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Executive Summary:

This project aimed to utilize biofilm for nitritation/denitritation in the effluent of UASB reactor treating municipal wastewater at temperatures relevant for wastewater streams in Central Europe (10 – 15 °C). Because the unit processes, i.e. nitritation in biofilm reactor and denitrification in reactor with granular biomass, have been demonstrated previously only at higher temperatures (above 25 °C), the main goals of this project were:

(1) to adapt the nitritation/denitritation process for low temperature (<15 °C),
(2) to characterize the biofilms developed in both reactors and
(3) to develop and operate a complex system for COD and nitrogen removal from municipal wastewater including solving technological issues connected to operation of the SBR (retention tank etc.).
(1) Adaptation of the nitritation/denitritation process for low temperature

a. Operation of Nitritation Moving Bed Biofilm Reactors (MBBR) at various temperatures applying varying technological parameters

The application of nitrification-denitrification over nitrates (nitritation-denitritation) with municipal (i.e. diluted and could) wastewater can substantially improve the energy balance of municipal wastewater treatment plants. For the accumulation of nitrite, it is crucial to inhibit Nitrite Oxidizing Bacteria (NOB) with simultaneous proliferation of Ammonium Oxidizing Bacteria (AOB). Our results show the effect of the influent total ammonium nitrogen (TAN) concentration on AOB and NOB activity in two moving bed biofilm reactors (MBBR) operated as Sequencing Batch Reactors (SBR) at 15 °C (SBR I) and 21 °C (SBR II). The nitritation reactors were not successfully optimized for the given conditions (low temperature, low TAN) yet. We achieved stable nitritation with influent concentration of 150 mg TAN/L (at 15 °C) and 300 mg TAN/L (at 22 °C), which are concentrations significantly higher than those found in municipal wastewater (50 mg/L). However, it was shown that by decreasing Dissolved Oxygen (DO) concentration to below 1 mg/L. This is the research direction that is being further investigated.

The above mentioned reactors were operated each for more than 1 year in order to describe the long-term changes in their performance. The operation of MBBRs was complemented with operation of rectors containing suspended biomass under conditions identical with the MBBR. Interestingly, the results were very similar.

b. Operation of reactors for denitritation and Anaerobic Ammonia Oxidation (ANOMMOX)

An alternative to the nitritation-denitritation process is Anammox process. Therefore, a 1-step Anammox biofilm reactor has been set-up in order to treat the effluent from the psychrophilic UASB. This reactor has been operated for more than 100 days. So far, it achieved 95% removal of Total Ammonium Nitrogen (TAN) and 75% removal of total nitrogen. Currently, the reactor is still operated in order to gradually increase its efficiency and nitrogen loading rate.

(2) Characterization of the biofilms developed in nitritation reactors

a. Characterisation of the development of microbial population in the denitritation biofilm using Fluorescence In Situ Hibridization (FISH).

FISH analysis was performed in order to determine the presence and relative amount of NOB and AOB in the biofilm grown in the reactors. The results show significant AOB prevalence in biofilm at the outer part of biofilm with most of NOB growing in the inner parts of biofilm. AOB growth was favoured on the edges of biofilm because of the lower affinity constant for oxygen than that for NOB and high initial FA content in medium. Stratification of bacteria allowed for proliferation of NOB inside the biofilm and subsequent cooperation of AOB and NOB resulting in complete nitrification and resistance to NOB inhibiting conditions in the medium.

b. Application of Magnetic Resonance Imaging (MRI) for characterization of biofilm

This research was so far focused on anaerobic granular biofilm and on the transport processes in this biofilm. Also, it was concern with the methodology. Applying MRI, we were able to relate the transport pattern with the bioavailability of trace nutrients in anaerobic biofilm.

(3) Development and operation of a complex system for COD and nitrogen removal from municipal wastewater

a. Operation of Anaerobic Up-Flow Sludge Blanket (UASB) reactor in combination with digester for solids removal at low temperature

A 2L psychrophilic UASB-Digester system has been set-up and operated so far for 150 days. It achieved about 70% removal of suspended organics (as Chemical Oxygen Demand – COD), but only negligible removal of soluble COD was observed. However, soluble COD present in the effluent was successfully utilized for denitritation.

In conclusion, the UASB reactor has proven to be only functioning as separation step for suspended COD. Therefore, currently we are developing technology for enhanced solids removal in the UASB reactor using bioflocculation process.

b. Denitritation in the UASB effluent

Firstly, batch experiments for denitritation were repeatedly performed and continuously operated reactors start in these days. It was shown that the organic material contained in the effluent of the psychrophilic UASB reactor treating municipal wastewater is available as substrate for denitritation and can supply enough electrons for reduction of at least 80% of nitrite that can be present in typical municipal wastewater after nitritation (up to 50 mg/L). The denitritation period has been recently implemented into the SBR cycle of the nitritation reactor in order to demonstrate these results in continuous operation.

All the important information about the project is available at