Aquaculture is a sustainable means of generating high-quality fish to satisfy the growing global fish consumption. Flatfish are attractive species for aquaculture due to their high market value and consumer demand. However, abnormal development during metamorphosis is often a problem and has hampered the development of successful flatfish aquaculture. Hence, it is critical to understand the specific molecular signalling pathways associated with abnormal metamorphosis. Larval metamorphosis has been studied using histological sections and gene expression profiling. However, the first requires sacrificing fish and the latter uses pool of larvae. Hence, there is an increasing demand for 3D imaging tools to directly visualise the mechanisms of metamorphosis in whole live flatfish. This offers a unique opportunity for optical imaging (OI) modalities, which are becoming popular for imaging targeted biological processes in model organisms (e.g. zebrafish). However, their full potential for biology and non-model organisms is being overlooked. The vision for this fellowship is to develop a non-invasive OI tool for real-time 3D in vivo imaging, for studying the molecular mechanisms regulating flatfish metamorphosis. The project combines recent advances in the mathematics of forward and inverse problems, marine molecular biology and modern OI technologies, including selective plane illumination microscopy and optical projection tomography. The developed OI tools will provide clear high-resolution pictures from the inside of live flatfish. Accurate light propagation models and image reconstruction methods will be developed to obtain high-quality images of biological significance. Moreover, novel acquisition schemes will be implemented to enable real-time 3D imaging. Finally, the OI tools will be used for in vivo flatfish imaging to obtain markers of normal and abnormal metamorphosis. This is essential to establish a sustainable and profitable flatfish aquaculture industry.

Rhodolith beds are one of the most extensive benthic ecosystems along the Atlantic coasts and key environments to continental shelf resilience. Besides providing substrate and habitat for numerous other algae and sessile invertebrates, their ability to calcify, their high abundance and biomass, makes rhodoliths major carbonate producers. Recent empirical estimations suggest that the carbonate marine deposits generated by these organisms represent a total potential carbon sink of 0.4 x 109 t C yr-1. Hence, giving the increasing role of marine ecosystems in the storage of blue carbon, rhodolith beds may represent a not yet considered significant carbon store. Regarding carbon sequestration, studies on rhodolith bed community metabolism are scarce and so far only available for two temperate beds that indicate that they can act both as CO2 source and organic carbon sink. As many marine ecosystems, rhodolith beds are currently under threat related to global climate change (GCC), with local impacts due to increasing coastal urbanization, potentially lowering even further their resilience. Thus, by using a physiological approach, this project will provide much needed information on the basic mechanistic understanding of rhodolith metabolism (photosynthesis, calcification), rhodolith responses to global and local stressors, and rhodolith bed community metabolism and carbon storage along a latitudinal gradient. Taken together, this information will allow assessing the importance of rhodolith beds as natural carbon sinks, thus, help ascertain whether these ecosystems meet the requirements to be integrated into climate mitigation policy, and will further allow quantifying the effects of GCC on their carbon sequestration and storage ability. In addition, it will help recognizing potential interactions between global and local stressors, hence, aid in the development of effective local conservation and management strategies.

The occurrence of toxins typical from tropical environments in European Union (EU) temperate coastal waters has been sporadically reported in the past years, usually preceded by human intoxication episodes. Therefore, in order to avoid public health impacts, there is a need for adequate monitoring programs, establishing appropriate legislation, and for optimizing effective methods for the analysis of these toxins. However, due to the very limited quantitative data and the lack of validated analytical methodology, risk associated with the exposure to emergent toxins in EU is currently not possible to determine. In ETOXPT, we will assess the current situation on the incidence and routes of exposure of emergent toxins (i.e. ciguatoxins, CTXs; tetrodotoxin, TTX; palytoxins, PLTXs; and ß-methylamino-L-alanine, BMAA) in Portuguese coastal waters using reliable LC-MS/MS methods. Additionally, the relationship of emergent toxins concentrations and profiles within trophic levels will be evaluated. These field data will be complemented with a mesocosm experiment, in which the potential of CTXs transfer, biotransformation and biodegradation along the food web will be predicted. We will also evaluate the cytotoxicity of individual PLTXs and CTXs analogues. Concurrently, we will develop innovative approaches to analyze the emergent toxins using nanocontainers. ETOXPT will be useful to advance understanding of emergent toxins kinetics, both within individuals and among trophic levels and could support the development of predictive models of risk assessment and contribute to the integration of emergent toxins in shellfish monitoring programs.

The aim of FISHODOR is to establish the identity of neural network components of the recently discovered tilapia reproductive pheromone. It will identify pheromone olfactory receptor(s) and their intracellular transduction pathways thereby increasing our understanding of olfactory perception and signal integration of fish pheromones. The project will determine the neuronal cell type (i.e. ciliated, microvillous or crypt-type olfactory receptor neurons) detecting the pheromone, it will establish the transduction pathways (i.e. AC/cAMP or PLC/IP3) involved in its olfactory detection and will identify the olfactory receptor(s) that binds the pheromone. If enough time is available it will attempt to map the neuronal network involved in the neuroendocrine response. The combination of electrophysiology, transcriptomics and molecular biology will bring high degree of novelty to explain the mechanism of action of olfactory receptor and neuronal signal integration.