Eberhard Morgenroth

Combined Nitritation–Anammox: Advances in Understanding Process Stability

Efficient nitrogen removal from wastewater containing high concentrations of ammonium but little organic substrate has recently been demonstrated by several full-scale applications of the combined nitritation-anammox process. While the process efficiency is in most cases very good, process instabilities have been observed to result in temporary process failures. In the current study, conditions resulting in instability and strategies to regain efficient operation were evaluated. First, data from full-scale operation is presented, showing a sudden partial loss of activity followed by recovery within less than 1 month. Results from laboratory-scale experiments indicate that these dynamics observed in full scale can be caused by partial inhibition of the ammonia oxidizing bacteria (AOB), while anammox inhibition is a secondary effect due to temporarily reduced O(2) depletion. Complete anammox inhibition is observed at 0.2 mg O(2) · L(-1), resulting in NO(2)(-) accumulation. However, this inhibition of anammox is reversible within minutes after O(2) depletion. Thus, variable AOB activity was identified as the key to reactor stability. With appropriate interpretation of the online NH(4)(+) signal, accumulation of NO(2)(-) can be detected indirectly and used to signal an imbalance of O(2) supply and AOB activity (no suitable online NO(2)(-) electrode is currently available). Second, increased abundance of nitrite-oxidizing bacteria (NOB; competing with anammox for NO(2)(-)) is known as another cause of instability. Based on a comparison of parallel full-scale reactors, it is suggested that an infrequent and short-term increased O(2) supply (e.g., for maintenance of aerators) that exceeds prompt depletion of oxygen by AOB may have caused increased NOB abundance. The volumetric air supply as a proxy for O(2) supply thus needs to be linked to AOB activity. Further, NOB can be washed out of the system during regular operation if the system is operated at a sludge age in the range of 45 days and by controlling the air supply according to the NO(3)(-) concentration in the treated effluent. Early detection of growing NOB abundance while the population is still low can help guide process operation and it is suggested that molecular methods of quantifying NOB abundance should be tested.

Mainstream partial nitritation and anammox: long-term process stability and effluent quality at low temperatures

The implementation of autotrophic anaerobic ammonium oxidation processes for the removal of nitrogen from municipal wastewater (known as “mainstream anammox”) bears the potential to bring wastewater treatment plants close to energy autarky. The aim of the present work was to assess the long-term stability of partial nitritation/anammox (PN/A) processes operating at low temperatures and their reliability in meeting nitrogen concentrations in the range of typical discharge limits below 2 mgNH4-N·L−1 and 10 mgNtot·L−1. Two main 12-L sequencing batch reactors were operated in parallel for PN/A on aerobically pre-treated municipal wastewater (21 ± 5 mgNH4-N·L−1 and residual 69 ± 19 mgCODtot·L−1) for more than one year, including over 5 months at 15 °C. The two systems consisted of a moving bed biofilm reactor (MBBR) and a hybrid MBBR (H-MBBR) with flocculent biomass. Operation at limiting oxygen concentrations (0.15–0.18 mgO2·L−1) allowed stable suppression of the activity of nitrite-oxidizing bacteria at 15 °C with a production of nitrate over ammonium consumed as low as 16% in the MBBR. Promising nitrogen removal rates of 20–40 mgN·L−1·d−1 were maintained at hydraulic retention times of 14 h. Stable ammonium and total nitrogen removal efficiencies over 90% and 70% respectively were achieved. Both reactors reached average concentrations of total nitrogen below 10 mgN·L−1 in their effluents, even down to 6 mgN·L−1 for the MBBR, with an ammonium concentration of 2 mgN·L−1 (set as operational threshold to stop aeration). Furthermore, the two PN/A systems performed almost identically with respect to the biological removal of organic micropollutants and, importantly, to a similar extent as conventional treatments. A sudden temperature drop to 11 °C resulted in significant suppression of anammox activity, although this was rapidly recovered after the temperature was increased back to 15 °C. Analyses of 16S rRNA gene-targeted amplicon sequencing revealed that the anammox guild of the bacterial communities of the two systems was composed of the genus “Candidatus Brocadia”. The potential of PN/A systems to compete with conventional treatments for biological nutrients removal both in terms of removal rates and overall effluent quality was proven.

Predation influences the structure of biofilm developed on ultrafiltration membranes

This study investigates the impact of predation by eukaryotes on the development of specific biofilm structures in gravity-driven dead-end ultrafiltration systems. Filtration systems were operated under ultra-low pressure conditions (65 mbar) without the control of biofilm formation. Three different levels of predation were evaluated: (1) inhibition of eukaryotic organisms, (2) addition of cultured protozoa (Tetrahymena pyriformis), and (3) no modification of microbial community as a control. The system performance was evaluated based on permeate flux and structures of the biofilm. It was found that predation had a significant influence on both the total amount and also the structure of the biofilm. An open and heterogeneous structure developed in systems with predation whereas a flat, compact, and thick structure that homogeneously covered the membrane surface developed in absence of predation. Permeate flux was correlated with the structure of the biofilm with increased fluxes for smaller membrane coverage. Permeate fluxes in the presence or absence of the predators was 10 and 5 L m(-2) h(-1), respectively. It was concluded that eukaryotic predation is a key factor influencing the performance of gravity-driven ultrafiltration systems.

Activity of metazoa governs biofilm structure formation and enhances permeate flux during Gravity-Driven Membrane (GDM) filtration.

The impact of different feed waters in terms of eukaryotic populations and organic carbon content on the biofilm structure formation and permeate flux during Gravity-Driven Membrane (GDM) filtration was investigated in this study. GDM filtration was performed at ultra-low pressure (65 mbar) in dead-end mode without control of the biofilm formation. Different feed waters were tested (River water, pre-treated river water, lake water, and tap water) and varied with regard to their organic substrate content and their predator community. River water was manipulated either by chemically inhibiting all eukaryotes or by filtering out macrozoobenthos (metazoan organisms). The structure of the biofilm was characterized at the meso- and micro-scale using Optical Coherence Tomography (OCT) and Confocal Laser Scanning Microscopy (CLSM), respectively. Based on Total Organic Carbon (TOC) measurements, the river waters provided the highest potential for bacterial growth whereas tap water had the lowest. An increasing content in soluble and particulate organic substrate resulted in increasing biofilm accumulation on membrane surface. However, enhanced biofilm accumulation did not result in lower flux values and permeate flux was mainly influenced by the structure of the biofilm. Metazoan organisms (in particular nematodes and oligochaetes) built-up protective habitats, which resulted in the formation of open and spatially heterogeneous biofilms composed of biomass patches. In the absence of predation by metazoan organisms, a flat and compact biofilm developed. It is concluded that the activity of metazoan organisms in natural river water and its impact on biofilm structure balances the detrimental effect of a high biofilm accumulation, thus allowing for a broader application of GDM filtration. Finally, our results suggest that for surface waters with high particulate organic carbon (POC) content, the use of worms is suitable to enhance POC removal before ultrafiltration units.

Sulfidation Kinetics of Silver Nanoparticles Reacted with Metal Sulfides

Recent studies have documented that the sulfidation of silver nanoparticles (Ag-NP), possibly released to the environment from consumer products, occurs in anoxic zones of urban wastewater systems and that sulfidized Ag-NP exhibit dramatically reduced toxic effects. However, whether Ag-NP sulfidation also occurs under oxic conditions in the absence of bisulfide has not been addressed, yet. In this study we, therefore, investigated whether metal sulfides that are more resistant toward oxidation than free sulfide, could enable the sulfidation of Ag-NP under oxic conditions. We reacted citrate-stabilized Ag-NP of different sizes (10-100 nm) with freshly precipitated and crystalline CuS and ZnS in oxygenated aqueous suspensions at pH 7.5. The extent of Ag-NP sulfidation was derived from the increase in dissolved Cu(2+) or Zn(2+) over time and linked with results from X-ray absorption spectroscopy (XAS) analysis of selected samples. The sulfidation of Ag-NP followed pseudo first-order kinetics, with rate coefficients increasing with decreasing Ag-NP diameter and increasing metal sulfide concentration and depending on the type (CuS and ZnS) and crystallinity of the reacting metal sulfide. Results from analytical electron microscopy revealed the formation of complex sulfidation patterns that seemed to follow preexisting subgrain boundaries in the pristine Ag-NP. The kinetics of Ag-NP sulfidation observed in this study in combination with reported ZnS and CuS concentrations and predicted Ag-NP concentrations in wastewater and urban surface waters indicate that even under oxic conditions and in the absence of free sulfide, Ag-NP can be transformed into Ag2S within a few hours to days by reaction with metal sulfides.

Influence of shear on the production of extracellular polymeric substances in membrane bioreactors

Shear is used to control fouling in membrane bioreactor (MBR) systems. However, shear also influences the physicochemical and biological properties of MBR biomass. The current study examines the relationship between the level of shear and extracellular polymeric substance (EPS) production in MBRs. Two identical MBRs were operated in parallel where the biomass in one reactor was exposed to seven times greater shear forces. The concentrations of floc-associated and soluble EPS were monitored for the duration of the experiment. The stickiness of extracted floc-associated EPS from each reactor was also characterized using atomic force microscopy. A mathematical model of floc-associated and soluble EPS production was applied to quantitatively describe changes in EPS production with shear. Biomass grown in a high shear environment has lower floc-associated EPS production compared to biomass grown in a lower shear environment. This decrease in floc-associated EPS production also corresponds to a decrease in soluble EPS production, which can be explained by both the lower concentration of floc-associated EPS and the production of stickier floc-associated EPS that is more erosion resistant in the high shear reactor. This research suggests that mechanical stresses can have a significant impact on the production rates of floc-associated and soluble EPS-key parameters governing membrane fouling in MBRs.

Activity and growth of anammox biomass on aerobically pre-treated municipal wastewater.

Direct treatment of municipal wastewater (MWW) based on anaerobic ammonium oxidizing (anammox) bacteria holds promise to turn the energy balance of wastewater treatment neutral or even positive. Currently, anammox processes are successfully implemented at full scale for the treatment of high-strength wastewaters, whereas the possibility of their mainstream application still needs to be confirmed. In this study, the growth of anammox organisms on aerobically pre-treated municipal wastewater (MWWpre-treated), amended with nitrite, was proven in three parallel reactors. The reactors were operated at total N concentrations in the range 5–20 mgN∙L−1, as expected for MWW. Anammox activities up to 465 mgN∙L−1∙d−1 were reached at 29 °C, with minimum doubling times of 18 d. Lowering the temperature to 12.5 °C resulted in a marked decrease in activity to 46 mgN∙L−1∙d−1 (79 days doubling time), still in a reasonable range for autotrophic nitrogen removal from MWW. During the experiment, the biomass evolved from a suspended growth inoculum to a hybrid system with suspended flocs and wall-attached biofilm. At the same time, MWWpre-treated had a direct impact on process performance. Changing the influent from synthetic medium to MWWpre-treated resulted in a two-month delay in net anammox growth and a two to three-fold increase in the estimated doubling times of the anammox organisms. Interestingly, anammox remained the primary nitrogen consumption route, and high-throughput 16S rRNA gene-targeted amplicon sequencing analyses revealed that the shift in performance was not associated with a shift in dominant anammox bacteria (“Candidatus Brocadia fulgida”). Furthermore, only limited heterotrophic denitrification was observed in the presence of easily biodegradable organics (acetate, glucose). The observed delays in net anammox growth were thus ascribed to the acclimatization of the initial anammox population or/and the development of a side population beneficial for them. Additionally, by combining microautoradiography and fluorescence in situ hybridization it was confirmed that the anammox organisms involved in the process did not directly incorporate or store the amended acetate and glucose. In conclusion, these investigations strongly support the feasibility of MWW treatment via anammox.

The influence of aeration intensity on predation and EPS production in membrane bioreactors.

Shear, in the form of vigorous aeration, is used to control fouling in membrane bioreactor (MBR) systems. However, shear also influences the physicochemical and biological properties of MBR biomass. The current study examines the relationship between the aeration intensity and extracellular polymeric substance (EPS) production in MBRs. Two identical submerged MBRs were operated in parallel but the aeration rate was three times greater in one of the MBRs. The concentrations of floc-associated and soluble EPS were monitored for the duration of the experiment. Microscopic images and floc-size measurements were also collected regularly. The membrane fouling potential of the biomass was quantified using the flux-step method. Increased aeration did not have a direct effect on soluble or floc-associated EPS production in the microfiltration MBRs. However, aeration intensity had a significant effect on predatory organisms. Large aquatic earthworms, Aeolosoma hemprichi, proliferated under lower shear conditions but were never observed in the high shear reactor. Predation by A. hemprichi resulted in increased floc-associated and soluble EPS production. Thus, the mixing conditions in the low shear MBR indirectly resulted in increased soluble EPS concentrations and higher fouling potential. This research suggests that predation can have a significant impact on the production rates of floc-associated and soluble EPS--key parameters driving membrane fouling in MBRs.

Evaluating operating conditions for outcompeting nitrite oxidizers and maintaining partial nitrification in biofilm systems using biofilm modeling and Monte Carlo filtering

In practice, partial nitrification to nitrite in biofilms has been achieved with a range of different operating conditions, but mechanisms resulting in reliable partial nitrification in biofilms are not well understood. In this study, mathematical biofilm modeling combined with Monte Carlo filtering was used to evaluate operating conditions that (1) lead to outcompetition of nitrite oxidizers from the biofilm, and (2) allow to maintain partial nitrification during long-term operation. Competition for oxygen was found to be the main mechanism for displacing nitrite oxidizers from the biofilm, and preventing re-growth of nitrite oxidizers in the long-term. To maintain partial nitrification in the model, a larger oxygen affinity (i.e., smaller half saturation constant) for ammonium oxidizers compared to nitrite oxidizers was required, while the difference in maximum growth rate was not important for competition under steady state conditions. Thus, mechanisms for washout of nitrite oxidizing bacteria from biofilms are different from suspended cultures where the difference in maximum growth rate is a key mechanism. Inhibition of nitrite oxidizers by free ammonia was not required to outcompete nitrite oxidizers from the biofilm, and to maintain partial nitrification to nitrite. But inhibition by free ammonia resulted in faster washout of nitrite oxidizers.

Changes in the structure and function of microbial communities in drinking water treatment bioreactors upon addition of phosphorus.

ABSTRACT Phosphorus was added as a nutrient to bench-scale and pilot-scale biologically active carbon (BAC) reactors operated for perchlorate and nitrate removal from contaminated groundwater. The two bioreactors responded similarly to phosphorus addition in terms of microbial community function (i.e., reactor performance), while drastically different responses in microbial community structure were detected. Improvement in reactor performance with respect to perchlorate and nitrate removal started within a few days after phosphorus addition for both reactors. Microbial community structures were evaluated using molecular techniques targeting 16S rRNA genes. Clone library results showed that the relative abundance of perchlorate-reducing bacteria (PRB) Dechloromonas and Azospira in the bench-scale reactor increased from 15.2% and 0.6% to 54.2% and 11.7% after phosphorus addition, respectively. Real-time quantitative PCR (qPCR) experiments revealed that these increases started within a few days after phosphorus addition. In contrast, after phosphorus addition, the relative abundance of Dechloromonas in the pilot-scale reactor decreased from 7.1 to 0.6%, while Zoogloea increased from 17.9 to 52.0%. The results of this study demonstrated that similar operating conditions for bench-scale and pilot-scale reactors resulted in similar contaminant removal performances, despite dramatically different responses from microbial communities. These findings suggest that it is important to evaluate the microbial community compositions inside bioreactors used for drinking water treatment, as they determine the microbial composition in the effluent and impact downstream treatment requirements for drinking water production. This information could be particularly relevant to drinking water safety, if pathogens or disinfectant-resistant bacteria are detected in the bioreactors.