MICROBIAL AND BIOMOLECULAR HETEROGENEITY OF AEROBIC GRANULES WITHIN A SINGLE BIOREACTOR ?

Aerobic granulation is considered as a promising alternative to conventional activated sludge in biological wastewater treatment. Instead of forming flocs the bacterial communities aggregate to dense granular biofilms. In the present study, granules were cultivated in bubble column reactors operated in sequencing batch mode. The operating cycle consisted of 4 phases: i) an anaerobic feeding phase, ii) a long aeration phase where the sludge was mixed, iii) a short settling phase and finally iv) a withdrawal phase where the treated water was taken out of the reactor. Feeding with synthetic wastewater was done from the bottom of the reactor in plug-flow mode without a mixing of the settled sludge. We made two observations which let us conclude that all the granules were not equally provided with substrate: 1) We measured the carbon consumption rate and calculated that only the lower 15-25% of the sludge bed got carbon in each cycle. If all granules would have had the same settling properties, this heterogeneity would have been cancelled over time. But 2) we observed a clear gradient in granule size within the sludge bed. Larger granules settled faster than smaller ones. Sludge from the bottom of the reactor was composed up to 80% by granules with diameter larger than 3.5 mm, whereas over the whole reactor the threshold diameter for 80% of the sludge volume was only at 2.4 mm. The heterogeneity in carbon accessibility is expected to influence the composition of bacterial communities and the wastewater treatment performances. For example, dephosphatation and denitrification are directly depending on the presence of bacteria containing internal carbon reserve in form of polyhydroxyalkanoates. We are presently investigating possible differences between “feast” and “famine” granules.

Elimination of micropollutants in wastewater treatment plants: Ozonation or activated carbon? Conclusions of a one-year pilot project

David Andrew Barry, Nathalie Chèvre Rossi, Luiz Felippe De Alencastro, Anoys Magnet, Jonas Margot, Luca Rossi

Many organic micropollutants present in wastewater, such as pharmaceuticals and pesticides, are poorly removed in conventional wastewater treatment plants (WWTP). To reduce the release of these substances into the aquatic environment, advanced treatments are necessary. Two advanced treatments were tested in pilot systems over more than one year at the municipal WWTP of Lausanne, Switzerland. The first pilot involves the ozonation of the effluent to oxidize organic substances. It is followed by sand filtration (SF) to remove the readily biodegradable reaction products. The second pilot consists of a powdered activated carbon (PAC) addition into the effluent to sorb micropollutants. It is followed either by a membrane filtration (ultrafiltration) or a sand filtration to separate the PAC from the treated water. A selection of 58 potentially problematic substances (pharmaceuticals, pesticides, endocrine disruptors) were regularly measured at different stages of treatment by ultra performance liquid chromatography coupled to a tandem mass spectrometer, and a large battery of ecotoxicological tests (16 in vitro and 9 in vivo assays with different organisms) were performed to evaluate the toxicity of the effluents. The results showed that most compounds were removed over 80% with an average ozone dose of about 5.5 mgO3/l or a PAC dose between 10 and 20 mg/l. Only some single compounds, such as X-ray contrast media, were only partially eliminated in both cases. A clear reduction in toxicity was also observed after the two treatments in most of the cases. These two processes (ozonation and PAC addition) are therefore effective to reduce the release of micropollutants into surface waters. Ozonation-SF and PAC-SF proved to be feasible in terms of implementation and operation at large-scale in WWTP, for relatively similar investment and operation costs (about 0.1 to 0.15 € per m3 treated based on local costs). Each technique has its advantages and disadvantages. Therefore the method selection should be made case by case for each WWTP depending on the local constraints (e.g. space, security, energy costs, existing treatment processes, sludge disposal process, need for disinfection, etc.), the quality and the quantity of incoming water and the desired output water quality

2011

Bacterial Resource Management for Nutrient Removal in Aerobic Granular Sludge Wastewater Treatment Systems

David Gregory Weissbrodt

The approach of wastewater treatment has shifted towards a holistic view in order to achieve sustainability in addition to environmental protection. The aerobic granular sludge (AGS) technology, which relies on the use of fast-settling granular biofilms called granules, is progressively becoming a new standard for high-rate biological nutrient removal (BNR) and secondary clarification in single sequencing batch reactors (SBR). The AGS intensive process has been related with definite savings in land area, construction, and operation costs. In an economical analysis, theoretical savings of CHF 0.45 per m3 (1 Swiss franc CHF ≈ 0.83 €) were computed for a Swiss wastewater treatment plant (WWTP) of 200’000 capita removing all nutrients biologically. Besides scale-up, fundamental research was required to understand and tailor the structure of underlying bacterial communities that form granules and remove nutrients. Mechanisms of bacterial selection were investigated in a systems approach to propose strategic axes for optimal management of the bacterial resource for efficient process performances. This research led to the following advances. After having designed a flexible reactor infrastructure for AGS research, a mathematical modeling methodology was developed to understand the hydraulic and biological processes involved during plug-flow transport of wastewater across the settled AGS bed during the feeding phase. BNR relies on the preferential selection of polyphosphate-accumulating organisms (PAO) by proper uncoupled supply of electron donors and electron acceptors. A bed height to diameter ratio of 6 was deduced to be optimal for full bed loading and efficient anaerobic acetate uptake under reference simulation conditions (20°C, pH 7.0). A bioinformatics methodology called PyroTRF-ID was developed for the identification of bacterial relatives involved in BNR processes by combination of terminal-restriction fragment length polymorphism (T-RFLP) and pyrosequencing data. This procedure provided high resolution of bacterial community structures and dynamics of AGS systems. Electrical conductivity and polyphosphatase assays were proposed for rapid and low-cost assessment of the fractions of active PAO and of the dephosphatating potential of activated sludge and granular sludge. Positive linear correlations were obtained between the fraction of PAO, the biomass specific rate of conductivity evolution measured in anaerobic metabolic batch tests, and the polyphosphate-hydrolyzing enzymatic activity of cell extracts. Wash-out conditions used to stimulate granule formation were shown to exert a selection pressure not only on physical properties of early-stage granules, but also on the underlying bacterial community composition. Operation with constant volumetric organic loading rates (OLR) during wash-out resulted in the formation of slow-settling fluffy granules dominated by filamentous Burkholderiales affiliates formed under low aeration (< 2 cm s-1) and high mesophilic temperature (30°C), and of fast-settling dense granules dominated by Zoogloea spp. under high aeration (4 cm s-1) and with an inoculum originating from a BNR-WWTP. These exopolysaccharide producers were considered as model organisms for granulation. However, their proliferation over PAO and nitrifiers correlated with poor BNR. PAO estab-lished at mature stage as soon as sufficient AGS was present to fully remove volatile fatty acids (VFA) during the anaerobic feeding phase. Without purge of excess sludge, glycogen-accumulating organisms (GAO) competed with PAO at a later stage of reactor operation. Granulation was shown to occur in stirred-tank SBRs operated under steady-state condi¬tions to cultivate PAO and GAO enrichments. The Accumulibacter- and Competibacter-dominated communities displayed granulation potential without involvement of Zoogloea spp. Fast-settling nuclei formed despite initial operation with a settling time as high as 60 min, and evolved towards granules after decrease of settling time to 10 min. Dynamic control of the OLR, anaerobic contact time, and sludge retention time (SRT) selected for active PAO in early-stage granules. Confocal laser scanning microscopy (CLSM) revealed that granula¬tion mechanisms depend on the predominant organisms involved. Fast-growing Zoogloea spp. formed smooth biofilm continuum matrices. The slower-growing PAO and GAO formed het¬erogeneous granular structures by proliferation in compact colonies around flocs. Fluctuations in operation variables were shown by multivariate statistics to impact on process performance and bacterial community structures in anaerobic-aerobic AGS-SBRs. The size of granules impacted on nitrification and dephosphatation by affecting oxygen mass transfer. Efficient BNR was obtained with at least 500 mgCOD L-1of acetate, 30 gCOD gP-1, and 10 gCOD gN-1. Although clades of PAO can denitrify, nitrogen removal was thus not restricted only to PAO. A broad denitrifying community was identified. The bacterial community continuum comprised two major opponent clusters, composed of members of the core microbiome of full-scale BNR-WWTPs. The first mainly comprised Accumulibacter, Nitrospira, Xanthomonadaceae, and Aminobacter affiliates selected by conditions of efficient BNR. The second mainly comprised Competibacter, Sphingobacteriales, Cytophaga, and Tetrasphaera affiliates correlating with periods of low BNR. Bacterial selection in AGS systems was mainly impacted by pH, according to multifactorial experiments. Accumulibacter and BNR were favored with alkaline pH, low mesophilic temperatures, and in the presence of propionate. Competibacter proliferated with acidic pH, temperature close to 30°C, and only acetate. Tetrasphaera spp., potential major PAO of certain full-scale plants, withstood Competibacter-selective conditions. Enhanced BNR was obtained under non-limiting conditions with food-to-microorganism ratios above 25 mgCODs gCODx-1. With fixed 2-h starvation, dephosphatation was only complete with full aeration. Since alternating aerobic-anoxic starvation conditions led to partial conversions, the length of starvation phases should be controlled in function of the redox conditions applied. Overall, optimal bacterial resource management in AGS systems requires full control of system behavior by taking advantage of the flexibility of the SBR technology. A methodology is proposed to this end, comprising milestones for wastewater characterization, SBR design, “anaerobic selector” design, control of the starvation phase length, shaving of fluctuations in operation variables, purge of excess sludge, proper selection for a stable, active and cooperative bacterial community, as well as efficient BNR. As key research perspective, investigations with real wastewater should validate the knowledge gained on bacterial behaviors under low-complexity conditions. Significant differences in predominant bacterial relatives were detected between lab-scale and full-scale BNR sludge. Metagenomics will provide a broader view on key organisms and functions of BNR and AGS microbiomes.

EPFL2013

Wastewater treatment processes modelling for anaerobic digestion effluents

Miriel Steudler

The general topic of this research project is the modelling of wastewater treatment processes for anaerobic digestion effluents, which are characterized by high nitrogen concentrations. In order to avoid eutrophication of lakes and rivers caused by nutrient enrichment as it happened in Switzerland a few decades ago, the implementation of efficient nitrogen removal processes for those effluents remains nowadays of importance. Moreover nitrogen removal will even become a pre-requisite for efficient micropollutants techniques to be implemented. The first objective of this master project is the modelling of the biological nitrogen removal process Anammox in the software gPROMS. This process converts ammonium and nitrate (close to 1:1 influent concentration ratio) into dinitrogen and nitrate by Anammox bacteria. In order to obtain an influent with the proper concentration ratio, this process can be coupled to a Sharon reactor operated for partial nitrification (two stages operation). In this study, the Anammox biological developed model has been integrated into a continuous stirred tank reactor (CSTR) combined with a particles separator. The two modeled reactors, Sharon and Anammox, have then been run in series and steady-state results have been obtained. Through a comparison with literature results (lab experience and pilot-plant), the steady-state simulation results in terms of nitrogen removal efficiency and effluent concentrations have been considered as reliable. However, a calibration of the model with real data still needs to be investigated. The second objective of the project is the study of key influencing parameters (pH and temperature) on the operation of the two stages Sharon-Anammox process. A sensitivity analysis showed that temperature and pH are influencing the nitrite to ammonium ratio Sharon effluents and consequently affecting the efficiency of the Anammox reactor; the growth and selection of bacteria in the Sharon reactor are determined by those two parameters (chemostat principle). The optimal range of operation for the Sharon reactor in order to maximize the efficiency of the Anammox reactor are found to be between 306K and 308K and between 7.8 and 8.0 for the pH according to the steady-state simulations; those results are coherent with literature findings and operation of full-scale Sharon reactors. The final objective of the project is the comparison of the Sharon-Anammox process with an activated sludge system in terms of energy consumption and nitrogen removal efficiency. By minimizing in both cases the mmonium effluent concentration and playing with the aeration needs, savings of 75% in benefit of the Sharon-Anammox system have been observed; the order of magnitude obtained by optimization is coherent with the stoichiometry implemented in the model. The biological Anammox model itself will be implemented in other types of reactors models (one stage technologies involving biofilms) and the models predictions will be compared with operational data (at pilot or full-scale). Modelling wastewater treatment processes is a help to better understand the processes themselves, their interactions, their optimal operation conditions both by taking into consideration the energy consumption and the water quality through optimization techniques. The challenge of modelling will however still remain the modelling of bacteria interaction and inhibition, as well as a deep understanding and quantification of various mass transfer processes involved. This will need a close collaboration with expert scientists in order to better understand these phenomena. Modelling is a good support for experimental studies, as it can help to plan efficient experimental campaign by minimizing the required experimentations run.

2014