The project main objective is to characterize the risk of Ciguatera Poisoning (CP) in Europe including several specific objectives: to determine the incidence of ciguatera in Europe and the epidemiological characteristics of cases; to assess the presence of ciguatoxin in food and the environment in Europe and to develop and validate methods for the detection, quantification and confirmation of the presence of ciguatoxin contaminated specimens.In order to estimate the incidence and to describe the epidemiological characteristics of ciguatera in the Europe, a protocol for a harmonized ciguatera surveillance in the European Union (EU)/Economic European Area (EEA) was elaborated. This protocol includes a ciguatera case definition and two questionnaires for collecting information on ciguatera cases or outbreak. Around half of the countries answered the data call for reporting cases. Four countries reported 34 outbreaks from 2012 to 2019. Spain and Portugal reported outbreaks due to consumption of fish captured in the Canary Islands and Madeira (autochthonous outbreaks), mainly due to consumption of Seriola spp. and Epinephelus spp. In more than half of these outbreaks, the fish was captured by sport fishing. Germany and France reported outbreaks due to consumption of fish imported from outside the EU (imported outbreaks); mainly Lutjanus spp. Spain, Germany and France reported outbreaks in travellers to tropical endemic territories (travel related outbreaks). The ciguatera cases and outbreaks presented neurological symptoms, most of them had also gastrointestinal symptoms and few outbreak cases mentioned cardiovascular symptoms. Six countries (Austria, France, Germany, Portugal, Spain and Switzerland) reported 34 single cases. As a main result, the incidence rate in the EU/EEA was very low (0.0054 cases per 100,000 inhabitants per year). However, Canary Islands constitute by far the area representing the highest risk with an incidence rate of 0.47 cases/100,000 inhabitants. Related to the study of the presence of ciguatoxins (CTXs) in the environment, the main objective was to evaluate microalgae associated to the production of CTXs and the evaluation of CTXs in seafood for the risk assessment of CP. The effort consisted on the evaluation of toxicity in 104 strains of microalgae and 1174 fish corresponding to 77 species. Sampling areas included the Canary Islands, Madeira and Selvagens Islands, Balearic Islands, Cyprus and Crete. Particularly, in the Canary Islands the presence of several dinoflagellates of the genus Gambierdiscus as the causative organism covered the whole archipelago, and the toxicity of the species, particularly G. excentricus, indicate their potential as source of CTX-like compounds. As for fish, according to the data on CTX toxicity, there is quite a high incidence of toxic fish (14% of a total of n=746). Regarding Madeira and Selvagens Islands, the genus Gambierdiscus was detected in both areas. Toxicity of fish has been identified in 42 fish out of 128 fish (33%). The toxic fish identified with the cell-based assay (CBA), defined as primary reference material was transferred to University of Vigo (UVIGO) to continue the characterization of the toxins present. From the eastern Mediterranean Sea, a great diversity of six taxa of Gambierdiscus and Fukuyoa with relatively low cell toxicity was detected. The first fish CTX-like positive by Neuroblastoma Cell-Based Assay (Neuro-2a CBA) from the Mediterranean has been detected in Cyprus with a low toxicity. From the Balearic Island, Gambierdiscus was identified for the first time in 2017. Up to date, only Gambierdiscus australes and Fukuyoa paulensis have been identified. From this area, no fish showed CTX-like toxicity. Efforts in Macaronesia should be centred for a better prediction of ciguatera poisoning cases, and link these to the ecology of ciguatera involving microalgae and fish. Efforts in the Mediterranean should be addressed to better understand the dynamics of toxic microalgal populations and tackle the potential presence of toxins in fish. In order to characterize the risk associated to CP in the EU a sensitive methodology of LC-MS/MS was developed. On the other hand, the preparation of reference materials including the main CTXs responsible for the contamination was considered the secondary objective in order to facilitate the implementation of the Liquid Chromatography-tandem Mass Spectrometry (LC-MS/MS) methods in the EU laboratories to characterize this emerging risk. Caribbean ciguatoxin 1 (C-CTX1) has been identified by LC-MS/MS and further confirmed by LC-HRMS as the main responsible for the CTX toxicity in the samples from the areas selected for this study. The low concentration levels of CTXs found on the samples evaluated has been a key limitation on the completion of the objectives, being necessary to establish contingency plans, not only to overcome the problems of sensitivity that might compromise the confirmation of the toxic profiles, but also the accomplishment of the task of preparing reference materials. The contingency plans involved the development of two complimentary LC-MS/MS approaches, as well as a methodological approach involving LC-MS/MS, Neuroblastoma cell assay and chromatographic fractionations (HPLC and GPC) to be able to characterize the toxins involved in the contaminated extracts. This approach has been also used on the preparation of reference materials to confirm the presence of the C-CTX1 in both pure solutions of C-CTX1 and Fish Tissue Reference Materials (FTRM) containing C-CTX1. The LC-MS/MS analysis of dinoflagellates samples (Gambierdiscus and Fukuyoa spp.) allowed to confirm the lack of correlation with the CTXs contamination of the fish samples from the areas were these dinoflagellates were collected. In fact, the Gambierdiscus toxicity was attributed to several Maitotoxins (MTXs) analogues as well as gambieric acids C and D, Gambierone, 44-methyl gambierone, gambieric acids (or their analogues) and gambieroxide which were identified by LC-HRMS with varying degrees of confirmation in strains of Gambierdiscus and Fukuyoa from the Mediterranean Sea and North East Atlantic Ocean. The EuroCigua project, as seen above, has provided extensive scientific contributions in the fields of public health and epidemiology, environmental assessment of toxins and the associated producing organisms or chemical characterization of CTXs. Hence EuroCigua has contributed significantly to a better characterization of ciguatera in Macaronesia and the Mediterranean Sea. Nonetheless some important issues such as the underreporting of CFP cases, the space and temporal distribution of toxin producing microalgae in some areas, the quantification of their communities, the characterization of the full CTX profiles involved in the CP contamination would require further work for which the availability of reference materials with adequate toxin concentrations is strictly necessary. Capacity building programmes, harmonization of methods, predictive work on ciguatera including modelling, especially focused on the identification of major drivers that may influence ciguatera and better understanding of the toxins involved will improve the assessment and prediction of future scenarios of ciguatera in Europe.