Quantifying Elastic Properties of Environmental Biofilms using Optical Coherence Elastography.

Biofilms are complex biomaterials comprising a well-organized network of microbial cells encased in self-produced extracellular polymeric substances (EPS). This paper presents a detailed account of the implementation of optical coherence elastography (OCE) measurements tailored for the elastic characterization of biofilms. OCE is a non-destructive optical technique that enables the local mapping of the microstructure, morphology, and viscoelastic properties of partially transparent soft materials with high spatial and temporal resolution. We provide a comprehensive guide detailing the essential procedures for the correct implementation of this technique, along with a methodology to estimate the bulk Young's modulus of granular biofilms from the collected measurements. These consist of the system setup, data acquisition, and postprocessing. In the discussion, we delve into the underlying physics of the sensors used in OCE and explore the fundamental limitations regarding the spatial and temporal scales of OCE measurements. We conclude with potential future directions for advancing the OCE technique to facilitate elastic measurements of environmental biofilms.

Partial Nitritation/Anammox and biological phosphorus removal integration in a single bioreactor under mainstream conditions.

Anammox-based nitrogen removal and enhanced biological phosphorus removal (EBPR) are increasingly applied for nutrient removal from wastewater, but are typically operated in separate reactors. Here, a novel process for integrated partial nitritation/anammox (PN/A) and EBPR in a single reactor employing integrated fixed film activated sludge was tested. The reactor was fed with mainstream municipal wastewater (5.4 ± 1.3 g COD/g N) at 20 °C for 243 days. Robust ammonium, total inorganic nitrogen, and orthophosphate removal efficiencies of 94 ± 4 %, 87 ± 7 % and 92 ± 7 % were achieved. Nitrite-oxidizing organisms suppression and ammonia-oxidizing organisms retention were achieved via solids retention time control, intermittent aeration, and suspended versus attached biomass population segregation. The contribution of anammox to nitrogen removal increased from 24 % to 74 %. In parallel, a substantial enrichment of Tetrasphaera polyphosphate accumulating organisms was observed. This work demonstrates a novel intensified bioprocess coupling PN/A and EBPR in the same reactor for efficient nutrient removal from wastewater.

Impact of operational strategies on a sidestream enhanced biological phosphorus removal (S2EBPR) reactor in a carbon limited wastewater plant.

Water resource recovery facilities are faced with stringent effluent phosphorus limits to reduce nutrient pollution. Enhanced biological phosphorus removal (EBPR) is the most common biological route to remove phosphorus; however, many facilities struggle to achieve consistent performance due to limited carbon availability in the influent wastewater. A promising process to improve carbon availability is through return activated sludge (RAS) fermentation via sidestream EBPR (S2EBPR). In this study, a full-scale S2EBPR pilot was operated with a sidestream plus carbon configuration (SSRC) at a carbon-limited facility. A model based on the pilot test was developed and calibrated in the SUMO platform and used to explore routes for improving orthophosphate (OP) effluent compliance. Modeling results showed that RAS diversion by itself was not sufficient to drive OP removal to permit limits of 1 mg L-1, therefore, other strategies were evaluated. Supplemental carbon addition of MicroC® at 1.90 L min-1 and controlling the phosphorus concentration below 3.5 mgP L-1 in the primary effluent (PE) proved to be valid supplemental strategies to achieve OP removal below 1 mg L-1 most of the time. In particular, the proposed supplemental carbon flow rate would result in an improvement of the rbCOD:P ratio from 17:1 to 26:1. The synergistic approach of RAS diversion and supplemental carbon addition increased the polyphosphate accumulating organisms (PAO) population while minimizing the supplemental carbon needed to achieve consistent phosphorus removal. Overall, this pilot and modeling study shows that joint strategies, including RAS diversion, carbon addition and PE control, can be effective to achieve optimal control of OP effluent.

Tailoring polyvinyl alcohol-sodium alginate (PVA-SA) hydrogel beads by controlling crosslinking pH and time.

Hydrogel-encapsulated catalysts are an attractive tool for low-cost intensification of (bio)-processes. Polyvinyl alcohol-sodium alginate hydrogels crosslinked with boric acid and post-cured with sulfate (PVA-SA-BS) have been applied in bioproduction and water treatment processes, but the low pH required for crosslinking may negatively affect biocatalyst functionality. Here, we investigate how crosslinking pH (3, 4, and 5) and time (1, 2, and 8 h) affect the physicochemical, elastic, and process properties of PVA-SA-BS beads. Overall, bead properties were most affected by crosslinking pH. Beads produced at pH 3 and 4 were smaller and contained larger internal cavities, while optical coherence tomography suggested polymer cross-linking density was higher. Optical coherence elastography revealed PVA-SA-BS beads produced at pH 3 and 4 were stiffer than pH 5 beads. Dextran Blue release showed that pH 3-produced beads enabled higher diffusion rates and were more porous. Last, over a 28-day incubation, pH 3 and 4 beads lost more microspheres (as cell proxies) than beads produced at pH 5, while the latter released more polymer material. Overall, this study provides a path forward to tailor PVA-SA-BS hydrogel bead properties towards a broad range of applications, such as chemical, enzymatic, and microbially catalyzed (bio)-processes.

Enhancing resource recovery via cranberry syrup waste at the Wisconsin Rapids WRRF: An experimental and modeling study.

The Wisconsin Rapids Wastewater Treatment Plant (WRWWTP) is faced with a more stringent effluent phosphorus requirement that will drive capital investment between 2020 and 2025. The facility will need to achieve a monthly average value of 0.36 mg L-1 of total phosphorus (TP). While the facility has sufficient influent carbon to drive a conventional enhanced biological phosphorus removal (EBPR) configuration, the existing infrastructure makes the addition of influent selector zones cost prohibitive. Underutilized aeration basin capacity was repurposed for testing return activated sludge (RAS) fermentation. The WRWWTP began pilot testing of RAS fermentation in April 2021. The facility moved through a series of operational setpoints to optimize phosphorus removal in a sidestream RAS (SSR) configuration, including RAS diversion, decrease of DO in aeration basins and chemical dosing shutoff. One of the key implementations was the addition of cranberry syrup waste to provide additional carbon for RAS fermentation, converting the process to a SSR plus carbon (SSRC) configuration. By the end of the testing period, effluent total phosphorus was averaging less than 0.4 mg L-1 with no chemical addition. A model was developed in the SUMO platform and was used to capture orthophosphate trends during the testing period. The model investigated microbial population dynamics and found that the operational changes including RAS diversion, chemical dosing shutoff and cranberry syrup waste addition impacted the enrichment of phosphorus accumulating organisms (PAO). After performing a sensitivity analysis on hydrolysis parameters, the predicted hydrolysis rate around 1.8-1.9 mg COD g VSS-1 hr-1 was found to match the batch rate testing data. This is the first study where cranberry syrup waste was used to successfully enhance EBPR performance, resulting in 90% TP removal. While further research is needed regarding the composition of the waste matrix and the microbial community composition, this expands the routes for resource recovery in the field of wastewater treatment.

Lab-scale data and microbial community structure suggest shortcut nitrogen removal as the predominant nitrogen removal mechanism in post-aerobic digestion (PAD).

Implementing an aerobic digestion step after anaerobic digestion, referred to as "post aerobic digestion" (PAD), can remove ammonia without the need for an external carbon source and destroy volatile solids. While this process has been documented at the lab-scale and full-scale, the mechanism for N removal and the corresponding microbial community that carries out this process have not been established. This research gap is important to fill because the nitrogen removal pathway has implications on aeration requirements and carbon demand, that is, short-cut N-removal requires less oxygen and carbon than simultaneous nitrification-denitrification. The aims of this research were to (i) determine if nitrite (NO2 - ) or nitrate (NO3 - ) dominates following ammonia removal and (ii) characterize the microbial community from PAD reactors. Here, lab-scale PAD reactors were seeded with biomass from two different full-scale PAD reactors. The lab-scale reactors were fed with biomass from full-scale reactors and operated in batch mode to quantify nitrogen species concentrations (ammonia, NH4 + , NO2 - , and NO3 - ) over time. Experimental results revealed that NO2 - production rates were several orders of magnitude greater than NO3 - production rates. Indeed, nitrite accumulation rate (NAR) was greater than 90% at most temperatures, confirming that shortcut nitrogen removal was the dominant NH4 + removal mechanism in PAD. Microbial community analysis via 16S rRNA sequencing indicated that ammonia oxidizing bacteria (AOB) were much more abundant than nitrite oxidizing bacteria (NOB). Overall, this study suggests that aeration requirements for post-aerobic digestion should be based on NO2 - shunt and not complete simultaneous nitrification denitrification. PRACTITIONER POINTS: AOB are a key feature of PAD microbial communities NOB are present, but in much lower abundance than AOB High nitrite accumulation ratio suggests shortcut nitrite as the main mechanism for nitrogen removal Nitritation in PAD reactors is sustained at temperatures as high as 40°C No ammonia oxidation occurred at 50°C implying different mechanisms of nitrogen removal including ammonia stripping.

Simultaneous nitrification-denitrification in biofilm systems for wastewater treatment: Key factors, potential routes, and engineered applications.

Simultaneous nitrification-denitrification (SND) is an advantageous bioprocess that allows the complete removal of ammonia nitrogen through sequential redox reactions leading to nitrogen gas production. SND can govern nitrogen removal in single-stage biofilm systems, such as the moving bed biofilm reactor and aerobic granular sludge system, as oxygen gradients allow the development of multilayered biofilms including nitrifying and denitrifying bacteria. Environmental and operational conditions can strongly influence SND performance, biofilm development and biochemical pathways. Recent advances have outlined the possibility to reduce the carbon and energy consumption of the process via the "shortcut pathway", and simultaneously remove both N and phosphorus under specific operational conditions, opening new possibilities for wastewater treatment. This work critically reviews the factors influencing SND and its application in biofilm systems from laboratory to full scale. Operational strategies to enhance SND efficiency and hints to reduce nitrous oxide emission and operational costs are provided.

Layered viscoelastic properties of granular biofilms.

Granular biofilms are dense spherical complex biological systems composed mainly of multi-microbial cells, water, and extracellular polymeric substances (EPS). They facilitate efficient purification and settling of activated sludge in wastewater treatment processes. The viscoelastic properties of these complex biofilm systems are important characteristics that control their growth and dictate how they respond to hydrodynamic forces and chemical stimuli. However, the viscoelastic properties of granular biofilms are poorly understood. In this paper, we study granular biofilms' viscoelastic properties using optical coherence elastography (OCE), a nondestructive method that integrates optical coherence tomography (OCT) with elastic wave propagation. While quantitative viscoelastic characterization of granular biofilms is challenging due to their heterogeneous properties, we show that elastic waves are suitable for this purpose. First, we employ guided elastic waves in a thin section of a granular biofilm to reveal a two-layered profile for the viscoelastic properties. Next, we utilize circumferential elastic waves that propagate near the surface of a non-sectioned spherical biofilm to quantify the layered system's viscoelastic properties. To the best of our knowledge, this work is the first quantitative study that characterizes the layered viscoelastic properties of granular biofilms. The measurement approach may provide a platform to study the interplay between the viscoelastic properties and other characteristics of granular biofilms such as the complex microbial system, morphology, and oxygen distribution.

Functionalizing a Polyelectrolyte Complex with Chitosan Derivatives to Tailor Membrane Surface Properties.

Polyelectrolyte complex (PEC) materials show promise in the development of tunable membranes for aqueous and organic solvent separations, as well as in the creation of surface layers for fouling control. In this study, we developed a polyelectrolyte complex (PEC) functionalized by negatively charged carboxymethyl chitosan (CMC-) and positively charged quaternized chitosan (QC+) to tailor its surface properties and antibacterial efficacy. CMC- and QC+ were prepared and characterized using FT-IR and 1H NMR, which confirmed the presence of the carboxymethyl group and trimethylammonium group in CMC- and QC+ with 65.6% and 83.9% substitution, respectively. The CMC- functionalized PEC (CMC-/PEC) and QC+ functionalized PEC materials (QC+/PEC) were evaluated for their stability in water, resistance to organic and inorganic adsorption, and antibacterial action against a model microorganism, Pseudomonas putida. The results showed no release of chitosan derivatives after adsorption, and CMC-/PEC and QC+/PEC exhibited charge-based, selective repulsion of model organic and inorganic substances. Moreover, the functionalized PEC surfaces displayed lower bacterial attachment due to their smoother surfaces as compared to the bare ceramic membrane and their antimicrobial properties. Among the PEC samples, CMC-/PEC had the lowest cell attachment, while QC+/PEC showed the highest attachment due to electrostatic attraction. The ceramic and bare PEC surfaces were negligibly bactericidal, while cell viability decreased to 34.4 ± 10.2% and 30.6 ± 8.2% with the CMC-/PEC and QC+/PEC surfaces, respectively. In the filtration experiments, the unmodified PEC and CMC-/PEC showed lower rates of flux decline due to organic fouling than did the bare ceramic or QC+/PEC due to electrostatic repulsion. Furthermore, PECs as protective layers promoted much higher flux recoveries than simply backwashing the uncoated membranes. This surface tunability, then, enhances the potential of PECs either as fouling resistant materials or as a method to create a sacrificial, protective layer on surfaces that once fouled can be dissolved and re-established.

Pushing the limits of solids retention time for enhanced biological phosphorus removal: process characteristics and Accumulibacter population structure.

Reducing the solids retention time (SRT) of the enhanced biological phosphorus removal (EBPR) process can increase organic carbon diversion to the sidestream for energy recovery, thereby realizing some of the benefits of the high rate activated sludge (HRAS) process. Determining the washout (i.e. minimum) SRT of polyphosphate accumulating organisms (PAOs), therefore, allows for simultaneous phosphorus and carbon diversion for energy recovery from EBPR systems. However, few studies have investigated the washout SRT of PAOs in real wastewater, and little is known of the diversity of PAOs in high rate EBPR systems. Here we demonstrate efficient phosphorus removal (83% orthophosphate removal) in a high rate EBPR sequencing batch reactor fed real primary effluent and operated at 20 °C. Stable operation was achieved at a total SRT of 1.8 ± 0.2 days and hydraulic retention time of 3.7-4.8 hours. 16S rRNA gene sequencing data demonstrated that Accumulibacter were the dominant PAO throughout the study, with a washout aerobic SRT between 0.8 and 1.4 days. qPCR targeting the polyphosphate kinase gene revealed that Accumulibacter clades IIA, IIB and IID dominated the PAO community at low SRT operation, while clade IA was washed out at the lowest SRT values.

In situ groundwater remediation with bioelectrochemical systems: A critical review and future perspectives.

Groundwater contamination is an ever-growing environmental issue that has attracted much and undiminished attention for the past half century. Groundwater contamination may originate from both anthropogenic (e.g., hydrocarbons) and natural compounds (e.g., nitrate and arsenic); to tackle the removal of these contaminants, different technologies have been developed and implemented. Recently, bioelectrochemical systems (BES) have emerged as a potential treatment for groundwater contamination, with reported in situ applications that showed promising results. Nitrate and hydrocarbons (toluene, phenanthrene, benzene, BTEX and light PAHs) have been successfully removed, due to the interaction of microbial metabolism with poised electrodes, in addition to physical migration due to the electric field generated in a BES. The selection of proper BESs relies on several factors and problems, such as the complexity of groundwater and subsoil environment, scale-up issues, and energy requirements that need to be accounted for. Modeling efforts could help predict case scenarios and select a proper design and approach, while BES-based biosensing could help monitoring remediation processes. In this review, we critically analyze in situ BES applications for groundwater remediation, focusing in particular on different proposed setups, and we identify and discuss the existing research gaps in the field.

Towards mechanical characterization of granular biofilms by optical coherence elastography measurements of circumferential elastic waves.

Microbial granular biofilms are spherical, multi-layered aggregates composed of communities of bacterial cells encased in a complex matrix of hydrated extracellular polymeric substances (EPS). While granular aggregates are increasingly used for applications in industrial and municipal wastewater treatment, their underlying mechanical properties are poorly understood. The challenges of viscoelastic characterization for these structures are due to their spherical geometry, spatially heterogeneous properties, and their delicate nature. In this study, we report a model-based approach for nondestructive characterization of viscoelastic properties (shear modulus and shear viscosity) of alginate spheres with different concentrations, which was motivated by our measurements in granular biofilms. The characterization technique relies on experimental measurements of circumferential elastic wave speeds as a function of frequency in the samples using the Optical Coherence Elastography (OCE) technique. A theoretical model was developed to estimate the viscoelastic properties of the samples from OCE data through inverse analysis. This work represents the first attempt to explore elastic waves for mechanical characterization of granular biofilms. The combination of the OCE technique and the theoretical model presented in this paper provides a framework that can facilitate quantitative viscoelastic characterization of samples with curved geometries and the study of the relationships between morphology and mechanical properties in granular biofilms.

Using kinetics and modeling to predict denitrification fluxes in elemental-sulfur-based biofilms.

Elemental sulfur (S0 ) can serve as an electron donor for water and wastewater denitrification, but few researchers have addressed the kinetics of S 0 -based reduction of nitrate (NO 3- ), nitrite (NO 2- ), and nitrous oxide (N 2 O). In addition, S 0 -based denitrifying biofilms are counter-diffusional. This is because the electron donor (S 0 ) is supplied from the biofilm attachment surface while the acceptor, for example, NO 3- , is supplied from the bulk liquid. No existing mathematical model for S 0 -based denitrification considers this behavior. In this study, batch tests were used to determine the kinetic parameters for the reduction of NO 3- , NO 2- , and N 2 O. Additionally, a biofilm model was developed to explore the effects of counter-diffusion on overall fluxes, that is, the mass of NO 3- or NO 2- removed per unit biofilm support area per unit time. The maximum specific substrate utilization rates ( qˆ ) for NO 3- , NO 2- , and N 2 O were 3.54, 1.98, and 6.28 g N g COD -1 ·d -1 , respectively. The maximum specific growth rates ( µË† ) were 0.71, 1.21, and 1.67 d -1 for NO 3- to NO 2- , NO 2- to N 2 O, and N 2 O to N 2 , respectively. Results suggest that the observed NO 2- accumulation during S 0 -based denitrification results from a low qˆ for NO 2- relative to that for NO 3- . The high qˆ for N 2 O, relative to that for NO 3- and NO 2- , suggest that little N 2 O accumulation occurs during denitrification. A counter-diffusional biofilm model was used to predict trends for NO 3- fluxes, and confirmed NO 2- accumulation in S 0 -based denitrification biofilms. It also explains the observed detrimental effects of biofilm thickness on denitrification fluxes. This study allows a more accurate prediction of NO 3- , NO 2- , and N 2 O transformations in S 0 -based denitrification.

Comammox Nitrospira are the dominant ammonia oxidizers in a mainstream low dissolved oxygen nitrification reactor.

Recent findings show that a subset of bacteria affiliated with Nitrospira, a genus known for its importance in nitrite oxidation for biological nutrient removal applications, are capable of complete ammonia oxidation (comammox) to nitrate. Early reports suggested that they were absent or present in low abundance in most activated sludge processes, and thus likely functionally irrelevant. Here we show the accumulation of comammox Nitrospira in a nitrifying sequencing batch reactor operated at low dissolved oxygen (DO) concentrations. Actual mainstream wastewater was used as influent after primary settling and an upstream pre-treatment process for carbon and phosphorus removal. The ammonia removal rate was stable and exceeded that of the treatment plant's parallel full-scale high DO nitrifying activated sludge reactor. 16S rRNA gene sequencing showed a steady accumulation of Nitrospira to 53% total abundance and a decline in conventional ammonia oxidizing bacteria to <1% total abundance over 400 + days of operation. After ruling out other known ammonia oxidizers, qPCR confirmed the accumulation of comammox Nitrospira beginning around day 200, to eventually comprise 94% of all detected amoA and 4% of total bacteria by day 407. Quantitative fluorescence in-situ hybridization confirmed the increasing trend and high relative abundance of Nitrospira. These results demonstrate that comammox can be metabolically relevant to nitrogen transformation in wastewater treatment, and can even dominate the ammonia oxidizing community. Our results suggest that comammox may be an important functional group in energy efficient nitrification systems designed to operate at low DO levels.

Nondestructive characterization of soft materials and biofilms by measurement of guided elastic wave propagation using optical coherence elastography.

Biofilms are soft multicomponent biological materials composed of microbial communities attached to surfaces. Despite the crucial relevance of biofilms to diverse industrial, medical, and environmental applications, the mechanical properties of biofilms are understudied. Moreover, most of the available techniques for the characterization of biofilm mechanical properties are destructive. Here, we detail a model-based approach developed to characterize the viscoelastic properties of soft materials and bacterial biofilms based on experimental data obtained using the nondestructive dynamic optical coherence elastography (OCE) technique. The model predicted the frequency- and geometry-dependent propagation velocities of elastic waves in a soft viscoelastic plate supported by a rigid substratum. Our numerical calculations suggest that the dispersion curves of guided waves recorded in thin soft plates by the dynamic OCE technique are dominated by guided waves, whose phase velocities depend on the viscoelastic properties and plate thickness. The numerical model was validated against experimental measurements in agarose phantom samples with different thicknesses and concentrations. The model was then used to interpret guided wave dispersion curves obtained by the OCE technique in bacterial biofilms developed in a rotating annular reactor, which allowed the quantitative characterization of biofilm shear modulus and viscosity. This study is the first to employ measurements of elastic wave propagation to characterize biofilms, and it provides a novel framework combining a theoretical model and an experimental approach for studying the relationship between the biofilm internal physical structure and mechanical properties.

A Serial Biofiltration System for Effective Removal of Low-Concentration Nitrous Oxide in Oxic Gas Streams: Mathematical Modeling of Reactor Performance and Experimental Validation.

Wastewater treatment plants (WWTPs) are among the major anthropogenic sources of N2O, a major greenhouse gas and ozone-depleting agent. We recently devised a zero-energy zero-carbon biofiltration system easily applicable to activated sludge-type WWTPs and performed lab-scale proof-of-concept experiments. The major drawback of the system was the diminished performance observed when fully oxic gas streams were treated. Here, a serial biofiltration system was tested as a potential improvement. A laboratory system with three serially positioned biofilters, each receiving a separate feed of artificial wastewater, was fed N2O-containing gas streams of varied flow rates (200-2000 mL·min-1) and O2 concentrations (0-21%). Use of the serial setup substantially improved the reactor performance. Fed fully oxic gas at a flow rate of 1000 mL·min-1, the system removed N2O at an elimination capacity of 0.402 ± 0.009 g N2O·m-3·h-1 (52.5% removal), which was approximately 2.4-fold higher than that achieved with a single biofilter, 0.171 ± 0.024 g N2O·m-3·h-1. These data were used to validate the mathematical model developed to estimate the performance of the N2O biofiltration system. The Nash-Sutcliffe efficiency indices ranged from 0.78 to 0.93, confirming high predictability, and the model provided mechanistic insights into aerobic N2O removal and the performance enhancement achieved with the serial configuration.

Nitrous oxide emissions from biofilm processes for wastewater treatment.

This paper discusses the microbial basis and the latest research on nitrous oxide (N2O) emissions from biofilms processes for wastewater treatment. Conditions that generally promote N2O formation in biofilms include (1) low DO values, or spatial DO transitions from high to low within the biofilm; (2) DO fluctuations within biofilm due to varying bulk DO concentrations or varying substrate concentrations; (3) conditions with high reaction rates, which lead to greater formation of intermediates, e.g., hydroxylamine (NH2OH) and nitrite (NO2-), that promote N2O formation; and (4) electron donor limitation for denitrification. Formation of N2O directly results from the activities of ammonia-oxidizing bacteria (AOB), ammonia-oxidizing archaea (AOA), and heterotrophic denitrifying bacteria. More research is needed on the roles of AOA, comammox, and specialized denitrifying microorganisms. In nitrifying biofilms, higher bulk ammonia (NH3) concentrations, higher nitrite (NO2-) concentrations, lower dissolved oxygen (DO), and greater biofilm thicknesses result in higher N2O emissions. In denitrifying biofilms, N2O accumulates at low levels as an intermediate and at higher levels at the oxic/anoxic transition regions of the biofilms and where COD becomes limiting. N2O formed in the outer regions can be consumed in the inner regions if COD penetrates sufficiently. In membrane-aerated biofilms, where nitrification takes place in the inner, aerobic biofilm region, the exterior anoxic biofilm can serve as a N2O sink. Reactors that include variable aeration or air scouring, such as denitrifying filters, trickling filters, or rotating biological contactors (RBCs), can form peaks of N2O emissions during or following a scouring or aeration event. N2O emissions from biofilm processes depend on the microbial composition, biofilm thickness, substrate concentrations and variability, and reactor type and operation. Given the complexity and difficulty in quantifying many of these factors, it may be difficult to accurately predict emissions for full-scale treatment plants. However, a better understanding of the mechanisms and the impacts of process configurations can help minimize N2O emission from biofilm processes for wastewater treatment.

Mechanisms of nitrous oxide (N2 O) formation and reduction in denitrifying biofilms.

Nitrous oxide (N2 O) is a potent greenhouse gas that can be formed in wastewater treatment processes by ammonium oxidizing and denitrifying microorganisms. While N2 O emissions from suspended growth systems have been extensively studied, and some recent studies have addressed emissions from nitrifying biofilms, much less is known about N2 O emissions from denitrifying biofilm processes. This research used modeling to evaluate the mechanisms of N2 O formation and reduction in denitrifying biofilms. The kinetic model included formation and consumption of key denitrification species, including nitrate (NO3-), nitrite (NO2-), nitric oxide (NO), and N2 O. The model showed that, in presence of excess of electron donor, denitrifying biofilms have two distinct layers of activity: an outer layer where there is net production of N2 O and an inner layer where there is net consumption. The presence of oxygen (O2 ) had an important effect on N2 O emission from suspended growth systems, but a smaller effect on biofilm systems. The effects of NO3- and O2 differed significantly based on the biofilm thickness. Overall, the effects of biofilm thickness and bulk substrate concentrations on N2 O emissions are complex and not always intuitive. A key mechanism for denitrifying biofilms is the diffusion of N2 O and other intermediates from one zone of the biofilm to another. This leads to zones of N2 O formation or consumption transformations that would not exist in suspended growth systems.

Predicting N2O emissions from nitrifying and denitrifying biofilms: a modeling study.

Wastewater treatment plants can be significant sources of nitrous oxide (N2O), a potent greenhouse gas. While our understanding of N2O emissions from suspended-growth processes has advanced significantly, less is known about emissions from biofilm processes. Biofilms may behave differently due to their substrate gradients and microbial stratification. In this study, we used mathematical modeling to explore the mechanisms of N2O emissions from nitrifying and denitrifying biofilms. Our ammonia-oxidizing bacteria biofilm model suggests that N2O emissions from biofilm can be significantly greater than from suspended-growth systems. The driving factor is the diffusion of hydroxylamine, a nitrification intermediate, from the aerobic to the anoxic regions of the biofilm. The presence of nitrite-oxidizing bacteria further increased emissions. For denitrifying biofilms, our results suggest that emissions are generally greater than for suspended-growth systems. However, the magnitude of the difference depends on the bulk dissolved oxygen, chemical oxygen demand, and nitrate concentrations, as well as the biofilm thickness. Overall, the accumulation and diffusion of key intermediates, i.e. hydroxylamine and nitrite, distinguish biofilms from suspended-growth systems. Our research suggests that the mechanisms of N2O emissions from biofilms are much more complex than suspended-growth systems, and that emissions may be higher in many cases.

Hydroxylamine diffusion can enhance N2O emissions in nitrifying biofilms: a modeling study.

Wastewater treatment plants can be significant sources of nitrous oxide (N2O), a potent greenhouse gas. However, little is known about N2O emissions from biofilm processes. We adapted an existing suspended-growth mathematical model to explore N2O emissions from nitrifying biofilms. The model included N2O formation by ammonia-oxidizing bacteria (AOB) via the hydroxylamine and the nitrifier denitrification pathways. Our model suggested that N2O emissions from nitrifying biofilms could be significantly greater than from suspended growth systems under similar conditions. The main cause was the formation and diffusion of hydroxylamine, an AOB nitrification intermediate, from the aerobic to the anoxic regions of the biofilm. In the anoxic regions, hydroxylamine oxidation by AOB provided reducing equivalents used solely for nitrite reduction to N2O, since there was no competition with oxygen. For a continuous system, very high and very low dissolved oxygen (DO) concentrations resulted in lower emissions, while intermediate values led to higher emissions. Higher bulk ammonia concentrations and greater biofilm thicknesses increased emissions. The model effectively predicted N2O emissions from an actual pilot-scale granular sludge reactor for sidestream nitritation, but significantly underestimated the emissions when the NH2OH diffusion coefficient was assumed to be minimal. This numerical study suggests an unexpected and important role of hydroxylamine in N2O emission in biofilms.