A modeling pipeline to relate municipal wastewater surveillance and regional public health data.

As COVID-19 becomes endemic, public health departments benefit from improved passive indicators, which are independent of voluntary testing data, to estimate the prevalence of COVID-19 in local communities. Quantification of SARS-CoV-2 RNA from wastewater has the potential to be a powerful passive indicator. However, connecting measured SARS-CoV-2 RNA to community prevalence is challenging due to the high noise typical of environmental samples. We have developed a generalized pipeline using in- and out-of-sample model selection to test the ability of different correction models to reduce the variance in wastewater measurements and applied it to data collected from treatment plants in the Chicago area. We built and compared a set of multi-linear regression models, which incorporate pepper mild mottle virus (PMMoV) as a population biomarker, Bovine coronavirus (BCoV) as a recovery control, and wastewater system flow rate into a corrected estimate for SARS-CoV-2 RNA concentration. For our data, models with BCoV performed better than those with PMMoV, but the pipeline should be used to reevaluate any new data set as the sources of variance may change across locations, lab methods, and disease states. Using our best-fit model, we investigated the utility of RNA measurements in wastewater as a leading indicator of COVID-19 trends. We did this in a rolling manner for corrected wastewater data and for other prevalence indicators and statistically compared the temporal relationship between new increases in the wastewater data and those in other prevalence indicators. We found that wastewater trends often lead other COVID-19 indicators in predicting new surges.

Trace Organic Contaminant Removal from Municipal Wastewater by Styrenic ß-Cyclodextrin Polymers.

Trace organic contaminants (TrOCs) present major removal challenges for wastewater treatment. TrOCs, such as perfluoroalkyl and polyfluoroalkyl substances (PFAS), are associated with chronic toxicity at ng L-1 exposure levels and should be removed from wastewater to enable safe reuse and release of treated effluents. Established adsorbents, such as granular activated carbon (GAC), exhibit variable TrOC removal and fouling by wastewater constituents. These shortcomings motivate the development of selective novel adsorbents that also maintain robust performance in wastewater. Cross-linked ß-cyclodextrin (ß-CD) polymers are promising adsorbents with demonstrated TrOC removal efficacy. Here, we report a simplified and potentially scalable synthesis of a porous polymer composed of styrene-linked ß-CD and cationic ammonium groups. Batch adsorption experiments demonstrate that the polymer is a selective adsorbent exhibiting complete removal for six out of 13 contaminants with less adsorption inhibition than GAC in wastewater. The polymer also exhibits faster adsorption kinetics than GAC and ion exchange (IX) resin, higher adsorption affinity for PFAS than GAC, and is regenerable by solvent wash. Rapid small-scale column tests show that the polymer exhibits later breakthrough times compared to GAC and IX resin. These results demonstrate the potential for ß-CD polymers to remediate TrOCs from complex water matrices.

Microplastic accumulation in riverbed sediment via hyporheic exchange from headwaters to mainstems.

In rivers, small and lightweight microplastics are transported downstream, but they are also found frequently in riverbed sediment, demonstrating long-term retention. To better understand microplastic dynamics in global rivers from headwaters to mainstems, we developed a model that includes hyporheic exchange processes, i.e., transport between surface water and riverbed sediment, where microplastic retention is facilitated. Our simulations indicate that the longest microplastic residence times occur in headwaters, the most abundant stream classification. In headwaters, residence times averaged 5 hours/km but increased to 7 years/km during low-flow conditions. Long-term accumulation for all stream classifications averaged ~5% of microplastic inputs per river kilometer. Our estimates isolated the impact of hyporheic exchange processes, which are known to influence dynamics of naturally occurring particles in streams, but rarely applied to microplastics. The identified mechanisms and time scales for small and lightweight microplastic accumulation in riverbed sediment reveal that these often-unaccounted components are likely a pollution legacy that is crucial to include in global assessments.

Critical Capability Needs for Reduction of Transmission of SARS-CoV-2 Indoors.

Coordination of efforts to assess the challenges and pain points felt by industries from around the globe working to reduce COVID-19 transmission in the indoor environment as well as innovative solutions applied to meet these challenges is mandatory. Indoor infectious viral disease transmission (such as coronavirus, norovirus, influenza) is a complex problem that needs better integration of our current knowledge and intervention strategies. Critical to providing a reduction in transmission is to map the four core technical areas of environmental microbiology, transmission science, building science, and social science. To that end a three-stage science and innovation Summit was held to gather information on current standards, policies and procedures applied to reduce transmission in built spaces, as well as the technical challenges, science needs, and research priorities. The Summit elucidated steps than can be taken to reduce transmission of SARS-CoV-2 indoors and calls for significant investments in research to enhance our knowledge of viral pathogen persistence and transport in the built environment, risk assessment and mitigation strategy such as processes and procedures to reduce the risk of exposure and infection through building systems operations, biosurveillance capacity, communication form leadership, and stakeholder engagement for optimal response. These findings reflect the effective application of existing knowledge and standards, emerging science, and lessons-learned from current efforts to confront SARS-CoV-2.

Standardizing data reporting in the research community to enhance the utility of open data for SARS-CoV-2 wastewater surveillance.

SARS-CoV-2 RNA detection in wastewater is being rapidly developed and adopted as a public health monitoring tool worldwide. With wastewater surveillance programs being implemented across many different scales and by many different stakeholders, it is critical that data collected and shared are accompanied by an appropriate minimal amount of metainformation to enable meaningful interpretation and use of this new information source and intercomparison across datasets. While some databases are being developed for specific surveillance programs locally, regionally, nationally, and internationally, common globally-adopted data standards have not yet been established within the research community. Establishing such standards will require national and international consensus on what metainformation should accompany SARS-CoV-2 wastewater measurements. To establish a recommendation on minimum information to accompany reporting of SARS-CoV-2 occurrence in wastewater for the research community, the United States National Science Foundation (NSF) Research Coordination Network on Wastewater Surveillance for SARS-CoV-2 hosted a workshop in February 2021 with participants from academia, government agencies, private companies, wastewater utilities, public health laboratories, and research institutes. This report presents the primary two outcomes of the workshop: (i) a recommendation on the set of minimum meta-information that is needed to confidently interpret wastewater SARS-CoV-2 data, and (ii) insights from workshop discussions on how to improve standardization of data reporting.

Gathering at the top? Environmental controls of microplastic uptake and biomagnification in freshwater food webs.

Microplastics are ubiquitous in the environment, with high concentrations being detected now also in river corridors and sediments globally. Whilst there has been increasing field evidence of microplastics accumulation in the guts and tissues of freshwater and marine aquatic species, the uptake mechanisms of microplastics into freshwater food webs, and the physical and geological controls on pathway-specific exposures to microplastics, are not well understood. This knowledge gap is hampering the assessment of exposure risks, and potential ecotoxicological and public health impacts from microplastics. This review provides a comprehensive synthesis of key research challenges in analysing the environmental fate and transport of microplastics in freshwater ecosystems, including the identification of hydrological, sedimentological and particle property controls on microplastic accumulation in aquatic ecosystems. This mechanistic analysis outlines the dominant pathways for exposure to microplastics in freshwater ecosystems and identifies potentially critical uptake mechanisms and entry pathways for microplastics and associated contaminants into aquatic food webs as well as their risk to accumulate and biomagnify. We identify seven key research challenges that, if overcome, will permit the advancement beyond current conceptual limitations and provide the mechanistic process understanding required to assess microplastic exposure, uptake, hazard, and overall risk to aquatic systems and humans, and provide key insights into the priority impact pathways in freshwater ecosystems to support environmental management decision making.

Knowledge, attitudes, intentions, and behavior related to green infrastructure for flood management: A systematic literature review.

Green infrastructure (GI), which mimics natural hydrological systems, is a promising solution for flood management at the intersection of urban built infrastructure and natural systems. However, it has not yet achieved widespread uptake, due in part to insufficient understanding of human dimensions of the broader socio-ecological-technical system. We therefore conducted a multidisciplinary systematic literature review to synthesize research on people's existing knowledge about flood risk and GI, and how that shapes their attitudes and motivation to adopt new solutions. We systematically screened 21,207 studies on GI for flood management; 85 met our inclusion criteria. We qualitatively analyzed these studies to extract results on knowledge, attitudes, intentions, and behavior relating to GI for flood management. Overall, knowledge of GI was low across the 44 studies in which it was evaluated. Seventy studies assessed attitudes about GI, including the functional, aesthetic, health and safety, recreational, conservation, financial, and cultural value of GI, albeit their measurement was inconsistent. Willingness to implement or pay for GI varied considerably across 55 studies in which it was measured. Twenty studies measured and documented behavior relating to GI use, and these found low rates of adoption. Few studies systematically assessed the role of demographic, socio-economic, or geographic characteristics that could influence individuals' knowledge, attitudes, intentions or behavior, and thereby the success of GI programs. We recommend that researchers should more systematically capture data on human dimensions of GI (i.e. knowledge, attitudes, intentions, and behavior) across diverse settings to improve program design and uptake, especially among vulnerable populations. Greater attention to the social component of the socio-ecological-technical system will help ensure that GI programs are equitable, inclusive, and sustainable.

Soil hydrology drives ecological niche differentiation in a native prairie microbiome.

While the impacts of soil moisture on soil microbiome diversity and composition are well characterized, the influence of hydrological regime has been overlooked. As precipitation patterns are altered by climate change, understanding the impact of soil hydrology on community structure and function is critical. In this work, water level was continuously monitored for over a year in a Midwestern prairie-wetland at 10 cm depth increments up to a depth of 120 cm in 10 locations. We analyzed microbiome composition and edaphic factors in soil cores collected from this unique spatially distributed, longitudinal data set. We demonstrate that the fraction of time that each sample was inundated explains more variability in diversity and composition across this site than other commonly assessed edaphic factors, such as soil pH or depth. Finally, we show that these compositional changes influence abundance of ammonia oxidizers. The observed patterns in community composition and diversity are fundamentally regulated by the interaction of water with a structured landscape, particularly an elevated sand ridge characterized by drier conditions and a lower-lying wetland with more clayey soils. Similar processes are generally expected to influence the biogeography of many terrestrial environments, as morphology, hydrology and soil properties generally co-vary.

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.

A systematic review of the human health and social well-being outcomes of green infrastructure for stormwater and flood management.

BACKGROUND: The increase in frequency and intensity of urban flooding is a global challenge. Flooding directly impacts residents of industrialized cities with aging combined sewer systems, as well as cities with less centralized infrastructure to manage stormwater, fecal sludge, and wastewater. Green infrastructure is growing in popularity as a sustainable strategy to mimic nature-based flood management. Although its technical performance has been extensively studied, little is known about the effects of green stormwater infrastructure on human health and social well-being. METHODS: We conducted a multidisciplinary systematic review of peer-reviewed and gray literature on the effects of green infrastructure for stormwater and flood management on individuals', households', and communities' a) physical health; b) mental health; c) economic well-being; and d) flood resilience and social acceptance of green infrastructure. We systematically searched databases such as PubMed, Web of Science, and Scopus; the first 300 results in Google Scholar; and websites of key organizations including the United States Environmental Protection Agency. Study quality and strength of evidence was assessed for included studies, and descriptive data were extracted for a narrative summary. RESULTS: Out of 21,213 initial results, only 18 studies reported health or social well-being outcomes. Seven of these studies used primary data, and none allowed for causal inference. No studies connected green infrastructure for stormwater and flood management to mental or physical health outcomes. Thirteen studies were identified on economic outcomes, largely reporting a positive association between green infrastructure and property values. Five studies assessed changes in perceptions about green infrastructure, but with mixed results. Nearly half of all included studies were from Portland, Oregon. CONCLUSIONS: This global systematic review highlights the minimal evidence on human health and social well-being relating to green infrastructure for stormwater and flood management. To enable scale-up of this type of infrastructure to reduce flooding and improve ecological and human well-being, widespread acceptance of green infrastructure will be essential. Policymakers and planners need evidence on the full range of benefits from different contexts to enable financing and implementation of instfrastructure options, especially in highly urbanized, flood-prone settings around the world. Therefore, experts in social science, public health, and program evaluation must be integrated into interdisciplinary green infrastructure research to better relate infrastructure design to tangible human outcomes.

Effects of vertical hydrodynamic mixing on photomineralization of dissolved organic carbon in arctic surface waters.

Photomineralization, the transformation of dissolved organic carbon (DOC) to CO2 by sunlight, is an important source of CO2 in arctic surface waters. However, quantifying the role of photomineralization in inland waters is limited by the understanding of hydrologic controls on this process. To bridge this gap, this study evaluates mixing limitations, i.e., whether and by how much vertical mixing limits the depth-integrated photomineralization rate, in freshwater systems. We developed a conceptual model to qualitatively assess mixing limitations across the range of light attenuation and hydrologic conditions observed in freshwaters. For the common case of exponential light attenuation over depth, we developed a mathematical model to quantify mixing limitation, and used this model to assess a range of arctic freshwater systems. The results demonstrate that mixing limitations are important when there is significant light attenuation by suspended sediment (SS), which is the case in some arctic, boreal and temperate waters. Mixing limitation is pronounced when light attenuation over depth is strong and when the photomineralization rate at the water surface exceeds the vertical mixing rate. Arctic streams and rivers have strong vertical mixing relative to surface photomineralization, such that model results demonstrate no mixing limitation regardless of how much SS is present. Our analysis indicates that well-mixed assumptions used in prior work are valid in many, but not all, arctic surface waters. The effects of mixing limitations in reducing the photomineralization rate must be considered in arctic lakes with high SS concentrations.

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.

Effects of resuspension on the mobility and chemical speciation of zinc in contaminated sediments.

Identifying and quantifying the processes governing the mobilization of metals during resuspension events is key to assessing long-term metals efflux from sediments and associated ecological impacts. We investigated the effects of sediment resuspension on the mobilization and chemical speciation of zinc in two-week-long batch experiments using metal-contaminated sediments from Lake DePue (IL, USA). Measurements of dissolved zinc and sulfate allowed us to characterize the kinetics of metal sulfide dissolution and the resulting net release of zinc to the aqueous phase. X-ray absorption spectroscopy (XAS) provided direct insights into the chemical speciation of iron and zinc and their dynamic transformations during resuspension. While ZnS rapidly oxidized during resuspension, dissolved zinc increased only after two days of resuspension. We proposed a kinetic model to explain changes in the chemical speciation of zinc during these experiments as constrained by the dissolved species concentrations and chemical speciation as informed by XAS. Only 15% of the zinc mobilized was released to the aqueous phase while the remaining fraction repartitioned the solid phase either as a carbonate precipitate or as a sorbed species. Our results show that zinc sorption onto particle surfaces and reprecipitation of zinc minerals limit zinc solubility during resuspension of metal-sulfide sediments.

Morphological analysis of pore size and connectivity in a thick mixed-culture biofilm.

Morphological parameters are commonly used to predict transport and metabolic kinetics in biofilms. Yet, quantification of biofilm morphology remains challenging because of imaging technology limitations and lack of robust analytical approaches. We present a novel set of imaging and image analysis techniques to estimate internal porosity, pore size distributions, and pore network connectivity to a depth of 1 mm at a resolution of 10 µm in a biofilm exhibiting both heterotrophic and nitrifying activities. Optical coherence tomography (OCT) scans revealed an extensive pore network with diameters as large as 110 µm directly connected to the biofilm surface and surrounding fluid. Thin-section fluorescence in situ hybridization microscopy revealed that ammonia-oxidizing bacteria (AOB) distributed through the entire thickness of the biofilm. AOB were particularly concentrated in the biofilm around internal pores. Areal porosity values estimated from OCT scans were consistently lower than those estimated from multiphoton laser scanning microscopy, though the two imaging modalities showed a statistically significant correlation (r = 0.49, p < 0.0001). Estimates of areal porosity were moderately sensitive to gray-level threshold selection, though several automated thresholding algorithms yielded similar values to those obtained by manually thresholding performed by a panel of environmental engineering researchers (±25% relative error). These findings advance our ability to quantitatively describe the geometry of biofilm internal pore networks at length scales relevant to engineered biofilm reactors and suggest that internal pore structures provide crucial habitat for nitrifier growth.

Interplay between flow and bioturbation enhances metal efflux from low-permeability sediments.

Understanding the interplay effects between processes such as hydrodynamic forcing, sediment resuspension, and bioturbation is key to assessment of contaminated sediments. In the current study, effects of hydrodynamic forcing, sediment resuspension, and bioturbation by the marine polychaete Nereis virens were evaluated both independently and together in a six-month flume experiment. The results show that hydrodynamic forcing without resuspension or worm action slightly enhanced efflux of dissolved Cu to the water column, sediment resuspension released considerable amounts of dissolved Cu, and interactions between hydrodynamics and worm burrowing further enhanced Cu efflux. In non-bioturbated sediments, fine particles were only resuspended to the overlying water under the highest imposed shear stress, 0.58Pa. However, bioturbated sediments were resuspended under all shear stresses tested (0.11-0.58Pa), indicating that bioturbation destabilized the sediment bed. Further, increases in fluid shear following bioturbation caused rapid releases of dissolved Cu to the overlying water within a few hours. Cu efflux under fluid shears of 0.47Pa and 0.58Pa were 360× and 15× greater after the introduction of worms compared with the same flow conditions without their presence. Overall, our results indicate that the release of metals from low-permeability sediments is greatly enhanced by interactions between flow and bioturbation.

Comparison of biofilm cell quantification methods for drinking water distribution systems.

Drinking water quality typically degrades after treatment during conveyance through the distribution system. Potential causes include biofilm growth in distribution pipes which may result in pathogen retention, inhibited disinfectant diffusion, and proliferation of bad tastes and odors. However, there is no standard method for direct measurement of biofilms or quantification of biofilm cells in drinking water distribution systems. Three methods are compared here for quantification of biofilm cells grown in pipe loops samplers: biofilm heterotrophic plate count (HPC), biofilm biovolume by confocal laser scanning microscopy (CLSM) and biofilm total cell count by flow cytometry (FCM) paired with Syto 9. Both biofilm biovolume by CLSM and biofilm total cell count by FCM were evaluated for quantification of the whole biofilms (including non-viable cells and viable but not culturable cells). Signal-to-background ratios and overall performance of biofilm biovolume by CLSM and biofilm total cell count by FCM were found to vary with the pipe material. Biofilm total cell count by FCM had a low signal-to-background ratio on all materials, indicating that further development is recommended before application in drinking water environments. Biofilm biovolume by CLSM showed the highest signal-to-background ratio for cement and cast iron, which suggests promise for wider application in full-scale systems. Biofilm biovolume by CLSM and Syto 9 staining allowed in-situ biofilm cell quantification thus elimination variable associated with cell detachment for quantification but had limitations associated with non-specific staining of cement and, to a lesser degree, auto-fluorescence of both cement and polyvinyl chloride materials. Due to variability in results obtained from each method, multiple methods are recommended to assess biofilm growth in drinking water distribution systems. Of the methods investigated here, HPC and CLSM and recommended for further development towards application in full-scale systems. HPC is a sample and widely applied method that quantifies viable culturable cells. CLSM analysis allows the elimination of experimental variables associated with cell detachment and affords the opportunity to evaluate biofilm components such as extracellular polymeric substances through the addition of specific probes. These two methods can be applied together to assess biofilms known to degrade treated water quality during conveyance in full-scale drinking water treatment systems. The significance of improved biofilm assessment methods for drinking water distribution systems lies in advancing understanding of biofilm growth and control mechanisms that may lead to improved water quality during conveyance and at the tap for greater public health protection.

A Process-Based Model for Bioturbation-Induced Mixing.

Bioturbation refers to the transport processes carried out by living organisms and their physical effects on soils and sediments. It is widely recognized as an important mixing mechanism, particularly at the sediment-water interface in many natural systems. In order to quantify its impact on mixing, we propose a process-based model based on simple assumptions about organism burrowing behavior. Specifically, we consider burrowing events to be stochastic but memoryless, leading to exponential inter-burrow waiting times and depths. We then explore the impact of two different transport mechanisms on the vertical concentration distributions predicted by the model for a conservative (inert) tracer. We compare the results of our model to experimental data from a recent laboratory study of bioturbation by the freshwater oligochaete worm Lumbriculus variegatus, and find good quantitative agreement.

Microbial diversity in an intensively managed landscape is structured by landscape connectivity.

Intensively managed land increases the rate of nutrient and particle transport within a basin, but the impact of these changes on microbial community assembly patterns at the basin scale is not yet understood. The objective of this study was to investigate how landscape connectivity and dispersal impacts microbial diversity in an agricultural-dominated watershed. We characterized soil, sediment and water microbial communities along the Upper Sangamon River basin in Illinois-a 3600 km2 watershed strongly influenced by human activity, especially landscape modification and extensive fertilization for agriculture. We employed statistical and network analyses to reveal the microbial community structure and interactions in the critical zone (water, soil and sediment media). Using a Bayesian source tracking approach, we predicted microbial community connectivity within and between the environments. We identified strong connectivity within environments (up to 85.4 ± 13.3% of sequences in downstream water samples sourced from upstream samples, and 44.7 ± 26.6% in soil and sediment samples), but negligible connectivity across environments, which indicates that microbial dispersal was successful within but not between environments. Species sorting based on sample media type and environmental parameters was the dominant driver of community dissimilarity. Finally, we constructed operational taxonomic unit association networks for each environment and identified a number of co-occurrence relationships that were shared between habitats, suggesting that these are likely to be ecologically significant.

Nitrosomonas europaea biofilm formation is enhanced by Pseudomonas aeruginosa.

Biofilms are useful in biotechnology applications such as wastewater treatment, where aggregation of cells on surfaces can increase retention of slow-growing organisms such as ammonia-oxidizing bacteria (AOB). The formation and morphological development of polymicrobial biofilms including AOB are not thoroughly understood. Here, we investigated the formation of Nitrosomonas europaea AOB biofilms in flow cell systems. Nitrosomonas europaea developed substantially greater biovolume in co-culture with heterotrophic Pseudomonas aeruginosa than when cultured alone. In single-species biofilms, N. europaea formed thin, dispersed layers of cells. Contrastingly, when N. europaea was added to flow cells containing pre-established P. aeruginosa biofilms, N. europaea associated closely with P. aeruginosa, resulting in dual-species clusters with greater quantities of N. europaea. These results indicate that P. aeruginosa enhances the formation of N. europaea in biofilms. This favorable association of N. europaea with heterotrophic biofilms is expected to facilitate development of improved strategies for retention of N. europaea and other slow-growing AOB in engineered bioreactors.