Shortage of potable water is a problem that affects many nations in the world and it will aggravate in a near future if pertinent actions are not carried out. Decrease in consumption, improvements in water distribution systems to avoid losses and more efficient water treatment processes are some actions that can be implemented to attack this problem. Membrane technology and biological processes are used in wastewater treatment to achieve high water quality standards. Some other technologies, besides water treatment, attempt to obtain energy from organic wastes present in water. In this study, a proof-of-concept was accomplished demonstrating that a Microbial Electrolysis Cell can be fully integrated with a Membrane Bioreactor to achieve wastewater treatment and harvest energy. Conductive hollow fiber membranes made of nickel functioned as both filter material for treated water reclamation and as a cathode to catalyze hydrogen production reaction. The produced hydrogen was subsequently converted into methane by hydrogenotrophic methanogens. Organic removal was 98.9% irrespective of operation mode. Maximum volumetric hydrogen production rate was 0.2 m3/m3d, while maximum current density achieved was 6.1 A/m2 (based on cathode surface area). Biofouling, an unavoidable phenomenon in traditional MBRs, can be minimized in this system through self-cleaning approach of hybrid membranes by hydrogen production. The increased rate of hydrogen evolution at high applied voltage (0.9 V) reduces the membrane fouling. Improvements can be done in the system to make it as a promising net energy positive technology for the low strength wastewater treatment.

Microbial electrolysis cell (MEC) system is a potential technology that could treat wastewater while simultaneously generating H2 (green energy). MEC's electroactive bacteria (EAB) are essential microbes responsible for oxidizing organic pollutants (such as acetate) in wastewater using an electrogenesis process. Since EABs comprise the core of MECs, they are essential for maintaining functional stability (Coulombic efficiency (CE), current density, and pollutant removal) of MECs. The cause of EAB becoming dominant at the anode of MECs fed with wastewater is still unclear. Furthermore, efficient EAB are typically not detected in wastewater, and when they are present their abundance is low, which affects their early colonization on the anode and subsequent growth into a mature biofilm. This study investigated bioaugmentation as a strategy to drive the assembly of functionally redundant anode EAB biofilms to improve MEC performance. Two bioaugmentation strategies (Conditions 2 and 3) with known EABs (G. sulfurreducens and D. acetexigens) were tested during the startup of MECs. Meanwhile, control MEC reactors (Condition 1) were operated with only wastewater as the sole source of inoculum to compare the anodic biofilm assembly and system performance with the bioaugmented reactors. Equal number of G. sulfurreducens and D. acetexigens cells were added to the wastewater-fed MEC (10% inoculum at 2.1E+07 live cells/mL). In Condition 3, anodic-biofilm colonized G. sulfurreducens and D. acetexigens was used as anode in wastewater fed MECs. Using single-chambered MEC reactors, the bioaugmented MECs (Condition 2 and 3) performed more efficiently than the non-bioaugmented (Condition 1) MECs. Current generation, CE and gas production were different between the three conditions tested (Condition 3 > Condition 2 > Condition 1). Analysis of 16S rRNA gene sequencing of anodic biofilm indicates revealed that the bacterial communities was not affected between the tested conditions. However, the relative abundance of EABs, mainly G. sulfurreducens and D. acetexigens, was markedly influenced by bioaugmentation compared to the control reactor. The highest peak current generation (~ 1500 mA/m2), CE (70.3 ± 9%), and gas production (0.04 m3/m3/day) was observed in Condition 3. Collectively, these results provide a framework for engineering the anode microbial communities in MECs for wastewater treatment.

Microbial electrochemical systems (MESs) have attracted much research attention in recent years due to their promising applications in renewable energy generation, bioremediation, and wastewater treatment. In a MES, microorganisms interact with electrodes via electrons, catalyzing oxidation and reduction reactions at the anode and the cathode. The bacterial community of a high power mixed consortium MESs (maximum power density is 6.5W/m��) was analyzed by using denature gradient gel electrophoresis (DGGE) and 16S DNA clone library methods. The bacterial DGGE profiles were relatively complex (more than 10 bands) but only three brightly dominant bands in DGGE results. These results indicated there are three dominant bacterial species in mixed consortium MFCs. The 16S DNA clone library method results revealed that the predominant bacterial species in mixed culture is Geobacter sp (66%), Arcobacter sp and Citrobacter sp. These three bacterial species reached to 88% of total bacterial species. This result is consistent with the DGGE result which showed that three bright bands represented three dominant bacterial species. Exoelectrogenic bacterial strain SX-1 was isolated from a mediator-less microbial fuel cell by conventional plating techniques with ferric citrate as electron acceptor under anaerobic conditions. Phylogenetic analysis of the 16S rDNA sequence revealed that it was related to the members of Citrobacter genus with Citrobacter sp. sdy-48 being the most closely related species. The bacterial strain SX-1 produced electricity from citrate, acetate, glucose, sucrose, glycerol, and lactose in MFCs with the highest current density of 205 mA/m�� generated from citrate. Cyclic voltammetry analysis indicated that membrane associated proteins may play an important role in facilitating electron transfer from the bacteria to the electrode. This is the first study that demonstrates that Citrobacter species can transfer electrons to extracellular electron acceptors. Citrobacter strain SX-1 is capable of generating electricity from a wide range of substrates in MFCs. This finding increases the known diversity of power generating exoelectrogens and provids a new strain to explore the mechanisms of extracellular electron transfer from bacteria to electrode. The wide range of substrate utilization by SX-1 increases the application potential of MFCs in renewable energy generation and waste treatment. Anode properties are critical for the performance of microbial electrolysis cells (MECs). Inexpensive Fe nanoparticle modified graphite disks were used as anodes to preliminarily investigate the effects of nanoparticles on the performance of Shewanella oneidensis MR-1 in MECs. Results demonstrated that average current densities produced with Fe nanoparticle decorated anodes were up to 5.9-fold higher than plain graphite anodes. Whole genome microarray analysis of the gene expression showed that genes encoding biofilm formation were significantly up-regulated as a response to nanoparticle decorated anodes. Increased expression of genes related to nanowires, flavins and c-type cytochromes indicate that enhanced mechanisms of electron transfer to the anode may also have contributed to the observed increases in current density. The majority of the remaining differentially expressed genes were associated with electron transport and anaerobic metabolism demonstrating a systemic response to increased power loads. The carbon nanotube (CNT) is another form of nano materials. Carbon nanotube (CNT) modified graphite disks were used as anodes to investigate the effects of nanostructures on the performance S. oneidensis MR-1 in microbial electrolysis cells (MECs). The current densities produced with CNT decorated anodes were up to 5.6-fold higher than plain graphite anodes. Global transcriptome analysis showed that cytochrome c genes associated with extracellular electron transfer are up-expressed by CNT decorated anodes, which is the leading factor to contribute current increase in CNT decorated anode MECs. The up regulated genes encoded to flavin also contribute to current enhancement in CNT decorated anode MECs. / Graduation date: 2013

Certains procédés d'électrolyse industriels, comme ceux de production de fluor et d'aluminium, génèrent du gaz sur des électrodes peu mouillées par l'électrolyte. Dans ces procédés, la convection est en grande partie induite par le mouvement des bulles. Le temps de résidence du gaz n'étant pas maitrisé, la modélisation de ces électrolyses se heurte à de nombreux défis. Ce travail propose un état des connaissances nécessaires à la modélisation de ce type de cellules. Après avoir mis au point une méthode permettant de réaliser des électrodes fortement hydrophobes, nous avons étudié le dégagement gazeux que celles-ci produisent. Nous avons observé que les bulles qui s'y forment croissent par coalescence et ce phénomène affecte la vitesse de croissance et la taille au détachement. Par la suite, nous avons démontré qu'en localisant le dépôt hydrophobe il est possible d'obtenir une population de bulles dont le nombre, la taille et la position sont maitrisés. Nous avons constaté que l'émission acoustique est un outil efficace pour mesurer, sans observation visuelle, la taille et la vitesse d'ascension des bulles produites par électrolyse. La fréquence la plus énergétique du spectre des salves d'émission acoustique des cellules d'électrolyse produisant du gaz permet, quand la fraction volumique de gaz est faible, de mesurer la taille de toutes les bulles dans la cellule. Quand les bulles sont nombreuses et denses, cette même fréquence est liée aux dimensions de la cellule, à la densité de courant, à la taille des bulles et à la fraction volumique en gaz dans le panache. Nous avons proposé une explication à cette observation basée sur l'hypothèse d'un couplage acoustique des bulles. En l'absence d'agitation vigoureuse, l'activité acoustique d'une cellule d'électrolyse produisant des bulles traduit l'importance du transport de matière aux électrodes. La mesure de la répartition de la densité de courant par la méthode des électrodes segmentées dans une cellule en convection libre a été réalisée. Cette étude a été menée dans le but de tester un outil de simulation numérique des cellules d'électrolyse produisant du gaz. L'effet de la mouillabilité de l'électrode sur la répartition du courant a été évalué. / Molten salts electrochemical processes, like fluorine and aluminium production processes produce bubbles on electrodes slightly wetted by an electrolyte. In those cells, the electrolyte convection is mainly induced by the motion of bubbles. Then, the residence time of gas is not under control and the modelling of such electrolysis still faces many challenges. This work begins with a collection of the information required to model gas-producing cells. We developed an electrochemical composite coating technique to achieve the production of highly hydrophobic electrodes. Then we studied the gas evolution obtained with these electrodes, we observed that coalescences determines the bubbles growth rate and sizes at detachment. When the hydrophobic deposit is local, it is possible to obtain a bubble population whose number, size and position are controlled. We found that the acoustic emission is an effective tool to measure the size and rising speed of bubbles produced by electrolysis without having to observe them visually. When the volume fraction of gas is low, the most energetic frequency of the spectrum of acoustic emission bursts allows measuring the size of all bubbles in the cell. When bubbles are numerous, the most energetic frequency of the acoustic emission spectrum is related to the dimensions of the cell, current density, sizes of the bubbles and gas volume fraction in the plume. We have proposed an explanation for this observation based on the assumption of an acoustic coupling of the bubbles. When bubbles are produced, the acoustic activity of an electrolysis cell is correlated with the mass transport at electrodes. We propose a set of measurements of the distribution of current density at the electrodes in a cell with electrolyte free convection. These measurements have been carried out to benchmark a numerical simulation tool. The effect of the wettability of the electrode on the current distribution was evaluated.

abstract: Electro-Selective Fermentation (ESF) combines Selective Fermentation (SF) and a Microbial Electrolysis Cell (MEC) to selectively degrade carbohydrate and protein in lipid-rich microalgae biomass, enhancing lipid wet-extraction. In addition, saturated long-chain fatty acids (LCFAs) are produced via β-oxidation. This dissertation builds understanding of the biochemical phenomena and microbial interactions occurring among fermenters, lipid biohydrogenaters, and anode respiring bacteria (ARB) in ESF. The work begins by proving that ESF is effective in enhancing lipid wet-extraction from Scenedesmus acutus biomass, while also achieving “biohydrogenation” to produce saturated LCFAs. Increasing anode respiration effectively scavenges short chain fatty acids (SCFAs) generated by fermentation, reducing electron loss. However, the effectiveness of ESF depends on biochemical characteristics of the feeding biomass (FB). Four different FB batches yield different lipid-extraction performances, based on the composition of FB’s cellular structure. Finally, starting an ESF reactor with a long solid retention time (SRT), but then switching it to a short SRT provides high lipid extractability and volumetric production with low lipid los. Lipid fermenters can be flushed out with short a SRT, but starting with a short SRT fails achieve good results because fermenters needed to degrading algal protective layers also are flushed out and fail to recover when a long SRT is imposed. These results point to a potentially useful technology to harvest lipid from microalgae, as well as insight about how this technology can be best managed. / Dissertation/Thesis / Doctoral Dissertation Civil, Environmental and Sustainable Engineering 2019

Anaerobic ammonium oxidation (anammox) by anammox bacteria contributes significantly to the global nitrogen cycle and plays a major role in sustainable wastewater treatment. To date, autotrophic nitrogen removal by anammox bacteria is the most efficient and environmentally friendly process for the treatment of ammonium in wastewaters; its application can save up to 60% of the energy input, nearly 100% elimination of carbon demand and 80% decrease in excess sludge compared to conventional nitrification/denitrification process. In the anammox process, ammonium (NH4+) is directly oxidized to dinitrogen gas (N2) using intracellular electron acceptors such as nitrite (NO2–) or nitric oxide (NO). In the absence of NO2– or NO, anammox bacteria can couple formate oxidation to the reduction of metal oxides such as Fe(III) or Mn(IV). Their genomes contain homologs of Geobacter and Shewanella cytochromes involved in extracellular electron transfer (EET). However, it is still unknown whether anammox bacteria have EET capability and can couple the oxidation of NH4+ with transfer of electrons to extracellular electron acceptors. In this dissertation, I discovered by using complementary approaches that in the absence of NO2–, freshwater and marine anammox bacteria couple the oxidation of NH4+ with transfer of electrons to carbon-based insoluble extracellular electron acceptors such as graphene oxide (GO) or electrodes poised at a certain potential in microbial electrolysis cells (MECs). Metagenomics, fluorescence in-situ hybridization and electrochemical analyses coupled with MEC performance confirmed that anammox electrode biofilms were responsible for current generation through EET-dependent oxidation of NH4+. 15N-labelling experiments revealed the molecular mechanism of the EET-dependent anammox process. NH4+ was oxidized to N2 via hydroxylamine (NH2OH) as intermediate when electrode was used as the terminal electron acceptor. Comparative transcriptomics analysis supported isotope labelling experiments and revealed an alternative pathway for NH4+ oxidation coupled to EET when electrode was used as electron acceptor. The results presented in my dissertation provide the first experimental evidence that marine and freshwater anammox bacteria can couple NH4+ oxidation with EET, which is a significant breakthrough that is promising in the context of implementing EET-dependent anammox process for energy-efficient treatment of nitrogen using bioelectrochemical systems.

京都大学 / 0048 / 新制・課程博士 / 博士(工学) / 甲第19730号 / 工博第4185号 / 新制||工||1645(附属図書館) / 32766 / 京都大学大学院工学研究科物質エネルギー化学専攻 / (主査)教授 江口 浩一, 教授 安部 武志, 教授 陰山 洋 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DGAM

Wastewater treatment has developed as a mature technology over time. However, conventional wastewater treatment is a very energy-intensive process. Bioelectrochemical system (BES) is an emerging technology that can treat wastewater and also recover resources such as energy in the form of electricity/hydrogen gas and nutrients such as nitrogen and phosphorus compounds. Microbial electrolysis cell (MEC) is a type of BES that, in the presence of an additional voltage, can treat wastewater and generate hydrogen gas. This is a promising approach for wastewater treatment and value-added product generation, though it may not be sustainable in the long run, as it relies on fossil fuels to provide that additional energy. Thus, it is important to explore alternative renewable resources that can provide energy to power MEC. Waste heat is one such resource that has not been researched extensively, particularly at the low-temperature spectrum. This was utilized as a renewable resource by converting waste heat to electricity using a device called thermoelectric generator (TEG). TEG converted simulated waste heat from an anaerobic digester to power an MEC. The feasibility of TEG to act as a power source for an MEC was investigated and its performance compared to the external power source. Various cold sources were analyzed to characterize TEG performance. To explore this integrated TEG-MEC system further, a hydraulic connection was added between the two systems. Wastewater was used as a cold source for TEG and it was recirculated to the anode of the MEC. This system showed improved performance with both systems mutually benefitting each other. The operational parameters were analyzed for the optimization of the system. The integrated system could generate hydrogen at a rate of 0.36 ± 0.05 m3 m-3 d-1 for synthetic domestic wastewater treatment. For the practical application, it is necessary to estimate the cost and narrow the focus on the functions of the system. Techno-economic analysis was performed for MEC with cost estimation and net present value model to understand the economic viability of the technology. The application niche of the BES was described and directions for addressing the challenges towards a full-scale operation were discussed. The present system provides a sustainable method for wastewater treatment and resource recovery which can play an important role in human health, social and economic development and a strong ecosystem. / Doctor of Philosophy / An average person produces about 50-75 gallons of wastewater every day. In addition to the households, wastewater is generated from industries and agricultural practices. As the population increases, the quantity of wastewater production will inevitably increase. To keep our rivers and oceans clean and safe, it is essential to treat the wastewater before it is discharged to the water bodies. However, the conventional wastewater treatment is a very energy (and thus cost) intensive process. For low-income and developing parts of the world, it is difficult to adapt the technology everywhere in its present form. Furthermore, as the energy is provided mostly by fossil fuels, their limited reserves and harmful environmental effects make it critical to find alternative methods that can treat the wastewater at a much lower energy input. For a circular and sustainable economy, it is important to realize wastewater as a resource which can provide us energy, nutrients, and water, rather than discard it as a waste. Bioelectrochemical systems (BES) is an emerging technology that can simultaneously treat wastewater and recover resources in the form of electricity/hydrogen gas, and nitrogen and phosphorus compounds. Microbial electrolysis cell (MEC) is a type of BES that is used to treat wastewater and generate hydrogen gas. An additional voltage is supplied to the MEC for producing hydrogen. In the long run, this may not be sustainable as it relies on fossil fuels to provide that additional energy. Thus, it is important to explore alternative renewable resources that can provide energy to power MEC. Waste heat is a byproduct of many industrial processes and widely available. This was utilized as a renewable resource by converting waste heat to electricity using a device called thermoelectric generator (TEG). TEG converted simulated waste heat from an anaerobic digester to power an MEC. The mutual benefit for MEC and TEG was also explored by connecting the system electrically and hydraulically. Cost-estimation of the system was performed to understand the economic viability and functions of the system were developed. The present system provides a sustainable method for wastewater treatment and resource recovery which can play an important role in human health, social and economic development and a strong ecosystem.

La cellule d’électrolyse microbienne (CEM) permet la conversion de la biomasse en dihydrogène via un apport théorique en énergie électrique 10 fois moindre que celui de l’électrolyse de l’eau. Un tel procédé fonctionne via la technologie des bioanodes, qui permet la catalyse de l’oxydation de la biomasse en CO2 par l’intermédiaire d’un biofilm électro-actif. Les travaux exposés dans ce manuscrit de thèse ont pour but l’optimisation des performances de la bioanode, la compréhension des mécanismes de catalyse et la réalisation d’un prototype à échelle laboratoire. L’optimisation par l’étude de paramètres opératoires en montage 3 électrodes a montré que l’utilisation de sédiments de salins comme source de micro-organismes avec électrode en feutre de carbone polarisée à + 0,1 V / ECS à une température de 40°C permet la formation de bioanodes capables de débiter jusqu’à 85 A.m-2 pour une conductivité de 10,4 S.m-1. A 30°C, le pyroséquençage ADN a mis en lumière l’émergence des genres bactériens Desulfuromonas et Marinobacter. La conception et l’exploitation d’un modèle de voltammétrie cyclique a montré que le transport des électrons au sein du biofilm était environ 100 fois plus lent que le métabolisme bactérien. L’utilisation de la spectroscopie d’impédance électrochimique montre que la résistance au transfert de charge à l’interface électrode/solution baisse de 24 kΩ.cm2 à 64 Ω.cm2 lors de la formation du biofilm. Un taux de production maximum de 2,85 LH2.L-1.j-1 ainsi qu’une durée de vie de plus de 50 jours du procédé ont été obtenus lors de la conduite d’un prototype laboratoire de CEM. / Microbial electrolysis cell (MEC) is a recent and promising bioelectrochemical process that converts biomass onto hydrogen thanks to an amount of electrical energy 10 times smaller than for water electrolysis. The operation of the process is making possible by the bioanode technology which catalyses the biomass combustion onto CO2 through an electro-active biofilm. The purpose of the present work consists on the optimisation of the bioanode, the understanding of the catalysis mechanism and a scaling-up by the designing of a MEC prototype. Using a three-electrode device and sediment of salt marshes as inoculum, the study of the experimental parameters demonstrated that carbon felt poised at + 0.1 V /ECS at 40°C led to the formation of bioanode able to generate up to 85 A.m-2 at a solution conductivity of 10,4 S.m-1. For a temperature of 30°C, DNA pyrosequencing denoted the presence of the two bacterial genera Desulfuromonas and Marinobacter. The development and the exploitation of a cyclic voltammetric model showed that electron transfer within the biofilm ran almost 100 times slower than bacterial metabolism. Electrochemical impedance spectroscopy during biofilm formation revealed a decreasing of the charge transfer resistance at the electrode/solution interface from 24 kΩ.cm2 to 64 Ω.cm2. Designing and first experiments with a 6L CEM prototype led to a hydrogen production rate of 2.85 LH2.L-1.j-1 and a process life time of up to 50 days. Those performances were achieved in a reproducible way.

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