Recovering valuable resources from wastewater will transform wastewater management from a treatment focused to sustainability focused strategy, and creates the need for new technology development. An innovative treatment concept - osmotic bioelectrochemical system (OsBES), which is based on cooperation between bioelectrochemical systems (BES) and forward osmosis (FO), has been introduced and studied in the past few years. An OsBES can accomplish simultaneous treatment of wastewater and recovery of resources such as nutrient, energy, and water (NEW). The cooperation can be accomplished in either an internal (osmotic microbial fuel cells, OsMFC) or external (microbial electrolysis cell-forward osmosis system, MEC-FO) configuration. In OsMFC, higher current generation than regular microbial fuel cell (MFC) was observed, resulting from the lower resistance of FO membrane. The electricity generation in OsMFC could greatly inhibit the reverse salt flux. Besides, ammonium removal was successfully demonstrated in OsMFC, making OsMFCs a promising technology for "NEW recovery" (NEW: nutrient, energy and water). For the external configuration of OsBES, an MEC-FO system was developed. The MEC produced an ammonium bicarbonate draw solute via recovering ammonia from synthetic organic solution, which was then applied in the FO for extracting water from the MEC anode effluent. The system has been advanced with treating landfill leachate. A mathematical model developed for ammonia removal/recovery in BES quantitatively confirmed that the NH4+ ions serve as effective proton shuttles across cation exchange membrane (CEM). / Ph. D.

Partial nitritation combined with the anaerobic ammonium oxidation (Anammox) process offers a way of replacing the conventional nitrogen removal process of nitrification-denitrification, lowering the need for oxygen and chemical input, as well as reducing the production of sludge. However, as a by-product of the biochemical reaction driven by anammox bacteria, it produces nitrate-nitrogen (NO3- - N) (16-26% nitrogen removed), which is problematic. Microbial desalination cells (MDCs) are a promising technology capable of converting biodegradable organics into electricity (by electroactive bacteria), providing for simultaneous desalination, and wastewater treatment. Despite being a promising technology, MDCs have limitations. The first-proof of-concept of MDC was demonstrated using acetate as the organic source, expensive platinum as a catalyst, and ferricyanide as an electron acceptor in the cathode that makes MDC costly, environmentally unfriendly, and unsustainable. This research investigated the integration of the anammox and nitration processes in MDCs as a long-term biocatalyst/biocathode for sustainable and energy-efficient nitrogen removal and electricity generation. A series of experiments were designed and performed to evaluate the performance of the anammox process as a biocatalyst in MDCs. The results concluded that the anammox process can be used as a biocatalyst to accept electrons in MDCs producing 444 mW/m3 of power density and 84% of ammonium nitrogen removal. Furthermore, the concept of using a one-stage nitritation anammox process as a biocathode in MDC was evaluated and produced a maximum power output of 1007 mW/m3. Two configurations of anammox MDCs (anaerobic-anammox cathode MDC (AnAmmoxMDC) and nitritation-anammox cathode MDC (NiAmoxMDC) were compared with an air cathode MDC (CMDC), operated in fed-batch mode. The NiAmoxMDC showed better performance in terms of power production and nitrogen removal. The co-existence of aerobic ammonium oxidizing bacteria (AOB) and anammox bacteria in the same biocathode of single-stage NiAmoxMDC concluded the resource-efficient wastewater treatment. Furthermore, two-stage nitritation anammox as a biocathode in MDC was evaluated and proved to be energy-efficient bioelectrochemical wastewater treatment by producing 1500 mW/m3 (300 mW/m2) of maximum power output. This research provides the first proof of concept that nitritation-anammox biocathode can provide a sustainable and energy-efficient nitrogen removal along with desalination and bioelectricity generation.

There is a growing awareness of the valuable nutrients (nitrogen and phosphorus) being lost in conventional wastewater treatment systems. Although the removal of these nutrients has been well addressed, efforts for nutrient recovery have seen little development. As the emphasis on sustainability in the wastewater treatment industry increases, conventional wastewater treatment processes are being re-evaluated and new treatment systems developed. A possible nutrient recovery mechanism is the precipitation of magnesium ammonium phosphate hexahydrate (MgNH4PO4·6H2O), commonly known as struvite. Human urine has been identified as a rich source of nutrients in wastewater; hence the separate collection of urine is considered a viable method of enabling struvite recovery. Since dilution of urine to a certain degree is inevitable, reconcentration of urine beyond the solubility limit of struvite is critical. Currently available methods for reconcentration (e.g., evaporation, freeze-thaw, reverse osmosis and electrodialysis) are relatively expensive with high energy demand. Thus, the research here aims to demonstrate nutrient reconcentration from diluted urine and simultaneous organic removal by using the principles of microbial desalination cells (MDCs), where energy released from organic oxidation is partially used for the separation of nutrient ions. With reduced energy demand, a sustainable method for the utilization of source-separated urine is examined. The performance of bioelectrochemical systems relies on the activity of exoelectrogenic bacteria to transfer electrons to the anode. An examination of exoelectrogen sensitivity at various wastewater treatment conditions (i.e. ammonia and oxygen) is an important component of this research. Methanogenesis is considered the greatest challenge in achieving practical applications in anaerobic bioelectrochemical systems. An electrolytic oxygen production method is suggested for effective control of methanogenesis in a feasible and cost-effective manner. / Thesis / Master of Applied Science (MASc)

Bioelectrochemical systems couple the oxidation of an electron donor at the anode with the reduction of an electron acceptor at the cathode, using microorganisms to catalyse one or both reactions. When the overall reaction is exergonic, a power output is generated and the system is referred to as microbial fuel cell (MFC); when power is added to the system and hydrogen is produced at the cathode through electrolysis of water, the system is referred to as microbial electrolysis cell (MEC). This PhD thesis is principally focused on the microbial fuel cells technology. Microbial fuel cells are regarded as a sustainable technology for electric energy generation from the oxidation of organic substrates contained in wastewater. The rising need for renewable energy sources and sanitation has encouraged intense research in this novel technology. Nevertheless, up untill now the interest has been primarily focused on the anodic oxidation of organic matter contained in wastewater. However, in addition to organics, wastewater also contains other pollutants, such as soluble nitrogen compounds, for which specific treatment is required. In conventional wastewater treatment systems, the organics available in the wastewater are typically used as electron donor during denitrification. However, a considerable fraction (>50%) of the chemical oxygen demand (COD) is still oxidized aerobically due to the large recirculation flows from the nitrification to the denitrification stages required in anoxic/aerobic configurations to allow for low nitrate levels in the final effluent. This increased COD demand is normally fulfilled by supplementary COD addition, with consequent increase of treatment costs. Alternatively, microorganisms can use inorganic carbon substrates and inorganic electron donors such as hydrogen for denitrification. However, the use of compressed hydrogen is hampered by its low solubility. As a solution, electrochemical hydrogen production permits in situ delivery of the electron donor and is advantaged by simplified control and dissolution of H2. The energy requirements to provide reducing power for denitrification can be decreased if bacteria use the electrode directly as electron donor without intermediate hydrogen production in bioelectrochemical systems. However, fundamental knowledge on bioelectrochemical denitrification is still lacking, therefore, this PhD thesis aims to fill some of these knowledge gaps and to solve some of the bottlenecks of the use of biocathodes. In particular, the goals of this work are: (i) to produce a suitable microbial community able to use the cathode as the sole electron donor during denitrification; (ii) to engineer a bioelectrochemical system able to couple the cathodic denitrification with the oxidation of organics at the anode; (iii) to characterize and quantify the electron losses during anodic and cathodic processes; (iv) to develop a bioelectrochemical system that maximises the nitrogen removal by integrating the nitrification stage into the cathode; finally, (v) to provide an insight into the structural properties of the biofilm performing nitrogen removal at the cathode. The results reveal that microbes can effectively utilize the electrode as electron donor for nitrate reduction to gaseous nitrogen at a redox potential that excludes intermediate production of hydrogen. Measurements revealed that acetoclastic methanogenesis and bacterial growth were responsible for causing the major electron losses at the anode. Adjusting the anodic potential did not achieve a significant overall reduction of the electron losses. At the cathode, the charge transfer efficiencies were instead very high, with the losses only due to the generation of nitrous oxide. Moreover, adjustments of the cathode potential resulted in higher efficiency. High carbon and nitrogen removal was obtained with a COD demand for denitrification as low as 2.4 g per g nitrogen denitrified, which is much lower than typically observed in heterotrophic–based nitrogen removal technologies (>7 g g 1). Nitrogen was removed at rates up to 0.256 kg N m-3 d-1, which is comparable to other autotrophic denitrification processes. Simultaneous nitrification and denitrification was observed in a combined system with cathodic aeration, at bulk dissolved oxygen (DO) levels up to 5 mg L-1, which is considerably higher than normally considered feasible for the process. Confocal laser scanning microscope analysis revealed the existence of a structured biofilm where putative nitrifying organisms occupied the outer layers in contact with the aerated bulk liquid, and putative denitrifying organisms occupy the layers closer to the electrode. These findings are significant in the field of bioelectrochemical systems as they help to unravel some of the complex questions relating to biocathodes. Additionally, the system provides an attractive option to achieve a very high level of nitrogen removal from wastewater with low COD/N ratios due to the selective utilisation of the COD for the denitrification reaction via the electrical transfer of reducing equivalents from the anode to the cathode. However, this research creates new questions, particularly regarding the mechanisms of electron transfer at the cathode. Also a number of practical design and optimisation challenges need to be overcome before wider applications can be considered.

This thesis presents studies that examine microbial extracellular electron transfer that an emphasis characterizing how environmental conditions influence electron flux between microbes and a solid-phase electron donor or acceptor. I used bioelectrochemical systems (BESs), fluorescence and electron microscopy, chemical measurements, 16S rRNA analysis, and qRT-PCR to study these relationships among chemical, physical and biological parameters and processes. / Engineering and Applied Sciences

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.

Heavy metal contamination in water is a serious environmental and human health issue. Lead (Pb2+) and cadmium (Cd2+) are strictly regulated in wastewater effluent due to their high toxicity at low concentrations. Heavy metals are difficult to remove in conventional biological wastewater treatment because they are water soluble and non-biodegradable. Advanced treatment, such as tight membrane filtration and ion exchange, can be applied but they often require a high electrical energy input and a large amount of chemicals for pre- or post-treatment. Microbial electrolysis cells (MECs) can be used to treat wastewater while simultaneously recovering energy in the form of hydrogen gas. Additionally, MECs were proven to be effective for heavy metal removal. The commonly investigated removal mechanism for heavy metals in MECs is reduction at the cathode where heavy metal ions are reduced to metallic solids. The research presented in this thesis examined the effectiveness of cathodic reduction and other heavy metal removal mechanisms in MECs over a wide range of metal concentrations (10 μg/L-12 mg/L). Lab-scale MEC operation demonstrated successful removal of both Pb2+ and Cd2+ under different electric conditions, operation times, and initial metal concentrations. In addition to cathodic reduction, heavy metal removal in MECs was demonstrated through chemical precipitation at the cathode and electrochemical reduction and biosorption at the bioanode. The results of this research also confirmed the importance of microbial activity at the bioanode to efficiently drive the removal mechanisms in MECs. / Thesis / Master of Applied Science (MASc)

The treatment of water and wastewater is energy intensive, and there is an urgent need to develop new approaches to address the water-energy challenges. Bioelectrochemical systems (BES) are energy-efficient technologies that can treat wastewater and simultaneously achieve multiple functions such as energy generation, hydrogen production and/or desalination. The objectives of this dissertation are to understand the fundamental microbiology of BES, develop cost-effective cathode catalysts, optimize the process engineering and identify the application niches. It has been shown in Chapter 2 that electrochemically active bacteria can take advantage of shuttle-mediated EET and create optimal anode salinities for their dominance. A novel statistical model has been developed based on the taxonomic data to understand and predict functional dynamics and current production. In Chapter 3, 4 and 5, three cathode catalyst (i.e., N- and S- co-doped porous carbon nanosheets, N-doped bamboo-like CNTs and MoS2 coated on CNTs) have been synthesized and showed effective catalysis of oxygen reduction reaction or hydrogen evolution reaction in BES. Chapter 6, 7 and 8 have demonstrated how BES can be combined with forward osmosis to enhance desalination or achieve self-powered hydrogen production. Mathematical models have been developed to predict the performance of the integrated systems. In Chapter 9, BES have been used as a research platform to understand the fate and removal of antibiotic resistant genes under anaerobic conditions. The studies in this dissertation have collectively demonstrated that BES may hold great promise for energy-efficient water and wastewater treatment. / Ph. D.

A sustainable and low-cost system, namely a photo-bioelectrochemical system (photo-BES), based on the natural blueprint of photosynthetic microorganisms was studied. The aim of this research work is to improve the efficiency of electron transfer of the microorganisms for bioelectricity generation. The first strategy adopted was the evaluation of the exoelectrogenic activity of oxygenic photosynthetic cyanobaterium, Synechococcus elongatus PCC 7942, in biophotovoltaic (BPV) platforms through a comparative performance analysis of bioanode materials. The second approach involved improving the performance of anoxygenic photosynthetic bacterium, Rhodopseudomonas palustris ATCC® 17001™, by varying the ratio of nitrogen to carbon sources (N:C) to maximise both biohydrogen production and exoelectrogenesis for conversion into bioelectricity in photosynthetic microbial fuel cells (photoMFCs). A linear correlation was obtained between average surface roughness/surface area and maximum power density of ITO-coated and graphene/ITO-coated substrates. Graphene/ITO-coated PET bioanodes produced the highest maximum power output of 29±4 μW m-2 in a single chamber BPV device due to improved biofilm formation and improved electrochemical activity. XG Leaf®, also known as graphene paper, helped to bridge the shortcomings of carbon fibres in terms of wettability. The most hydrophilic, 240 μm thick graphene paper, produced the highest maximum power output of 393±20 μW m-2 in a membrane electrode assembly (MEA)-type BPV device, mainly due to reduced electrochemical polarisation. A proof of concept study compared the performance of screen-printed graphene onto a membrane separator against 3D-printed bioanodes coated with carbon nanotubes. One mm thick 3D-printed bioanode was better performing as its structures promoted a much denser biofilm with extensive fibrous extracellular matrix. Using a ratio of N:C=0.20 resulted in higher biohydrogen production and higher exoelectrogenic activity, generating a maximum power output of 361±157 mW m-2 and 2.39±0.13 mW m-2, respectively. This study provided additional insight in improving the electron transfer efficiency, which could be used to further optimise photo-BESs as part of future research and development for sustainable technologies.

A long-held practice is to mix agricultural soil with a soil amendment to improve growing conditions in crops. A common soil amendment is biosolids produced from both municipal and dairy wastewater due to the macro- and micronutrients within it. Both the agricultural and wastewater industries are participating in the Circular Economy concept (CEC). Two experiments explored using expended bioelectrochemical systems (BES) that treated either synthetic dairy wastewater (DWW) or synthetic municipal wastewater (SWW) as soil amendments to improve corn plant growth when treated with three different nutrient treatments: 100%- 50%- and 0% Hoagland Nutrient Solutions. Biochar and used terracotta clay were used as soil amendments too. Additionally, the DWW and SWW soil amendments are being invested to see if soil moisture can be retained during simulated drought conditions. The experiments took place in the late fall and winter of 2021 and summer of 2022 in Starkville, Mississippi.