Usability of the international HAVNet hepatitis A virus database for geographical annotation, backtracing and outbreak detection.

BackgroundHAVNet is an international laboratory network sharing sequences and corresponding metadata on hepatitis A virus in an online database. Aim: We give an overview of the epidemiological and genetic data and assess the usability of the present dataset for geographical annotation, backtracing and outbreak detection. Methods: A descriptive analysis was performed on the timeliness, completeness, epidemiological data and geographic coverage of the dataset. Length and genomic region of the sequences were reviewed as well as the numerical and geographical distribution of the genotypes. The geographical signal in the sequences was assessed based on a short common nt stretch using a 100% identity analysis. Results: The 9,211 reports were heterogeneous for completeness and timeliness, and for length and genomic region of the sequences. Some parts of the world were not represented by the sequences. Geographical differences in prevalence of HAV genotypes described previously could be confirmed with this dataset and for a third (1,075/3,124) of the included sequences, 100% identity of the short common sequence coincided with an identical country of origin. Conclusion: Analysis of a subset of short, shared sequences indicates that a geographical annotation on the level of individual countries is possible with the HAVNet data. If the current incompleteness and heterogeneity of the data can be improved on, HAVNet could become very useful as a worldwide reference set for geographical annotation and for backtracing and outbreak detection.

Surveillance of antimicrobial use in Dutch long-term care facilities.

Objectives: : Residents living in a long-term care facility (LTCF) are more susceptible to infections. Treatment with antimicrobials is sometimes necessary; however, antibiotic use is considered one of the most important drivers of the development of antibiotic resistance. Surveillance data on antibiotic use in these LTCFs are necessary to get more insight into these patterns. The objective of this study was to describe antibiotic use in LTCFs in the Netherlands. Methods: : One hundred and seventy-seven LTCFs in the Netherlands were contacted and asked to participate in a study concerning antibiotic resistance and antibiotic use. Associated pharmacies were asked to provide data about systemic antibiotic use for each participating LTCF location over 1 year. Results on antibiotic use are reported here. Results: : Antibiotic use data from 96 LTCFs were collected from the pharmacies, and 68 of these LTCFs completed additional questionnaires on general characteristics of their location. Mean total use of systemic antimicrobials was 73 DDDs/1000 residents per day (range 2-197 DDDs/1000 residents per day). Co-amoxiclav (23 DDDs/1000 residents/day, range 0-70) was used the most, followed by nitrofurantoin derivatives (12 DDDs/1000 residents/day, range 0-38) and fluoroquinolones (12 DDDs/1000 residents/day, range 0-52). Statistical analysis revealed no significant correlations between the LTCF characteristics and the level of antibiotic use. Conclusions: There was a high use of broad-spectrum antimicrobials, with a large variation in total antibiotic use between individual locations. Further analysis of more in-depth data and possible influencing factors is needed.

Carriage of antimicrobial-resistant commensal bacteria in Dutch long-term-care facilities.

OBJECTIVES: The objective of this study was to assess carriage of antimicrobial-resistant commensal microorganisms, i.e. Escherichia coli and Staphylococcus aureus, and its predictors in long-term-care facilities (LTCFs). METHODS: Nasal swabs and/or urine or incontinence samples were collected from participating residents in 111 LTCFs and tested for the presence of S. aureus and/or E. coli, respectively. Antimicrobial resistance to eight antimicrobials was linked to antimicrobial usage in the year preceding sampling and to LTCF characteristics. Using multilevel logistic regression, predictors of carriage of ESBL-producing E. coli in LTCFs were identified. RESULTS: S. aureus was identified in 1269/4763 (26.6%) nasal swabs, including 13/4763 (0.3%) MRSA carriers in 9/107 (8%) LTCFs. Of the 5359 urine/incontinence samples, 2934 (55%) yielded E. coli, including 123 (4.2%) producing ESBL, which were found in 53/107 locations (range 1%-33%). For all but one antimicrobial (i.e. nitrofurantoin) >20% of isolated E. coli were resistant. Multilevel multivariable logistic regression identified two predictors of carriage of ESBL-producing E. coli: (i) antimicrobial usage (OR 1.8, 95% CI 1.1-3.0 for each extra 50 DDD/1000 residents/day); and (ii) presence of MRSA carriers in the LTCFs (OR 2.4, 95% CI 1.0-5.6). CONCLUSIONS: The low proportion of 4.2% ESBL-producing E. coli and the low prevalence of 0.3% MRSA carriage found in LTCF residents suggest that Dutch LTCFs are not yet an important reservoir of MDR potential pathogens. Nevertheless, the large variation between LTCFs warrants close monitoring of antimicrobial resistance in LTCFs. Integrated surveillance, i.e. linking data sources on antimicrobial usage, microbiological testing, clinical background data and epidemiological data, is needed.

International linkage of two food-borne hepatitis A clusters through traceback of mussels, the Netherlands, 2012.

This report describes an outbreak investigation starting with two closely related suspected food-borne clusters of Dutch hepatitis A cases, nine primary cases in total, with an unknown source in the Netherlands. The hepatitis A virus (HAV) genotype IA sequences of both clusters were highly similar (459/460 nt) and were not reported earlier. Food questionnaires and a case-control study revealed an association with consumption of mussels. Analysis of mussel supply chains identified the most likely production area. International enquiries led to identification of a cluster of patients near this production area with identical HAV sequences with onsets predating the first Dutch cluster of cases. The most likely source for this cluster was a case who returned from an endemic area in Central America, and a subsequent household cluster from which treated domestic sewage was discharged into the suspected mussel production area. Notably, mussels from this area were also consumed by a separate case in the United Kingdom sharing an identical strain with the second Dutch cluster. In conclusion, a small number of patients in a non-endemic area led to geographically dispersed hepatitis A outbreaks with food as vehicle. This link would have gone unnoticed without sequence analyses and international collaboration.

Norovirus genotype profiles associated with foodborne transmission, 1999-2012.

Worldwide, noroviruses are a leading cause of gastroenteritis. They can be transmitted from person to person directly or indirectly through contaminated food, water, or environments. To estimate the proportion of foodborne infections caused by noroviruses on a global scale, we used norovirus transmission and genotyping information from multiple international outbreak surveillance systems (Noronet, CaliciNet, EpiSurv) and from a systematic review of peer-reviewed literature. The proportion of outbreaks caused by food was determined by genotype and/or genogroup. Analysis resulted in the following final global profiles: foodborne transmission is attributed to 10% (range 9%%-11%) of all genotype GII.4 outbreaks, 27% (25%-30%) of outbreaks caused by all other single genotypes, and 37% (24%%-52%) of outbreaks caused by mixtures of GII.4 and other noroviruses. When these profiles are applied to global outbreak surveillance data, results indicate that ≈14% of all norovirus outbreaks are attributed to food.

Underdiagnosis of foodborne hepatitis A, The Netherlands, 2008-2010(1.).

Outbreaks of foodborne hepatitis A are rarely recognized as such. Detection of these infections is challenging because of the infection's long incubation period and patients' recall bias. Nevertheless, the complex food market might lead to reemergence of hepatitis A virus outside of disease-endemic areas. To assess the role of food as a source of infection, we combined routine surveillance with real-time strain sequencing in the Netherlands during 2008-2010. Virus RNA from serum of 248 (59%) of 421 reported case-patients could be sequenced. Without typing, foodborne transmission was suspected for only 4% of reported case-patients. With typing, foodborne transmission increased to being the most probable source of infection for 16%. We recommend routine implementation of an enhanced surveillance system that includes prompt forwarding and typing of hepatitis A virus RNA isolated from serum, standard use of questionnaires, data sharing, and centralized interpretation of data.

Unrecognized norovirus infections in health care institutions and their clinical impact.

Noroviruses (NoVs) have emerged as the leading cause of acute viral gastroenteritis (GE) in humans. Although diagnostic facilities have greatly improved, significant underdiagnosis of NoV in hospitals may still occur, thereby increasing clinical burden and nosocomial spread. We evaluated the underdiagnosis of sporadic NoV infections in a tertiary care hospital and estimated its clinical impact. From December 2008 until July 2009, fecal samples specifically referred for bacterial but not viral examination were retrospectively tested for NoV by real-time PCR. The clinical and virological data from patients with undiagnosed NoV infection (missed patients) were evaluated and compared with those from patients with recognized NoV. During the study period, 45 patients with undiagnosed NoV were detected, whereas 50 patients were regularly diagnosed. The missed NoV cases were more frequently adults than children (80% versus 46%; P < 0.001). The viral load levels did not differ between the diagnosed and missed patients, but missed patients more frequently presented without diarrhea (20% versus 4%; P < 0.07). The newly admitted missed NoV cases with GE underwent more diagnostic imaging (24% versus 4%; P < 0.01) and tended to be hospitalized longer. When missed-NoV patients were included, the number of nosocomial clusters doubled and missed patients were index cases in 5 of the 6 clusters. These data indicate that NoV infections are frequently missed despite routine laboratory testing and demonstrate that underdiagnosis of NoV patients is associated with costly abdominal imaging and nosocomial clustering. Awareness of NoV infection in adult patients and education about the importance of viral GE should be increased.

Quantifying transmission of norovirus during an outbreak.

BACKGROUND: Healthcare workers are thought to play a role in nosocomial transmission of norovirus, but the level and direction of norovirus transmission between patients and healthcare workers in sustaining transmission during an outbreak have not been quantified. METHODS: We developed a method for finding plausible transmission trees of who acquired their infection from whom. We applied the method to data from an outbreak of norovirus in 4 wards of a psychiatric institution in the Netherlands in 2008. The simulated transmission trees were based on serial intervals for time between symptom onsets, weighted for the number of days that healthcare workers were present. The obtained transmission trees were linked to the Barthel Index, a measure of patient reliance on healthcare in their basic daily activities. RESULTS: The dominant recognized transmission route was from patient to patient (64%), followed by patient to healthcare worker (29%). The overall estimated reproduction number for healthcare workers was low compared with patients (0.25 vs. 1.20; mean difference = 0.95 [95% confidence interval (CI) = 0.60 to 1.30]). The average number of all subsequent cases attributable to the downstream branch of one single infected healthcare worker in the transmission tree was 4.4 compared with 6.5 for cases attributable to one single infected patient (mean difference = 2.1 [95% CI = -4.7 to 8.9]). In the ward with patients requiring the highest level of care from healthcare workers, the attack rate among healthcare workers was highest. CONCLUSION: This approach provides a framework to quantify the magnitude and direction of transmission between healthcare workers and patients during a norovirus outbreak. The utility of this method in outbreaks of other infections and in different settings should be explored.

Selection of a phylogenetically informative region of the norovirus genome for outbreak linkage.

The recognition of a common source norovirus outbreak is supported by finding identical norovirus sequences in patients. Norovirus sequencing has been established in many (national) public health laboratories and academic centers, but often partial and different genome sequences are used. Therefore, agreement on a target sequence of sufficient diversity to resolve links between outbreaks is crucial. Although harmonization of laboratory methods is one of the keystone activities of networks that have the aim to identify common source norovirus outbreaks, this has proven difficult to accomplish, particularly in the international context. Here, we aimed at providing a method enabling identification of the genomic region informative of a common source norovirus outbreak by bio-informatic tools. The data set of 502 unique full length capsid gene sequences available from the public domain, combined with epidemiological data including linkage information was used to build over 3,000 maximum likelihood (ML) trees for different sequence lengths and regions. All ML trees were evaluated for robustness and specificity of clustering of known linked norovirus outbreaks against the background diversity of strains. Great differences were seen in the robustness of commonly used PCR targets for cluster detection. The capsid gene region spanning nucleotides 900-1,400 was identified as the region optimally substituting for the full length capsid region. Reliability of this approach depends on the quality of the background data set, and we recommend periodic reassessment of this growing data set. The approach may be applicable to multiple sequence-based data sets of other pathogens.

High prevalence of acute hepatitis E virus infection in pigs in Dutch slaughterhouses.

Hepatitis E is caused by hepatitis E virus (HEV), one of the causes of acute viral hepatitis. Domestic pigs are considered as the main reservoir of HEV-3. The recently reported high prevalence of HEV in liver- and meat products on the Dutch market warranted a cross-sectional prevalence study on HEV infection among 5-6 months old pigs slaughtered in the Netherlands (n = 250). For this, liver, caecum content and blood samples were analyzed for the presence of genomic HEV RNA by RT-PCR. In addition, a serological test was performed to detect HEV IgG. Background information was retrieved on the corresponding farms to evaluate potential risk factors for HEV at pig slaughter age. HEV IgG was detected in sera from 167 pigs (67.6 %). HEV RNA was detected in 64 (25.6 %) caecum content samples, in 40 (16.1 %) serum samples and in 25 (11.0 %) liver samples. The average level of viral contamination in positive samples was log10 4.6 genome copies (gc)/g (range 3.0-8.2) in caecum content, log10 3.3 gc/ml (range 2.4-5.9) in serum and log10 3.2 gc/0.1 g (range 1.7-6.2) in liver samples. Sequence analyses revealed HEV-3c only. Ten times an identical strain was detected in two or three samples obtained from the same pig. Each animal in this study however appeared to be infected with a unique strain. The presence of sows and gilts and welfare rating at the farm of origin had a significant effect (p < 0.05) on the distribution over the four groups representing different stages of HEV infection based on IgG or RNA in caecum and/or serum. The observed proportion of tested pigs with viremia (16 %) was higher than in other reported studies and was interestingly often observed in combination with a high number of HEV genome copies in liver and caecum content as detected by RT-qPCR. Data provided will be useful for risk assessment for food safety of pork products, will provide baseline data for future monitoring of HEV infections in pigs and new thoughts for mitigation strategies.

An integrated approach to identifying international foodborne norovirus outbreaks.

International foodborne norovirus outbreaks can be difficult to recognize when using standard outbreak investigation methods. In a novel approach, we provide step-wise selection criteria to identify clusters of outbreaks that may involve an internationally distributed common foodborne source. After computerized linking of epidemiologic data to aligned sequences, we retrospectively identified 100 individually reported outbreaks that potentially represented 14 international common source events in Europe during 1999-2008. Analysis of capsid sequences of outbreak strains (n = 1,456), showed that ≈7% of outbreaks reported to the Foodborne Viruses in Europe database were part of an international event (range 2%-9%), compared with 0.4% identified through standard epidemiologic investigations. Our findings point to a critical gap in surveillance and suggest that international collaboration could have increased the number of recognized international foodborne outbreaks. Real-time exchange of combined epidemiologic and molecular data is needed to validate our findings through timely trace-backs of clustered outbreaks.

Year-round prevalence of norovirus in the environment of catering companies without a recently reported outbreak of gastroenteritis.

Food handlers play an important role in the transmission of norovirus (NoV) in food-borne outbreaks of gastroenteritis (GE). In a year-round prevalence study, the prevalence of NoV in catering companies without recently reported outbreaks of GE was investigated and compared to the observed prevalence in catering companies with recently reported outbreaks. Swab samples were collected from surfaces in the kitchens and (staff) bathrooms in 832 randomly chosen companies and analyzed for the presence of NoV RNA. In total, 42 (1.7%) out of 2,496 environmental swabs from 35 (4.2%) catering companies tested positive. In contrast, NoV was detected in 147 (39.7%) of the 370 samples for 44 (61.1%) of the 72 establishments associated with outbreaks of gastroenteritis. NoV-positive swabs were more frequently found in winter, in specific types of companies (elderly homes and lunchrooms), and in establishments with separate bathrooms for staff. We found a borderline association with population density but no relation to the number of employees. Sequence analysis showed that environmental strains were interspersed with strains found in outbreaks of illness in humans. Thus, the presence of NoV in catering companies seemed to mirror the presence in the population but was strongly increased when associated with food-borne GE. Swabs may therefore serve as a valuable tool in outbreak investigations for the identification of the causative agent, although results should be interpreted with care, taking into account all other epidemiological data.

Use of norovirus genotype profiles to differentiate origins of foodborne outbreaks.

Because secondary transmission masks the connection between sources and outbreaks, estimating the proportion of foodborne norovirus infections is difficult. We studied whether norovirus genotype frequency distributions (genotype profiles) can enhance detection of the sources of foodborne outbreaks. Control measures differ substantially; therefore, differentiating this transmission mode from person-borne or food handler-borne outbreaks is of public health interest. Comparison of bivalve mollusks collected during monitoring (n = 295) and outbreak surveillance strains (n = 2,858) showed 2 distinguishable genotype profiles in 1) human feces and 2) source-contaminated food and bivalve mollusks; genotypes I.2 and I.4 were more frequently detected in foodborne outbreaks. Overall, approximately 21% of all outbreaks were foodborne; further analysis showed that 25% of the outbreaks reported as food handler-associated were probably caused by source contamination of the food.

Selection tool for foodborne norovirus outbreaks.

Detection of pathogens in the food chain is limited mainly to bacteria, and the globalization of the food industry enables international viral foodborne outbreaks to occur. Outbreaks from 2002 through 2006 recorded in a European norovirus surveillance database were investigated for virologic and epidemiologic indicators of food relatedness. The resulting validated multivariate logistic regression model comparing foodborne (n = 224) and person-to-person (n = 654) outbreaks was used to create a practical web-based tool that can be limited to epidemiologic parameters for nongenotyping countries. Non-genogroup-II.4 outbreaks, higher numbers of cases, and outbreaks in restaurants or households characterized (sensitivity = 0.80, specificity = 0.86) foodborne outbreaks and reduced the percentage of outbreaks requiring source-tracing to 31%. The selection tool enabled prospectively focused follow-up. Use of this tool is likely to improve data quality and strain typing in current surveillance systems, which is necessary for identification of potential international foodborne outbreaks.

Norovirus on swabs taken from hands illustrate route of transmission: a case study.

Recently, environmental swabs from kitchen and bathroom surfaces have been described as an additional tool for the detection of norovirus in outbreak settings. This article describes an outbreak investigation in response to the reporting of gastroenteritis in three unrelated groups of 6, 12, and 13 adults approximately 30 h after having meals in the same restaurant. Fecal samples were collected from 13 patients and six food handlers, and environmental swabs were taken from the soap dispenser, working bench, doorknobs of cupboards, and the grip of a knife in the kitchen and in bathrooms as well as from the hands of each of three employees on the day of inspection. Clinical and environmental samples were analyzed separately in time and location for the presence of norovirus by real-time reverse transcription PCR. Structured interviews revealed that all staff members had suffered from gastroenteritis, one after the other. Norovirus RNA (GGI.6) was detected in 17 of 19 fecal samples as well as in 4 environmental samples, including a swab sample from the hands of a staff member who was preparing ready-to-eat food. Sequences obtained from clinical and environmental samples showed an identity of 100% (235 nucleotides). To our knowledge, this is the first case study to directly demonstrate the presence of norovirus RNA on a food handler's hands in an outbreak setting. This finding provides direct evidence for the feasibility of transmission of norovirus by a food handler to food. Education of food handlers on the infectivity of norovirus and updating of hygienic codes are strongly recommended.

Chronic sequelae and severe complications of norovirus infection: A systematic review of literature.

Norovirus causes an estimated 18% of all cases of acute gastroenteritis worldwide and is found to be associated with mortality. To create a first overview of severe complications and chronic sequelae of norovirus infections, a systematic review of literature was performed. Of 3928 individual hits, 176 publications remained for data extraction. Study periods varied between 1974 and 2017, though strongly skewed towards the last decade (n = 122, 70%). Countries of studies were worldwide, though Africa, and Carribean, Central and South America were underrepresented. Strong evidence was found for chronic diarrhea in immunocompromised patients, affecting 9%-100% of investigated cohorts. The duration of chronic diarrhea varied from four weeks up to nine years, leading to either wasting, weight loss or failure to thrive in a third of the reported cases (224). Other complications with significant evidence were necrotizing enterocolitis (NEC) in preterm infants associated with norovirus infection (8 papers), and benign infantile convulsions with gastroenteritis (BICG; 19 papers). Studies on norovirus infection and inflammatory bowel disease (IBD) mostly concluded against this association (5 of 7). The remaining papers mentioned a large variety of possible sequelae or complications. Based on the available literature, chronic norovirus diarrhea is the major sequela of norovirus infection in primary immune deficient, oncologic and transplant patients. Norovirus infection - like other gastrointestinal pathogens - can cause a range of sequelae and complications, and should be considered in the differential diagnosis of these manifestations.

Outbreak detection and secondary prevention of Legionnaires' disease: a national approach.

BACKGROUND: To stop a possible outbreak of Legionnaires' disease (LD) at an early stage an outbreak detection programme was installed in The Netherlands. METHODS: The programme consisted of sampling and controlling of potential sources to which LD patients had been exposed during their incubation period. Potential sources were considered to be true sources of infection if two or more LD patients (cluster) had visited them, or if available patients' isolates and environmental Legionella spp. were indistinguishable by amplified fragment length polymorphism genotyping. RESULTS: Rapid sampling and genotyping as well as cluster recognition helped to target control measures. Despite these measures, two small outbreaks were only stopped after renewal of the water system. The combination of genotyping and cluster recognition lead to 29 of 190 (15%) patient-source associations. CONCLUSION: Systematic sampling and cluster recognition can contribute to outbreak detection and lead to cost-effective secondary prevention of Legionnaires' disease.

Risk of hepatitis A decreased among Dutch travelers to endemic regions in 2003 to 2011.

We divided the number of travel-related hepatitis A cases notified in Dutch surveillance (2003-2011) by travel data obtained from an annual holiday survey to estimate the risk of hepatitis A among Dutch travelers. Of the 2,094 cases notified, 931 (44%) were imported. Morocco (n = 272, 29%), Turkey (n = 98, 11%), and Egypt (n = 87, 9%) accounted for the largest proportion of cases. Attack rates in returnees from high or intermediate endemic regions declined from 7.5 per 100,000 travelers (95% CI 6.7-8.4) in 2003-2005 to 3.5 (95% CI 3.0-4.0) in 2009-2011 (p < 0.01). Despite the decrease in risk, vaccination remains important, and routine risk monitoring should also be considered.

Domestic exposure to legionellae for Dutch Legionnaires' disease patients.

The source of infection for travelers who develop Legionnaires' disease (LD) shortly after a journey abroad is difficult to ascertain. Infection is likely to have occurred abroad, but could also have occurred at the patient's own residence. The authors conducted a case-control study to determine risk for acquiring LD at home in the Netherlands after traveling abroad. They compared homes of 44 traveling LD patients with 44 homes of nontraveling LD patients, using logistic regression models. Geographic distribution was confounding the association between traveling and presence of Legionella spp. in residences; adjustment was necessary. In traveler's homes, legionellae were present more often, with crude and adjusted OR (95% CI) being 1.6 (0.5-5.0) and 1.4 (0.4-4.4), respectively. The authors' findings indicate that the patient's residence can be a potential source of infection after traveling.

Global prevalence of norovirus in cases of gastroenteritis: a systematic review and meta-analysis.

BACKGROUND: Despite substantial decreases in recent decades, acute gastroenteritis causes the second greatest burden of all infectious diseases worldwide. Noroviruses are a leading cause of sporadic cases and outbreaks of acute gastroenteritis across all age groups. We aimed to assess the role of norovirus as a cause of endemic acute gastroenteritis worldwide. METHODS: We searched Embase, Medline, and Global Health databases from Jan 1, 2008, to March 8, 2014, for studies that used PCR diagnostics to assess the prevalence of norovirus in individuals with acute gastroenteritis. We included studies that were done continuously for 1 year or more from a specified catchment area (geographical area or group of people), enrolled patients who presented with symptoms of acute gastroenteritis, and used PCR-based diagnostics for norovirus on all stool specimens from patients with acute gastroenteritis. The primary outcome was prevalence of norovirus among all cases of gastroenteritis. We generated pooled estimates of prevalence by fitting linear mixed-effect meta-regression models. FINDINGS: Of 175 articles included, the pooled prevalence of norovirus in 187 336 patients with acute gastroenteritis was 18% (95% CI 17-20). Norovirus prevalence tended to be higher in cases of acute gastroenteritis in community (24%, 18-30) and outpatient (20%, 16-24) settings compared with inpatient (17%, 15-19, p=0·066) settings. Prevalence was also higher in low-mortality developing (19%, 16-22) and developed countries (20%, 17-22) compared with high-mortality developing countries (14%, 11-16; p=0·058). Patient age and whether the study included years of novel strain emergence were not associated with norovirus prevalence. INTERPRETATION: Norovirus is a key gastroenteritis pathogen associated with almost a fifth of all cases of acute gastroenteritis, and targeted intervention to reduce norovirus burden, such as vaccines, should be considered. FUNDING: The Foodborne Disease Burden Epidemiology Reference Group (FERG) of WHO and the Government of the Netherlands on behalf of FERG.