that multiple introductions were responsible for the outbreak of 2012-2013 in Denmark, and suggests that poorly disinfected vehicles crossing the border into Denmark were potentially the source of the outbreak. Low levels of single nucleotide polymorphisms (SNPs) differences (0-4 SNPs) can be observed between clonal strains isolated from different organs of the same animal. Proper disinfection of livestock vehicles and improved quality control of livestock feed could help to prevent future spread of S. Choleraesuis or other more serious infectious diseases such as African swine fever (ASF) in the European pig production system.

Long-Term Risk of Hospitalization Among Five-Year Survivors of Childhood Leukemia in the Nordic Countries.

BACKGROUND: Adverse effects from childhood leukemia treatment may persist or present years after cure from cancer. We provide a comprehensive evaluation of subsequent hospitalization in five-year survivors of childhood acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), and chronic myeloid leukemia (CML). METHODS: In the Adult Life after Childhood Cancer in Scandinavia Study, we identified 4003 five-year survivors diagnosed with childhood leukemia 1970-2008 in Denmark, Sweden, Iceland, and Finland. Survivors and 129 828 population comparisons were followed for first-time nonpsychiatric hospitalizations for 120 disease categories in the hospital registries. Standardized hospitalization rate ratios and absolute excess rates were calculated. All statistical tests were two-sided. RESULTS: Survivors of ALL (n = 3391), AML (n = 389), and CML (n = 92) had an increased overall hospitalization rate compared with population comparisons. The rate ratio for any hospitalization was 1.95 (95% confidence interval [CI] = 1.83 to 2.07) in ALL, 3.09 (95% CI = 2.53 to 3.65) in AML, and 4.51 (95% CI = 3.03 to 6.00) in CML survivors and remained increased even 20 years from leukemia diagnosis. Corresponding absolute excess rates per 1000 person-years were 28.48 (95% CI = 24.96 to 32.00), 62.75 (95% CI = 46.00 to 79.50), and 105.31 (95% CI = 60.90 to 149.72). CONCLUSION: Leukemia survivors have an increased rate of hospitalization for medical conditions. We provide novel insight into the relative and absolute rate of hospitalization for 120 disease categories in survivors of ALL, AML, and CML, which are likely to be informative for both survivors and healthcare providers.

Use of multiple-locus variable-number of tandem repeats analysis (MLVA) to investigate genetic diversity of Salmonella enterica subsp. enterica serovar Typhimurium isolates from human, food, and veterinary sources.

Salmonella enterica subspecies enterica serovar Typhimurium is the most common zoonotic pathogen in Bulgaria. To allow efficient outbreak investigations and surveillance in the food chain, accurate and discriminatory methods for typing are needed. This study evaluated the use of multiple-locus variable-number of tandem repeats analysis (MLVA) and compared results with antimicrobial resistance (AMR) determinations for 100 S. Typhimurium strains isolated in Bulgaria during 2008-2012 (50 veterinary/food and 50 human isolates). Results showed that isolates were divided into 80 and 34 groups using MLVA and AMR, respectively. Simpson's index of diversity was determined to 0.994 ± 0.003 and 0.945 ± 0.012. The most frequently encountered MLVA profiles were 3-11-9-NA-211 (n = 5); 3-12-9-NA-211 (n = 3); 3-12-11-21-311 (n = 3); 3-17-10-NA-311 (n = 3); 2-20-9-7-212 (n = 3); and 2-23-NA-NA-111 (n = 3). No clustering of isolates related to susceptibility/resistance to antimicrobials, source of isolation, or year of isolation was observed. Some MLVA types were found in both human and veterinary/food isolates, indicating a possible route of transmission. A majority (83%) of the isolates were found to be resistant against at least one antimicrobial and 44% against ≥4 antimicrobials. Further studies are needed to verify MLVA usefulness over a longer period of time and with more isolates, including outbreak strains.

Investigation of Outbreaks of Salmonella enterica Serovar Typhimurium and Its Monophasic Variants Using Whole-Genome Sequencing, Denmark.

Whole-genome sequencing is rapidly replacing current molecular typing methods for surveillance purposes. Our study evaluates core-genome single-nucleotide polymorphism analysis for outbreak detection and linking of sources of Salmonella enterica serovar Typhimurium and its monophasic variants during a 7-month surveillance period in Denmark. We reanalyzed and defined 8 previously characterized outbreaks from the phylogenetic relatedness of the isolates, epidemiologic data, and food traceback investigations. All outbreaks were identified, and we were able to exclude unrelated and include additional related human cases. We were furthermore able to link possible food and veterinary sources to the outbreaks. Isolates clustered according to sequence types (STs) 19, 34, and 36. Our study shows that core-genome single-nucleotide polymorphism analysis is suitable for surveillance and outbreak investigation for Salmonella Typhimurium (ST19 and ST36), but whole genome-wide analysis may be required for the tight genetic clone of monophasic variants (ST34).

Investigating Salmonella Eko from Various Sources in Nigeria by Whole Genome Sequencing to Identify the Source of Human Infections.

Twenty-six Salmonella enterica serovar Eko isolated from various sources in Nigeria were investigated by whole genome sequencing to identify the source of human infections. Diversity among the isolates was observed and camel and cattle were identified as the primary reservoirs and the most likely source of the human infections.

Whole-genome Sequencing Used to Investigate a Nationwide Outbreak of Listeriosis Caused by Ready-to-eat Delicatessen Meat, Denmark, 2014.

BACKGROUND: Listeriosis is a serious foodborne infection. Outbreaks of listeriosis occur rarely, but have often proved difficult to solve. In June 2014, we detected and investigated a listeriosis outbreak in Denmark using patient interviews and whole-genome sequencing (WGS). METHODS: We performed WGS on Listeria monocytogenes isolates from patients and available isolates from ready-to-eat foods and compared them using single-nucleotide polymorphism (SNP) analysis. Case patients had L. monocytogenes with ≤3 SNPs (the outbreak strain) isolated in September 2013-December 2014. Through interviews, we established case patients' food and clinical histories. Food production facilities were inspected and sampled, and we performed trace-back/trace-forward of food delivery chains. RESULTS: In total, 41 cases were identified; 17 deaths occurred (41%). An isolate from a delicatessen meat (spiced meat roll) from company A was identical to the outbreak strain. Half of the patients were infected while hospitalized/institutionalized; institutions were supplied food by company A. The outbreak strain was repeatedly isolated from further samples taken within this company and within companies in its distribution chain. Products from company A were traced and recalled from >6000 food establishments, after which the outbreak ended. CONCLUSIONS: Ready-to-eat spiced meat roll from a single production facility caused this outbreak. The product, served sliced and cold, is popular among the elderly; serving it at hospitals probably contributed to the high case-fatality rate. WGS used for patient isolates and isolates from food control inspections, coupled with routine epidemiological follow-up, was instrumental in swiftly locating the source of infections, preventing further illnesses and deaths.

Application of Molecular Typing Results in Source Attribution Models: The Case of Multiple Locus Variable Number Tandem Repeat Analysis (MLVA) of Salmonella Isolates Obtained from Integrated Surveillance in Denmark.

Salmonella is an important cause of bacterial foodborne infections in Denmark. To identify the main animal-food sources of human salmonellosis, risk managers have relied on a routine application of a microbial subtyping-based source attribution model since 1995. In 2013, multiple locus variable number tandem repeat analysis (MLVA) substituted phage typing as the subtyping method for surveillance of S. Enteritidis and S. Typhimurium isolated from animals, food, and humans in Denmark. The purpose of this study was to develop a modeling approach applying a combination of serovars, MLVA types, and antibiotic resistance profiles for the Salmonella source attribution, and assess the utility of the results for the food safety decisionmakers. Full and simplified MLVA schemes from surveillance data were tested, and model fit and consistency of results were assessed using statistical measures. We conclude that loci schemes STTR5/STTR10/STTR3 for S. Typhimurium and SE9/SE5/SE2/SE1/SE3 for S. Enteritidis can be used in microbial subtyping-based source attribution models. Based on the results, we discuss that an adjustment of the discriminatory level of the subtyping method applied often will be required to fit the purpose of the study and the available data. The issues discussed are also considered highly relevant when applying, e.g., extended multi-locus sequence typing or next-generation sequencing techniques.

Outbreak of Salmonella enterica serovar Typhimurium phage type DT41 in Danish poultry production.

Salmonella enterica subspecies enterica serovar Typhimurium (S. Typhimurium) is one of the most prevalent serovars in Europe - where both poultry and poultry related products are common sources of human salmonellosis. Due to efficient control programs, the prevalence of S. Typhimurium in Danish poultry production is very low. Despite this, during the past decades there has been a reoccurring problem with infections with S. Typhimurium phage type DT41 in the Danish poultry production without identifying a clear source. In the end of 2013 and beginning of 2014 an increased isolation of S. Typhimurium DT41 was noted mainly in this production, but also in other samples. To investigate this is in more detail, 47 isolates from egg layers (n=5, 1 flock), broilers (n=33, 13 flocks), broiler breeding flocks and hatches (n=5; 2 flocks and 1 environmental hatchery sample), feed (n=1), poultry slaughter house (n=3, environmental sample and meat) were typed with multi locus variable number of tandem repeat analysis (MLVA) and pulsed-field gel electrophoresis (PFGE) to investigate the epidemiology of the outbreak. Based on PFGE results isolates were divided into four groups (Simpson's index of diversity (DI)=0.24±0.15). Due to the low DI, PFGE was not sufficient to provide information to unravel the outbreak. Based on MLVA typing the DT41 (42/47 isolates) and the RDNC isolates (5/47) were split into nine groups (DI=0.65±0.14). When a maximum divergence at one locus was permitted these could be gathered into four groups. Using this criterion, combined with epidemiological information, a spread of one type from broiler breeders to broilers and further to the poultry slaughter house was plausible. In conclusion, although it could be concluded that a spread within the broiler production pyramid had taken place the source of the sudden increase of S. Typhimurium DT41 remains unclear. To investigate this in more detail, further studies using whole genome sequencing to obtain a higher discriminatory strength and including isolates from a longer period of time and from various sources are in progress.

Reappearance of Salmonella serovar Choleraesuis var. Kunzendorf in Danish pig herds.

Salmonella enterica serovar Choleraesuis is a porcine adapted serovar which may cause serious outbreaks in pigs. Here we describe outbreaks of salmonellosis due to S. Choleraesuis in four Danish pig farms in 2012-2013 by clinic, serology, and microbiology and compare the isolates to those of a previous outbreak in 1999-2000. The infection was in some herds associated with high mortality and a moderate to high sero-prevalence was found. In 2012-2013 the disease contributed to increased mortality but occurred concomitant with other disease problems in the herds, which likely delayed the diagnosis by up to several months. Nine isolates from the four farms in 2012-2013 and 14 isolates obtained from the outbreak in Denmark in 1999-2000 were subjected to typing using pulsed-field gel electrophoresis (PFGE). Seven isolates were selected for whole genome sequencing (WGS). The PFGE results of 23 isolates displayed five different profiles. The isolates from 2012 to 2013 revealed two distinct profiles, both different from the isolates recovered in 1999-2000. Two of the 2012-2013 farms shared PFGE profiles and had also transported pigs between them. The profile found in the two other 2012-2013 farms was indistinguishable but no epidemiological connection between these farms was found. Analysis of the number of single nucleotide polymorphisms (SNPs) from the WGS data indicated that the isolates from the farms in 2012-2013 were more closely related to each other than to isolates from the outbreak in 1999. It was therefore concluded that the infection was a new introduction and not a persistent infection since the outbreak in 1999. It may further be suggested that there were two or three independent rather than a single introduction. The re-introduction of S. Choleraesuis in Denmark emphasizes the importance of strict hygiene measures in the herds. Further investigations using WGS are now in progress on a larger collection of isolates to study clonality at European level and trace the origin of the infections.