Periodic Reporting for period 3 - EuroBioTox (European programme for the establishment of validated procedures for the detection and identification of biological toxins) Berichtszeitraum: 2019-12-01 bis 2021-05-31 Zusammenfassung vom Kontext und den Gesamtzielen des Projekts The EuroBioTox project focuses on biological toxins of potential bioterrorism risk integrating 63 expert institutions from 23 countries. Previous studies showed that there is a lack of robustness in European preparedness for biotoxin incidents. In this context recent incidents in France and Germany highlighted that extremists take an interest in the production of biotoxins, and that they even succeeded in the production of toxic material. Therefore, the European capabilities urgently need to be strengthened to cope with a potential intentional release of biotoxins. The toxins in the scope of EuroBioTox comprise selected large protein toxins (ricin, abrin, botulinum neurotoxins [BoNT], staphylococcal enterotoxins [SE]) as well as small molecule biotoxins (saxitoxin [STX]). The project is implemented by 13 core partners in collaboration with 50 outer network partners who have access to training, reagents, protocols, and proficiency testing.EuroBioTox focuses on four main objectives:1) Evaluate currently available analytical tools, methods, reagents, and matrices using the expertise of the 13 core partners.2) Improve validation and testing capabilities for biological toxin analysis across the EU.3) Enable laboratories across the EU to further improve their analytical skills in detecting toxins.4) Enable the replacement of animal experiments for the detection of BoNT. Arbeit, die ab Beginn des Projekts bis zum Ende des durch den Bericht erfassten Berichtszeitraums geleistet wurde, und die wichtigsten bis dahin erzielten Ergebnisse The objective of WP1 is the development of certified reference materials (CRM) for different biotoxins. Until month 48, five candidate CRMs (SEB, BoNT/A, BoNT/B, ricin, abrin) were produced and filled. Comprehensive molecular characterisation of the highly pure materials including homogeneity and stability testing demonstrated the required material properties and will enable the assignment of reference values in the near future. The CRMs shall be made available to authorised laboratories.In WP2 proprietary tools and reagents for the detection of toxins were produced and made available in a EuroBioTox repository accessible for all network partners. In a comprehensive validation study, selected detection methods based on reagents from the repository were evaluated in detail.Within WP3, seven proficiency tests (PTs) on different biotoxins were organised, targeting either laboratory-based detection, identification and quantification of biotoxins or in-situ detection with up to 29 participating laboratories. The exercises highlighted technical capabilities of EuroBioTox network partners and demonstrated further room for technical advancement. Four more exercises are in preparation.WP4, dedicated to the implementation of quality procedures, determined a set of criteria to prioritise applicants for different training programmes based on an online survey. A standardised PT protocol containing guidelines for PT organisation was produced, along with validation protocols for immunological and mass-spectrometry methods.WP5 is focused on the organisation and evaluation of 19 training courses on ricin, abrin, STX, BoNT and SE toxins. Until now, 14 training courses on different methods, toxins and levels were successfully conducted including distance teaching courses, supporting capability building within the EuroBioTox network.Based on a comprehensive threat analysis, forensic scenarios were developed in WP6 and information on European organisations with the ability to train first responders was compiled. Procedural guidelines for first responders on toxin sampling, detection and decontamination were developed and demonstrated at a workshop for first responders. Additionally, the development of a forensic peptide database is ongoing. With respect to animal replacement methods, a recovery study for the evaluation of five different methods for BoNT detection against the current gold standard, the mouse bioassay, has been initiated in WP7. A panel of BoNT sero- and subtypes was produced and comprehensively characterised as basis for the comparison of different in vitro and ex vivo detection methods.WP8 aims at the dissemination of information within the network and to relevant decision makers and scientists. To this end, a project website was established counting more than 35000 visitors so far, and three annual newsletters were released. 46 oral and 6 poster presentations, three scientific publications, a EuroBioTox brochure, two explanatory movies as well as different e-learning materials on detection methods for biotoxins were delivered. WP9 is dedicated to the general coordination and management of the project, including all administrative and financial issues. This task covered, among others, the organisation of three face-to-face meetings and a series of virtual meetings so far.In WP10 a number of documents highlighting relevant ethics issues were provided and information on ethics issues are continuously documented. Fortschritte, die über den aktuellen Stand der Technik hinausgehen und voraussichtliche potenzielle Auswirkungen (einschließlich der bis dato erzielten sozioökonomischen Auswirkungen und weiter gefassten gesellschaftlichen Auswirkungen des Projekts) EuroBioTox successfully consolidated an expert network working on biological toxins by linking professionals from the disaster management sector, first responders and industrial partners with expert laboratories in the security, health and food sectors. Progress beyond state of the art was already achieved by production of five candidate CRMs that were produced, filled and comprehensively characterised. Since there is no other certified reference material available worldwide for those toxins, the reagents will be highly useful for expert laboratories and practitioners, having a great potential for further exploitation beyond the lifetime of EuroBioTox. The creation of a European repository with proprietary toxin-specific tools is a prerequisite for the harmonisation of detection methods and helps to spread excellence among network partners. The EuroBioTox training programme broadens technical expertise within EU in the analysis of biotoxins relevant to investigations of alleged use. The implemented 14 training courses were beyond state of the art in that they were tailored to the maximum benefit of the attendees. In that respect, the courses foster multidisciplinary discussions integrating expertise from different fields. The seven proficiency tests organised by EuroBioTox so far offered an opportunity for participants to self-evaluate their technical capabilities. This is important, since there is almost no other option for regular self-assessment for the detection of the toxins in scope of EuroBioTox. With respect to the specific needs of first responders, conceptual guidelines on sampling, detection and decontamination of biotoxins were established and presented at a first responder workshop in 2020. With respect to animal replacement methods, a comprehensive evaluation study for five in vitro methods for BoNT detection against the mouse bioassay has been initiated. Provided it will be successful, the study will directly support implementation requirements specified in Directive 2010/62/EU on the protection of animals.Overall, EuroBioTox aims at implementing a comprehensive mechanism of training, method sharing, improvement of quality assurance measures and proficiency testing. It is expected that the spreading of good analytical practices will improve preparedness and response planning at national and international level. Along this line, EuroBioTox will contribute to minimise potential health and security threats in the EU and will increase resilience of civil society by sound capacity building and technical improvement.