Bioactivity of extracts from extremophile microalgae Coccomyxa sp. on chloride cells in operculum tissue of Fundulus heteroclitus

Microalgae have been studied because of their great potential as a source of new compounds with important value for biotechnology and to understand their strategies of survival in extreme environments. The microalgae Coccomyxa sp., studied in this thesis, is a poly-extremophile witch was isolated from the acid mine drainage of S. Domingos mine. This environment is characterized by low pH (<3) and high concentration of metals, such as copper and iron. The main purpose of the present work was to evaluate the potential bioactivity in an ex-vivo animal model (Fundulus heteroclitus), and expression on selected genes, of cellular extracts obtained from cultures of Coccomyxa sp. at pH 7 without or with exposure to copper (0.6mM Cu²+). The extracts of Coccomyxa sp. cultured at pH 7 exposed to copper show a great potential to be used as epithelial NKCC inhibitors, revealing their potential use as diuretics, but did not show significant effects on gene expression. Coccomyxa sp. could be a good source of cellular extracts with a great potential to be used in pharmaceutical and biotechnology industries.

Modeling, characterization and microalgal cultivation in an airlift panel photobioreactors.

Microalgae cultures are attracting great attentions in many industrial applications. However, one of the technical challenges is to cut down the capital and operational costs of microalgae production systems, with special difficulty in reactor design and scale-up. The thesis work open with an overview on the microalgae cultures as a possible answer to solve some of the upcoming planet issues and their applications in several fields. After the work offers a general outline on the state of the art of microalgae culture systems, taking a special look to the enclosed photobioreactors (PBRs). The overall objective of this study is to advance the knowledge of PBRs design and lead to innovative large scale processes of microalgae cultivation. An airlift flat panel photobioreactor was designed, modeled and experimentally characterized. The gas holdup, liquid flow velocity and oxygen mass transfer of the reactor were experimentally determined and mathematically modeled, and the performance of the reactor was tested by cultivation of microalgae. The model predicted data correlated well with experimental data, and the high concentration of suspension cell culture could be achieved with controlled conditions. The reactor was inoculated with the algal strain Scenedesmus obliquus sp. first and with Chlorella sp. later and sparged with air. The reactor was operated in batch mode and daily monitored for pH, temperature, and biomass concentration and activity. The productivity of the novel device was determined, suggesting the proposed design can be effectively and economically used in carbon dioxide mitigation technologies and in the production of algal biomass for biofuel and other bioproducts. Those research results favored the possibility of scaling the reactor up into industrial scales based on the models employed, and the potential advantages and disadvantages were discussed for this novel industrial design.

Struttura filogenetica della comunità batterica associata ad Ostreopsis cf. Ovata in colture batch e dinamiche di crescita

Negli ultimi 10 anni i blooms attribuibili alla dinoflagellata bentonica Ostreopsis cf. ovata sono aumentati in termini di frequenza ed intensità lungo le coste del Mediterraneo, avendo ripercussioni negative sulla salute umana e forti impatti sulle comunità marine bentoniche, ciò a seguito della produzione di potenti tossine (composti palitossina-simili) da parte della microalga. Tra i fattori ecologici che innescano o regolano le dinamiche dei bloom tossici le interazioni tra microalghe e batteri sono in misura sempre maggiore oggetto di ricerca. In questo studio è stata analizzata la struttura filogenetica della comunità batterica associata ad O. cf. ovata in colture batch e valutate le dinamiche successionali della stessa in relazione alle differenti fasi di crescita della microalga (oltre che in relazione alle dinamiche di abbondanza virale). Lo studio filogenetico è stato effettuato tramite l’ausilio di metodiche molecolari di sequenziamento di next generation (Ion Torrent). Le abbondanze dei batteri e delle particelle virali sono state determinate tramite microscopia ad epifluorescenza; l’abbondanza cellulare algale è stata stimata tramite metodo Uthermohl. Il contributo della frazione batterica ad elevata attività respiratoria è stato determinato tramite doppia colorazione con coloranti DAPI e CTC. Dai dati emersi si evince che la comunità batterica attraversa due fasi di crescita distinte, una più marcata e concomitante con la fase esponenziale di O. cf. ovata, l'altra quando la microalga è in fase media stazionaria. Per quanto concerne la composizione filogenetica della comunità sono stati rilevati 12 phyla, 17 classi e 150 generi, sebbene i dati ottenuti abbiano rilevato una forte dominanza del phylum Proteobacteria con la classe Alphaproteobacteria, seguita dal phylum Bacteroidetes con la classe Sphingobacteria. Variazioni nella struttura filogenetica della comunità batterica, a livello di generi, tra le diverse fasi di crescita della microalga ha permesso di evidenziare ed ipotizzare particolari interazioni di tipo mutualistico e di tipo competitivo.

Outdoor production of Isochrysis galbana (T-iso) in industrial scale photobioreactors and modelling of its photosynthesis and respiration rate

Isochrysis galbana is a widely-used strain in aquaculture in spite of its low productivity. To maximize the productivity of processes based on this microalgae strain, a model was developed considering the influence of irradiance, temperature, pH and dissolved oxygen concentration on the photosynthesis and respiration rate. Results demonstrate that this strain tolerates temperatures up to 35ºC but it is highly sensitive to irradiances higher than 500 µE·m-2·s-1 and dissolved oxygen concentrations higher than 11 mg·l-1. With the researcher group of the “Universidad de Almeria”, the developed model was validated using data from an industrial-scale outdoor tubular photobioreactor demonstrating that inadequate temperature and dissolved oxygen concentrations reduce productivity to half that which is maximal, according to light availability under real outdoor conditions. The developed model is a useful tool for managing working processes, especially in the development of new processes based on this strain and to take decisions regarding optimal control strategies. Also the outdoor production of Isochrysis galbana T-iso in industrial size tubular photobioreactors (3.0 m3) has been studied. Experiments were performed modifying the dilution rate and evaluating the biomass productivity and quality, in addition to the overall performance of the system. Results confirmed that T-iso can be produced outdoor at commercial scale in continuous mode, productivities up to 20 g·m-2·day-1 of biomass rich in proteins (45%) and lipids (25%) being obtained. The utilization of this type of photobioreactors allows controlling the contamination and pH of the cultures, but daily variation of solar radiation imposes the existence of inadequate dissolved oxygen concentration and temperature at which the cells are exposed to inside the reactor. Excessive dissolved oxygen reduced the biomass productivity to 68% of maximal, whereas inadequate temperature reduces to 63% of maximal. Thus, optimally controlling these parameters the biomass productivity can be duplicated. These results confirm the potential to produce this valuable strain at commercial scale in optimally designed/operated tubular photobioreactors as a biotechnological industry.