In Ethiopia industrial effluents containing high contents of organic matter, nitrogen and heavy metals are discharged into inland surface waters with little or no pre-treatment. Significant pollution concerns related to these effluents include dissolved oxygen depletion, toxicity and eutrophication of the receiving waters. This has not only forced the government to formulate regulations and standards for discharge limits but also resulted in an increasing interest and development of methods and systems by which wastewater can be recycled and used sustainably. The need for technologies for environmentally friendly treatment of industrial wastes such as tannery wastewaters is therefore obvious. Biological processes are not only cost effective but also environmentally sound alternatives to the chemical treatment of tannery wastewaters. The aim of the research presented in this thesis was to develop and optimize processes for biological nitrogen removal from tannery wastewaters and to identify the most efficient denitrifying organisms in tannery wastewaters laden with toxic substances. A pilot plant consisting of a predenitrification anoxic system, aerated nitrification compartment and a sedimentation tank (clarifier) all arranged in series was developed and installed on the premises of Addis Ababa University, Ethiopia. In spite of high influent chromium and sulphide perturbations over the successive feeding phases, the performance of the pilot plant was encouraging. The overall removal efficiency of the pilot plant over the experimental feeding phases varied between 82-98% for total nitrogen, 95-98% for COD, 96-98% for BOD5, 46-95% for ammonia nitrogen, 95-99% for sulphide and 93-99% for trivalent Chromium. Six isolates from over 1000 pure cultures were identified as the most efficient denitrifying bacteria. From both cellular fatty acid profiles and 16S rRNA gene sequencing, the six selected strains were phylogenetically identified as Brachymonas denitrificans in the β-subdivision of the Proteobacteria. All the six strains contain cd1-type nitrite reductase. The efficient isolates characterized in this study are of great value because of their excellent denitrifying properties and high tolerance to the concentrations of toxic compounds prevailing in tannery wastewaters. Bio-augmentation of the pilot plant with this bacterium showed a clear correlation between in situ denitrifying activities measured by nitrate uptake rate, population dynamics of the introduced B.denitrificans monitored by fluorescent in situ hybridization and the pilot plant performance, suggesting that the strategy of introducing this species for enhancing process performance has potential applications. Moreover, the nitrate-reducing, sulphur-oxidizing bacteria (NR-SOB) were also found in the pilot plant in abundance with steady sulphide removal efficiency during the study period. This could provide opportunities for the application of biologically mediated simultaneous removal of sulphide and nitrogen from tannery effluents. In addition to enriching high consortia of denitrifiers in the anoxic system to attain high denitrification efficiency and also improving the overall nitrification efficiency of the system, the predenitrification-nitrification pilot process plant stimulated the activity of indigenous NR-SOB to simultaneously remove sulphide from the system. Thus, the pilot plant was found to be operationally efficient for the removal of nitrogen, organic matter and other pollutants from tannery wastewaters. Keywords: Biological nitrogen and sulphide removal, denitrifying bacteria, nitrate-reducing, sulphur-oxidizing bacteria, nitrate uptake rate, fluorescent in situ hybridization, pollution, tannery effluents.

<p>In Ethiopia industrial effluents containing high contents of organic matter, nitrogen and heavy metals are discharged into inland surface waters with little or no pre-treatment. Significant pollution concerns related to these effluents include dissolved oxygen depletion, toxicity and eutrophication of the receiving waters. This has not only forced the government to formulate regulations and standards for discharge limits but also resulted in an increasing interest and development of methods and systems by which wastewater can be recycled and used sustainably. The need for technologies for environmentally friendly treatment of industrial wastes such as tannery wastewaters is therefore obvious. Biological processes are not only cost effective but also environmentally sound alternatives to the chemical treatment of tannery wastewaters.</p><p>The aim of the research presented in this thesis was to develop and optimize processes for biological nitrogen removal from tannery wastewaters and to identify the most efficient denitrifying organisms in tannery wastewaters laden with toxic substances. A pilot plant consisting of a predenitrification anoxic system, aerated nitrification compartment and a sedimentation tank (clarifier) all arranged in series was developed and installed on the premises of Addis Ababa University, Ethiopia. In spite of high influent chromium and sulphide perturbations over the successive feeding phases, the performance of the pilot plant was encouraging. The overall removal efficiency of the pilot plant over the experimental feeding phases varied between 82-98% for total nitrogen, 95-98% for COD, 96-98% for BOD5, 46-95% for ammonia nitrogen, 95-99% for sulphide and 93-99% for trivalent Chromium. Six isolates from over 1000 pure cultures were identified as the most efficient denitrifying bacteria. From both cellular fatty acid profiles and 16S rRNA gene sequencing, the six selected strains were phylogenetically identified as Brachymonas denitrificans in the β-subdivision of the Proteobacteria. All the six strains contain cd1-type nitrite reductase. The efficient isolates characterized in this study are of great value because of their excellent denitrifying properties and high tolerance to the concentrations of toxic compounds prevailing in tannery wastewaters. Bio-augmentation of the pilot plant with this bacterium showed a clear correlation between in situ denitrifying activities measured by nitrate uptake rate, population dynamics of the introduced B.denitrificans monitored by fluorescent in situ hybridization and the pilot plant performance, suggesting that the strategy of introducing this species for enhancing process performance has potential applications.</p><p>Moreover, the nitrate-reducing, sulphur-oxidizing bacteria (NR-SOB) were also found in the pilot plant in abundance with steady sulphide removal efficiency during the study period. This could provide opportunities for the application of biologically mediated simultaneous removal of sulphide and nitrogen from tannery effluents. In addition to enriching high consortia of denitrifiers in the anoxic system to attain high denitrification efficiency and also improving the overall nitrification efficiency of the system, the predenitrification-nitrification pilot process plant stimulated the activity of indigenous NR-SOB to simultaneously remove sulphide from the system. Thus, the pilot plant was found to be operationally efficient for the removal of nitrogen, organic matter and other pollutants from tannery wastewaters.</p><p><b>Keywords:</b> Biological nitrogen and sulphide removal, denitrifying bacteria, nitrate-reducing, sulphur-oxidizing bacteria, nitrate uptake rate, fluorescent in situ hybridization, pollution, tannery effluents.</p>

This master thesis is based on own studies. A four-month study was performed at Hammarby Sjostad Research Station, which is located in Stockholm. One-stage deammonification process was evaluated in two different system configurations in pilot plant scale. The theoretical background for this thesis works is presented in the first part and where is presented negative impacts of nitrogen compounds in environment and requirements for purified wastewater in European Union (Sweden and Poland). In the next part of the thesis the nitrogen cycle is described and with focus on biological reactions for nitrogen removal. Especially, nitrification/denitrification and anammox processes are described with special focus on parameters affecting the anammox process performance and its advantages and disadvantages of using this process. Experimental results from the four-month study and evaluation of the microbial activity are described in the last part.

Partial Nitritation/Anammox process (deammonification process), by which occurs oxidation of ammonium to nitrogen gas by autotrophic bacteria in anaerobic conditions, considered to be cost-effective and environmentally friendly method of nitrogen removal. Present research work focuses on achieving a high nitrogen removal degree, thanks to Anammox bacteria, while providing the best performance of the ongoing process. Integrated fixed-film activated sludge (IFAS) reactor was supplied with the main stream of the wastewater after UASB reactor, characterized by low concentration of nitrogen and organic matter. The bacteria ability to accommodate, were tested in the biofilm and in the activated sludge, depending on the different stages in which the process were being conducted. Batch test, such as Specific Anammox Activity (SAA), Nitrate Uptake Rate (NUR) and Oxygen Uptake Rate (OUR), were used for the evaluation of activity of various groups of bacteria. On the basis of laboratory analysis verified the values obtained from the batch tests. It was determined that a high degree of nitrogen removal (92% of NH4-N) was achieved thanks to the dominant activity of the Anammox bacteria, with low participation of other groups of bacteria. It was also proved, that Anammox bacteria activity were overwhelming in the biofilm. Dominant role of Ammonium Oxidizing Bacteria (AOB) was associated with high activity of Anammox bacteria, which together satisfyingly out-competed Nitrite Oxidizing Bacteria (NOB) and heterotrophic bacteria. It has been shown that Anammox bacteria quickly adapt to the new conditions and they are able to assume a dominant role, even in the case of inoculation of the reactor with the sludge from SBR. This allows conclude, that in the case of operational problems, the reactor can be supplied from another source, in order not to inhibit the process.