Posters

Abstract: The rate of translocation of ingested nanoparticles (NPs) and how the uptake is affected by a food matrix are key aspects of health risk assessment. In this study, female Sprague Dawley rats (N=4/group) received 0, 1.4, or 13 mg of cerium oxide (CeO2 NM-212) NPs/rat/day by gavage or in a chocolate spread snack 5 days/week for 1 or 2 weeks followed by 2 weeks of recovery. A dose and time-dependent uptake in the liver and spleen of 0.1–0.3 and 0.004–0.005 parts per million (ng/mg) of the total administered dose was found, respectively. There was no statistically significant difference in cerium concentration in the liver or spleen after gavage compared to snack dosing. Microscopy revealed indications of necrotic changes in the liver and decreased cellularity in white pulp in the spleen. The snack provided precise administration and a more human-relevant exposure of NPs and could improve animal welfare as alternative to gavage.   read more...   read less

Abstract: The presence of micro- and nanoplastics in marine and terrestrial ecosystems is causing environmental, ecological, and human exposure concerns, resulting in a recent increase in publications, news articles and reports. Many comprehensive reviews (1-5) and reports (6-10) on this subject identified significant knowledge gaps in analytical methods and methodologies for the isolation, separation, identification and quantification of micro- and nanoplastics mixtures from real-world samples. Publicly available databases for identifying unknown mixtures isolated from the environment are lacking. Here, we report our initial work towards the development of a comprehensive analytical database for the identification, characterization and quantitation of the most common polymers found in the environment, utilizing a hyphenated Thermogravimetric Analyzer (TGA)-coupled to Fourier Transformed Infrared Spectrometer (FTIR)- and Gas Chromatograph Mass Spectrometer (GCMS). Various factors such as TGA temperature ramp rate, polymer particle size, molecular weight, composition of mixtures, and sample mass were investigated and optimized for each polymer. A database of the most common polymer compositions of micro- and nanoplastics (12 different types of polymers, 35 individual polymers) has been developed and will be updated. The TGA-FTIR-GCMS system could separate and identify additives present in the polymer matrix based on thermal degradation profile as well as identify compositions in a range of consumer products. There are several advantages of this hyphenated system compared to Pyrolysis GCMS. These data serve as a baseline to identify and quantify polymers in micro- and nanoplastics mixtures and will be added to a collaborative comprehensive and curated database, to be identified, to make it publicly available. Disclaimer: The views expressed are those of the presenter and should not be construed to represent FDA, NIEHS, or NTP’s views or policies.   read more...   read less

Abstract: Early childhood exposure to heavy metals like arsenic (As), cadmium (Cd), and lead (Pb) through baby foods unfolds many concerns about their toxic effects on growth and health. In this study, occurrence and dietary intake of As, Cd, and Pb in stage 1 infant formula (0–6 months), stage 2 infant formula (7–12 months), cereal-based meals, and biscuits were estimated. First, the levels of As, Cd, and Pb were determined with ICP-MS, followed by the calculation of estimated daily intake (EDI), target hazard quo- tient (THQ), and hazard index (HI) for As and Cd, and margin of exposure (MoE) for Pb. Mean levels of As, Cd, and Pb were the highest in cereal-based meals and biscuits as 15.5–11.1, 5.18–8.76, and 35.2–53.8 μg/kg, respectively. Newborns to 6 months old infants were estimated to be the highest exposed population to Cd and Pb (0.08 and 0.36 μg/kg bw/day), while infants aged 7–12 months old were exposed the highest to As. Based on the THQ, HI, and MoE findings, the current exposure levels from the selected baby foods to As, Cd, and Pb pose low potential chronic risks to both infant age groups. This research provides a roadmap for future investigations in chemical contaminants often detected in baby foods consumed regularly by Saudi infants. KEYWORDS arsenic, cadmium, infant nutrition, lead, Saudi Arabia, toxic elements PUBLICATION LINK: https://onlinelibrary.wiley.com/doi/full/10.1002/fsn3.3485   read more...   read less

Abstract: Introduction: Following the recent success of lipid nanoparticles (LNPs) for the delivery of mRNA COVID-19 vaccines, there is an anticipated increase in demand for such lipid-based therapeutics and vaccines. Accordingly, there is incentive to develop suitable reference materials. Such materials would provide significant benefits to regulatory agencies and industry for product development, quality control, and technology transfer. Our laboratory has developed a continuous manufacturing platform that can produce monodisperse liposomes and LNPs with precise control over particle size1. The objective of this work was to investigate long-term stability and characterization of candidate liposomal reference materials under different storage conditions. Methods: Candidate liposomal reference formulations were prepared using coaxial turbulent jet in co-flow via the ethanol injection method1. Fluid flow rates and other processing conditions were varied to produce monodispersed liposomes at 50, 100, 150, 200 d.nm with 5% charged lipid, as well as 100 d.nm particles with 0%, 2.5%, 5% PEG-lipid. All formulations were characterized via DLS (particle size), pH, and UPLC (lipid components and degradation products), over 18 months at 25 °C and 4 °C storage conditions (monthly triplicate sampling). Results: All samples remained stable and homogeneous over the 18 month study period (below 2-5% error in particle size and polydispersity index values below 0.13). The pH was stable for all samples, although 2.5%, 5% PEG had consistently lower pH stored at 25°C compared to 4°C. Zeta potential remained as expected with the 5% charged lipid formulations around -40 mV and the 2.5% and 5% PEG-lipid around -15 mV. Beyond 18 months, UPLC results showed no cholesterol degradation while PG, DSPE-mPEG2000, and HSPC produced ~ 5.5% total degradation products mainly in the form of Lyso-18 and to a lesser extent Lyso-16 and stearic acid. Conclusions/Impact: Their long-term stability, monodispersity and precise control of particle size, indicates that liposomes produced via continuous manufacturing have potential as reference standards that also have biological significance. Acknowledgements: Funding for this project was made possible by a U.S. Food and Drug Administration grant (75F40120C002-01). This abstract reflects the views of the authors and should not be construed to represent FDA’s views or policies.   read more...   read less

Abstract: In the last few decades, with the global health awareness on the rise, increased interest in health effects associated with cheese consumption has been recognised. Recently, studies have emphasized cheese as the source of bioactive compounds demonstrating antioxidant, antimicrobial, anti-inflammatory, immunomodulatory and analgesic activity. The aim of this article was to analyse pesticide residues and heavy metal content cheeses produced in Croatia. Sample consisted of 80 cheses of different ruminant origin (sheep, cow, goat, mixed), as well as geographical origin. Cheeses were tested for 509 pesticides and for lead (Pb), nikal (Ni), manganese (Mn), cadmium (Cd) and chromium (Cr) as a havy metal representatives. Pesticide residues were analyzed using liquid chromatography (UPLC-MS/MS) and gas chromatography (GC-MS/MS) with triple quadrupole mass spectrometry. In only 2 samples (2.5 %), pesticide residues were found and, in both cases, piperonyl butoxide was found in concentration 0.038 mg/kg and 0.043 mg/kg. For piperonyl butoxide there is no maximum residue limit set. Heavy metals were analyzed on ion coupled plasma with mass spectrometer ICP-MS. Metal residues were found in range min-max (average), <0.005-0.012 mg/kg (0.01 mg/kg), <0.02 - 0.84 mg/kg (0.18 mg/kg), 0.03 - 1.13 mg/kg (0.36 mg/kg), <0.03 - 0.67 mg/kg (0.19 mg/kg), <0.01 - 0.12 mg/kg (0.033 mg/kg), for Cd, Cr, Mn, Ni and Pb, respectively. Only 3 samples had Cd residues above the limit of quantification. Without metal residues 21, 30 and 16 samples were for Cr, Ni and Pb, respectively. Mn were found in all analysed samples. Results showed that cheeses with higher fat and protein content had higher heavy metal content. Keywords: cheese, pesticides residue, heavy metals, ICP-MS, LC-MS/MS, GC-MS/MS adela.krivohlavek@stampar.hr ACKNOWLEDGMENT This work was carried out within the project “Food Safety and Quality Center” (KK.01.1.1.02.0004). The project is co-financed by the European Union from the European Regional Development Fund.   read more...   read less

Abstract: Antimicrobial resistance (AMR) in livestock is a public health threat due to the risk of zoonotic transmission to humans and its negative consequences on animal health and welfare when diseases cannot be treated. Antimicrobial treatment options for post- weaning diarrhoea in pigs are increasingly limited because of planned restrictions in the use of zinc oxide and colistin, two current choices for prevention/treatment of post- weaning diarrhoea caused by enterotoxic E. coli (ETEC). Faecal filtrate treatment (FFT) was developed during AVANT, a H2020 funded European project, as a promising intervention. In a project financed by the Independent Research Foundation Denmark and SEGES-Innovation (#8022-00188B), the treatment of piglets with faecal filtrate transplantation showed significant reduction of diarrhoea prevalence after weaning and reduction of mortality in the weaning period.   read more...   read less

Abstract: The dependence of cancer cells on the essential amino acid methionine has triggered extensive investigations of methionine γ-lyase (MGL) - a bacterial pyridoxal 5’-phosphate-dependent enzyme that degrades methionine - as a therapeutic tool capable of inhibiting cancer cells growth through methionine starvation. Our most recent research on the MGL-mediated depletion of methionine is devoted on the development of a novel delivery platform for MGL - based on enzyme adsorption on spherical citrate-capped gold nanoparticles (AuNPs) - to overcome the numerous challenges related to enzyme-based therapeutics (i.e. protein stability, bioavailability and immunogenicity) and boost clinical development and applicability. A combination of multiple biochemical and biophysical techniques allowed an extensive characterization of MGL-AuNP conjugates in terms of particle size and size distribution, protein loading yield, impact of functionalization on protein structure and enzymatic activity.   read more...   read less

Abstract: Here, a systematic approach (‘Early4AdMa’) is proposed to identify and describe potential safety, sustainability and regulatory issues of advanced materials at the early stages of their development or use. The approach described here is based on a system developed by the RIVM (NL), BfR (DE), BAuA (DE) and UBA (DE), and adapted within the OECD WPMN Steering Group on Advanced Materials. The outcome of the approach can inform decision makers, policy makers, risk assessors, and regulators and could facilitate regulatory preparedness and making timely decisions to avoid or reduce safety and/or sustainability impacts. As such, the Early4AdMa can be considered as a pre-regulatory and an anticipatory risk governance tool. Thereby, the system could be implemented in the development, production, use, and end-of-life of safer and more sustainable advanced materials and the products into which they are incorporated. This would allow their exploitation to be safer and more sustainable, facilitating the delivery on solutions for societal and environmental challenges via application of advanced materials.   read more...   read less

Abstract: Based on the recommendations of the EFSA Roadmap on NAMs, EFSA has funded projects to promote the use of NAM-based IATA to fill data gaps in the risk assessment practice, integrating the available toxicological information with newly generated NAM-based studies developing proof-of-concept case studies. An example in the field of nanomaterials is the EFSA NAMs4NANO Project, established thought partnership with EU Member State organisations. The new EFSA NAMs4NANO Project aims to develop a “Qualification system for NAMs for EFSA” and, in parallel, a number of risk assessment and methodological proof-of-concept case studies.   read more...   read less

Abstract: The JRC has led the work towards the development and adoption of the OECD TG for VSSA of nanomaterials (TG 124). The main steps undertaken are described. These include : i) preparatory steps with experts for the selection of testing materials, the information exchange on existing protocols and drafting a standard operating procedure (SOP) ii) organisation and evaluation of an interlaboratory study to gain information on the domain applicability and method transferability and iii) drafting and review of the test guideline until its final adoption by the OECD Working Group of the National Coordinators for the Test Guidelines Programme (WP-MNT). This TG will be taken up by the EU Test Methods Regulation.   read more...   read less

Abstract: Intestinal organoids mimic the structure, function and physiology of human intestine in vitro. Thanks to these advantages, intestinal human organoids (iHO) are suitable for various applications from drug discovery to translational research in biomedical field. These benefits make intestine human organoids (iHO) attractive for a range of applications, including drug discovery and toxicity assessments. There are currently no standards in place to monitor whether iPSCs successfully differentiate into healthy or sick tissue-like standards. In this study, human induced pluripotent stem cells (iPSCs) were used to create an iHO three-dimensional culture. After differentiating in vitro, iHO exhibit the intricate crypt-villus architecture of the intestine as well as the presence of enterocytes, goblet cells, and enteroendocrine cells, which are typical cell types of the intestinal tissue. The study will suggest some quality criteria demonstrating the successful differentiation into intestinal cell types that can be used for in vitro toxicity testing. iPSCs pluripotency and iHO formation has been identified by a first morphological observation in culture and then by analysis of specific marker expression by immunofluorescence staining. The analysis confirmed that iPSCs showed the typical colony morphology and expressed the pluripotency-associated proteins (Oct4, Nanog, Sox2 and TRP-1). iPSCs were cultured for several passages in vitro maintaining their pluripotency and differentiation ability. After differentiation, iHO showed a heterogenous morphology based of their maturations stage: iHO with spheroid shape (immature) and iHO with a crypt-villus architectures (mature). Then, the efficacy of their differentiation has been confirmed by the expression of specific marker of human intestinal development including LGR5 (intestinal stem cell), VILLIN (enterocyte), MUCIN 2 (goblet cells), CHROMOGRANIN A (enteroendocrine cells) and lysozyme (paneth cells). The study proved the biological relevance of intestinal organoids in assessing the absorption and toxicity of nanomaterials. Additional research is required to demonstrate the toxicological relevance using chosen representative materials.   read more...   read less

Abstract: The current understanding on potential hazards associated with titanium dioxide in nanoforms is still not fully conclusive. In the presented study, we investigated the capability of different cellular models to internalise and accumulate TiO2. The use of radioactive labelled TiO2 allowed us to quantify the accumulated nanomaterial in cells even after 30 days in culture. Such in vitro platforms can support the investigation of long-term effects of nanomaterial with biological systems but require an in depth characterisation of the in vitro model, in particular when sophisticated and long-term exposure protocols are used with the aim to investigate cell type specific impairments that are going beyond the modulation of classical and well described biological pathways.   read more...   read less

Abstract: Reference particle models are essential to obtain an accurate perspective of how environmental nanoplastics behave in natural systems and to generate data on their environmental fate and impact on living organisms. However, although the current available models are particularly important for filling the initial knowledge gaps on nanoplastics, they do not demonstrate enough diversity and/or accuracy to represent the actual heterogeneity of the physical and chemical properties of environmental nanoplastics. In this framework, the Joint Research Centre (JRC) and the National Institute of Standards and Technology (NIST) are collaborating on a challenging project, which aims to identify a novel strategy for the production of aged nanoplastics. These new test materials are designed to become environmentally relevant models made by the combination of mechanical abrasion and artificial UV exposure for ageing. The objective is to obtain nanoplastics, which mimic naturally weathered ones in size distribution and surface chemistry. The scalable production of these nanoplastics is investigated to make them available to stakeholders. This test material will have applications for the development of new analytical methodology to detect nanoplastics, to better understand their role in carrying other pollutants and increase knowledge related to their behaviour and fate in the environment.   read more...   read less

Abstract: Based on the recommendations of EFSA “Guidance on risk assessment of nanomaterials to be applied in the food and feed chain”, the NAMS4NANO project was launched to establish proof of concept case studies addressing nano specific concerns when using new approach methodologies (NAMs). Notably, there is a need to address nano considerations of possible uptake by cells at the intestinal level and potential adversities following the intracellular uptake using NAMs. Adverse Outcome Pathways (AOPs) is a conceptual framework that defines the causal chain of perturbed key events (KEs) leading from an exposure to a stressor to an adverse outcome. AOPs are believed to be highly instrumental in comprehending the biological and toxicological significance of NAMs. However, despite the numerous AOPs already created for food safety evaluations, AOPs for nanomaterials in food are still very limited. In that context, we aimed here to build AOPs focusing on cellular uptake and translocation of nanomaterials at the level of the intestinal barrier. As the intestinal epithelium comprises different cell types with tissue-specific functions (enterocytes, goblet cells, M-cells and stem cells), we envisioned to develop a series of AOPs relevant to each cell type. Based on the literature and available knowledge, we proposed to evaluate if cellular uptake of nanomaterials at the intestinal level can lead to endocytosis machinery dysfunction with accumulation in the lysosome compartment eventually leading to mitochondrial dysregulation, cell death, intestinal barrier disruption, decrease of mucus production and/or inflammation depending on the cell type. These AOPs will then permit to anchor relevant NAMs (in vitro assays) to the appropriate KEs in a mechanistic-based approach of risk assessment. Thus, in line with EFSA NAMs roadmap, building AOPs relevant to nanomaterials in food along with in vitro testing might facilitate the regulatory use and integration of data from NAM methods for assessing the toxicity of nanomaterials.   read more...   read less

Abstract: Overview about activities of JRC’s Nanobiotechnology Lab, including (a) Fast adaptation to emerging health threats; (b) Response to emerging pollutants, i.e. micro(nano)plastics; (c) Support to EU policies to promote Safe & Sustainable Innovation for advanced materials; (d) Open Access to JRC Infrastructure including training & capacity building.

Abstract: The JRC has led the work towards the development and adoption of the OECD TG for VSSA of nanomaterials (TG 124). The main steps undertaken are described. These include : i) preparatory steps with experts for the selection of testing materials, the information exchange on existing protocols and drafting a standard operating procedure (SOP) ii) organisation and evaluation of an interlaboratory study to gain information on the domain applicability and method transferability and iii) drafting and review of the test guideline until its final adoption by the OECD Working Group of the National Coordinators for the Test Guidelines Programme (WP-MNT). This TG will be taken up by the EU Test Methods Regulation.   read more...   read less

Abstract: In the last few years, EFSA started dedicated projects on New Approach Methodologies (NAMs) to promote their implementation in regulatory risk assessment. Information from animal studies is frequently available, but often presents data gaps that create uncertainties and inconclusive risk assessment. The main objective of the EFSA NAMs projects is to promote a comprehensive chemical risk assessment to fill data gaps in the context of EFSA’s remit (i.e. food and feed area) while fostering the transition towards a mechanistic-based risk assessment. Different pilot projects have been launched to investigate the use NAMs for the hazard and exposure assessment of the pesticide Tebufenpyrad, for the hazard assessment of dietary nanofibers, to explore the immunotoxicity of the contaminants PFASs and to explore interspecies metabolic differences on essential oils as feed additives. Additionally, EFSA has launched a dedicated project to explore the use of Artificial Intelligence for extracting and integrating NAM-based data for chemical risk assessment.   read more...   read less

Abstract: In the EU context of regulated food and feed products, when an application concerns a material that meets the definition of engineered nanomaterial set out in Regulation (EU) 2015/2283, the application must follow the EFSA Scientific Committee Guidance on Risk Assessment of Nanomaterials, published in 2018 and updated in 2021. This Guidance covers the application areas within EFSA’s remit, including novel foods, food contact materials, food/feed additives, and pesticides, in the context of human and animal health risk assessment. The Guidance proposes a structured pathway for carrying out the safety assessment of nanomaterial and provides practical suggestions for the types of testing needed and the methods that can be used. However, nano-specific considerations for risk assessment may be required also for conventional materials that contain a fraction of small particles, but do not meet the definition of engineered nanomaterial. For this reason, following a mandate from the European Commission, EFSA has developed and published in 2021 its new Guidance on Technical Requirements to establish the presence of Nanoparticles, which sets out the information requirements for applications in the regulated food and feed product areas and establishes criteria for assessing the presence of a fraction of small particles, including particles requiring specific assessment at the nanoscale, in conventional materials which do not meet the definition of engineered nanomaterial. Both Nano Guidances should be considered as complementary, and their application should be integrated into the risk assessment of relevant sectoral frameworks.   read more...   read less

Abstract: Current approaches for the assessment of environmental and human health risks due to exposure to chemical substances have served their purpose reasonably well so far. Nevertheless, the assessment systems in place for different chemical regulations are faced with several challenges, ranging from a growing number of chemicals to changes in the types of chemicals and materials produced. This has triggered awareness of the need for a paradigm shift, notably appearing in the CSS, requiring new concepts for chemical risk assessment. As a result, new approach methods (NAM) and next-generation risk assessment (NGRA) are generally regarded as the way forward. However, incorporating new scientific insights and innovative approaches into hazard assessment in such a way that regulatory needs are adequately met has appeared to be challenging. The European Partnership for the Assessment of Risks from Chemicals (PARC) has been designed to address various challenges associated with innovating chemical risk assessment. Its overall goal is to consolidate and strengthen the European research and innovation capacity for chemical risk assessment to protect human health and the environment. With around 200 participating organisations from all over Europe, including three European agencies, and a total budget of over 400 million euro, PARC is one of the largest projects of its kind. It has a duration of seven years and is coordinated by ANSES, the French Agency for Food, Environmental and Occupational Health & Safety. The WP hazard assessment focusses on closing data gaps for substances where data are lacking, development of new approach methods as well as increasing their regulatory robustness.   read more...   read less

Abstract: There are several stages in the supply chain of a mass manufactured herbal medicine. Provision of the raw material, refinement and processing, receiving FDA approvals, distributing the goods through authorized channels of distribution, and ultimately reaching the consumer, are all important components of such an endeavor. How raw ingredients are supervised and regulated is a crucial aspect of herbal medicine mass production that ensures product efficacy and safety for consumers. Naturally, the Food and Drug Administration needs to improve regulatory standards and the ability to track such raw materials in order to combat counterfeiting. By tracing crude plants, extracts or essential oils that were imported or produced locally as raw material form, all the way to the finished product the consumer receives, this study attempted to improve raw ingredient tracking capabilities. Considering the vast spectrum of herbal crops that should be monitored in farmlands, but fall outside the FDA’s supervision jurisdiction, the study initialized its monitoring approach only towards prepared raw materials. In the venture to accomplish this goal, it was essential to apply digital and contemporary technical solutions, such as Blockchain, which helped with data transmission and improved the accuracy of authenticity detection. Through our effort and a way that was carefully considered, we established a method to improve and develop such tracking systems. Corresponding author: Faezeh Vahidi Motlagh   read more...   read less

Abstract: Metagenomic analysis has proven to be an invaluable method for determining the taxonomic breakdown of the microbes in an environment or a host. This has led to an interest in the use of metagenomics in public health for pathogen monitoring. However, most computational tools for metagenomic analysis have the general purpose of classifying sequence reads into different taxa, rather than the specific goal of identifying particular pathogens, and distinguishing them from closely or more distantly related taxa. We have developed an in silico benchmark dataset, which addresses the problem of distinguishing pathogen reads from related reads in a short read metagenome, introducing decoy reads from a closely related strain. The benchmark takes five exemplar bacterial pathogens and simulates, in silico, the pathogen and decoy taxon reads, against a simulated background of other species representative of a metagenome. Here, we describe the benchmark dataset, provide the code used to generate it and other potential benchmark datasets, and discuss the issues around the application of this benchmark to different pathogen identification approaches. The goal is that this benchmark dataset should be used in the assessment and development of metagenomic analysis workflows for pathogen detection, and form the basis of an expanding suite of various benchmarks addressing different aspects of the pathogen identification problem.   read more...   read less

Abstract: Several states are putting drastic policies in place to phase out animal use in biomedical research and regulatory testing. As understood from arguments for and against animal research, these political and regulatory moves result from two major unresolved issues: one, the ethical aspects of experimenting on nonhuman animals and second, the robustness of animal research as a methodology of scientific investigation of human physiology. Whilst these issues remain, the contribution of research on animals to human and veterinary medicine as well as other relevant aspects of our livelihood, including agricultural production and environmental health, cannot be downplayed. By experimenting on animals, curious scholars of old gained knowledge on biological processes which we still apply today. Despite the massive biomedical transformations witnessed today, the many health problems of the human race incite me to conclude that we still have a long way to go. We have understood well enough but there is still much more to understand about our biological processes in order to address our ever increasing health problems. For now, the novel replacement technologies holding out the promise of hope for total replacement of animal research are generally based on what we have understood so far about animal and human biology. Experts on one side wonder: what about what we do not know yet about the “black box” animal/human physiologies? How well would novel animal replacement technologies give perfect replication of animal/human physiologies? What about the need to unravel the yet hidden explanations of biological processes we are not yet aware of? Those on the other side also wonder: would animal usage in the past always justify its continuous usage in the present and future despite the ethical and technical challenges it poses? Is total phase-out in sight? Is policy considerate enough of all the complexities above? Instead of rushed policies which might seem “one-sided”, we should consider all aspects of the dilemmas involved. In this paper, I suggest taking a more consensual and pragmatic approach to address the two fundamental issues underlying the call for total phase-out whilst addressing the questions all parties ask. Keywords: animal research methods, replacement technologies, ethics, policy, science regulation   read more...   read less

Abstract: Chemical risk assessment is currently undergoing a paradigm shift driven by ethical considerations, regulatory action and the need to ensure the safety of chemicals using efficient, cost-effective and robust methods. Non-animal approaches represent a powerful alternative to improve safety assessments by using more human-relevant tools providing a good coverage of key biological targets. Next-generation risk assessment (NGRA) provides a framework integrating data coming from diverse so called new approach methodologies (NAMs) into the decision-making process, allowing for safety assessments to be conducted without the use of animal data. The assumption behind the use of NGRA is, that if the exposure level of a chemical in humans is below the concentration needed for exhibiting any biological effect, it is unlikely it would trigger any toxicity. Whitin this context, estimates of exposure can be obtained using physiologically based kinetic (PBK) models while potential biological effects are assessed by means of points of departure (PODs) from a range of in vitro assays. These assays are selected to both provide a good biological coverage and to detect very early biological perturbations before the onset of any adverse effect. Then, the PODs and exposure estimates can be combined into a single metric often referred to as margin of safety. Here we present a NGRA framework under development at Nestlé, built to support the safety assessment of food chemicals and food contact materials without the need of animal data. The whole approach relies on the chemical mode of action (MoA). Depending on whether the MoA is known or not, a defined panel of in silico and in vitro assays will be used to estimate internal exposures and in vitro PoDs. The derived margins of safety will be then used to define the risk. The current framework will undergo validation by running multiple case studies to assess accuracy and reliability with the final goal of being implemented as a routine approach for safety assessment.   read more...   read less

Abstract: Objective: Evaluating the impact of microbial pollution through artificial intelligence methods. To build a more feasible Campylobacter cross-contamination model for poultry slaughter in China. To proposed corresponding suggestions for improving the slaughter and process operations. Highlights: An estimation model for the key coefficient of cross- contamination during the slicing process of broilers was established using artificial intelligence methods. This model uses genetic algorithm(GA) to search for large-scale coefficient space and indirectly estimate coefficients, trying to make the model distribution fit the actual distribution as much as possible by this algorithm and further by gradient descent(GDA). In the process of optimization, the fitness function resorts to the accuracy evaluation method of Kolmogorov–Smirnov (K-S) test, together with unbiased standard deviation and unbiased mean value.   read more...   read less

Abstract: Purpose: The Food Safety Commission of Japan (FSCJ) is a risk assessment organization for science-based assessment of food safety risks to human health in the Cabinet Office of Japanese Government. In 2019, FSCJ published “the Guidance on the Use of the Benchmark Dose (BMD) Approach in Risk Assessment by FSCJ (FSCJ BMD Guidance (https://www.fsc.go.jp/english/what_we_do.data/For_HP_revision_BMD_approach_guidance.pdf))”, which provides basic stance and procedures for the use of the BMD approach for deriving a suitable point of departure (POD) in the risk assessment of chemicals in food. In recent years, BMD approach based on Bayesian estimation has been gaining underway used for decision making at regulatory frameworks in various nations. Hence, FSCJ Working Group (WG) on assessment methodology development has discussed the adaption of Bayesian estimation to the current FSCJ BMD Guidance. Results: The main objective of revision of the FSCJ BMD Guidance was to introduce the use of Bayesian estimation. In WG, the points to consider for using Bayesian estimation in BMD approach and the issues need to be clarified for the use were discussed. Based on results of the discussion, the points of the concept using Bayesian estimation on BMD approach were summarized as the appendix in the revised FSCJ BMD Guidance. Discussion: WG considered that it would be difficult to promptly shift statistical paradigm of BMD approach from the existing frequentist to the bayesian. WG also, however, agreed that the effort to introduce of Bayesian BMD modeling should be executed. Such experiences would promote understanding of the technical advantages of Bayesian BMD approach for deriving a suitable POD in food risk assessment. The public consultation of the revised guidance will end in early Aug 2023. The finalization is planned to be in autumn 2023.   read more...   read less

Abstract: The EFSA Pesticide Residue Intake Model (PRIMo) has, since its first version in 2007, become the standard tool used at the EU level to estimate the acute and chronic dietary exposure to pesticide residues. The PRIMo is a deterministic model, primarily used as a prospective screening tool for dietary risk assessment in the framework of Regulation (EC) No 396/2005 and Regulation (EU) No 1107/2009. The aim of the project was to develop a new revision of the tool and transform the former Excel-based calculation spreadsheet to an online web-application with updated features and new functionalities. One of the main novelties is that the model uses individual consumption data from the Comprehensive Food Consumption Database of EFSA, where the foods reported as consumed were disaggregated to their ingredients and converted to their raw primary commodity (RPC) equivalents by means the EFSA RPC model. The use of this consumption database allows the integration of additional EU countries and harmonized population classes in the assessment compared to the previous version. The food consumption and occurrence data considered in the assessments are coded both according to the pesticide legislation categories and in the EFSA FoodEx2 classification system. The innovations allow to have more detailed exposure assessment results, including results for minor products and processed foods. Another important new feature of PRIMo 4 is that in addition to the mean exposure, calculation of the highest reliable percentile (HRP) for both the chronic and acute estimation provides information on highly exposed subjects. Exposure assessment results are exportable to different formats including graphical outputs and MS Excel tables which allows for further data processing and re-use of the data in future sessions. The beta version of the PRIMo4 tool was subject to a public consultation and its final version is planned to be published by end of 2023.   read more...   read less

Abstract: In vitro tests, like embryonic stem (ES) cell-based assays, provide advantages for teratology research, such as speed, cost-effectiveness, scalability, and mechanistic investigations. To enhance their regulatory application in teratogenic chemical exposure assessment, we aimed to define the applicability domain of the mouse embryoid body test (EBT) model. We cultivated mouse embryonic stem cells (mESCs) in a feeder-free environment, successfully forming embryoid bodies (EBs) using the 384-ultra low attachment (ULA) well format. The morphological changes in the EBs were readily discernible through high-content screening machine analysis. To define the applicability domain of the EBT model, 39 compounds (33 from ICH Reference Compound List, 6 from commonly assessed compounds) were tested. These compounds covered categories: Channel modulator, DNA modifier, enzyme modulator, kinase modulator, nucleoside modulator, receptor modulator, transcription modulator, and others. The EBT effectively detected the teratogenic potential of kinase modulators, nucleoside modulators, and transcription modulators. However, several limitations have been identified in previous predictions using EBT models. One of them is that chemicals without detectable ID50 and IC50 values in EB, mESC, and 3T3 cells at the highest soluble concentration hindered the predictions. Most teratogens exhibit effects of inducing general toxicity leading to cell death and also possess mutagenic properties that cause genetic alterations. The mechanism of action employed by teratogens can disrupt various cellular pathways involved in embryonic development. In this study, we intentionally induced a reduction in EB size using the well-known teratogen 5-fluorouracil to investigate the specific genes impacted by this perturbation. By employing RNA sequencing, we have successfully identified several genes associated with the teratogenicity induced by 5-fluorouracil in EBs. These identified genes exhibit promising potential as candidates for the development of more precise reporter gene assays, expanding the applicability domain of the assay and enabling enhanced prediction of compound teratogenicity.   read more...   read less

Abstract: Currently, risk assessment is largely based on animal experimentation. These studies do not reflect human biology or involve mechanistic toxicodynamic information. However, accurate risk assessment using human cells is hampered by limitations in the understanding of the population variability in toxicodynamics. The TD-TRAQ project aims to understand population variability of critical toxicity-related pathways to derive uncertainty factors (UFs) for toxicodynamics. We seek to prove that systematic transcriptomics-based analysis of toxicodynamics properties of diverse chemical entities in combination with statistical modelling approaches will provide a scientific basis for setting toxicodynamics UFs for implementation in risk assessment. Adverse outcomes are marked by modulation of (stress response) cellular signalling pathways that drive the altered expression of pathway specific genes. Therefore, quantitative mapping of the concentration-response of diverse chemicals with specific mode-of-action representing critical adverse outcome pathway (AOP) responses using transcriptomics technologies will allow qualification and quantification of such signalling responses. Application of this approach in a large panel of freshly isolated peripheral blood mononuclear cells (PBMCs) that are derived from a large group of healthy volunteers with different gender, sex and ethnicity will allow the discovery of the population variability in pathway activation. Statistical modelling of the large dataset will subsequently allow quantification of the population variance as well as calculation of UFs related to toxicodynamics. We will integrate various technological approaches. After isolation of blood leukocytes, we apply both high-content imaging and flow cytometry to assess the functional readouts related to cytotoxicity endpoints. In parallel we use the cells for high-throughput transcriptomics approaches. The latter data are used to establish gene co-expression networks for biological interpretation. Bioinformatics is used to derive the concentration response effects at the gene and pathway level of the various chemical exposures for all the different individual donor cells. These quantitative datasets are then used for population modelling to derive UFs with the ultimate aim to refine current chemical risk assessment practice. Supported by the TD-TRAQ project funded by European Food Safety Authority (OC/EFSA/SCER/2021/03)   read more...   read less

The Global Coalition for Regulatory Science Research (GCRSR) was established in 2013 under the leadership of the US-FDA. The mission of GCRSR is to foster the uptake of emerging technologies by engaging regulatory agencies in the global context. This is an international coalition with the objectives of facilitating education, scientific training and scientific exchanges in the field of regulatory science. It focuses on research to support regulatory decision making by identifying and promoting best practices to understand and interpret data from innovative technologies such as genomics. To date, GCRSR discussions have been focused on (1) defining the role of global research collaborations in advancing regulatory science and its impact on public health, (2) exploring the future of regulatory science research as a tool for advancing regulatory science in the areas of food safety and medical products, and (3) developing strategies for training of regulatory scientists in a global setting. Consequently, its main activities involve (1) holding workshops and scientific meetings, including the annual Global Summit on Regulatory Science, to discuss the current development of new technologies and their potential utility in the regulatory settings; (2) exchanging scholars and students for the purpose of providing education and training courses; and (3) enhancing the development and use of regulatory science principles. To achieve these goals, the annual GSRS meetings have been instituted. GSRS conferences provide a venue where regulators and researchers can meet and develop collaborations to address the challenges and needs in the interest of advancing regulatory science. Eight meetings have been conducted so far, with the first (GSRS2011) taking place in the US (Slikker et al., 2012), GSRS12 in China (Miller, et al., 2013), GSRS13 in the US (Howard, et al.), GSRS14 in Canada (Tong, et al., 2015), GSRS15 in Italy (Healy et al., 2016), GSRS16 in the US (Patri, et al., 2016), GSRS17 in Brazil (Slikker et al., 2018) GSRS18 in China (Thakkar et al., 2019) GSRS20 virtual (Allan et al., 2020) GSRS21 virtual (Thakkar et al., 2021) GSRS22 was held in person in Singapore and the GSRS24 will be held in the U.S.A).   read more...   read less