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Organic waste-fueled carbonaceous resource recovery using approaches such as arrested anaerobic digestion generates economically attractive products such as volatile fatty acid (VFA). The production of VFA expands the applications of anaerobic biotechnologies beyond the traditionally produced biogas. Compared to biogas, VFA is produced and recovered in a concentrated form in the aqueous phase, which is more conducive to direct utilization in downstream bioplastic, biodiesel production, and nitrogen/phosphorus removal in water resource-recovery facilities. However, this application is limited by the variability in VFA yield and composition as obtained from different complex solids streams. Additionally, the lack of understanding of the nexus between the performance-structure-function of the microbial community within the arrested anaerobic digestion process leads to the massive gap between the optimized engineering regulations and the high-throughput VFA production. Consequently, this dissertation aimed to unlock the potential of VFA production with maximized yield and regulated composition through the manipulation of the operational parameter (hydraulic retention time (HRT)) and the feedstock condition (thermal hydrolysis pretreatment (THP)). In response, meta-omics-derived approaches were applied to elucidate the dynamic changes of microbial structure, potential, and extant functionality in terms of the two processes (hydrolysis and acidification) within arrested anaerobic digestion of food waste. Specifically, the objectives were (1) Performance: Evaluate the hydrolysis and acidification performance changes including hydrolysis yield, VFA yield, VFA composition, methane yield, etc. under different HRTs and feeding THP or non-THP food waste. (2) Microbial structure: characterize and compare the significance of HRT and feedstock condition in shaping microbial structures. (3) Functional analysis: Interpret the community-level dynamic changes of potential and extant functions within the (3.1) customized acidification metabolic networks and the (3.2) carbohydrate hydrolysis niches. The highlighted findings are as follows: (1) Performance of the arrested anaerobic digestion (including hydrolysis and acidification processes): Neither the hydrolysis yield nor the VFA yield was improved by the extended HRT from 4 to 8 days (P > .05). The inclusion of THP on feedstock didn’t improve the hydrolysis yield (P > .05) while the VFA yield was significantly decreased (P = .003). Among all conditions, the methane production was less than 5% of the chemical oxygen demand (COD) and a propionic acid-dominant type product was robustly formed. (2) Microbial structures in the arrested anaerobic digestors (including core hydrolyzers and acidification microbial communities): Both HRT and the inclusion of THP on feedstock shaped distinct microbial structures in the arrested anaerobic digestors (P = .02 and .01). Although the extension of HRT didn’t change the Shannon diversity Index (P > .05), it was significantly decreased after feeding with THP food waste (P = .03), which might stem from the reduced indigenous microbes in the initial food waste feedstock. Prevotella was always the most abundant genus under all conditions, which might contribute to the dominantly produced propionic acid among all conditions. The successfully suppressed growth of methanogenic archaea was reflected in terms of the low relative abundance (<1.5%) among all conditions. (3.1) Functional analysis of the customized acidification metabolic networks: Under the two selected HRTs, the potential and extant functions of acidification were unchanged between the two reactors (P > .05), which indicated a community-level redundancy in convergent potential and extant acidification functions even under a completely shifted microbial structure. However, the inclusion of THP diminished the potential and extant functions of acidification, in the meantime, shifting the main producer of butyric acid from Bacteroides to Prevotella through the expression of gene buk2. Among all conditions, the highest potential and extant functions in propionic acid production corresponded to the propionic acid-dominant acid profile in all reactors. The prevalently enriched Prevotella contributed to the stable propionic acid-dominant production via the acryloyl-CoA to propionyl phosphate to the propionic acid pathway. (3.2) Functional analysis of the carbohydrate hydrolysis niches: The extension of HRT from 4 days to 8 days didn’t impact the potential and extant functions of carbohydrate-activated enzymes (CAZys) and the hydrolysis of polysaccharides. Only two intermediate steps (gene malQ and lplD) during the hydrolysis of starch and pectin were enhanced with higher absolute transcriptional activities (mRNA/DNA RPKM) under HRT 8 days. The abundance ratio of the two main hydrolysis phyla Firmicutes: Bacteroidetes was unchanged between the two HRTs. When feeding with THP feedstock, the potential and extant functions of CAZys were both enhanced. All steps within the hydrolysis of cellulose (polysaccharides) exhibited increased absolute transcriptional activities (mRNA/DNA RPKM). The abundance ratio of Firmicutes: Bacteroidetes was decreased after the inclusion of THP on feedstock, which corresponded to the increased hydrolysis of polysaccharides- cellulose. Although the carbohydrate hydrolysis functions were improved after feeding with THP food waste, the total hydrolysis yield was not enhanced. The hydrolysis of other compounds such as proteins and lipids could also contribute to the total hydrolysis yield. The taxonomic analysis revealed that in all four conditions, the genus Prevotella presented with the highest potential functions in CAZys, while the genus Pararhodospirillum exhibited the highest extant functions in CAZys. This indicated that distinct bacteria were endowed with different functional potentials of CAZys and mobilized these functions differently. Overall, this research provides practical suggestions for engineering designs to maximize the VFA production profits from arrested anaerobic digestion of food waste: (1) A properly controlled HRT enables a long-term high-throughput production of VFA with stable yield and the unchanged dominant acid type (2) The inclusion of THP to the feedstock was not suggested to be applied to maximize the VFA yield even the dominant acid type may not change. (3) The dominantly produced propionic acid could be targeted by enriching the Prevotella genus to produce the propionic acid through the acryloyl-CoA to propionyl phosphate to the propionic acid pathway. Besides the engineering aspect, this research also specifically elucidates the long-time lumped and simplified acidification and carbohydrate hydrolysis processes with the extended metabolic databases including each reaction, key intermediates, enzymes, and corresponding genes. This expanded database served as an essential upstream process, which could be integrated into the current anaerobic digestion model. Additional applications could be extended to the human digestion systems' microbiome and be exploited commercially for other mixed-culture biosynthesis processes such as bioplastic and biodiesel production. Finally, the application of meta-omics-derived methodology revealed the functional redundancy and the potential discrepancy between the most abundant group and the most actively functional group underlying the formed black box of VFA production performance. This discussion of the nexus of performance-structure-function suggested the importance of applying meta-omics approaches in engineering practice, especially when feeding the mixed-culture community with real complex solid streams. The targeted VFA profiles cannot be reached without identifying the actual functional bacteria under selected engineering conditions.

We analyze a simplified mathematical model of the complete degradation of monochlorophenol. The model takes form of a system of six ordinary differential equations, the dynamics of which can be reduced to the dynamics of a three-dimensional system on the invariant set. We extend the previous analysis by considering multiple substrate inflow. We also focus on the bifurcations occurring in the system and their biological meaning. / Thesis / Master of Science (MSc)

Anaerobic digestion (AD) is an essential process in wastewater treatment to stabilize waste organic solids and produce biogas. This research is comprised of two projects in the discipline of anaerobic digestion. First, the effect of high sulfate concentration on anaerobic digestion of wastewater sludge was investigated. Secondly, the performance of acetoclastic methanogens Methanosaeta spp. and Methanosarcina spp. were investigated under intensified AD operation conditions (i.e., elevated acetate concentrations, vigorous mixing, etc.). In the sulfate experiments, the cumulative biogas and methane production decreased linearly with increasing initial sulfate doses (0 – 3,300 mg S L-1) and the correlation between the sulfate dose and methane production was verified with theoretical predictions, indicating complete reduction of sulfate to sulfide in AD. The examined sulfate concentrations resulted in no clear negative effects on the COD (chemical oxygen demand) removal or VSS (volatile suspended solids) destruction of the wastewater sludge, indicating that previous findings on sulfide toxicity might have been attributed to potential COD overestimation of digested sludge with high levels of sulfide. To avoid potential misinterpretation of AD performance on sulfide toxicity effects, we proposed a new method for COD correction for digested sludge. In the second project focused on acetoclastic methanogens, vigorous mixing conditions substantially decreased Methanosarcina spp. growth and methane production, and the decreased methanogenesis was more pronounced at higher acetate concentrations. Methanosarcina spp. prefer to grow in clusters and the vigorous mixing can disrupt cluster formation; as a result, reduced chances for cluster formation limited the growth of Methanosarcina spp.. While Methanosarcina spp. growth and methane production increased with the increasing acetate concentration, Methanosaeta spp. growth was unaffected by the examined vigorous mixing and soluble substrate conditions with negligible relative growth. Thus, rapid enrichment of Methanosarcina spp. is critical for successful operation intensified of AD processes under high organic loading conditions. / Thesis / Master of Applied Science (MASc)

Anaerobic digester design and operation influences the biomass degradation efficiency performed by complex and diverse microbial communities. Optimum anaerobic digester design and operational parameters in residential on-site wastewater treatment sites (OWTS) establishes physiochemical environments suitable for the growth and stability of the microbial communities responsible for organic waste degradation. A comparative study of the microbial communities and their functional profiles between different OWTS designs and operational parameters have not been done despite their functional importance in residential organic waste removal. Using whole-metagenome shotgun sequencing, microbial community compositions and functions were compared between two digester designs: conventional box septic tanks and septic tanks equipped with a novel closed-conduit tube called the InnerTubeTM. Wastewater was sampled along the length of each digester to explore the microbial community stratification during the anaerobic digestion treatment process. Additionally, the effect of effluent, aerobic recirculating-lines on the digester microbiome was also explored. Physiochemical characteristics in the form of oxygen demand, nitrogen and solids content was used as endpoints and correlated with microbial community and functional gene abundances to explore the microbes driving anaerobic digestion. Conventional digesters were characterized by syntrophic proprionate-oxidizing microbes and acetoclastic methanogens, while InnerTube™ digesters were characterized by syntrophic sulfate-reducing microbes and hydrogenotrophic methanogens. Recirculating digesters were enriched with denitrifying microbial consortia in syntrophy with hydrogenotrophic methanogens. Microbial communities were organized according to hydrolytic, acidogenic, acetogenic, and methanogenic groups along the digester treatment process. Insight into the core microbiome of OWTS can inform bioaugmentation and digester design and operation optimization strategies to improve the treatment of decentralized residential sewage sources. / Thesis / Master of Science (MSc) / Anaerobic digesters are used throughout North America to treat residential sewage. Despite their prevalence, the composition and function of the microbial communities driving sewage degradation in residential digesters has not been studied. We used DNA sequencing to compare the microbial communities and functional genes in different anaerobic digester designs across Southern Ontario. Our findings suggest there are successive microbial groups along the length of septic tanks and that different septic tank designs harbor characteristic sulfidogenic and methanogenic microbes. Characterization of these microbes could inform septic tank bioaugmentation, design and operational optimization strategies to improve sewage treatment performance.

Yes / A novel Euler-Lagrangian (EL) computational uid dynamics (CFD) nite volume-based model to simulate the gas mixing of sludge for anaerobic digestion is developed and described. Fluid motion is driven by momentum transfer from bubbles to liquid. Model validation is undertaken by assessing the ow eld in a labscale model with particle image velocimetry (PIV). Conclusions are drawn about the upscaling and applicability of the model to full-scale problems, and recommendations are given for optimum application.

Yes / For the first time, a set of Lattice-Boltzmann two-way coupling pointwise Euler-Lagrange models is applied to gas mixing of sludge for anaerobic digestion. The set comprises a local model, a “first-neighbour” (viz., back-coupling occurs to the voxel where a particle sits, plus its first neighbours) and a “smoothing-kernel” (forward- and back-coupling occur through a smoothed-kernel averaging procedure). Laboratory-scale tests display grid-independence problems due to bubble diameter being larger than voxel size, thereby breaking the pointwise Euler-Lagrange assumption of negligible particle size. To tackle this problem and thereby have grid-independent results, a novel data-scaling approach to pointwise Euler-Lagrange grid independence evaluation, based on an application of the Buckingham π theorem, is proposed. Evaluation of laboratory-scale flow patterns and comparison to experimental data show only marginal differences in between the models, and between numerical modelling and experimental data. Pilot-scale simulations show that all the models produce grid-independent, coherent data if the Euler-Lagrange assumption of negligible (or at least, small) particle size is recovered. In both cases, a second-order convergence was achieved. A discussion follows on the opportunity of applying the proposed data-scaling approach rather than the smoothing-kernel model.

Understanding the respiratory physiology and energy metabolism is important for establishing the dissolved oxygen (DO) requirements of freshwater mussels, determining the metabolite(s) indicative of environmental stress, and interpreting environmental conditions based on physiological indicators of mussels. Three studies were undertaken to address these questions. The first study was conducted with seven mussel species collected from reservoir and riverine habitats. The two objectives were to determine the diurnal patterns of valve gaping of freshwater mussels from different habitats, and to monitor heart rate changes of a mussel species that exhibited the diurnal gaping. The results showed that night gaping is evident for the mussels collected from lentic areas, but not for those collected from lotic areas. The heart rate of Pyganodon grandis increased when they gaped. The second study was conducted with nine species of freshwater mussels from different habitats. The three objectives were to determine the patterns (i.e., regulator and conformer) of oxygen consumption (OC) rate under declining DO, evaluate the effects of temperature on ability to regulate OC under declining DO, and finally to use this information to predict DO criteria for maintaining freshwater mussels in captivity. The results showed that mussels living in the habitats subjected to low DO have a better ability to regulate the OC and were more tolerant to hypoxia. The third study assessed three mussel species from different habitats with different abilities to regulate OC under low DO. The two objectives in this study were to identify the energetic metabolite changes under different DO levels and air exposure for the three species, and to determine the appropriate tissue(s) and metabolite(s) to use for estimating the stress in mussels. The results showed that different biochemical strategies were used by Villosa iris, Elliptio complanata, and Pyganodon grandis. Villosa iris had the lowest anaerobic capacity. Elliptio complanata had a more efficient anaerobic metabolism, and P. grandis reduced energy metabolism under low DO and air exposure. posterior adductor muscle, gill and mantle were good tissues for evaluating hypoxic stress. The mantle tissue had the highest glycogen store and was the best tissue for non-lethal study of physiological condition. / Ph. D.