A microbiological approach to improve the performance of single-chamber bioelectrochemical systems

Tese arquivada ao abrigo da Portaria nº 227/2017 de 25 julho.The need of renewable energy and the constant threat of global warming have motivated the development of emerging sustainable technologies. Among them, bioelectrochemical systems envisage a future where energy and other added value products, such as hydrogen, can be obtained from organic waste streams. Thus, bioelectrochemical systems fit perfectly in the new paradigm with respect to wastes, which should be valorized rather than treated. Bioelectrochemical systems, also known with the name of microbial electrochemical systems (MXCs), combine the electrochemistry with the metabolism of a particular group of microorganisms called exoelectrogens or anode respiring bacteria (ARB). The need of understanding the metabolic activity of ARB and their optimal growth conditions are of high importance to ensure the maximal performance of MXCs. In this frame, this thesis aimed to develop, to study and to upgrade the microbial community of MXCs in view of increasing their performance. For this purpose, ribosomal 16S gene targeted metagenomics study and quantitative real-time PCR (qPCR) techniques were implemented in this thesis. qPCR curves were built to study the evolution of ARB and methanogen microbial communities. Further, 454 pyrosequencing analysis were conducted in order to deepen the composition of microbial electrodes communities. Different experimental setups were designed to study operational conditions under which ARB could outcompete other microorganisms that can undermine MXCs performance, such as methanogens and homoacetogenic bacteria. The first step was the ARB enrichment at different external resistances in Microbial Fuel Cells (MFCs). This study showed that the anode inoculated under low external resistance (12 Ω) in MFC mode showed better performance in the posterior Microbial Electrolysis Cell (MEC) mode (i.e. gave higher current intensity and showed higher H2 production rate) than other MFCs inoculated under higher resistances (220 and 1000 Ω). Moreover, qPCR confirmed that the use of a low external resistance provides an MFC anodic biofilm with the highest content of Geobacter (the most usual ARB). A long term study of the most common methanogens chemical inhibitor: 2-bromoethanesulfonate (BES) was performed in two steps to underline its limitations. A long term acetate-fed MXCs showed BES ineffectiveness caused by Archaea (hydrogen-oxidizing genus Methanobrevibacter, of the Methanobacteriales order) resistance to high BES concentration (up to 200 mM) in MEC. In addition, BES degradation was demonstrated in MFC. Moreover, at higher BES concentration (200 mM), methanogenesis activity decreased but resulted in an increase of H2 recycling by homoacetogenesis due to the favorable conditions for these microorganisms. A MEC inoculated with Geoalkalibacter ferrihydriticus pure culture at high pH revealed high current intensity production (up to 10 A·m-2). These results suggested the possibility to use alkaline conditions with the objective to improve MXCs performance by creating a more selective environment. Then, a high pH community was selected from anaerobic sludge using alkaline medium and an online pH control. High performances were obtained in both MFC and MEC (around 50 mA·m-2 of current density and 2.6 LH2·L-1REACTOR·d -1). Alkalibacter genus was highly detected in an alkaline MFC (37% of the bacterial community) and it was identified as a potential ARB. The presence of Geoalkalibacter genus was confirmed at high pH (9.3) conditions and especially in MEC (43 %). Finally, a successful strategy was developed with the purpose of obtaining a cheese-whey fermentative-ARB syntrophic community. Cheese-whey was fermented to acetate mainly by lactic acid bacteria (Enterococcus genus was 22% of the total bacterial community) and other fermenting bacteria as Sphaerochaeta and Dysgonomonas genera. Exoelectrogenic activity was performed by Geobacter sp. (37%), that used the acetate as electron donor. This microbial community was able to degrade cheese whey to produce directly energy or H2 (0.6 LH2·L-1REACTOR·d-1). Moreover, cheese whey MXCs demonstrated the intrinsic ability to inhibit methanogenic activity without using other external inhibition strategies