Intensification refers to utilizing wastewater treatment processes that decrease chemical and energy demands, increase energy recovery, and reduce the process footprint (or increased capacity in an existing footprint) all while providing the same level of nutrient removal as traditional methods. Shortcut nitrogen removal processes; including nitrite shunt, partial nitritation/anammox, and partial denitrification/anammox, as well as low-carbon biological phosphorus removal, were critically-evaluated in this study with an overall objective of intensification of existing infrastructure. At the beginning of this study, granular sidestream deammonification was becoming well-established in Europe, but there was virtually no experience with startup or operation of these processes in North America. The experience gained from optimization of the sidestream deammonification moving bed biofilm reactor (MBBR) in this study, including the novel pH-based aeration control strategy, has influenced the startup procedure and operation of subsequent full-scale installations in the United States and around the world. Long startup time remains a barrier to the implementation of sidestream deammonification processes, but this study was the first to show the benefits of utilizing media with an existing nitrifying biofilm to speed up anammox bacteria colonization. Utilizing media with an established biofilm from a mature integrated fixed film activated sludge (IFAS) process resulted in at least five times greater anammox activity rates in one month than virgin media without a preliminary biofilm. This concept has not been testing yet in a full-scale startup, but has the potential to drastically reduce startup time. False dissolved oxygen readings were observed in batch scale denitrification tests, and it was determined that nitric oxide was interfering with optical DO sensors, a problem of which the sensor manufacturers were not aware. This led to at least one sensor manufacturer reevaluating their sensor design and several laboratories and full-scale process installations were able to understand their observed false DO readings. There is an industry-wide trend to utilize influent carbon more efficiently and realize the benefits of mainstream shortcut nitrogen removal. The A/B pilot at the HRSD Chesapeake Elizabeth Treatment provides a unique chance to study these strategies in a continuous flow system with real wastewater. For the first time, it was demonstrated that the presence of influent particulate COD can lead to higher competition for nitrite by heterotrophic denitrifying bacteria, resulting in nitrite oxidizing bacteria (NOB) out-selection. TIN removal was affected by both the type and amount of influent COD, with particulate COD (pCOD) having a stronger influence than soluble COD (sCOD). Based on these findings, an innovative approach to achieving energy efficient biological nitrogen removal was suggested, in which influent carbon fractions are tailored to control specific ammonia and nitrite oxidation rates and thereby achieve energy efficiency in the nitrogen removal goals downstream. Intermittent and continuous aeration strategies were explored for more conventional BNR processes. The effect of influent carbon fractionation on TIN removal was again considered, this time in the context of simultaneous nitrification/denitrification during continuous aeration. It was concluded that intermittent aeration was able to achieve equal or higher TIN removal than continuous aeration at shorter SRTs, whether or not the goal is nitrite shunt. It is sometimes assumed that converting to continuous aeration ammonia-based aeration control (ABAC) or ammonia vs. NOx (AvN) control will result in an additional nitrogen removal simply by reducing the DO setpoint resulting in simultaneous nitrification/denitrification (SND). This work demonstrated that lower DO did not always improve TIN removal and most importantly that aeration control alone cannot guarantee SND. It was concluded that although lower DO is necessary to achieve SND, there also needs to be sufficient carbon available for denitrification. While the implementation of full-scale sidestream anammox happened rather quickly, the implementation of anammox in the mainstream has not followed, without any known full-scale implementations. This is almost certainly because maintaining reliable mainstream NOB out-selection seems to be an insurmountable obstacle to full-scale implementation. Partial denitrification/anammox was proven to be easier to maintain than partial nitritation/anammox and still provides significant aeration and carbon savings compared to traditional nitrification/denitrification. There is a long-standing interest in combining shortcut nitrogen removal with biological phosphorus removal, without much success. In this study, biological phosphorus removal was achieved in an A/B process with A-stage WAS fermentation and shortcut nitrogen removal in B-stage via partial denitrification. / Doctor of Philosophy / When the activated sludge process was first implemented at the beginning of the 20th century, the goal was mainly oxygen demand reduction. In the past few decades, treatment goals have expanded to include nutrient (nitrogen and phosphorus) removal, in response to regulations protecting receiving bodies of water. The only practical way to remove nitrogen in municipal wastewater is via biological treatment, utilizing bacteria, and sometimes archaea, to convert the influent ammonium to dinitrogen gas. Orthophosphate on the other hand can either be removed via chemical precipitation using metal salts or by conversion to and storage of polyphosphate by polyphosphate accumulating organisms (PAO) and then removed in the waste sludge. Nitrification/denitrification and chemical phosphorus removal are well-established practices but utilize more resources than processes without nutrient removal in the form of chemical addition (alkalinity for nitrification, external carbon for denitrification, and metal salts for chemical phosphorus removal), increased reactor volume, and increased aeration energy. Intensification refers to utilizing wastewater treatment processes that decrease chemical and energy demands, increase energy recovery, and reduce the process footprint (or increased capacity in an existing footprint) all while providing the same level of nutrient removal as traditional methods. Shortcut nitrogen removal processes; including nitrite shunt, partial nitritation/anammox, and partial denitrification/anammox, as well as low-carbon biological phosphorus removal, were critically-evaluated in this study with an overall objective of intensification of existing infrastructure. Partial nitritation/anammox is a relatively new technology that has been implemented in many full-scale sidestream processes with high ammonia concentrations, but that has proven difficult in more dilute mainstream conditions due to the difficulty in suppressing nitrite oxidizing bacteria (NOB). Even more challenging is integrating biological phosphorus removal with shortcut nitrogen removal, because biological phosphorus removal requires the readily biodegradable carbon that is diverted. Partial denitrification/anammox provides a viable alternation to partial nitritation/anammox, which may be better suited for integration with biological phosphorus removal.

M.S.

The effects of effluent type, effluent loading rate, dosing interval, and temperature on denitrification in low pressure distribution, on-site wastewater treatment and disposal systems (OSWTDS) were evaluated in this study. The treatments were surface and subsurface soil horizons; nitrified and non-nitrified wastewaters; 0.5, 1.0, and 1.5 times the Virginia Department of Health (VDH 1989) recommended wastewater loading rate; 24 and 48 hour dosing intervals; and summer and winter temperatures. Surface and subsurface soil cores were collected from a Groseclose silt loam soil (clayey, mixed, mesic Typic Hapludult) and subjected to the various treatments. The effects of the treatments on denitrification were evaluated based on analyses of leachate from the cores, soil chemical analyses, and microcosm studies to estimate actual denitrification activity. A model was developed from the study that estimated the mean N₂O production for each combination of experimental treatments. The results of the study and the model indicate that denitrification can be enhanced in OSWTDS by the application of non-nitrified wastewater at one-half the VDH recommended loading rate, or 1.25 cm/day, for surface soil horizons (30 min inch⁻¹ percolation rate) using a 48 hour dosing interval. A field study was conducted on a Lowell silt loam soil (fine, mixed, mesic Typic Hapludalf). Denitrification was measured at this site using acetylene blocking and the results compared to those predicted by the denitrification model developed from the laboratory data. The field measurements of denitrification based on N₂O concentration in the soil atmosphere were three orders of magnitude higher than that predicted by the model. It was concluded that the laboratory techniques can be used to determine optimum method of operation for denitrification in a low pressure distribution system, but it cannot be used to determine the field design loading rates. / Ph. D.

In the recent past more and more attention has been given by sanitary engineers to the problem of nitrogen removal in sewage treatment. This attention is brought about by several problems associated with nitrogen. First, in some locations, such as our southwest United States, there is both an essentially constant supply of water and an increasing demand for water. This calls for water recycling. or reuse, as a means of meeting the demand for potable water. An accumulation of impurities, such as compounds of nitrogen, can limit the recycling of water. One such substance is nitrate nitrogen, which when in excess of ten parts per million can cause the disease methemo-globinemia in bottle-fed infants. / Master of Science

Landfill leachate characteristics vary depending on the operation type of the landfill and the age of the landfill. At landfills operated as bioreactors, where leachate recirculation is practiced, leachate ammonia nitrogen concentrations may accumulate to extremely higher levels than during single pass leaching, thereby requiring treatment before final discharge to a receiving system (Onay, 1998). Usually several physical/chemical wastewater treatment technologies are used to treat the leachate. In most cases the COD and BOD are treated, and then nitrification is performed in a separate sophisticated ex situ system. The additional costs of these systems can be very high. The use of a readily available media for in situ nitrification should be considered a prime objective to avoid extra costs. The possibility of removing ammonia nitrogen from bioreactor landfill leachate using trickling filter biofilm technology was studied in four laboratory scale reactors filled with four different types of packing media. The different packing media were examined to see which media is the most efficient at supporting ammonia removal biofilms. The highest efficiency was achieved by a packing media consisting of pine wood chips. The effects of varied concentration loading, varied hydraulic loading, and nitrification inhibitors were studied. Varied ammonia concentration did not have a huge impact on the ammonia removal rates (77-87%) in the reactor with pine wood media. The ammonia removal rates showed a strong dependence on hydraulic loading rate with the lowest loading rate producing the highest removal rates. Landfill leachate from the Middle Peninsula Landfill in Glens, Virginia was determined not to contain nitrifying inhibitors. Using a wood media filter chip and a low hydraulic loading rate was determined to be the best method to remove ammonia nitrogen from landfill bioreator leachate. / Master of Science

Data from Hampton Roads Sanitation District was used to calculate nitrification and denitrification rates for the A²/O mode (1987) and the VIP mode (1988) of operation. Nitrification and denitrification rates compared to literature values for similar sludge ages. The mean VIP nitrification rate was eight percent less compared to the A²/O mode. Denitrification varied with the amount of nitrate loading to the anoxic zone and the rate of total nitrate recycle. The amount of denitrification that occurred in each zone during the different operations was determined. Process mode variations caused different percentages in each zone. Anaerobic and anoxic denitrification was a linear function of the mass of nitrate recycled to the anoxic zone. Fifty to seventy-five percent of the denitrification took place in the aerobic basin during both process modes, but more aerobic denitrification occurred for the A²/O operation. Secondary clarifier nitrate varied inversely with the nitrate recycle similarly for both process modes. The differences in sludge production between the VIP and A²/O process could be explained by the differences in mean cell residence time. / Master of Science

Thesis (MScEng) -- Stellenbosch University, 2014. / ENGLISH ABSTRACT: Urine contains up to 80% of nitrogen, 50 % of phosphates and 90 % of potassium of the total load in domestic wastewater but makes up less than 1% of the total volume (Larsen et al., 1996). The source separation and separate treatment of this concentrated waste stream can have various downstream advantages on wastewater infrastructure and treated effluent quality. The handling of undiluted source separated urine however poses various challenges from the origin onward. The urine has to be transported to a point of discharge and ultimately has to be treated in order to remove the high loads of organics and nutrients. Wilsenach (2006) proposed onsite treatment of source separated urine in a sequencing batch reactor before discharging it into the sewer system. This study focused on the treatment of urine in a sequencing batch reactor (SBR) primarily for removal of nitrogen through biological nitrification-denitrification. The aim of the study was to determine nitrification and denitrification kinetics of undiluted urine as well as quantification of the stoichiometric reactions. A further objective was to develop a mathematical model for nitrification and denitrification of urine using experimental data from the SBR. The SBR was operated in 24 hour cycles consisting of an anoxic denitrification phase and an aerobic nitrification phase. The sludge age and hydraulic retention time was maintained at 20 days. pH was controlled through influent urine during volume exchanges. Undiluted urine for the study was obtained from a source separation system at an office at the CSIR campus in Stellenbosch. Conditions in the reactor were monitored by online temperature, pH and ORP probes. The OUR of the system was also measured online. One of the main challenges in the biological treatment of undiluted urine was the inhibiting effect thereof on nitrification rate. The anoxic mass fraction was therefore limited to 17 % in order to allow longer aerobic phases and compensate for the slow nitrification rates. Volume exchanges were also limited to 5% of the reactor volume in order to maintain pH within optimal range. Samples from the reactor were analysed for TKN, FSA-N, nitrite-N, nitrate-N and COD. From the analytical results it was concluded that ammonia oxidising organisms and nitrite oxidising organism were inhibited as significant concentrations of ammonia-N and nitrite-N were present in the effluent. It was also concluded that nitrite oxidising organisms were more severely inhibited than ammonia oxidising organisms as nitrate-N was present in very low concentrations in the effluent and in some instances not present at all. Ultimately the experimental system was capable of converting 66% of FSA-N to nitrite- N/nitrate-N of which 44% was converted to nitrogen gas. On average 48% of COD was removed. A mathematical model was developed in spreadsheet form using a time step integration method. The model was calibrated with measured online data from the SBR and evaluated by comparing the output with analytical results. Biomass in the model was devised into three groups, namely heterotrophic organisms, autotrophic ammonia oxidisers (AAO) and autotrophic nitrite oxidisers (ANO). It was found that biomass fractionation into these three groups of 40% heterotrophs, 30% AAO and 30% ANO produced best results. The model was capable of reproducing the general trends of changes in substrate for the various organism groups as well as OUR. The accuracy of the results however varies and nearexact results were not always achievable. The model has some imperfections and limitations but provides a basis for future work.

A ampla ocorrência de contaminação de águas por compostos de nitrogênio em concentrações superiores às recomendadas pela legislação tem suscitado interesse no desenvolvimento de tecnologias viáveis de remoção desses compostos. A remoção biológica de nitrogênio apresenta como principais vantagens os custos relativamente reduzidos e a possibilidade de maior eficiência. Compostos reduzidos de enxofre como sulfetos podem ser oxidados a enxofre elementar ou a sulfato por bactérias oxidantes de sulfeto que utilizam nitrato ou nitrito como receptor de elétrons. Esta desnitrificação reduz os requerimentos globais de carbono para a remoção de nutrientes, com menor produção de lodo, proporcionando grande economia. O objetivo desta pesquisa consistiu em contribuir para o conhecimento acerca dos aspectos microbiológicos do processo de desnitrificação com o uso de sulfeto. Foram avaliados os efeitos dos modos de operação dos reatores desnitrificantes sobre a biomassa em cada uma das diferentes configurações e monitorada a colonização microbiana por meio de técnicas de Biologia Molecular como PCR/DGGE, sequenciamento e análises filogenéticas. Os fragmentos do gene RNAr 16S foram relacionados aos gêneros Pseudomonas, Aeromonas, Acidobacteria, Chlroroflexi, Clostridium, Cupriavidus e Ralstonia. Filotipos dos clones para o sistema piloto foram associados a bactérias não cultiváveis e Firmicutes envolvidos na digestão anaeróbia em reatores tratando água residuária, Synergistetes, Deferribacteria e Proteobacteria. Foram identificados micro-organismos presentes nos reatores com reconhecida capacidade para desnitrificação: Pseudomonas, Desulfovibrio desulfuricans e Ralstonia. Amostras de ambos os reatores desnitrificantes apresentaram reações de amplificação positivas com primers específicos para bactérias semelhantes a Thiomicrospira associados a primers universais: 100% das amostras amplificaram com OST1F/1492R e 75% com EUB8F/OSTR1R. Para duas condições de operação do reator em escala de bancada foram identificados micro-organismos semelhantes a Sulfurimonas denitrificans, bactéria autotrófica redutora de nitrato e oxidadora de sulfeto. Chloroflexi também foram encontrados em digestores localizados em plantas de tratamento de águas residuárias recebendo essencialmente efluente doméstico e Propionibacterium foi associada a comunidade microbiana de reatores UASB tratando água residuária industrial. Bactérias em associações sintróficas com participação no ciclo do enxofre e/ou na digestão anaeróbia foram igualmente identificadas: Clostridium sulfidigenes e outros Firmicutes, Synergistetes, clones de bactérias de cultura de enriquecimento e bactérias do gênero Syntrophorhabdus. Os resultados deste trabalho proporcionaram associar a colonização microbiana com o desempenho e as características metabólicas de alguns micro-organismos com reatores desnitrificantes combinados para o tratamento de águas residuárias. / The widespread nitrate contamination in concentrations higher than recommended by legislation has raised interest in technologies for water and wastewater treatment. Biological nitrogen removal is relatively low cost and higher efficiency. Sulfide as electron donor can be oxidized to elemental sulfur or sulfate by sulfide oxidizing bacteria that can use nitrate or nitrite as electron acceptor. This type of denitrification reduces the overall requirements for removal of carbon nutrients and less sludge is produced. The aim of this research was to contribute to microbiological knowledge about denitrification using sulfide. The effects of operation conditions on the denitrifying biomass were monitored through molecular biology techniques such as PCR/DGGE, sequencing and phylogenetic analysis. 16S rRNA gene fragments were related to Pseudomonas, Aeromonas, Acidobacteria, Chlroroflexi, Clostridium, Cupriavidus and Ralstonia. Phylotypes of clones from samples of pilot-scale reactor were associated with non-cultivable bacteria and Firmicutes involved in anaerobic digestion of wastewater, Synergistetes, Deferribacteria and Proteobacteria. Microorganisms with ability to denitrification were identified in both reactors: Pseudomonas, Desulfovibrio desulfuricans and Ralstonia. Samples of denitrifying reactors showed positive amplification with specific primers for Thiomicrospira associated to universal primers: 100% of the samples amplified with OST1F/1492R and 75% EUB8F/OSTR1R. Sulfurimonas denitrificans-like were identified for two operational conditions of the bench scale reactor. Chloroflexi were also found in treatment plants digesters receiving domestic wastewater and Propionibacterium was associated with microbial community of UASB reactors treating industrial wastewater. Syntrophic bacteria participating in the sulfur cycle and/or anaerobic digestion were also identified: Clostridium sulfidigenes and other Firmicutes, Synergistetes, clone enrichment culture bacteria and Syntrophorhabdus. These results provide to associate this microbial colonization and metabolic characteristics of some microorganisms with performance of combined denitrifying reactors for treatment of wastewater.

O processo de nitrificação e desnitrificação simultâneas (NDS) permite alcançar a remoção combinada de matérias carbonácea e nitrogenada em uma única unidade. O reator de leito estruturado, com biomassa imobilizada e recirculação interna, apresenta características positivas para que estes processos envolvidos ocorram, tais como propiciar a formação de biofilme e evitar a colmatação do leito. Esta configuração tem sido estudada com êxito em reatores em escala de bancada para tratamento de esgoto. Nesta pesquisa foi utilizado um reator de leito estruturado em escala piloto com a finalidade de avaliar sua implantação, eficiência e estabilidade tratando esgoto doméstico em condições reais para futura aplicação em pequenas comunidades, condomínios residenciais entre outros como sistema descentralizado. O reator foi construído em fibra de vidro, de formato cilíndrico, com diâmetro interno de aproximadamente 0,80 m e 2,0 m de altura. O volume total foi de aproximadamente 0,905 m3 e o volume útil de 0,642 m3. A operação foi realizada sob condições de aeração contínua e intermitente e os tempos de detenção hidráulica (TDH) testados foram de 48, 36 e 24 horas. A remoção de DQO manteve-se acima de 90% com TDH de 48 e 36 horas. A melhor eficiência de remoção de nitrogênio total foi de 72,4 ± 6,4%, sob TDH de 48 horas e a aeração intermitente, com 2 horas de aeração e 1 hora não aerada. A concentração de oxigênio dissolvido (OD) média de 2,8 ± 0,5 mg.L-1 na fase aerada e temperatura média de 24,7 ± 1,0 °C. Nesse mesmo período, a eficiência média de remoção de DQO foi de 94 ± 4 %. Apesar das dificuldades apresentadas no controle da aeração, as eficiências das remoções obtidas indicaram que o reator de leito estruturado e aeração intermitente (LEAI) se apresenta como uma alternativa promissora em escala plena, requerendo ajustes para construção e incremento da estabilidade da NDS. / The simultaneous nitrification and denitrification (SND) process allow achieving a combined nitrogen and carbon removal in a single unit. The structured-bed reactor, with immobilized biomass and internal recirculation, presents positive characteristics for occurrence of these involved processes, such as providing biofilm formation and prevent clogging. This configuration have been study successfully in a bench scale for sewage treatment. In this research was used a structured-bed reactor in a pilot scale in order to evaluate its implantation, efficiency and stability treating sewage in real conditions to future application in small communities, residential condominium among others, as decentralized treatment. The reactor was constructed with cylindrical glass fiber modules, with internal diameter of 0.8 m and 2.0 m height. The total volume was about 0.905 m3 and 0.642 m3 working volume. It was worked under continuous and intermittent aeration conditions and the tested hydraulic retention times (HRT) were 48, 36 e 24 hours. The maximum total nitrogen removal was 72.4 ± 6.4% ; the HRT was 48 hours with intermittent aeration, in which 2 hours in aeration and 1-hour nonaeration. An average dissolved oxygen (DO) concentration of 2.8 ± 0.5 mg.L-1 in the aerated time and an average temperature of 24.7 ± 1.0°C. In the same period, the average COD removal was 94 ± 4%. Despite of the difficulties presented in aeration control, the obtained removals indicated the structured-bed reactor subjected to recirculation and intermittent aeration (SBRRIA) can be a promising alternative in full scale, demanding adjustments to improve the reactor construction and the SND stability.

This doctoral research provides an advanced molecular approach for the investigation of microbial structure and function in response to operational conditions of biological nitrogen removal (BNR) processes, including those leading to direct production of a major greenhouse gas, nitrous oxide (N₂O). The wastewater treatment sector is estimated to account with 3% of total anthropogenic N₂O emissions. Nevertheless, the contribution from wastewater treatment plants (WWTPs) is considered underestimated due to several limitations on the estimation methodology approach suggested by the Intergovernmental Panel on Climate Change (IPCC). Although for the past years efforts have been made to characterize the production of N₂O from these systems, there are still several limitations on fundamental knowledge and operational applications. Those include lack of information of N₂O production pathways associated with control of aeration, supplemental organic carbon sources and adaptation of the microbial community to the repeated operational conditions, among others. The components of this thesis, lab-scale investigations and full-scale monitoring of N₂O production pathways and emissions in conjunction with meta-omics approach, have a combined role in addressing such limitations. Lab-scale experiments imposing short-term anoxic-aerobic cycling on partial- and full-nitrification based processes were conducted to investigate the microbial response to N₂O production. Interestingly, it was determined that full-nitrification systems could be a higher contributor to N₂O production and emissions than partial-nitrification. While it has been reported in the literature a higher contribution from the latter when the microbial community is not subjected to oxygen cycling conditions. Following the knowledge obtained with a single anoxic-aerobic cycle imposed to nitrifying communities, long-term adaptation of the microbial community to continued anoxic-aerobic cycling and its impact on N₂O production were investigated through a meta-omics approach. Long-term studies are particularly significant regarding engineered systems, where the microorganisms are continually subjected to cycling conditions again and again. A microbial adaptation at the RNA level was identified on both autotroph and heterotroph organisms. The transcripts of the metabolic pathways related to NO and N₂O production (nir, nor) and consumption (nor, nos) were initially induced followed by a gradual decline, leading to a parallel reduction in gaseous emissions over time. Other pathways not typically interrogated in conjunction with the nitrogen metabolism, such as electron transport chain and carbon fixation were also investigated and revealed a mechanism to overcome the imbalance in electron flow and generation of proton motive force (increased transcription of terminal oxidase genes, cco and cox) to uphold carbon fixation during continued cycling. The second part of this thesis focuses on full-scale WWTPs, where it is crucial to determine specific nuances of the systems’ dynamics and of the different types of treatment that may contribute to increased production and emissions of N₂O. For that purpose, two distinct BNR systems not usually considered and studied in terms of N₂O production and emissions were chosen. First, a separate centrate treatment (SCT) process employing glycerol as the supplemental carbon source was monitored. Significantly, this system was found to have one of the highest levels of N₂O production and emission report thus far. Glycerol revealed to foster a microbial community (i.e. Burkholderiales, Rhodobacterales and Sphingomonadales) that stores internal carbon and promote partial denitrification, leading to accumulation of nitrite and N₂O [7-11]. Second, both fixed- and moving-bed biofilm BNR systems were investigated. The overall N₂O emission fractions for the Integrated Fixed-Film Activated Sludge (IFAS)(0.09 – 1.1% infl-TKN) and denitrification filters (0.11 – 1.4% infl-TN) were similar to the reported emissions from suspended growth activated sludge systems [4-6]. For the IFAS system, aqueous and gaseous N₂O profiles paralleled the diurnal variability on influent nitrogen load. The production of N₂O was significantly correlated with ammonia concentration (p<0.05, r=0.91), suggesting the production through hydroxylamine oxidation pathway. Denitrification filters displayed a very peculiar pattern on N₂O emissions associated with intermittent operational cycles (i.e. nitrogen release cycle and backwash). These intrinsic operations of the denitrification filters contributed to transient oxygen conditions and nearly the entire N₂O emissions through gaseous stripping and production by inhibition of denitrification. Similarly to suspended growth systems, process design and operations demonstrated to also play an important role in N₂O emissions from attached growth processes. Finally, aeration strategies for energy efficient conventional nitrification based on the microbial community development and its associated performance was investigated in lab-scale. It was demonstrated that using the same air supply rate, continuous and intermittent aeration resulted in completely different microbial structure. Consequently, distinct kinetics and nitrification performance were observed. The aeration rate could be minimized (resulting in reduction in energy consumption) for high ammonia removal efficiency and lower N₂O emissions, as long as the process is designed accordingly to the microbial ecology developed in such conditions. In sum, the microbial structure, function and connection of metabolic pathways of complex engineered microbial communities as applicable to BNR systems and its operations were investigated in detail. From an engineering perspective, this dissertation provides an advancement on the molecular approach to characterize structure and function of microbial responses to engineered operations beyond the business-as-usual target genes, which can eventually result in better design and control of engineered BNR processes. This study offers more than an improved scientific understanding of the complex microbial environment and direct engineering applications. It connects sanitation with water quality and the greenhouse gas effect by prioritizing concurrent enhanced biological nitrogen removal and mitigation of N₂O production and emission. Ultimately the implications of the result presented herein can provide economical, environmental, health benefits for the society.