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"Adult neurogenesis, i.e., the production of new neurons in the adult brain, has been studied intensively in the past years, both in humans and in animal models, as the understanding of this process can have major clinical implications. The study of neurogenesis in fish has been receiving more attention as, unlike mammals, they possess remarkably high levels of adult neurogenesis and a high capability for neuronal regeneration and replacement where neuronal death has occurred. Less is known, however, on the importance of adult neurogenesis for behavioural plasticity, i.e., for the capacity to change behaviour according to context. As a product of the brain, behaviour relies on functional neuronal networks and it may be expected that more permanent changes in behavioural states imply structural reorganization of neuronal circuits, with the integration of new neurons. Interestingly, the high level of brain plasticity of fish is paralleled by a high degree of behavioural plasticity, with many examples of species that change, either reversibly or irreversibly, their behavioural phenotype during their lifetime, as illustrated by species with functional sex-change and alternative reproductive phenotypes. Flexibility in behaviour may thus require a reorganization of neuronal networks underlying these behaviours with recruitment of new neurons. In this thesis, the link between brain and behavioural plasticity was studied in a small marine fish that inhabits the Mediterranean and adjacent Atlantic coasts, the peacock blenny Salaria pavo. In this species, males adopt nests in rock crevices and attract females into the nest for egg laying, with the male taking care of the eggs until hatching. In some populations, a scarcity of nest sites drives smaller and young males to adopt an alternative reproductive tactic to reproduce. These “sneaker” males mimic the females’ morphology and reproductive behaviour in order to illude the larger nesting males and parasitically fertilize eggs during mating events. Sneaker males later transition into the nesting male phenotype, and this major behavioural transformation in the same animal, first courting males and afterwards courting females, may imply significant reorganization of brain areas associated with reproductive behaviour. During the study, a brain atlas for the species was developed and the main cell proliferation regions, i.e. niches of stem cells birth that may differentiate into cells of the nervous system, characterized. Proliferative areas were observed throughout the whole brain and paralleled the pattern described for other teleosts. Proliferative cells were abundant namely in areas like the olfactory bulbs (granular and glomerular), the anterior subdivision of the dorsomedial telencephalon (DMa), the dorsal and ventral part of the ventral subdivision of the dorsomedial telencephalon (DMvd and DMvv), the dorsal part of the dorsal subdivision of the dorsomedial telencephalon (DMdd), the posterior subdivision of the dorsolateral telencephalon (DLp), the posterior zone of the dorsal telencephalic area (DP), the preoptic area (POA), the dorsal, supracommissural and ventral nucleus of the ventral telencephalic area (Vd, Vs and Vv), the optic tectum and its periventricular grey zone (TeO and PGZ), the ventral zone of the periventricular hypothalamus (Hv), the cerebellum, mainly the molecular layer (CCeM) and the caudal lobe (LCa). A study of the brain nuclei activated during female courtship events using immediate early-genes suggested that some of the areas of the social behaviour network (SBN), a set of brain nuclei underlying the expression of social behaviour across vertebrates, are implicated in female courtship, in particular nuclei in the ventral telencephalic regions. This was followed by an experiment to investigate the possible link between cell proliferation and male tactic switch. Nest availability was manipulated to allow a fraction of sneaker males to adopt a nest and start the transition to nesting males. Ten days after the experiment, some of the smaller males had indeed started switching into nesting males, adopting a nest and starting to develop male secondary sexual characters. The pattern of brain proliferation was studied in these fish to try to confirm that the irreversible behavioural transition would be associated with the reorganization of brain nuclei, assuming that cell proliferation relates to neurogenesis and structural reorganization. Transitional males had elevated cell proliferation levels, as compared to males that remained sneakers, in the dorsolateral anterior and posterior telencephalic regions, thought to be homologous to the hippocampus in mammals. Cell proliferation levels were generally elevated in ventral and ventromedial telencephalic nuclei in both sneakers and transitional males, as compared with nesting males and females, areas considered to be homologous to nuclei of the amygdaloid complex of mammals. There was large variation in proliferation levels within transitional males, and in particular one male more advanced in the transition had higher numbers of BrdU-positive cells than the others. This suggests that a longer time-window for detecting the peak in brain cell proliferation associated with tactic transition in some fish may have been needed. Overall, the study supports the hypothesis that behavioural transition in males of this species is paralleled by an increase in cell proliferation in nuclei potentially relevant for the expression of reproductive behaviours, and establishes the peacock blenny as a new relevant model for the study of neuronal plasticity in vertebrates."

According to the United Nations (UN) World Urbanization Prospects 2018, cities are growing in both size and number, posing challenges for sustainable development. With urbanization, the increase of impermeabilization and landscape fragmentation urges for territorial planning and resources management. To promote resilient and resourceful cites, strategic planning where nature-based solutions (NBS) are included into the built environment, counteracting the negative effects of urbanization through the provision of ecosystem services, should be considered. Green roofs (GR) are an example of NBS that provide a panoply of ecosystem services at the level of the building and city, besides contributing for climate change mitigation and adaptation. This NBS can be included in the urban planning agenda in the new building or as a retrofit solution in the existing built environment. Macao SAR (here mentioned as Macao) is the second more densely populated city in the world, according to the UN Department of Economic and Social Affairs Population Division. The present study aims to assess the inclusion of GR in Macao and assess the ecosystem services that they may provide in a holistic approach aligned with the city’s sustainable development. For this purpose, an extensive GR implemented in Taipa Island of Macao was monitored in terms of associated biodiversity and carbon cycle assessment. Further on, it was carried out a comparison between this GR and a conventional roof (CR) in terms of surface thermal buffering. The results of this study have shown to be very promising in terms of the applicability of GR in Macao and on the relevant ecosystem services that they can provide. First, data loggers were installed in the GR and CR to assess the impact on temperature mitigation. The GR had a significant effect in decreasing and buffering temperature at the surface, with temperatures of 35.6 ºC in the GR substrate comparing with temperatures of 57.7 ºC in the CR surface. The results show that GR can be a cost-efficiency strategy to reduce heating of buildings and their associated environmental and economic costs with cooling. Second, the biodiversity of plants and animals was assessed using a combination of methods. It was observed a high level of biodiversity under a low maintenance and low disturbance of GR vegetation, with 23 species of plants and 34 species of animals identified. This shows that GR can contribute to increase biodiversity in cities. Third, an experiment was run with leaf litter bags to investigate the role of macrofauna (e.g., insects) in the decomposition of organic matter. With macrofauna activity, the decomposition rate increased 17 % when compared to decomposition rates without macrofauna, suggesting that the animals harboured in GR, in particular insects, play a key role in organic matter decomposition and thus on the carbon cycle in cities. This work is, to our knowledge, the first empirical GR study in Macao. The results support the application of GR in Macao for temperature mitigation, biodiversity increase and carbon sequestration, promoting the integration of this NBS in the future urban planning and policy of the city. More generally, the work supports the use of GR as an important NBS to mitigate the impact of climate change in urban settings

Reproduction of the sea cucumber Apostichopus japonicus is critical for aquaculture production. Gonadal development is the basis of reproduction, and lipids, which are among the main nutrients required for gonadal development, directly affect reproduction. We investigated whether gonadal and intestinal lipid metabolism differed between male and female A. japonicus. Transcriptome analysis of the intestines of sexually mature male and female wild-caught individuals revealed differences in gene expression, with 27 and 39 genes being up-regulated in females and males, respectively. In particular, the expression of the fatty acid synthase gene was higher in males than in females. Metabolome analysis of the gonads identified 141 metabolites that were up-regulated and 175 metabolites that were down-regulated in the testes compared with the ovaries in the positive/negative mode of an LC-MS/MS analysis. A variety of polyunsaturated fatty acids were found at higher concentrations in the testes than in the ovaries. 16 s rDNA sequencing analysis showed that the composition and structure of the intestinal microbiota were similar between males and females. These results suggest that sex differences in intestinal metabolism of A. japonicus are not due to differences in the microbiota, and we speculate that gonadal metabolism may be related to intestinal morphology. This information might be useful in improving the reproductive efficiency of sea cucumbers in captivity.

Parental nutrient reserves are directly related to reproductive performance in sea cucumbers. This study focused on the lipid requirements of male and female sea cucumbers Apostichopus japonicus during the reproductive stage and analyzed their physiological responses to a high-fat diet (HFD). The intestinal lipid metabolites and microbiome profile changed significantly in animals fed with the HFD, as given by an upregulation of metabolites related to lipid metabolism and an increase in the predominance of Proteobacteria in the microbiome, respectively. The metabolic responses of male and female sea cucumbers to the HFD differed, which in turn could have triggered sex-related differences in the intestinal microbiome. These results suggest that the lipid content in diets can be differentially adjusted for male and female sea cucumbers to improve nutrition and promote reproduction. This data contributes to a better understanding of the reproductive biology and sex differences of sea cucumbers.

The demand for plastic has led to enormous plastic waste in the environment, which persist and negatively impact the ecosystems. Polyethylene terephthalate (PET) is one of the most common thermoplastic polymers available on the market. The concerns about plastic waste generated an interest in strategies to enhance its biodegradation and finding alternative polymers. In this work was investigated the possibility of using bacteria to degrade PET and to produce bioplastics (Polyhydroxyalkanoates, PHAs). Finally, the integration of the two processes was tested. Overall, the work aimed to investigate the potential to recycle PET into bioplastic using bacteria. The potential of bacterial consortia from various environmental samples to degrade PET granules in liquid matrix was investigated. . The results revealed maximum PET granules degradation of 1.1 % by one of the tested consortia. PET degradation intermediate terephthalic acid (TPA) was not detected at the end of 55 days. Fourier-transform infrared spectroscopy (FTIR) results showed major spectral peak shifts and bends on PET chemical structure compared to non-inoculated control. The biodegradation of PET films buried in the soil (A), with mangrove plants (B), and bioaugmented with a bacterial consortium (C) was also investigated. The experiments were conducted for 270 days at ambient conditions. The results revealed no difference between treatments in the degradation, with a maximum weight loss of 0.118 % in the bioaugmented treatment. Nevertheless, Scanning Electron Microscope (SEM) and FTIR results indicated significant surface changes, spectral peak shifts, and stretches in PET chemical structures. Bacterial consortia isolated from the soil of the experimental treatments were assessed for degradation of PET monomers, TPA and monoethylene glycol (MEG), and intermediate Bis(2-hydroxyethyl) terephthalate (BHET). The consortia were inoculated in flasks containing minimal media with 1000 mg/L TPA or BHET or1113 mg/L MEG as the sole carbon source. Results showed complete degradation of TPA and significant degradation of BHET (96.09%), and MEG (83.65%) by the consortia. In the second part of the study, bacteria were isolated from various environmental samples and screened for PHA production using Sudan Black B staining on colonies and smeared glass slides. Transmission Electron Microscope images were captured to confirm the intracellular PHA inclusions. A total of 35 isolates were screened for PHA, and 22 showed positive staining. The isolate showing higher levels of PHA synthesis (EC2-30-3) was identified based on 16S rRNA gene sequence as Bacillus sp. and selected for PET monomers degradation and fermentation cultures for PHA production. It was cultured in minimal (Moreira et al., 2013) media with 1000 mg/L TPA and 1113 mg/L MEG as the carbon source for eight days. The isolate grew better in media containing MEG, which was selected as a substrate model for PHA fermentation. To integrate PET monomers biodegradation and production of PHA, the isolate was cultured in 0.2 % MEG. A control with 0.2 % of glucose was prepared, and the cultures were incubated for 96 hours. Bacillus sp. EC2-30-3 showed higher PHA accumulation in media supplied with MEG (40.31%) than glucose (25.53%). This is the first report showing that Bacillus sp. uses PET monomer as carbon source to produce a biopolymer. FTIR results of the extracted PHA identified its functional units as C–H, CH3, C=O, and C–O groups. The absorption bands obtained are closely related to the structure of PHB. The study thus confirmed the ability of the isolated bacteria to degrade PET monomers and produce biopolymers. The results of this work open the possibility for upscaling the use of bacteria to mitigate the impact of PET on the environment while producing environmentally friendly bioplastics