This membrane aerated biofilm reactor (MABR) pilot project was performed at Ekeby wastewater treatment plant (WWTP) in Eskilstuna, Sweden. This plant is facing a future challenge of effluent TN < 10mg/L according to the new standard and the growing population, where higher treatment capacity is needed. The MABR as a newly invented technology, is chosen as a promising countermeasure towards the challenge, because of the simultaneous nitrification and denitrification of this technology. By the time of reporting, this project is still on-going, and more information will be reported later in separate report. The feed water comes from the secondary clarifier of full-scale plant. Dissolved oxygen (DO), processing air and flow rate was manually controlled to test different operational settings. However, there were a lot challenges during the testing period which makes it hard to evaluate the performance of this pilot. The ammonium removal efficiency is satisfying when the pilot was running smoothly. However, the TN removal efficiency did not comply with the expectation, achieved in average of 39,01%, due to the limitation of readily bio-degradable COD (rbCOD), which is a limitation of the biological process in general and is not specific to MABR. This technology is considered as promising by the end of the current testing period, since it can oxidize the ammonium effectively with smaller volume. / Detta pilotprojekt med membranluftad biofilmreaktor (MABR) utfördes på Ekeby avloppsreningsverk i Eskilstuna, Sverige. Denna anläggning står inför en framtida utmaning med utflöde-TN <10 mg / L enligt den nya standarden och den växande befolkningen, där högre behandlingskapacitet behövs. MABR som nyutvecklad teknik, väljs som en lovande motåtgärd för utmaningen på grund av den samtidiga nitrifikationen och denitrifikationen med denna teknik. Vid rapporteringstillfället är projektet fortfarande pågående och mer information kommer att rapporteras senare i separat rapport. Matarvattnet kommer från den sekundärfällningen i fullskaleanläggningen. Löst syre (DO), bearbetningsluft och flödeshastighet kontrollerades manuellt för att testa olika driftsinställningar. Det fanns emellertid många utmaningar under testperioden vilket gör det svårt att utvärdera prestanda för denna pilot. Ammoniumavlägsningsgraden var tillfredsställande när piloten körde smidigt. TN-avlägsningseffektiviteten som i genomsnitt uppnådde 39,01% TN-avlägsning motsvarade emellertid inte förväntan, på grund av begränsningen av lätt biologisk nedbrytbar COD (rbCOD), vilken är en begränsning av den biologiska processen i allmänhet och inte specifik för MABR. Denna teknik anses vara lovande vid slutet av den aktuella testperioden, eftersom den kan oxidera ammoniumen effektivt med en mindre volym.

Research was performed at a pilot-scale wastewater treatment plant operating an adsorption/bio-oxidation (A/B) process at 20C. The study compared B-Stage performance under DO Control, Ammonia Based Aeration Control (ABAC), and Ammonia vs. NOx (AvN) control. AvN in 1) fully-intermittent and 2) intermittently-aerated MLE configurations was compared to DO Control and ABAC, each with continuous aeration, in an MLE configuration. The study also examined operation of each aeration strategy with two different feed types: A-Stage effluent (ASE) and primary clarifier effluent (PCE). Operating modes were compared on the basis of nitrogen removal performance, COD utilization efficiency for denitrification, and alkalinity consumption. AvN was found to provide comparable nitrogen removal performance to DO Control and ABAC. The highest nitrogen removal performance was seen when operating DO Control (81.4 ± 1.2%) and ABAC (81.1 ± 1.2%) with PCE. High nitrogen removal efficiency (77.5 ± 6.1%) was seen when fully-intermittent AvN operation was fed ASE containing a high particulate COD fraction. A high effluent nitrite accumulation ratio (NAR = NO2-/(NO2-+NO3-)) was seen during this period (46 ± 15%) accompanied by the out-selection of Nitrospira. Feeding effluent from AvN control to an Anammox MBBR improved removal efficiency. Increased soluble COD loading resulted in greater nitrogen removal with strategies operating in an MLE configuration while particulate COD was found to be important for processes where removal was designed to occur in downstream reactors. Efficiency of COD for denitrification was found to vary based on the amount and type of influent COD; however AvN in an MLE configuration was found to use COD more efficiently than fully-intermittent AvN. In either configuration, AvN required less alkalinity addition than DO Control or ABAC. High sCOD concentrations in PCE led to increased nutrient removal as compared to ASE but increased heterotrophic growth and mixed liquor concentrations in the B-Stage making the A-Stage an attractive option for its ability to control the C/N ratio fed to BNR processes. / Master of Science

Deammonification is a widely used technology for side stream treatment with rich ammonium streams at relatively high temperatures, such as, the reject water coming from dewatering units in treatment of digested sludge and industrial wastewaters. The deammonification process has lower operational costs than conventional systems, consumes less energy, enables the increase of biogas production and it is easy to implement. However, this technology has not yet been applied in full- scale mainstream treatment due to its restrictions in coping with high C/N ratios, low temperatures, and the need for post-treatment processes. These conditions are allegedly negative to the growth and performance of anammox bacteria affecting the bacterial groups’ behavior in the process. This master thesis project aimed to evaluate the feasibility of using deammonification to remove nitrogen from mainstream wastewater, which was studied by monitoring the bacterial activity in a pilot scale reactor. The different bacterial groups involved (AOB, NOB, heterotrophs, and denitrifiers) were monitored by weekly measuring their activity in batch activity tests. The results allowed the evaluation of different operational scenarios and their impact by following up on the changes in the bacterial competition. The study was conducted for six months in a single-stage IFAS (integrated fixed-film activated sludge) pilot-scale reactor located in Stockholm and fed with pretreated (with a UASB) municipal wastewater. The different operational scenarios involved changes in temperature, aeration patterns, DO concentration, SRT, and HRT. The adjustment of these features was done in the interest of promoting AOB and anammox bacterial growth, leading to an improvement of the deammonification efficiency in future studies. However, the chosen operational conditions were to enhance bacterial competition and facilitate its visualization, not to maximize nitrogen removal. Thus, the most suitable scenario found during this study included DO concentration of 1.5 mg/L with 10 aeration-20 non-aeration pattern and ensured nitrogen removal rates within normal values while allowing the monitoring of all the bacterial groups. TN removal reached a value above 50% and NH4-N above 95%, whereas nitrogen Removal Rate (NRR) increased to 30g/N/m3-d and the system had an overall nitrogen removal efficiency of 75%. Nevertheless, it was proven that in the right environment, the necessary bacterial groups can be selectively accumulated and successfully perform deammonification and reduce nitrogen levels in mainstream wastewater. / Deammonifikation är en välanvänd teknik för rening av sidoströmmar med höga ammoniumkoncentrationer vid relativt hög temperatur, som till exempel rejektvatten från avvattning av rötslam eller industriellt avloppsvatten. Deammonifikationsprocessen har lägre driftkostnad än konventionella reningsprocesser, förbrukar mindre energi samt möjliggör högre biogasproduktion samtidigt som processen är enkel att implementera. Reningstekniken har dock ännu inte tillämpats i fullskala för rening av huvudströmmen på grund av den höga C/N-kvoten och de låga vattentemperaturerna i kommunalt avloppsvatten samt behovet av efterbehandling. Detta anses ha en negativ inverkan på anammoxbakteriernas tillväxthastighet och funktion vilket påverkar bakteriegruppens beteende i processen. Syftet med detta examensarbete var att utvärdera om det är praktiskt genomförbart att använda deammonifikation för att rena kväve från kommunalt avloppsvatten, vilket följdes upp genom att studera bakterieaktiviteten i en pilotskalereaktor. De involverade bakteriegrupperna (AOB, NOB, heterotrofer och denitrifierare) övervakades genom att mäta den mikrobiella aktiviteten varje vecka med hjälp av batch-tester. Resultaten användes till att utvärdera olika driftstrategier och deras effekt genom att följa förändringarna i mikrobiell aktivitet hos de konkurrerande bakteriegrupperna. Studien genomfördes i Stockholm under sex månader i en enstegs-IFAS-pilotskalereaktor (integrerad process med biofilm på fast bärarmaterial och aktivslam) som matades med kommunalt avloppsvatten som förbehandlats i en UASB-reaktor. De olika driftstrategierna omfattade olika temperaturer, luftningsstrategier, syrekoncentrationer, slamåldrar och hydrauliska uppehållstider. Syftet med driftstrategierna var att främja AOB- och anammoxbakteriers tillväxt för att i framtida studier kunna erhålla en förbättrad deammonifikationsprocess. Syftet i denna studie var dock i första hand att förbättra den bakteriella konkurrensen och göra den lättare att mäta, inte att uppnå bästa möjliga kväverening. Den driftstrategi som gav bäst resultat i denna studie innebar att hålla en syrehalt på 1,5 mg/l med 10 minuter luftning följt av 20 minuter utan luftning vilket säkerställde en normal kväveavskiljning och samtidigt möjliggjorde övervakning av samtliga fyra bakteriegrupper. Totalkväveavskiljningen var över 50 % och ammoniumavskiljningen över 95 % medan kvävereningsaktiviteten ökade till 30 g N/m3-d och systemet hade en övergripande effektivitet på 75 %. Studien visade att under rätt förutsättningar kan de nödvändiga bakteriegrupperna selekteras fram och deammonifikation av kommunalt avloppsvatten kan utföras på ett framgångsrikt sätt.

Mathematical models of a nitritation: anaerobic ammonia oxidation (anammox)-coupled biofilm in a counter-diffusion hollow fiber membrane bioreactor (HFMBR) and a nitritation: anammox-coupled biofilm in a co-diffusion rotating biological contactor (RBC) were developed and implemented using AQUASIM. Four different start-up scenarios on the nitritation: anammox-coupled biofilm in an HFMBR were investigated. The supply of oxygen was simulated with the flow through the lumen of the hollow fiber membrane. For the four scenarios, two scenarios investigated the start-up when nitrite was supplied in the feed while the other two scenarios investigated when the source of nitrite was through nitritation only. The results showed that the presence of nitrite in the feed facilitated the start-up of the reactor. In addition, the results also showed that increasing oxygen flux through the membrane up to a certain ratio of ammonia flux with oxygen flux affected reactor performance by improving nitrogen removal and reducing start up time. For the nitritation: anammox-coupled biofilm in an RBC, four different process options were investigated: the number of reactors, the initial anammox (AnAOB) biomass fraction, the bulk oxygen concentration and the maximum biofilm thickness. Modeling results revealed that the steady state total nitrogen removal in RBC reactors in series occurred primarily in the first and second reactors. It is concluded that the number of reactors in series dictates the effluent performance and, therefore, this number can be selected depending upon the desired total nitrogen removal. Simulation results also revealed that increasing the initial AnAOB biomass fraction from 0.01% to 1.0% had no effect in the steady state nitrogen removal but had an effect in the required time to reach the steady state total nitrogen removal and the maximum biofilm thickness. Modeling results of the third process option showed that increasing the bulk oxygen concentration in the reactor from 0.2 g/m3 to 5 g/m3 linearly increased the steady state total nitrogen removal and reduced the time to reach the maximum biofilm thickness. Beyond 5 g/m3, steady state total nitrogen removal decreased. In addition, simulation results revealed that the thicker biofilm clearly showed a more linear correlation between the increase in bulk oxygen concentration and the increase in the steady state total nitrogen removal within a range of bulk oxygen concentrations. The results showed that RBC performance could be controlled by several process options: the number of reactors in series, initial biomass fraction, the bulk oxygen concentration and the maximum biofilm thickness. The mathematical modeling results for the HFMBR and RBC have shown that both have potential as carriers for nitritation: anammox-coupled biofilms targeted at the removal of nitrogen in the wastewater. / Master of Science

Övergödning är ett utbrett miljöproblem i hav och sjöar, som yttrar sig i toxiska algblomningar och en minskad biodiversitet. Eftersom kväve är en betydande källa till övergödning, har flera avloppsreningsverk (ARV) krav på kväveavskiljning innan utsläpp i recipient. Tierp ARV omfattas [TA1] för närvarande inte av sådana utsläppskrav på kväve, men förväntas bli det i och med kommande lagstiftning. En utbyggnad av det biologiska reningssteget, som idag endast medför viss kväveavskiljning, kan därför bli nödvändigt för att åstadkomma fullständig kväverening.  Syftet med examensarbetet var att föreslå reningstekniker som kan åstadkomma tillräcklig kväveavskiljning vid Tierp ARV och bedöma vilken av dessa som är mest fördelaktig för implementering. Vidare utvärderades nuvarande reduktion av kväve och organiskt material samt rådande nitrifikation i det biologiska reningssteget. Massbalanser upprättades utifrån provtagning på dessa parametrar. Lämpliga kvävereningstekniker föreslogs med hänsyn till lokala krav och förutsättningar. Dessa tekniker jämfördes därefter i en multikriterieanalys med avseende på platsåtgång, växthusgasutsläpp, drift- och underhållskostnader, kapitalkostnader och arbetsinsats.  Massbalanserna uppvisade en reduktion på 31 % för totalkväve, 10 % för ammoniumkväve, 63 % för nitratkväve, 7 % för nitritkväve, 62 % för COD och 67 % för BOD7. Nuvarande nitrifikation bedömdes vara relativt låg, vilket troligtvis är resultatet av den låga temperaturen på avloppsvattnet, eventuell otillräcklig syretillförsel samt ogynnsamt pH. De reningstekniker som föreslås kunna minska kväveutsläppen från Tierp ARV är aktivslamprocess, membranbioreaktor (MBR) och hybridprocess med biofilmsbärare och aktivslam (IFAS). Utifrån en samlad bedömning av kriterierna i multikriterieanalysen bedömdes IFAS som mest fördelaktig för etablering och MBR som minst gynnsamt. / Eutrophication is a major environmental problem in oceans and lakes, which manifests itself intoxic algal blooms and reduced biodiversity. As nitrogen is a significant contributor toeutrophication, numerous wastewater treatment plants (WTPs) have requirements for nitrogenremoval before discharge into the recipient. Tierp WTP is not currently subject to suchregulation, but is expected to be so with forthcoming legislation. Consequently, an expansion ofthe biological treatment, which presently facilitates only partial nitrogen removal, may becomepertinent to achieve complete nitrogen removal. The objective of this thesis was to propose treatment methods capable of achieving adequatenitrogen removal at Tierp WTP and evaluate the most suitable option for implementation.Furthermore, the current nitrogen and organic material reduction, as well as present nitrificationin the biological treatment, were assessed. Mass balances were established based on sampling ofthese parameters. Suitable nitrogen purification techniques were proposed, considering localrequirements and conditions. These techniques were then compared through a multi-criteriaanalysis, taking into account space occupancy, greenhouse gas emissions, operation andmaintenance costs, capital costs and workload. The mass balances revealed reductions of 31% for total nitrogen, 10% for ammonium nitrogen,63% for nitrate nitrogen, 7% for nitrite nitrogen, 62% for COD and 67% for BOD7. The currentnitrification was assessed as relatively low, likely attributable to the low temperature of thewastewater, possibly insufficient oxygen supply and unfavourable pH. The treatment methodsproposed in the thesis to achieve sufficient nitrogen removal at Tierp WTP are the activatedsludge process, membrane bioreactor (MBR), and hybrid process with biofilm carriers andactivated sludge (IFAS). According to the multicriteria analysis, IFAS was deemed the mostadvantageous option for implementation, while MBR was evaluated as the least favourable.

Ökade utsläpp av växthusgaser är ett stort problem idag för den pågående klimatförändringen. Lustgas (N2O) är en mycket potent växthusgas och är relaterad till kväverening vid rening av avloppsvatten. I februari 2014 ansattes en separat rejektvattenreningsprocess på Bromma reningsverk som använder ANITA™Mox med anammox bakterier som reningsteknik. Denna masteruppsats undersöker samband och korrelationer mellan kväverening och N2O utsläpp i rejektvattenreningen på Bromma reningsverk, men syftet att presentera hur Stockholm Vatten och Avfall AB kan minska sina lustgasutsläpp. En manuell profilmätning genomfördes även för att utvärdera hur processen fungerade. Fyra olika scenarion testades under åtta veckor: intermittent luftning med olika fördelning mellan luftning och icke-luftning, olika löst syrekoncentrationer, ändringar i ammoniumbelastningen samt seriekörning av processen. Den manuella profilmätningen mätte pH, konduktivitet, löst syre, nitrit- och ammoniumkoncentrationen. Dessa mätningar togs i sju olika punkter, inkluderat rejektet direkt från centrifugerna och utgående rejekt. Resultaten påvisade att vid intermittent luftning gavs de högsta utsläppen vid 60 minuter luftning och 7,5 minuter paus (1,46% N2O-N/inkommande N-tot). Lägsta utsläppen var vid samma luftningstid men 15 minuter paus (0,44% N2O-N/inkommande N-tot).  Både höga och låga koncentrationer av löst syre gav ökade utsläpp. Med de högsta utsläppen vid 0,77 mg/L (0,88% N2O-N/inkommande N-tot ) och lägsta vid 0,34 mg/L (0,07% N2O-N/inkommande N-tot ). Ändringen av ammoniumbelastning (1,5 högre belastning) hav ingen signifikant skillnad gällande utsläpp, med utsläpp på 0,42–0,90% N2O-N/inkommande N-tot. Seriekörning av processen resulterade i driftproblem med höga ammoniumhalter, med utsläpp på 0,90-1,47% N2O-N/inkommande N-tot. Profilmätningarna visade på att processen är stabil och välutvecklad för att rena rejektet på kväve. Uppsatsen kan ge slutsatsen att rejektvattenreningsprocessen är komplex med många parametrar som påverkar varandra och N2O utsläppen. En ingående och bred förståelse om vad som påverkar utsläppen av lustgas är viktigt för att förstå hur utsläppen ska minskas. En positiv linjär korrelation mellan halten nitrit i processen och lustgasutsläpp kan verifieras, samt påverkan av ökade utsläpp vid både för låga och höga koncentrationer syre. Från profilmätningarna kan slutsatsen dras att det är viktigt att rengöra onlineinstrumenten regelbundet. / Greenhouse gas emissions, particularly nitrous oxide (N2O), are a significant environmental challenge linked to biological nitrogen removal in wastewater treatment. Bromma Wastewater Treatment Plant (WWTP) implemented a separate reject water treatment process in February 2017 using ANTIA™Mox with anammox bacteria. This Master's thesis investigates the relationship and correlations between nitrogen removal and N2O production in this process, aiming to assist Stockholm Vatten och Avfall AB in reducing emissions. A manual profile measurement was conducted to investigate the ongoing process. Over eight weeks, four operating strategies were tested: intermittent aeration with varying ratios between aeration and non-aeration, different dissolved oxygen (DO) concentrations, changes in ammonium load, and series configuration. Manual profile measurements of pH, conductivity, DO, nitrite, and ammonium were taken at seven points in the treatment process, including the supernatant and discharged reject. Findings indicate that intermittent aeration, particularly 60 minutes of aeration followed by a 7.5-minute pause, produced the highest N2O emissions (1.46% N2O-N/Influent N-tot load). The lowest emissions (0.44% N2O-N/Influent N-tot load) occurred with a 15-minute pause. The lowest emission (0.07% N2O-N/Influent N-tot load) was detected at 0.34 mg/L, and high DO concentrations (0.77 mg/L) had the highest emissions (0.88% N2O-N/Influent N-tot load). Variations in ammonium load (1.5 times higher) did not significantly affect emissions, averaging 0.42-0.90% N2O-N/Influent N-tot load. The series configuration faced operational challenges with elevated ammonium levels, leading to emissions of 0.90-1.47% N2O-N/Influent N-tot load. Profile measurements confirmed a stable nitrogen removal process. The study concludes that N2O emissions in reject water treatment are complex and influenced by multiple factors. A positive linear correlation was identified between nitrite concentration and N2O emissions, alongside the impact of not having too-low or too-high DO levels, which results in elevated. Regular maintenance of oxygen sensors is crucial for accurate measurements and effective N2O mitigation.

I takt med att Uppsala växer behöver kapaciteten för avloppsvattenrening på Kungsängsverket byggas ut. För att möta en framtida ökad belastning bedömer Uppsala Vatten & Avfall AB att en kolkälla kommer behöva tillsättas i den biologiska kvävereningen för att effektivisera den heterotrofa denitrifikationen på Kungsängsverket, som idag sker utan tillsats av kolkälla. Potentialen till förbättrad denitrifikation med olika kolkällor utvärderades genom satsvisa laboratorieförsök och litteraturstudier. Syftet var att bättre förstå de studerade kolkällornas funktion och prestanda i denitrifikation för att ge underlag inför en framtida fullskalig implementering av kolkälla. Det övergripande målet var att identifiera vilken eller vilka kolkällor som är mest fördelaktiga med avseende på reningseffektivitet, processpåverkan, doseringsbehov, ekonomi och miljöpåverkan. Triplikata försök genomfördes som denitrifikationstester med aktivt slam vid en genomsnittlig slamtemperatur på 14 ℃ och pH 7-8, där fem externa kolkällor (etanol, Brenntaplus VP1 och tre industriella restprodukter) samt försedimenterat avloppsvatten testades. I litteraturstudien inkluderades även metanol. Från försöksdata bestämdes specifika denitrifikationshastigheter, COD/N-kvoter och utbyteskoefficienter. Även bieffekter såsom nitritackumulering och fosforsläpp studerades. Därefter uppskattades doseringsbehov och kostnader baserat på erhållna resultat och antaganden om framtida produktionsmål för nitratkväve. En likartad prestanda erhölls med en av restprodukterna (RTP-vätska) och etanol som uppnådde högst denitrifikationshastigheter och reduktionsgrader (98 % respektive 97 %). Doseringsbehovet uppskattades vara 4 gånger högre med RTP-vätska jämfört med etanol. Med de två andra restprodukterna (dextrandrank och sackaroslösning) uppnåddes lägst denitrifikationseffektivitet och reduktionsgraderna uppgick till 79 % respektive 47 %. Vid test av sackaroslösning observerades dessutom ofullständig denitrifikation samt höga fosforsläpp. Dextrandranken uppträdde på liknande sätt. I egenskap av restprodukt är RTP-vätskan intressant för fortsatt utvärdering. Fullskalig implementering av RTP-vätska förutsätter att doseringsbehoven kan tillgodoses samt att lämplig distribuering och lagerhållning kan ordnas på Kungsängsverket. / The city of Uppsala is expanding and consequently enhanced capacity at the wastewater treatment plant of Kungsängen will be required in the future. As for the biological nitrogen removal process, Uppsala Vatten & Avfall AB expects an additional carbon source to be necessary in the future denitrification process. Currently, the nitrogen removal is employed without the addition of a carbon source. The potential of enhancing denitrification with different carbon sources was evaluated by conducting lab-scale batch tests and compiling literature data. The objective of this work was to better understand the performance of the chosen carbon sources as electron donors in heterotrophic denitrification and thereby provide groundwork for a future full-scale implementation of a carbon source. Based on information drawn from batch tests and literature, the carbon sources were evaluated with respect to removal efficiency, process compliance, quantitative dosing requirements, costs, and environmental sustainability. Lab-scale trials were conducted as denitrification tests (triplicate) at a mean sludge temperature of 14 ℃ and pH 7-8 with five external carbon sources (ethanol, Brenntaplus VP1, and three industrial waste products) and pretreated wastewater. In the literature review, methanol was included as well. Results obtained from the batch tests were used to determine kinetic parameters, mainly specific denitrification rates, COD/N ratios, and anoxic yield coefficients. Moreover, unwanted side effects due to addition of carbon sources were examined. Dosing requirements and costs were assessed based on previously determined kinetic parameters and supposed future production guidelines for effluent quality with respect to nitrate concentration. Similar performance was observed with one of the waste products (RTP liquid) and ethanol which achieved the highest denitrification rates and degree of removal (98 and 97 %, respectively). The estimated dosing requirement was 4 times higher with the RTP liquid compared to ethanol. The other two waste products, solutions of fructose (dextran) and sucrose, reached the lowest denitrification efficiency and removal degrees were 79 and 47 %, respectively. During tests with sucrose solution, incomplete denitrification and release of phosphorous were observed. The fructose solution showed somewhat similar behavior but to a lesser degree. Being a waste product, the RTP liquid is interesting for future evaluation. Full-scale implementation needs further considerations regarding dosing requirements, distribution, and storage conditions at the site of Kungsängsverket.

Esta pesquisa enfoca a avaliação do desempenho de nova configuração de unidade para tratamento biológico de esgoto sanitário por processo combinado, visando à remoção de nutrientes, especialmente nitrogênio. O sistema construído em escala piloto tem volume útil igual a 71,48 litros. Nesta nova configuração de reatores para tratamento terciário de esgoto sanitário, utilizaram-se três reatores sobrepostos, sendo: um reator UASB - “Upflow Anaerobic Sludge Blanket"; um reator de leito móvel e filme fixo aeróbio (com aplicação de oxigênio puro); e, um reator de leito móvel e filme fixo com ambiente anóxico. O material suporte utilizado nos reatores de leito móvel e filme fixo constituiu alternativa inédita no tratamento de efluentes líquidos. Foram utilizadas cavilhas “ranhuradas" de madeira, de dimensões aproximadas de 8,0 mm de diâmetro por 8,0 mm de comprimento. O desempenho geral do sistema quanto à remoção de matéria carbonácea e compostos nitrogenados, e as associações microbianas formadas nos três reatores são objetos principais de estudo deste trabalho. Foram obtidos resultados de remoção de DQO e de nitrogênio bastante satisfatórios, sobretudo quando o tempo de detenção hidráulica total esteve próximo a 20 horas, incluindo o compartimento de decantação (94,6 % para DQO; e 96,7 % para N-NTK, com formação de nitrato em concentrações inferiores a 10,0 mgN-NO3-/l). O TDH estudado variou entre 5 e 24 horas para os sistema completo. O estudo dos consórcios de microrganismos forneceu excelentes resultados quanto à quantidade, diversidade, e atividade das populações desenvolvidas nos diferentes ambientes, confirmando o bom desempenho do sistema e o fornecimento de ambiente adequado para o desenvolvimento das diferentes populações nos três reatores. A idealização desta configuração tem como finalidade realização de tratamento de esgoto sanitário até nível terciário em planta compacta, podendo ser utilizada por pequenas e grandes comunidades, devido à possibilidade de se construir diversos módulos do sistema proposto. / This research is focused on the performance evaluation of a new system configuration of combined biological reactors, treating domestic wastewater till tertiary level. The pilot scale proposed system had a total useful volume of 71,48 liters. This new configuration involves three types of superposed reactors: an Upflow Anaerobic Sludge Blanket reactor; an aerobic fixed film mobile bed reactor (fed with pure oxygen); and a fixed film mobile bed reactor at anoxic environment. The support material used inside the fixed film mobile bed reactors was wood made grooved dowels. The main purposes of this work are to evaluate the systems general performance regarding COD and nitrogen removal, and the microbial associations inside the reactors. It reached quite satisfactory results regarding COD and nitrogen removal when the total hydraulic retention time (HRT) was around 20 hours, including the clarification zone volume (94,6 % for the COD; and 96,7 % for the TKN-N, whit nitrate formation below 10,0 mgN-NO3-/l). HRTs from 5 to 20 hours were studied. The microbial associations were present in large number, with a great diversity and high specific activities at the different environments, confirming the good results obtained and the suitable environment provided for the growth of the different populations inside the three reactors. The idealization of this treatment system has the goal to treat domestic wastewater from small to large communities till tertiary level, in compact treatment plants, due to the possibility of constructing several modules of the system.

[Truncated abstract] The use of constructed wetlands for tertiary purification of municipal wastewater has received increasing attention around the world because direct discharge of secondary-treated municipal wastewater to water bodies has caused eutrophication. Plant species selection and vegetation management may enhance nutrient removal efficiency in constructed wetlands. However, there is a lack of knowledge on the relations between plant growth and nutrient removal efficiency in constructed wetlands. The objective of this study is to better understand how plant growth and resource allocation are influenced by nutrients in wastewater and how nutrient removal efficiencies are affected by plant species and vegetation management. The preliminary experiment was conducted to select macrophytes, especially ornamental species, to grow in the wastewater in the wetland microcosms. Ten plant species, comprising six ornamental species: Alocasia macrorrhiza, Canna indica, Iris louisiana, Lythrum sp., Zantedeschia aethiopica, Zantedeschia sp., and four sedge species: Baumea articulate, Baumea juncea, Carex tereticaulis and Schoenoplectus validus, were planted in the wetland microcosms and fed a simulated wastewater solution in the concentrations similar to the secondary-treated municipal wastewater. C. indica has shown vigorous and healthy growth, and a relatively high potential of rooting-zone aeration and nutrient removal efficiency. B. articulata and S. validus also showed relatively high nutrient removal efficiency. ... The high nutrient availability and optimum N/P ratio were required for stimulating plant growth, resulting in allocation of more resources to above-ground tissues compared to below-ground parts, and enhancing nutrient removal efficiency. Nutrient removal efficiencies were significantly influenced by growth of C. indica and S. validus, nutrient loading rates and N/P ratios in the wastewater. The nutrient uptake kinetics of C. indica and S. validus were investigated to elucidate the differences in nutrient uptake between species. Wetland plant species have shown differential nutrient uptake efficiency and different preferences for inorganic N source, with C. indica preferring NO3-N and S. validus preferring NH4-N. C. indica had greater capacity than S. validus to take up PO4-P when the concentration of PO4-P in the solution was relatively low, whereas S. validus was more capable than C. indica to take up NO3-N when the concentration of NO3-N in the solution was relatively low. The PO4-P uptake capacity was higher in younger than older plants. Overall, the study has suggested that different plant species have differential capacity to take up nutrients. In addition to nutrient uptake, plants have significant other roles in terms of nutrient removal from the wastewater (such as leaking oxygen into the rhizosphere in which oxidation of substances like ammonia can occur). The properly high nutrient availability and optimum N/P ratio are required to stimulate the plant growth, resulting in enhancing the treatment performance in the wetlands. These findings have important implications for improving our ability to engineer ecological solutions to the problems associated with nutrient-rich wastewater.

The aim of this thesis was to identify the denitrifying microbial communities in activated sludge from full-scale treatment plants and from small-scale reactors exposed to acetate or methanol as external carbon sources. Biological denitrification is currently the most widely used, sustainable and cost-effective process to remove nitrogen from wastewater. Increasingly strict effluent discharge standards are posing significant challenges to plant operators to reduce effluent NO3--N concentrations to levels as low as 2-3 mg L-1 or even lower. The lack of sufficient influent carbon in many municipal wastewater treatment plants makes it very difficult to achieve such low NO3--N concentrations in the effluent. An effective solution to the problem is to introduce additional external carbon sources to enhance denitrification. The selection of external carbon sources is not purely based on costs but is also dependent on the possible microbial transformations that these carbon sources may bring about in activated sludge. The most common carbon source used is methanol due to its low cost, but it has been found to cause long delays until an improvement in denitrification performance is observed. On the other hand, acetate has been found to improve denitrification almost instantaneously when added, but it has a significantly higher cost. In this study, methanol and acetate utilising denitrifiers were investigated in activated sludge with and without enrichment in laboratory scale bioreactors. The relevant denitrifiers were identified and evaluated in situ using culture independent methods particularly stable isotope probing (SIP), 16S rDNA cloning, fluorescence in situ hybridisation (FISH) and microautoradiography (MAR). Activated sludge collected from a biological nutrient removal plant exhibiting good denitrification was enriched in an anoxically-operated sequencing batch reactor (SBR) by feeding methanol as the sole carbon source and nitrate as the electron acceptor. The SBR was operated over a duration of 7 months and the SBR denitrification rate improved from 0.02 mg NO3--N mg mixed liquor volatile suspended solids (MLVSS)-1 h-1 to a steady-state value of 0.06 mg NO3-N mg MLVSS-1 h-1. At steady state operation the enriched biomass was subjected to SIP with 13C-methanol to biomark the denitrifiers capable of utilising methanol under anoxic conditions. The separated 12C-DNA and 13C-DNA fractions from the SIP experiment were individually subjected to full cycle rRNA analysis. The dominant 16S rRNA gene phylotype (Group-A clones) in the 13C-library was closely related to the obligate methylotrophs Methylobacillus and Methylophilus in the order Methylophilales of the Betaproteobacteria (96-97% sequence identities), while the most abundant clone groups in the 12C-library mostly belonged to the family Saprospiraceae in the Bacteroidetes phylum. Oligonucleotide probes were designed for FISH to target the Group-A clones and Methylophilales (probes DEN67 and MET1216, respectively) and the Saprospiraceae clones (probe SAP553). Application of these probes on SBR biomass over the enrichment period demonstrated a strong correlation between the level of SBR denitrification and relative abundance of DEN67-targeted bacteria in the SBR community. By contrast, no correlation was found between denitrification rate and the relative abundances of the well known denitrifying genera Hyphomicrobium and Paracoccus nor the Saprospiraceae-clones visualised by FISH in the SBR biomass. FISH combined with microautoradiography independently confirmed that the DEN67-targeted cells were the dominant bacterial group capable of anoxic [14C] methanol uptake in the enriched biomass. As observed in full-scale operations, the methanol-fed SBR experienced a lag period of several weeks before denitrification performance increased. Using FISH quantification, it was shown that this coincided with the lag phase in the growth of the DEN67-targeted denitrifying population. It was therefore concluded that the Methylophilales bacteria dominant in our SBR system are likely to be important in full-scale methanol-fed denitrifying sludges. The acetate utilising microbial consortium in activated sludge was investigated without prior enrichment using stable isotope probing (SIP). 13C-acetate was used in SIP to biomark the DNA of the denitrifiers. The extracted 13C-DNA fraction was subjected to a full cycle rRNA analysis. The dominant 16S rRNA gene phylotypes in the 13C-library were closely related to bacterial families Comamonadaceae and Rhodocyclaceae of class Betaproteobacteria (96-97% sequence identities). Seven oligonucleotide probes (DEN444, DEN220, DEN581, DEN441, DEN124, DEN220a and DEN1454) for use in FISH was designed to specifically target the identified phylotypes. Application of these probes on the sludge of a continuously fed denitrifying sequencing batch reactor (CFDSBR) operated over a duration of 16 days indicated a strong correlation between the level of CFDSBR denitrification and relative abundance of all probe-targeted bacteria in the CFDSBR community. FISH combined with microautoradiography (FISH-MAR) further confirmed that the DEN581- and DEN124-targeted cells dominating the CFDSBR were capable of taking up [14C] acetate under anoxic conditions. The initial occurrence of the DEN444- and DEN1454-targeted bacteria and the final dominance of DEN581- and DEN124-targeted bacteria in the CFDSBR community were likely related to the changing in-reactor nitrite concentrations during the first few days of CFDSBR operation. Hence, the DEN444- and DEN1454-targeted bacteria were hypothesised to have low affinities for nitrite while DEN124- and DEN581-targeted bacteria have higher nitrite affinities. However, it was clear that all probe-targeted bacteria were denitrifiers capable of utilising acetate as a carbon source. The rapid increase in numbers of the probe-targeted organisms positively correlates with the immediate increase in denitrification rates. The rapid response and community shifts observed when acetate was used to enhance denitrification suggest that an intermittent application of acetate is quite effective to temporarily enhance the denitrification capacity of a treatment plant. However, the importance of a bacterial impact assessment of activated sludge subjected to intermittent acetate supplementation is recommended prior to the wide use of acetate in the wastewater industry. The acetate utilising denitrifying microbial communities investigated in the previous chapter were characterised according to their eco-physiological properties using the r- and K-selection criteria. The electron donor (acetate) and acceptor (nitrite) affinities of these probe-identified denitrifiers were used as traits for this characterisation. The substrate to microorganism (S/M) ratio was manipulated to provide high and low substrate concentrations in the reactor to create conditions favourable for r- and K-strategists, respectively. Two factors, namely feeding regimes and sludge retention times, were studied to achieve the desired S/M ratios and enable r/K characterisation. The high substrate affinities and high specific growth rates of two probe-identified denitrifiers (DEN124 and DEN581) did not enable resolution of these two organisms with the feeding regimes used in this study. However, the application of different sludge retention times as a control strategy to maintain constant high and low in-reactor S/M ratios enabled characterisation of the two probe-targeted denitrifiers DEN124 and DEN581 as K- and r-strategists, respectively. The in-reactor S/M ratios applied in this study did not facilitate the characterisation of populations targeted by probes DEN444 and DEN1454. The minor fluctuations of the S/M ratios during a cycle in the SBR operation was considered as a drawback, but conclusive results could still be obtained from the study. A chemostat reactor operation with constant loading and variable flow rates is suggested as an alternative. Conclusively, this study was able to identify specific groups of denitrifying microorganisms in activated sludge when exposed to acetate and methanol. Unlike most previous studies, which relied on culture dependent methods, this study adopted a pure culture independent approach to identify microorganisms in relation to their function, i.e. denitrification. Moreover, acetate denitrifiers were in situ characterised based on eco-physiological properties. The identification of denitrifying communities in this study has paved the way to a larger research project on the optimisation of denitrification processes with external acetate, methanol and other carbon supplements. As such, this study has contributed significantly to the understanding of the denitrification processes by linking process data with microbial investigations.