Background Over the past 20 years, water suppliers have become increasingly interested in endocrine disrupting compounds (EDCs) and their negative environmental and health impacts. EDCs are a very large and diverse collection of chemical substances, which cause developmental disorders, such as sexual development problems, in aquatic organisms. The EU’s regulation on chemicals and their safe use, REACH, highlighted EDCs as substances of very high concern (SVHC), while the Water Framework Directive regarded them as main pollutants. A large number of studies have investigated the removal of EDCs, but little is known about how such removal would change their composition and biological activity and thus the impact on aquatic ecosystems and human health. No exhaustive list or regulatory thresholds have yet been defined for EDCs. Most urban wastewater treatment plants (WWTP) were designed 20 years ago before the identification of ‘emerging compounds’ shown to be present in surface water. As a result, these WWTP were not specifically designed to remove such compounds, including EDCs, and alternative water treatment is necessary. Wastewater treatment facilities make a significant contribution to the discharge of EDCs. Given that tighter legislations on EDCs are expected, there is a clear need to correctly measure endocrine disrupting effects and propose new technologies for their removal. Objectives The BIOTTOPE project aimed to develop and validate an automated tool to detect EDCs in wastewater, based on biological aquatic models (fish fry or tadpoles) genetically modified to become fluorescent in the presence of specific EDCs. This tool – the FrogBox – will enable EDCs to be monitored at WWTPs, and will enable additional treatments and strategies for reducing endocrine disrupting pollution to be assessed and subsequently implemented, in order to enhance the performance of combined wastewater treatment systems. The technology will be demonstrated at water treatment plants in two cities: Saint Thibault des Vignes (France) and Brussels (Belgium). Results The BIOTTOPE project firstly designed, constructed and validated an automated biological readout system, using transgenic fish fry or tadpoles continuously exposed to water samples, to evaluate the endocrine disruption impact (estrogenic and thyroidal effects) of wastewater from WWTPs. This system, called the FrogBox, is a biological tool that assesses the level of endocrine disruption of wastewater directly on site and in real time. This information thus allows treatments to be moderated to reduce the impact of effluent on ecosystems. The project then evaluated two existing wastewater treatment strategies for removing micro-pollutants that could cause estrogenic and thyroid disruptions. The first treatment system was a tertiary treatment process, using Actiflo(R)Carb technology, which is based on activated carbon adsorption and an accelerated settling step; the pilot Actiflo(R)Carb unit was tested in Brussels. The second involved optimising the operating conditions of a biological treatment process (a conventional WWTP) to improve the efficiency of ECD removal; this was demonstrated in Saint Thibault des Vignes. The project technologies were demonstrated on full-scale WWTPs in Saint Thibault des Vignes and Brussels (190 000 and 1 400 000 equivalent inhabitants, respectively), to avoid any upscaling problems further down the line. The technologies used in both facilities are standard for urban wastewater treatment. These characteristics would facilitate the ease of replicating the project process at other WWTPs. The main project innovation is the FrogBox, the biological tool for the analysis of endocrine disruption of wastewater directly on site. It enables levels of potential endocrine disruptors in the wastewater to be followed in real time. Consequently, treatments performed to reduce the impact of the effluent on ecosystems can be adapted according to the pollution. Thanks to this automated process for measuring biological effects, it will be possible to modify the instructions for operating all kinds of conventional WWTPs or to enable tertiary treatments to be applied only when the risk of endocrine disruption is present. The results can be accessed online, via an end-user interface, where the results are presented in an easily-understood graphical form. The project plan to commercialise their prototype technology in 2016 or 2017. The evaluation process confirmed the efficiency of the FrogBox, as well as the Actiflo(R)Carb process, for reducing EDCs in water discharged from WWTPs. The project demonstrated that the overall improvement in wastewater quality was greater than a 50% reduction in selected emerging pollutant emissions. By adding a tertiary treatment, such as the Actiflo(R)Carb process, the project showed that estrogen hormones and pharmaceuticals could be removed with an efficiency of 90-100% and 50-100%, respectively. As a result, water quality will be restored, due to fewer discharges of micro-pollutants into aquatic resources, and impacts on aquatic ecosystems and biodiversity will be reduced due to lower endocrine disruption activity. The project process also has direct benefits for human health. For WWTPs, the project technologies represent a potential reduction in investment in operation costs to improve the quality of effluents. It also requires less expenditure on the laboratory analysis (substance by substance) of water quality. By identifying and validating a new strategy to improve the quality of WWTP effluents, the project technology helps to achieve ‘good status’ for surface waters as required under the Water Framework Directive. Further information on the project can be found in the project's layman report and After-LIFE Communication Plan (see "Read more" section).