Phylogeny and physiology of bacteria important for floc structure and settling properties in activated sludge

Research output: PhD thesis

Abstract

The activated sludge process is the most common all over the world for treatment of domestic and industrial wastewaters. The success of the activated sludge process relies on the formation of good flocs to obtain a high effluent quality and a good dewaterability of the sludge. However, problems exist in wastewater treatment plants (WWTP), like deflocculation of the activated sludge flocs, which results in settling and dewatering problems. Another problem is the abundance of filamentous bacteria, which can result in bulking sludge or foaming. Therefore, it is of high importance to obtain knowledge about phylogeny and physiology of bacteria important for floc structure and settling properties in activated sludge, which possibly in the long term can optimise flocculation, settling and dewatering. The knowledge of floc-forming bacteria is very limited, while more filamentous bacteria are known, however, not those involved in floc-formation. The purpose of this Ph.D. study has been to investigate the identity, function and floc-forming properties of selected microcolony-forming bacteria in WWTP, and study the identity and function of important filamentous bacteria. Floc-strenght of different probe-defined populations was investigated by shear treatment combined with fluorescence in situ hybridization (FISH), and a large difference in the strength and colloid-chemical properties was found. It was shown that Betaproteobacteria formed the strongest microcolonies, and the strength seemed mainly related to the charge of the extracellular polymeric substances, and maybe entanglements of exopolymers, but not hydrophobicity. Alphaproteobacteria were weak microcolony-formers. Hitherto unknown bacteria related to the Aquaspirillum genus in the Betaproteobacteria were identified, and a specific oligonucleotide probe was designed after micromanipulation of FISH-labelled microcolonies and subsequently applying the 16S rRNA approach. These bacteria were shown to be very abundant and dominant floc-formers capable to denitrify only utilizing few amino acids of the substrates tested, indicating an important role in protein degradation in the WWTP. Thauera and Azoarcus, also belonging to the Betaproteobacteria and abundant denitrifying microcolony-formers in most treatment plants, were also studied, and were more diverse in their substrate utilities. Together these 3 groups of bacteria often constitute more than 40-50% of the biomass in WWTP with biological nitrogen and phosphorus removal. Surface properties and storage capabilities of the Aquaspirillum-related bacteria, Thauera, and Azoarcus were investigated, and they all had hydrophobic surfaces, possibly characteristic for Betaproteobacteria. Neither of them stored high amounts of polyphosphate under the in situ conditions tested, whereas the Aquaspirillum-related bacteria and Thauera were able to convert carbon sources into poly--hydroxyalkanoate granules. Furthermore, it was shown, that the potential for accumulating poly--hydroxyalkanoate granules was much higher for the Aquaspirillum-related bacteria and Thauera than observed under in situ conditions. Different exoenzymatic activities were tested, and esterase activity was observed by Thauera and Azoarcus, which correlated well with their ability to use fatty acids. Due to the dominance of these denitrifying Betaproteobacteria, they are of high importance for the floc-formation and properties. Some filamentous bacteria were also studied. Morphological identification is not suitable for their identification, and new oligonucleotide probes were developed. A filamentous bacterium often with epiphytic growth was targeted by the probe for Aquaspirillum-related bacteria. This is the first identified filamentous bacterium capable to perform full denitrification. Furthermore, Meganema perideroedes gen. nov., sp. nov., a new filamentous Alphaproteobacterium from activated sludge was identified and characterized in detail. Eikelbooms Type 0041 of which some belong to the TM7 phylum and its characteristic epiphytic growth was studied by MAR-FISH and found to be very versatile in substrate uptake pattern, which explain its abundance in many treatment plants. The results in this Ph.D. have increased the knowledge on phylogeny and physiology of dominant floc-forming bacteria and filamentous bacteria. A large fraction of denitrifyers from especially WWTP with biological nitrogen and phosphorus removal are hereby identified and described from a physiologically point of view. In addition our knowledge on filamentous bacteria with epiflora has been increased significantly. Future studies will resolve why the filamentous bacteria targeted by the new oligonucleotide probe for Aquaspirillum-related bacteria are able to utilize substrate under anaerobic conditions. If different subpopulations of Thauera and Azoarcus are responsible for different substrate utilizations need to be investigated, and furthermore it will be interesting to see if it is possible to selectively enrich for wanted or unwanted bacteria fractions in sludge using the shear experiments. This can hopefully contribute to increased knowledge on optimization of biological processes, flocculation, settling and dewatering in WWTP.
Original languageDanish
Place of PublicationUNIPRINT, Aalborg, Denmark
Publisher
Print ISBNs8790033361
Electronic ISBNs0909-6159
Publication statusPublished - 2004

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