Scientific Committee guidance on appraising and integrating evidence from epidemiological studies for use in EFSA's scientific assessments.

EFSA requested its Scientific Committee to prepare a guidance document on appraising and integrating evidence from epidemiological studies for use in EFSA's scientific assessments. The guidance document provides an introduction to epidemiological studies and illustrates the typical biases, which may be present in different epidemiological study designs. It then describes key epidemiological concepts relevant for evidence appraisal. This includes brief explanations for measures of association, exposure assessment, statistical inference, systematic error and effect modification. The guidance then describes the concept of external validity and the principles of appraising epidemiological studies. The customisation of the study appraisal process is explained including tailoring of tools for assessing the risk of bias (RoB). Several examples of appraising experimental and observational studies using a RoB tool are annexed to the document to illustrate the application of the approach. The latter part of this guidance focuses on different steps of evidence integration, first within and then across different streams of evidence. With respect to risk characterisation, the guidance considers how evidence from human epidemiological studies can be used in dose-response modelling with several different options being presented. Finally, the guidance addresses the application of uncertainty factors in risk characterisation when using evidence from human epidemiological studies.

How long can you store vitamins? Stability of tocopherols and tocotrienol during different storage conditions in broccoli and blueberries.

Differences between the stability of α-, ß-, γ-, and δ-tocopherol as well α-tocotrienol stored at -20 °C and -80 °C were studied in broccoli and blueberry samples. Before storage up to 28 days, they underwent different initializing processes such as freezing quickly with liquid nitrogen and freeze-drying, followed by homogenization. While α-tocopherol levels in blueberries did not significantly differ, levels in broccoli were substantially higher after homogenization of freeze-dried samples compared to fresh broccoli samples. This might be caused by higher extractability of α-tocopherol from the changed cell structure. Storage of fresh broccoli samples at -20 °C led to decreasing α-tocopherol levels. Nevertheless, the deviation between freeze-dried samples to the initial fresh samples and fresh samples frozen with liquid nitrogen stored at -20 °C for 7 days were in the same order of magnitude. In conclusion, storage up to 7 days for vitamin relevant samples before analysis seemed to be justifiable.

Collection of occurrence data in foods - The value of the BfR MEAL study in addition to the national monitoring for dietary exposure assessment.

Two different data sets of occurrence data are available in Germany at present: the German National Food Monitoring and the BfR MEAL Study. To determine the suitability of each data set for exposure assessment and to develop concepts for a target-oriented selection and application of data, possibilities, limitations and scope of substance as well as food selection is quantitatively compared. The National Monitoring data provides comprehensive information on the variability of substance levels. This enables short- and long-term exposure assessment and consumer-loyal scenarios. The BfR MEAL Study supplements the monitoring data set with > 100 substances or by complementing the food spectrum for substances already included in the National Monitoring. The study design benefits especially the long-term dietary exposure assessment for the German population including the total diet. Using both programmes enables case-dependent selection of the appropriate dataset and in combination both sets can contribute to enhanced consumer safety.

Results of the BfR MEAL Study: The food type has a stronger impact on calcium, potassium and phosphorus levels than factors such as seasonality, regionality and type of production.

The BfR MEAL Study aims to provide representative levels of chemical substances in foods consumed by the population in Germany for dietary exposure assessment. Calcium, potassium and phosphorus (Ca, K, P) are essential to obtain physiological functions in humans. Levels were investigated in 356 foods. Foods were purchased representatively, prepared as typically consumed and pooled before analysis. High mean levels were found in milk, dairy products, legumes, nuts, oilseeds and spices as well as chia seeds (Ca, K, P), chewing gum (Ca) and cocoa powder (K). Different levels comparing organically and conventionally produced foods were determined among others in cereal cracker (puffed), olives and tofu. Higher K levels were found in fried compared to boiled potatoes. Similar P levels were mainly found in regionally and seasonally sampled foods. These data provide a substantially improved basis to address dietary exposure assessment of the population in Germany for Ca, K and P.

Analysis of German BSE Surveillance Data: Estimation of the Prevalence of Confirmed Cases versus the Number of Infected, but Non-Detected, Cattle to Assess Confidence in Freedom from Infection.

Quantitative risk assessments for Bovine spongiform encephalopathy (BSE) necessitate estimates for key parameters such as the prevalence of infection, the probability of absence of infection in defined birth cohorts, and the numbers of BSE-infected, but non-detected cattle entering the food chain. We estimated three key parameters with adjustment for misclassification using the German BSE surveillance data using a Gompertz model for latent (i.e., unobserved) age-dependent detection probabilities and a Poisson response model for the number of BSE cases for birth cohorts 1999 to 2015. The models were combined in a Bayesian framework. We estimated the median true BSE prevalence between 3.74 and 0.216 cases per 100,000 animals for the birth cohorts 1990 to 2001 and observed a peak for the 1996 birth cohort with a point estimate of 16.41 cases per 100,000 cattle. For birth cohorts ranging from 2002 to 2013, the estimated median prevalence was below one case per 100,000 heads. The calculated confidence in freedom from disease (design prevalence 1 in 100,000) was above 99.5% for the birth cohorts 2002 to 2006. In conclusion, BSE surveillance in the healthy slaughtered cattle chain was extremely sensitive at the time, when BSE repeatedly occurred in Germany (2000-2009), because the entry of BSE-infected cattle into the food chain could virtually be prevented by the extensive surveillance program during these years and until 2015 (estimated non-detected cases/100.000 [95% credible interval] in 2000, 2009, and 2015 are 0.64 [0.5,0.8], 0.05 [0.01,0.14], and 0.19 [0.05,0.61], respectively).

Exposure assessment of methylmercury in samples of the BfR MEAL Study.

The BfR MEAL Study is the first German total diet study and will establish a representative and comprehensive database for dietary exposure assessment in Germany. The present study reports first results of the BfR MEAL Study regarding methylmercury in fish, seafood and mushrooms. In total, 34 MEAL foods were purchased nationally or regionally according to a defined sampling plan, prepared in a representative way for German households, pooled into 49 samples, homogenized and subjected to ICP-MS analysis. Dogfish, tuna, ocean perch, halibut and eel were the fish species with highest MeHg concentrations, while levels in mushrooms and mushroom products had markedly lower MeHg levels. Exposure was estimated by matching the present results with consumption data at appropriate levels of food group aggregation. MeHg exposure for adult high consumers (P 95) exceeded the tolerable weekly intake recommended by the European Food Safety Authority in two age groups (14-17 and 18-24 years). In children, no age group exceeded the recommended tolerable weekly intake. Regional samples differed only slightly in MeHg levels. The differences in exposure found in four regions of Germany were influenced by consumption habits rather than MeHg level in the investigated food.

Draft for internal testing Scientific Committee guidance on appraising and integrating evidence from epidemiological studies for use in EFSA's scientific assessments.

EFSA requested its Scientific Committee to prepare a guidance document on appraising and integrating evidence from epidemiological studies for use in EFSA's scientific assessments. The guidance document provides an introduction to epidemiological studies and illustrates the typical biases of the different epidemiological study designs. It describes key epidemiological concepts relevant for evidence appraisal. Regarding study reliability, measures of association, exposure assessment, statistical inferences, systematic error and effect modification are explained. Regarding study relevance, the guidance describes the concept of external validity. The principles of appraising epidemiological studies are illustrated, and an overview of Risk of Bias (RoB) tools is given. A decision tree is developed to assist in the selection of the appropriate Risk of Bias tool, depending on study question, population and design. The customisation of the study appraisal process is explained, detailing the use of RoB tools and assessing the risk of bias in the body of evidence. Several examples of appraising experimental and observational studies using a Risk of Bias tool are annexed to the document to illustrate the application of the approach. This document constitutes a draft that will be applied in EFSA's assessments during a 1-year pilot phase and be revised and complemented as necessary. Before finalisation of the document, a public consultation will be launched.

Comparison of Bayesian and frequentist methods for prevalence estimation under misclassification.

BACKGROUND: Various methods exist for statistical inference about a prevalence that consider misclassifications due to an imperfect diagnostic test. However, traditional methods are known to suffer from truncation of the prevalence estimate and the confidence intervals constructed around the point estimate, as well as from under-performance of the confidence intervals' coverage. METHODS: In this study, we used simulated data sets to validate a Bayesian prevalence estimation method and compare its performance to frequentist methods, i.e. the Rogan-Gladen estimate for prevalence, RGE, in combination with several methods of confidence interval construction. Our performance measures are (i) error distribution of the point estimate against the simulated true prevalence and (ii) coverage and length of the confidence interval, or credible interval in the case of the Bayesian method. RESULTS: Across all data sets, the Bayesian point estimate and the RGE produced similar error distributions with slight advantages of the former over the latter. In addition, the Bayesian estimate did not suffer from the RGE's truncation problem at zero or unity. With respect to coverage performance of the confidence and credible intervals, all of the traditional frequentist methods exhibited strong under-coverage, whereas the Bayesian credible interval as well as a newly developed frequentist method by Lang and Reiczigel performed as desired, with the Bayesian method having a very slight advantage in terms of interval length. CONCLUSION: The Bayesian prevalence estimation method should be prefered over traditional frequentist methods. An acceptable alternative is to combine the Rogan-Gladen point estimate with the Lang-Reiczigel confidence interval.

Analysis of consumer behavior for the estimation of the exposure to chemicals in personal care products.

In this study, the main objective was to implement an integrative modelling framework in order to support the prioritization and screening of chemicals present in personal care products (PCPs) regarding their potential to expose users across multiple possible pathways. Here, we implemented an exposure-based framework based on product intake fractions (PiFs) calculated using a two-compartment model reproducing the skin uptake and the competing volatilization of chemicals applied on skin during PCP use. The implemented framework enabled to simultaneously and comprehensively accommodate coupled chemical specific parameters (i.e. physical and chemical properties of the candidate chemicals), exposure information specific for product-chemical combinations, and survey data informing on consumer behavior. A case-study, based on the usage pattern data of 22 PCPs investigated among Swiss individuals (Garcia-Hidalgo et al., 2017a) and 113 candidate chemicals chosen for their suspected presence in the PCP categories of interest was defined to evaluate the applicability of the framework. Nonnegative matrix factorization (NMF) and hierarchical clustering were subsequently applied to identify chemicals with the highest exposure potential and to highlight most relevant mixtures of chemicals on the basis of the specific usage patterns of the considered survey individuals.

Use of meat juice and blood serum with a miniaturised protein microarray assay to develop a multi-parameter IgG screening test with high sample throughput potential for slaughtering pigs.

BACKGROUND: Serological screening of pig herds at the abattoir is considered a potential tool to improve meat inspection procedures and herd health management. Therefore, we previously reported the feasibility of a miniaturised protein microarray as a new serological IgG screening test for zoonotic agents and production diseases in pigs. The present study investigates whether the protein microarray-based assay is applicable for high sample throughput using either blood serum or meat juice. MATERIAL AND METHODS: Microarrays with 12 different antigens were produced by Abbott (formerly Alere Technologies GmbH) Jena, Germany in a previously offered 'ArrayTube' platform and in an 'ArrayStrip' platform for large-scale use. A test protocol for the use of meat juice on both microarray platforms was developed. Agreement between serum and meat juice was analysed with 88 paired samples from three German abattoirs. Serum was diluted 1:50 and meat juice 1:2. ELISA results for all tested antigens from a preceding study were used as reference test to perform Receiver Operating Characteristic analysis for both test specimens on both microarray platforms. RESULTS: High area under curve values (AUC > 0.7) were calculated for the analysis of T. gondii (0.87), Y. enterocolitica (0.97), Mycoplasma hyopneumoniae (0.84) and Actinobacillus pleuropneumoniae (0.71) with serum as the test specimen and for T. gondii (0.99), Y. enterocolitica (0.94), PRRSV (0.88), A. pleuropneumoniae (0.78) and Salmonella spp. (0.72) with meat juice as the test specimen on the ArrayStrip platform. Cohens kappa values of 0.92 for T. gondii and 0.82 for Y. enterocolitica were obtained for the comparison between serum and meat juice. When applying the new method in two further laboratories, kappa values between 0.63 and 0.94 were achieved between the laboratories for these two pathogens. CONCLUSION: Further development of a miniaturised pig-specific IgG protein microarray assay showed that meat juice can be used on microarray platforms. Two out of twelve tested antigens (T. gondii, Y. enterocolitica) showed high test accuracy on the ArrayTube and the ArrayStrip platform with both sample materials.

Refinement of dietary exposure assessment using origin-related scenarios.

Global sourcing of food may lead to variability in concentrations of contaminants or pesticide residues. It would be important to incorporate origin influences in dietary exposure assessment. To characterise uncertainties, substance concentrations from GFM (German Food Monitoring), chosen based on the highest CV (coefficient of variation), and food consumption from NVS II (German National Nutrition Survey II) were combined in standard scenarios. Averages or higher percentiles of non-grouped concentrations were used. Additional origin-related scenarios used concentrations grouped by origin. For bromide in tomatoes the most conservative origin-related scenario for Italian tomatoes resulted in the highest exposure of 0.015 mg/d/kg BW. The impact of origin was not covered by the conservative standard scenario (0.006 mg/d/kg BW). For ethephon in pineapples and aluminium in kiwifruits, the highest intake estimates were obtained with the conservative standard scenario resulting in 0.895 µg/d/kg BW and 0.023 mg/week/kg BW, respectively. In these two cases, standard scenarios cover origin influences but the conservative origin-related scenario based on origins with higher concentrations identifies lower exposures of 0.835 µg/d/kg BW for ethephon from African pineapples and 0.014 mg/week/kg BW for aluminium from non-EU kiwifruits. Hence, the inclusion of origin information can refine exposure assessment.

Aggregated aluminium exposure: risk assessment for the general population.

Aluminium is one of the most abundant elements in earth's crust and its manifold uses result in an exposure of the population from many sources. Developmental toxicity, effects on the urinary tract and neurotoxicity are known effects of aluminium and its compounds. Here, we assessed the health risks resulting from total consumer exposure towards aluminium and various aluminium compounds, including contributions from foodstuffs, food additives, food contact materials (FCM), and cosmetic products. For the estimation of aluminium contents in foodstuff, data from the German "Pilot-Total-Diet-Study" were used, which was conducted as part of the European TDS-Exposure project. These were combined with consumption data from the German National Consumption Survey II to yield aluminium exposure via food for adults. It was found that the average weekly aluminium exposure resulting from food intake amounts to approx. 50% of the tolerable weekly intake (TWI) of 1 mg/kg body weight (bw)/week, derived by the European Food Safety Authority (EFSA). For children, data from the French "Infant Total Diet Study" and the "Second French Total Diet Study" were used to estimate aluminium exposure via food. As a result, the TWI can be exhausted or slightly exceeded-particularly for infants who are not exclusively breastfed and young children relying on specially adapted diets (e.g. soy-based, lactose free, hypoallergenic). When taking into account the overall aluminium exposure from foods, cosmetic products (cosmetics), pharmaceuticals and FCM from uncoated aluminium, a significant exceedance of the EFSA-derived TWI and even the PTWI of 2 mg/kg bw/week, derived by the Joint FAO/WHO Expert Committee on Food Additives, may occur. Specifically, high exposure levels were found for adolescents aged 11-14 years. Although exposure data were collected with special regard to the German population, it is also representative for European and comparable to international consumers. From a toxicological point of view, regular exceedance of the lifetime tolerable aluminium intake (TWI/PTWI) is undesirable, since this results in an increased risk for health impairments. Consequently, recommendations on how to reduce overall aluminium exposure are given.

Refining animal research: The Animal Study Registry.

The Animal Study Registry (ASR; www.animalstudyregistry.org) was launched in January 2019 for preregistration of animal studies in order to increase transparency and reproducibility of bioscience research and to promote animal welfare. The registry is free of charge and is designed for exploratory and confirmatory studies within applied science as well as basic and preclinical research. The registration form helps scientists plan their study thoroughly by asking detailed questions concerning study design, methods, and statistics. With registration, the study automatically receives a digital object identifier (DOI) that marks it as intellectual property of the researcher. To accommodate the researchers concerns about theft of ideas, users can restrict the visibility of their registered studies for up to 5 years. The full content of the study becomes publicly accessible at the end of the embargo period. Because the platform is embedded in the infrastructure of the German Federal Government, continuity and data security are provided. By registering a study in the ASR, researchers can show their commitment to transparency and data quality to reviewers and editors, to third-party donors, and to the general public.

Dietary exposure assessment of aluminium and cadmium from cocoa in relation to cocoa origin.

Cocoa contains aluminium and cadmium as environmental contaminants while concentrations are supposed to be country of origin-related. Integrating origin in dietary exposure assessment could refine calculations. Averages or higher percentiles of concentrations in cocoa powder from German Food Monitoring (GFM) and cocoa consumption from the German National Nutrition Survey II (NVS II) were combined in standard scenarios. Additional origin-related scenarios used concentration data grouped into origin A (lower concentrations) and origin B (higher concentrations) as plausible origin information was rare. The most conservative standard scenario resulted in the highest intake estimates for aluminium and cadmium with 0.152 mg/week/kg BW and 0.363 µg/week/kg BW and covered the origin influence calculated in origin-related scenarios. Having plausible origin information would help to refine exposure assessment as it is exemplarily shown here that origin-related lower intake estimates are possible. Using the Eurostat database and the Mintel Global New Product Database (GNPD) generated more origin information for products available on the German market. For Germany, cocoa beans, cocoa powder and cocoa mass were mainly sourced in Côte d'Ivoire, while the Netherlands was the main distributor. Packages of cocoa powders were sourced from different origins.

The disease burden associated with Campylobacter spp. in Germany, 2014.

Bacteria of the genus Campylobacter are an important cause of human illness worldwide. Campylobacter infections are expressed as gastroenteritis and can lead to severe sequelae like reactive arthritis, Guillain-Barré syndrome, irritable bowel syndrome and inflammatory bowel disease. In Germany, Campylobacter-associated gastroenteritis cases are notifiable but there is no reporting obligation for the sequelaes and the disease burden is clearly underestimated. The aim of our study was to quantify reliably the current disease burden of all Campylobacter spp.-associated diseases for Germany with the method of disability-adjusted life years (DALYs). DALYs combine mortality and morbidity in a single summary measure, whereby one DALY represents the loss of one year in full health. For acute gastroenteritis, we estimated 967 DALYs of which only 484 DALYs were detected within the reporting system. Overall, we estimated that 8811 DALYs were caused by the campylobacter-related diseases known so far. 98% of the DALYs were associated with morbidity and 2% with mortality. Mortality was caused by the health outcomes Gastroenteritis and Guillain-Barré syndrome exclusively.

Development of a miniaturized protein microarray as a new serological IgG screening test for zoonotic agents and production diseases in pigs.

In order to monitor the occurrence of zoonotic agents in pig herds as well as to improve herd health management, the development of new cost-effective diagnostic methods for pigs is necessary. In this study, a protein microarray-based assay for the simultaneous detection of immunoglobulin G (IgG) antibodies against different zoonotic agents and pathogens causing production diseases in pigs was developed. Therefore, antigens of ten different important swine pathogens (Toxoplasma gondii, Yersinia enterocolitica, Salmonella spp., Trichinella spp., Mycobacterium avium, Hepatitis E virus, Mycoplasma hyopneumoniae, Actinobacillus pleuropneumoniae, the porcine reproductive and respiratory syndrome virus, Influenza A virus) were spotted and covalently immobilized as 'antigen-spots' on microarray chips in order to test pig serum for the occurrence of antibodies. Pig serum was sampled at three German abattoirs and ELISA tests for the different pathogens were conducted with the purpose of creating a panel of reference samples for microarray analysis. To evaluate the accuracy of the antigens on the microarray, receiver operating characteristic (ROC) curve analysis using the ELISA test results as reference was performed for the different antigens. High area under curve values were achieved for the antigens of two zoonotic agents: Toxoplasma gondii (0.91), Yersinia enterocolitica (0.97) and for three production diseases: Actinobacillus pleuropneumoniae (0.77), Mycoplasma hyopneumoniae (0.94) and the porcine reproductive and respiratory syndrome virus (0.87). With the help of the newly developed microarray assay, collecting data on the occurrence of antibodies against zoonotic agents and production diseases in pig herds could be minimized to one measurement, resulting in an efficient screening test.

[Health reporting of poisoning in Germany-a scientific investigation into the establishment of national poisoning monitoring in Germany : Background, preliminary work, requirements]. / Gesundheitsberichterstattung über Vergiftungen in Deutschland ­ Wissenschaftliche Untersuchung zur Einrichtung eines nationalen Monitorings von Vergiftungen in Deutschland : Hintergründe, Vorarbeiten, Anforderungen.

As a result of decentralized and non-harmonized documentation of poisoning cases within Germany, in comparison to other countries (e.g. the USA, Switzerland, Sweden, and the Netherlands), there is a substantial lack of harmonized national data, in particular for the early detection and risk assessment at the German Federal Institute for Risk Assessment (BfR).The establishment of a national monitoring system for poisonings in Germany is therefore an important step towards enabling faster and more efficient technical access to consolidated accumulated national poisoning data. Additionally, the monitoring would consolidate the publication of annual reports (modelled on the USA National Poison Data System reports) of the German Poison Information Centres and the BfR. Timely assessment of poisoning cases enables fast recognition and confirmation of acute as well as previously unrecognized poisoning hazards. Also relevant here is the safety of products and the associated protection, especially of children, against poisoning accidents.The continuous monitoring and reporting requirements of such a system necessitate additional and challenging work, suitable human resources and IT infrastructure and an annual budget that must be taken into consideration for overall as well as financial planning.

Design, statistical analysis and reporting standards for test accuracy studies for infectious diseases in animals: Progress, challenges and recommendations.

The quality of diagnostic accuracy studies (DAS) for infectious diseases of animals has improved over the last 20 years because of international educational efforts, use of design and reporting standards to guide researchers and test developers, and acceptance of the use of latent class models to account for imperfect reference tests. In this review, we focus on measurement of diagnostic sensitivity and specificity as a measure of clinical validity, describe the leadership role of the World Organisation of Animal Health (OIE) in setting standards for test validation in the context of fitness-for-purpose, and describe how design and reporting quality have facilitated the increased use of systematic reviews and meta-analysis of DAS. Ongoing challenges for design, conduct, analysis and reporting of DAS are identified; and we make recommendations for improvements in these areas for OIE-listed and non-listed infectious diseases.

[Systematic Review of the Burden of Campylobacter-associated Disease]. / Systematischer Review zur Krankheitslast durch Campylobacter spp.

Bacteria of the genus Campylobacter spp. are one of the most common causes of gastroenteritis and can lead to serious sequelae. Several studies have estimated the disease burden of Campylobacter spp. with the quantitative metric of disability-adjusted life years (DALY). The aim of this systematic review is to give an overview of the information available about different countries and periods for which DALYs were calculated and how the different results are comparable. One of the most important transmission pathways for Campylobacter spp. is food. Therefore, special attention was given to studies that only estimated the foodborne disease burden of Campylobacter bacteria. With a systematic search for the period 1/1996-6/2016, one worldwide and 21 country-specific publications of the WHO were identified. Because of the different methods and the quality of the different data sets, the estimated results of all Campylobacter health outcomes of the country-specific studies vary from 0.4 DALYs per 100000 people in France to 109 DALY per population in Poland. The calculation of the attributable foodborne disease burden was based on the estimations of the incidences of all Campylobacter health outcomes with the associated uncertainty for each result. So the estimations of the foodborne disease burden show a large range from 0.5 DALYs per 100000 people in Greek to 21.2 DALYs per 100000 people in New Zealand. This span can only be partially explained by the country-specific variability in the food production, the consumption behavior and the incidence of Campylobacter bacteria.