Epidemiology of gastrointestinal infections: lessons learned from syndromic testing, Region Zealand, Denmark.

The aim of this study was to investigate the value of syndromic diagnostic testing for a better understanding of the epidemiology of gastrointestinal infections in Denmark. Here we evaluated the QIAstat-Dx® Gastrointestinal (GI) Panel 1 assay on 18,610 fecal samples requested for analysis for enteric pathogens in Region Zealand, Denmark, in 1 year (October 1, 2021, to September 30, 2022). In total, 6905 (37%) samples were detected positive for one or more diarrhoeal bacteria, viruses, and protozoa. The most common bacterial, viral, and parasitic pathogens detected with the QIAstat-Dx® Gastrointestinal Panel 1 were EPEC (in patients ≥ 2 years of age) (n = 1420 (20.6%)), rotavirus (n = 948 (13.7%)), and Cryptosporidium spp. (n = 196 (2.84%)). We identified a large diversity in infections likely reflecting substantial differences in the epidemiology including origin of infections, mode of transmission, seasonality, age-dependent susceptibility to disease, severity, and travel history. All pathogens were detected as both single and coinfections. Viral infections peaked in March with a positive rate of 31.6%, and bacterial infections peaked in August with a positive rate of 35.3%. ETEC, Shigella/EIEC, EAEC, and P. shigelloides were most related to travel activity, and coinfections were frequent. The distribution of Ct values varied significantly between the pathogens, with the lowest Ct values (median 17-18) observed in astrovirus, adenovirus, and rotavirus. Our results highlight the value of providing extensive diagnostic testing on fecal samples for sufficient detection of relevant diarrhoeal pathogens for optimal clinical care.

Imported spring onions related to the first recorded outbreak of enteroinvasive Escherichia coli in Denmark, November to December 2021.

Between November and December 2021, the first ever recorded outbreak of enteroinvasive Escherichia coli in Denmark occurred at national scale. We describe the investigation of this outbreak, which was initially recognised in early December 2021. A total of 88 cases (58 female; 30 male) with a median age of 52 years (range: 0-91) were detected by PCR-based diagnostic methods. Case ascertainment was complicated by current culture-free diagnostic procedures, with only 34 cases confirmed by culture, serotyping and whole genome sequencing. Isolates from cases grouped into two serotypes (O136:H7 and O96:H19), which was supported by whole-genome-sequence-phylogeny, also yielding two clusters. Interviews of 42 cases and traceback investigation pointed towards consumption of ready-to-eat salads as the outbreak cause. While the ready-to-eat salads comprised different vegetables, imported spring onions were the only common ingredient and thus the likely source. Environmental investigations failed to recover outbreak strains. This report highlights the value of fast typing (here O-typing) to confirm cases in an outbreak situation. Timely communication and data sharing are also important, and were facilitated by the national collaboration between relevant laboratories, the public health institute and the veterinary and food administration. High hygiene standards for imported fresh vegetables intended for ready-to-eat products are essential.

The incidence of laboratory-confirmed cases of enteric pathogens in Denmark 2018: a national observational study.

BACKGROUND: Only a subset of enteric pathogens is under surveillance in Denmark, and knowledge on the remaining pathogens detected in acute gastroenteritis is limited. Here, we present the one-year incidence of all enteric pathogens diagnosed in Denmark, a high-income country, in 2018 and an overview of diagnostic methods used for detection. METHODS: All 10 departments of clinical microbiology completed a questionnaire on test methods and provided 2018-data of persons with positive stool samples with Salmonella species, Campylobacter jejuni/coli, Yersinia enterocolitica, Aeromonas species, diarrheagenic Escherichia coli (Enteroinvasive (EIEC), Shiga toxin-producing (STEC), Enterotoxigenic (ETEC), Enteropathogenic (EPEC), and intimin-producing/attaching and effacing (AEEC)), Shigella species., Vibrio cholerae, norovirus, rotavirus, sapovirus, adenovirus, Giardia intestinalis, Cryptosporidium species, and Entamoeba histolytica. RESULTS: Enteric bacterial infections were diagnosed with an incidence of 229.9 cases/100,000 inhabitants, virus had an incidence of 86/100,000 and enteropathogenic parasites of 12.5/100,000. Viruses constituted more than half of diagnosed enteropathogens for children below 2 years and elderly above 80 years. Diagnostic methods and algorithms differed across the country and in general PCR testing resulted in higher incidences compared to culture (bacteria), antigen-test (viruses), or microscopy (parasites) for most pathogens. CONCLUSIONS: In Denmark, the majority of detected infections are bacterial with viral agents primarily detected in the extremes of ages and with few intestinal protozoal infections. Incidence rates were affected by age, clinical setting and local test methods with PCR leading to increased detection rates. The latter needs to be taken into account when interpreting epidemiological data across the country.

Correlation of High Seawater Temperature with Vibrio and Shewanella Infections, Denmark, 2010-2018.

During 2010-2018 in Denmark, 638 patients had Vibrio infections diagnosed and 521 patients had Shewanella infections diagnosed. Most cases occurred in years with high seawater temperatures. The substantial increase in those infections, with some causing septicemia, calls for clinical awareness and mandatory notification policies.

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.

Verification of analytical bacterial spectrum of QIAstat-Dx® GI V2 and Novodiag® Bacterial GE+ V2-0 diagnostic panels.

BACKGROUND: Implementing multiplex PCR or syndromic panel-based testing platforms to detect microbial species that cause acute diarrhoea may guide patient management more effectively and efficiently. OBJECTIVES: To assess and compare the performance of two syndromic panel-based testing systems, QIAstat-Dx® Gastrointestinal Panel V2 (QGI) and the Novodiag® Bacterial GE+ V2-0 (NGE). METHODS: The QGI and NGE panels include 16 and 14 bacterial gastrointestinal pathogens, respectively. The performance of the panels was tested retrospectively using 141 positive clinical stool specimens, External Quality Assessment (EQA) panels and spiked faecal specimens. RESULTS: For Campylobacter jejuni and coli (n = 20), Salmonella (n = 24), Shigella (n = 13), Yersinia enterocolitica (non-1A biotypes) (n = 8), Clostridioides difficile (n = 24) and Vibrio parahaemolyticus (n = 2), QGI correctly verified 19/20, 20/24, 13/13, 8/8, 23/24 and 2/2, whereas NGE correctly verified 20/20, 17/24, 13/13, 8/8, 14/24 and 1/2. Among diarrhoeagenic Escherichia coli (n = 29), QGI reported one Shiga toxin-producing E. coli (STEC) stx1a O26:H11 as STEC serotype O157:H7 and NGE failed on one enteropathogenic E. coli, one enteroaggregative E. coli and one STEC (stx2e). Y. enterocolitica biotype 1A (non-pathogenic) (n = 6) were all positive in QGI, but negative in NGE. CONCLUSIONS: Both QGI and NGE testing panels can improve laboratory workflow and patient management by providing user-friendly platforms that can rapidly detect a number of targets with one specimen. QGI was significantly more sensitive in identifying C. difficile. Both methods had suboptimal detection of Salmonella and this needs to be examined further. The short hands-on time and turnaround time are of value for on-demand testing and use in a high-throughput setting.

Danish national guideline for the treatment of Clostridioides difficile infection and use of faecal microbiota transplantation (FMT).

Aim: This Danish national guideline describes the treatment of adult patients with Clostridioides (formerly Clostridium) difficile (CD) infection and the use of faecal microbiota transplantation (FMT). It suggests minimum standard for implementing an FMT service.Method: Four scientific societies appointed members for a working group which conducted a systematic literature review and agreed on the text and recommendations. All clinical recommendations were evalluated for evidence level and grade of recommendation.Results: In CD infection, the use of marketed and experimental antibiotics as well as microbiota-based therapies including FMT are described. An algorithm for evaluating treatment effect is suggested. The organisation of FMT, donor recruitment and screening, laboratory preparation, clinical application and follow-up are described.Conclusion: Updated evidence for the treatment of CD infection and the use of FMT is provided.

Whole genome sequencing data used for surveillance of Campylobacter infections: detection of a large continuous outbreak, Denmark, 2019.

BackgroundCampylobacter is one of the most frequent causes of bacterial gastroenteritis. Campylobacter outbreaks are rarely reported, which could be a reflection of a surveillance without routine molecular typing. We have previously shown that numerous small outbreak-like clusters can be detected when whole genome sequencing (WGS) data of clinical Campylobacter isolates was applied.AimTyping-based surveillance of Campylobacter infections was initiated in 2019 to enable detection of large clusters of clinical isolates and to match them to concurrent retail chicken isolates in order to react on ongoing outbreaks.MethodsWe performed WGS continuously on isolates from cases (n = 701) and chicken meat (n = 164) throughout 2019. Core genome multilocus sequence typing was used to detect clusters of clinical isolates and match them to isolates from chicken meat.ResultsSeventy-two clusters were detected, 58 small clusters (2-4 cases) and 14 large clusters (5-91 cases). One third of the clinical isolates matched isolates from chicken meat. One large cluster persisted throughout the whole year and represented 12% of all studied Campylobacter cases. This cluster type was detected in several chicken samples and was traced back to one slaughterhouse, where interventions were implemented to control the outbreak.ConclusionOur WGS-based surveillance has contributed to an improved understanding of the dynamics of the occurrence of Campylobacter strains in chicken meat and the correlation to clusters of human cases.

Randomised clinical trial: a 12-strain bacterial mixture versus faecal microbiota transplantation versus vancomycin for recurrent Clostridioides difficile infections.

BACKGROUND: A defined bacterial mixture could be a safer alternative to faecal microbiota transplantation (FMT). AIMS: To compare the efficacy of a 12-strain mixture termed rectal bacteriotherapy with either FMT or vancomycin for recurrent Clostridioides difficile infection (CDI) in an open-label 3-arm randomised controlled trial. METHODS: We screened all individuals positive for C difficile from May 2017 to March 2019. Persons with laboratory-confirmed recurrent CDI were included. Before FMT and rectal bacteriotherapy, we pre-treated with vancomycin for 7-14 days. Rectal bacteriotherapy was applied by enema on three consecutive days and FMT by enema once with possible repetition for two to three infusions within 14 days. The vancomycin group was treated for 14 days with additional five weeks of tapering for multiple recurrences. The primary outcome was clinical cure within 90 days. A secondary outcome was 180-day all-cause mortality. RESULTS: Participants in the FMT group (n = 34) were cured more often than participants receiving vancomycin (n = 31), 76% vs 45% (OR 3.9 (1.4-11.4), P < 0.01) or rectal bacteriotherapy (n = 31), 76% vs 52% (OR 3.0 (1.1-8.8), P = 0.04). Rectal bacteriotherapy and vancomycin performed similarly (P = 0.61). The mortality rate was 6% in the FMT group, 13% in the bacteriotherapy group and 23% in the vancomycin group. FMT tended to reduce mortality compared with vancomycin, OR 0.2 (0.04-1.12), P = 0.07. CONCLUSIONS: Rectal bacteriotherapy appears as effective as vancomycin but less effective than 1-3 FMTs. FMT by enema with 1-3 infusions is superior to vancomycin for treating recurrent C difficile infections and might reduce mortality.

Whole-Genome Sequencing to Detect Numerous Campylobacter jejuni Outbreaks and Match Patient Isolates to Sources, Denmark, 2015-2017.

In industrialized countries, the leading cause of bacterial gastroenteritis is Campylobacter jejuni. However, outbreaks are rarely reported, which may reflect limitations of surveillance, for which molecular typing is not routinely performed. To determine the frequency of genetic clusters among patients and to find links to concurrent isolates from poultry meat, broiler chickens, cattle, pigs, and dogs, we performed whole-genome sequencing on 1,509 C. jejuni isolates from 774 patients and 735 food or animal sources in Denmark during 2015-2017. We found numerous clusters; 366/774 (47.3%) clinical isolates formed 104 clusters of >2 isolates. A total of 41 patient clusters representing 199/366 (54%) patients matched a potential source, primarily domestic chickens/broilers. This study revealed serial outbreaks and numerous matches to concurrent food and animal isolates and highlighted the potential of whole-genome sequencing for improving routine surveillance of C. jejuni by enhancing outbreak detection, source tracing, and potentially prevention of human infections.

Establishing a donor stool bank for faecal microbiota transplantation: methods and feasibility.

Faecal microbiota transplantation (FMT) is a promising treatment, but donor selection and implementation in clinical practice are difficult. Here, we describe the establishment of a donor stool bank based on the Tissue Act. Stool donors were recruited among blood donors and asked to donate five times in a month. A screening questionnaire, a medical interview and testing of blood and stool were conducted before and after donations. Donations were made at home and transported to the lab, where 50 g of stool was suspended and filtered in saline and 20-mL glycerol (final concentration of 10%) to a volume of 170 mL. The processed stool was assigned a batch number, frozen within 2 h after defecation and stored at - 80 °C for up to 1 year. All steps were documented and cross-checked before donor stool were released for clinical use. Thirteen donors were eligible at the first interview and started donations. Two donors were excluded due to a positive Helicobacter pylori test, two withdrew consent and one was lost to follow-up. One donor took a single dose of NSAIDs 2 days prior to a donation, which was discarded. There were no other excluding findings at the second interview or testing. Eight of the 13 donors were approved as stool donors. All donated five times with each donation yielding 1-6 portions. Eighty-four portions were released for clinical use. Recruiting stool donors among blood donors is safe and effective. The Tissue Act yields an appropriate regulative framework for FMT.

Disease-Specific Enteric Microbiome Dysbiosis in Inflammatory Bowel Disease.

Inflammatory Bowel disease (IBD) is traditionally divided into Crohn's disease (CD) and ulcerative colitis (UC). UC is a relapsing non-transmural inflammatory disease that is restricted to the colon and is characterized by flare-ups of bloody diarrhea. CD is a chronic, segmental localized granulomatous disease that can affect any part of the entire gastrointestinal tract. Ileo-anal pouch is a procedure restoring functionality of the rectum after a colectomy. IBD is a multifactorial disease and flares of IBD are probably triggered by changes in the intestinal microbiota followed by an abnormal immune response. In this study, we aim to analyze the intestinal bacterial diversity in IBD patients during various stages of disease compared with healthy controls. Permission for human experiments and recruitment of participants was obtained from the Ethic Committee for Copenhagen County hospitals (Permission no. KA-03019, Permission no. KA-20060159). Stools from 26 healthy controls, 42 CD, 38 UC and 18 pouch patients were collected. Stool DNA extraction was performed using Qiagen, DNA mini stool kit Denmark. DGGE-PCR amplifying the V2-V3 region of 16S-rDNA gene of the bacteria was amplified by universal primers HDA1 and HDA2. Analysis of DGGE was performed blinded using BioNumerics version 7.5. After normalization, a DGGE gel band matching was performed. The similarities between profiles were calculated with a ranked Pearson correlation coefficient based on the band matching results using band intensities. Simpson's index of diversity and Pielou's species evenness were calculated. Based on the Shannon Diversity Index, UC patients had lower species diversity and bacterial evenness in comparison to healthy persons, p < 0.05. However, only CD and disease pouch patients had lower species diversity compared to those with inactive disease and healthy controls. Well-functioning pouch patients had decreased species evenness in comparison to diseased pouch patients and control group. During the active disease stage in CD and pouch, the patients have a low bacterial diversity in their gut when compared to the inactive stage. In UC patients, a generally low diversity was observed at all stages of the disease compared to healthy controls.

Characterization of Diarrheagenic Enteroaggregative Escherichia coli in Danish Adults-Antibiotic Treatment Does Not Reduce Duration of Diarrhea.

Enteroaggregative Escherichia coli (EAEC) is frequently isolated from sporadic cases of diarrhea and in outbreaks of gastroenteritis in several regions of the world. The pathophysiology of EAEC continues to be enigmatic, and the efficacy of antibiotic treatment in EAEC-associated diarrhea has been discussed. Since the level of antibiotic resistance is increasing, it is essential to restrict the use of antibiotics to prevent further resistance development. We aimed to investigate EAEC strains in adult Danish patients suffering from diarrhea and from healthy controls. We examined the antibiotic resistance in EAEC strains, the clinical response to antibiotic treatment in EAEC diarrheal cases, and the distribution of virulence genes in diarrheal cases. The EAEC strains were collected from patients suffering from diarrhea in a Danish multicenter study. A medical doctor interviewed the patients by using a questionnaire regarding gastrointestinal symptoms, exposures, and use of antibiotic and over-the-counter antidiarrheal drugs. Follow-up was performed after 3-5 months to inquire about differential diagnosis to gastrointestinal disease. A multiplex polymerase chain reaction characterized virulence genes in diarrheal cases. Finally, the level of antibiotic resistance was examined by using the disc diffusion method. Asymptomatic carriage of EAEC in the adult Danish population was rare, in contrast to findings in healthy Danish children. The duration of diarrhea was not shortened by antibiotic treatment, specifically ciprofloxacin treatment, or by over-the-counter antidiarrheal drugs. Follow-up revealed no pathology in diarrheal patients apart from irritable bowel syndrome in two patients. A high number of patients suffered from long-term diarrhea, which was associated with the enterotoxin EAST-1 and a high virulence factor score. A high level of antibiotic resistance was observed and 58% of the EAEC strains were multidrug resistant. Multidrug resistance was most pronounced in cases of travelers' diarrhea, and it was seen that antibiotic treatment did not reduce the duration of diarrhea.

Genetic Virulence Profile of Enteroaggregative Escherichia coli Strains Isolated from Danish Children with Either Acute or Persistent Diarrhea.

Enteroaggregative Escherichia coli (EAEC) is frequently found in diarrheal stools worldwide. It has been associated with persistent diarrhea, weight loss, and failure to thrive in children living in developing countries. A number of important EAEC virulence genes are identified; however, their roles in acute and persistent diarrhea have not been previously investigated. The aim of this study was to identify specific EAEC virulence genes associated with duration and type of diarrhea in Danish children. We aimed to improve the current diagnostics of EAEC and enable targeting of strains with an expected severe disease course. Questionnaires answered by parents provided information regarding duration of diarrhea and presence of blood or mucus. A total of 295 EAEC strains were collected from children with acute (≤7 days) and persistent diarrhea (≥14 days) and were compared by using multiplex PCR targeting the genes sat, sepA, pic, sigA, pet, astA, aatA, aggR, aaiC, aap, agg3/4C, ORF3, aafA, aggA, agg3A, agg4A, and agg5A. Furthermore, the distribution of EAEC genes in strains collected from cases of bloody, mucoid, and watery diarrhea was investigated. The classification and regression tree analysis (CART) was applied to investigate the relationship between EAEC virulence genes and diarrheal duration and type. Persistent diarrhea was associated with strains lacking the pic gene (p = 0.002) and with the combination of the genes pic, sat, and absence of the aggA gene (p = 0.05). Prolonged diarrhea was associated with the combination of the genes aatA and astA (p = 0.03). Non-mucoid diarrhea was associated with strains lacking the aatA gene (p = 0.004). Acute diarrhea was associated with the genes aggR, aap, and aggA by individual odds ratios. Resistance toward gentamicin and ciprofloxacin was observed in 7.5 and 3% of strains, respectively. Multi-drug resistance was observed in 38% of strains. Genetic host factors have been associated with an increased risk of EAEC-associated disease. Therefore, we investigated a panel of risk factors in two groups of children-EAEC-positive and EAEC-negative-to identify additional factors predisposing to disease. The duration of breastfeeding was positively correlated with the likelihood of belonging to the EAEC-negative group of children.

National Automated Surveillance of Hospital-Acquired Bacteremia in Denmark Using a Computer Algorithm.

BACKGROUND In 2015, Denmark launched an automated surveillance system for hospital-acquired infections, the Hospital-Acquired Infections Database (HAIBA). OBJECTIVE To describe the algorithm used in HAIBA, to determine its concordance with point prevalence surveys (PPSs), and to present trends for hospital-acquired bacteremia SETTING Private and public hospitals in Denmark METHODS A hospital-acquired bacteremia case was defined as at least 1 positive blood culture with at least 1 pathogen (bacterium or fungus) taken between 48 hours after admission and 48 hours after discharge, using the Danish Microbiology Database and the Danish National Patient Registry. PPSs performed in 2012 and 2013 were used for comparison. RESULTS National trends showed an increase in HA bacteremia cases between 2010 and 2014. Incidence was higher for men than women (9.6 vs 5.4 per 10,000 risk days) and was highest for those aged 61-80 years (9.5 per 10,000 risk days). The median daily prevalence was 3.1% (range, 2.1%-4.7%). Regional incidence varied from 6.1 to 8.1 per 10,000 risk days. The microorganisms identified were typical for HA bacteremia. Comparison of HAIBA with PPS showed a sensitivity of 36% and a specificity of 99%. HAIBA was less sensitive for patients in hematology departments and intensive care units. Excluding these departments improved the sensitivity of HAIBA to 44%. CONCLUSIONS Although the estimated sensitivity of HAIBA compared with PPS is low, a PPS is not a gold standard. Given the many advantages of automated surveillance, HAIBA allows monitoring of HA bacteremia across the healthcare system, supports prioritizing preventive measures, and holds promise for evaluating interventions. Infect Control Hosp Epidemiol 2017;38:559-566.

Evaluation of Xpert MRSA Gen 3 and BD MAX MRSA XT for meticillin-resistant Staphylococcusaureus screening in a routine diagnostic setting in a low-prevalence area.

Screening and pre-emptive isolation of at-risk patients are important aspects of the Danish approach to the prevention of meticillin-resistant Staphylococcusaureus (MRSA) infection, but screening with conventional culture can take up to 3 days for results to become available with attendant costs and disadvantages of prolonged isolation. We sought to evaluate the accuracy, time to availability of results and potential economic benefits of two next-generation MRSA screening assays, Xpert MRSA Gen 3 (GX MRSA) and BD MAX MRSA XT, in a setting of a consolidated laboratory serving a number of hospitals with a low prevalence of MRSA and using enrichment culture as a reference method. Four hundred and forty-seven screening samples together with 49 previously positive MRSA samples were evaluated. Xpert MRSA Gen 3 demonstrated sensitivity, specificity, positive predictive value and negative predictive value of 88.2, 97.9, 62.5 and 99.5 %, respectively, and for BD MAX MRSA XT, they were 88.2, 97.4, 57.7 and 99.5 %, respectively. Hands-on time was 8.8 and 21.6 min, respectively, for the Xpert MRSA Gen 3 and BD MAX MRSA XT PCR assays when five samples were handled simultaneously. The mean laboratory turnaround time was 2.9 (1-6) hours for the Xpert MRSA Gen 3 assay, 6.5 (2-46) hours for BD MAX MRSA XT and 49.6 (42-122) hours for enriched culture. Despite laboratory costs being higher for the rapid PCR assays, when the costs of isolation are taken into account, the assays offer the potential for significant cost savings.

Novel multiplex format of an extended multilocus variable-number-tandem-repeat analysis of Clostridium difficile correlates with tandem repeat sequence typing.

Subtyping of Clostridium difficile is crucial for outbreak investigations. An extended multilocus variable-number tandem-repeat analysis (eMLVA) of 14 variable number tandem repeat (VNTR) loci was validated in multiplex format compatible with a routine typing laboratory and showed excellent concordance with tandem repeat sequence typing (TRST) and high discriminatory power.

The application of new molecular methods in the investigation of a waterborne outbreak of norovirus in Denmark, 2012.

In December 2012, an outbreak of acute gastrointestinal illness occurred in a geographical distinct area in Denmark covering 368 households. A combined microbiological, epidemiological and environmental investigation was initiated to understand the outbreak magnitude, pathogen(s) and vehicle in order to control the outbreak. Norovirus GII.4 New Orleans 2009 variant was detected in 15 of 17 individual stool samples from 14 households. Norovirus genomic material from water samples was detected and quantified and sequencing of longer parts of the viral capsid region (>1000 nt) were applied to patient and water samples. All five purposely selected water samples tested positive for norovirus GII in levels up to 1.8×10(4) genomic units per 200 ml. Identical norovirus sequences were found in all 5 sequenced stool samples and 1 sequenced water sample, a second sequenced water sample showed 1 nt (<0.1%) difference. In a cohort study, including 256 participants, cases were defined as residents of the area experiencing diarrhoea or vomiting onset on 12-14 December 2012. We found an attack rate of 51%. Being a case was associated with drinking tap-water on 12-13 December (relative risk = 6.0, 95%CI: 1.6-22) and a dose-response relation for the mean glasses of tap-water consumed was observed. Environmental investigations suggested contamination from a sewage pipe to the drinking water due to fall in pressure during water supply system renovations. The combined microbiological, epidemiological and environmental investigations strongly indicates the outbreak was caused by norovirus contamination of the water supply system.

Psychometric scores and persistence of irritable bowel after Campylobacter concisus infection.

OBJECTIVE: Gastroenteritis with Campylobacter concisus is an emerging infection, but the risk of irritable bowel syndrome (IBS) following it is unknown. MATERIAL AND METHODS: In a prospective, community-based study of gastroenteritis with C. concisus and C. jejuni/coli, we invited adult patients to participate in a questionnaire study, including IBS symptoms and psychometric scores, at baseline and at 6 months. We estimated adjusted RR (RRadj) (for age, sex and comorbidity) for IBS as the primary outcome. RESULTS: The development of IBS symptoms at 6 months was reported in 26/106 (25%) patients with C. concisus infection, and in 30/162 (19%) of C. jejuni/coli patients. The baseline predictors for IBS in C. concisus infection were high anxiety scores (RRadj 2.0; 95% CI 1.1-3.6, p<0.05), chills (RRadj 1.9; 95% CI 1.0-3.6, p<0.05), headache (RRadj 2.5; 95% CI 1.1-6.0, p<0.05), dizziness (RRadj 2.6; 95% CI 1.2-5.8, p<0.05) and muscle ache (RRadj 3.6; 95% CI 1.4-8.9, p<0.01). For all Campylobacter patients (n=268), we confirmed previous reports of anxiety (RRadj 2.0; 95% CI 1.3-3.1), depression (RRadj 2.3; 95% CI 1.3-4.0) and high somatization scores (RRadj 3.0; 95% CI 1.5-6.0) as predictors for post-infectious IBS (PI-IBS). CONCLUSIONS: Gastroenteritis with C. concisus carries a 25% risk of IBS at 6-month follow-up. The risk factors for IBS are chills, headache, dizziness and muscle ache in the acute stage, as well as preexisting high psychometric scores for anxiety. Our findings suggest that psychological factors play a role in the development of PI-IBS.

Comparison of polycarbonate and cellulose acetate membrane filters for isolation of Campylobacter concisus from stool samples.

One thousand seven hundred ninety-one diarrheic stool samples were cultivated for Campylobacter spp. We found a high prevalence of Campylobacter concisus with use of a polycarbonate filter (n = 114) compared to a cellulose acetate filter (n = 79) (P < .0001). The polycarbonate filter is superior to the commonly used cellulose acetate filter for detection of C. concisus.