Wastewater surveillance for bacterial targets: current challenges and future goals.

Wastewater-based epidemiology (WBE) expanded rapidly in response to the COVID-19 pandemic. As the public health emergency has ended, researchers and practitioners are looking to shift the focus of existing wastewater surveillance programs to other targets, including bacteria. Bacterial targets may pose some unique challenges for WBE applications. To explore the current state of the field, the National Science Foundation-funded Research Coordination Network (RCN) on Wastewater Based Epidemiology for SARS-CoV-2 and Emerging Public Health Threats held a workshop in April 2023 to discuss the challenges and needs for wastewater bacterial surveillance. The targets and methods used in existing programs were diverse, with twelve different targets and nine different methods listed. Discussions during the workshop highlighted the challenges in adapting existing programs and identified research gaps in four key areas: choosing new targets, relating bacterial wastewater data to human disease incidence and prevalence, developing methods, and normalizing results. To help with these challenges and research gaps, the authors identified steps the larger community can take to improve bacteria wastewater surveillance. This includes developing data reporting standards and method optimization and validation for bacterial programs. Additionally, more work is needed to understand shedding patterns for potential bacterial targets to better relate wastewater data to human infections. Wastewater surveillance for bacteria can help provide insight into the underlying prevalence in communities, but much work is needed to establish these methods.IMPORTANCEWastewater surveillance was a useful tool to elucidate the burden and spread of SARS-CoV-2 during the pandemic. Public health officials and researchers are interested in expanding these surveillance programs to include bacterial targets, but many questions remain. The NSF-funded Research Coordination Network for Wastewater Surveillance of SARS-CoV-2 and Emerging Public Health Threats held a workshop to identify barriers and research gaps to implementing bacterial wastewater surveillance programs.

The effect of Paenibacillus on IDEXX Enterolert results from freshwater stream environments.

Enterolert, a fluorogenic substrate test, is used as a quantitative method for determining freshwater concentrations of Enterococcus for water quality indicators. However, there is some evidence from recent studies suggesting that Enterolert may not suppress false positives due to pollution sources in waterbodies. In this study, we evaluated this method by analyzing field water and sediment samples from four freshwater streams. We also performed a laboratory microcosm study from two of the stream sediments. The Enterolert method was investigated by phenotypic and genomic analyses for accuracy of isolating and quantifying Enterococcus and/or Streptococcus. Additionally, we tested isolates from Enterolert panels for antibiotic resistance. Results from the field and microcosm studies from initial to final time points indicated that false positives were predominantly Paenibacillus spp. and other non-fecal indicator bacteria. Furthermore, the microcosm study indicated shifts from lactic acid to non-lactic acid bacteria between initial to final time points, but Enterococcus concentrations from Enterolert panels remained stable for the duration of the study for both stream sediments. Antibiotic resistance indicated no distinct pattern of resistance or susceptibility to a suite of antibiotics. However, all isolates tested were resistant to bacitracin and nalidixic acid. In conclusion, we found that Enterolert was not exclusively selective for Enterococcus from freshwater environments and that sediment and polluted waterbodies have the potential to skew the presumed concentrations. More research is needed to evaluate the effectiveness and selectivity of the medium used for the fluorogenic substrate test for Enterococcus enumeration.

Field studies of pollutant removal from nursery and greenhouse runoff by constructed wetlands.

Plant nursery runoff commonly contains pesticides and nutrients that often threaten aquatic ecosystems. Constructed wetlands could be a tool to remove pesticides and nutrients from nursery runoff but have not been extensively studied in this setting. Two field-scale constructed wetlands (one subsurface-flow constructed wetland [SFCW] and one free-surface constructed wetland [FSCW]) were implemented and monitored for water quality improvement. The SFCW demonstrated significant mass reduction of 78% or greater for nitrate, orthophosphate, total nitrogen, total phosphorus, and total suspended solids. The SFCW also demonstrated significant mass reduction of 79% or greater for 10 of the 12 pesticide compounds detected in over half of the collected samples. The FSCW demonstrated significant mass reduction of 46% or greater for all nonpesticide analytes except total nitrogen. Loading rate and actual storage volume compared with inflow volume likely affected performance. Reduced size and increased loading rate of the FSCW likely reduced its ability to effectively reduce pesticides. Results from this study indicate that constructed wetlands are likely an effective tool for nursery runoff management. When designing and implementing constructed wetlands, it is important for practitioners to consider the tradeoff between system size (additional cost and land otherwise dedicated to production) and performance.

Urban stormwater characterization, control, and treatment.

This review summarizes over 280 studies published in 2019 related to the characterization, control, and management of urban stormwater runoff. A summary of quantity and quality concerns is provided in the first section of the review, serving as the foundation for the following sections which focus on the control and treatment of stormwater runoff. Finally, the impact of stormwater control devices at the watershed scale is discussed. Each section provides a self-contained overview of the 2019 literature, common themes, and future work. Several themes emerged from the 2019 literature including exploration of substrate amendments for improved water quality effluent from stormwater controls, the continued study of the role of vegetation in green infrastructure practices, and a call to action for the development of new models which generate reliable, computationally efficient results under the physical, chemical, biological, and social complexity of stormwater management. PRACTITIONER POINTS: Over 280 studies were published in 2019 related to the characterization, control, and treatment of urban stormwater. Studies on bioretention and general stormwater characteristics represented the two most common subtopics in 2019. Trends in 2019 included novel substrate amendments, studies on the role of vegetation, and advancements in computational models.

Urban stormwater characterization, control, and treatment.

This review summarizes over 250 studies published in 2018 related to the characterization, control, and management of urban stormwater runoff. The review covers three broad themes: (a) quantity and quality characterization of stormwater, (b) control and treatment of stormwater runoff, and (c) implementation and assessment of watershed-scale green stormwater infrastructure (GSI). Each section provides an overview of the 2018 literature, common themes, and future work. Several themes emerged from the 2018 literature including exploration of contaminants of emerging concern within stormwater systems, characterization and incorporation of vegetation-driven dynamics in stormwater control measures, and the need for interdisciplinary perspectives on the implementation and assessment of GSI. PRACTITIONER POINTS: Over 250 studies were published in 2018 related to the characterization, control, and treatment of stormwater. Studies cover general stormwater characteristics, control and treatment systems, and watershed-scale assessments. Trends in 2018 include treatment trains, vegetation dynamics, and interdisciplinary perspectives.

Urban Stormwater Characterization, Control and Treatment.

A review of literature related to stormwater runoff characterization and its subsequent management and treatment from 2017 was conducted. The 250 articles summarized herein are organized along three central themes: (1) stormwater quality and quantity characteristics, (2) site-scale stormwater management practices, with a focus on low impact development (LID) and green infrastructure (GI), and (3) watershed-scale performance of stormwater controls. Within each section, common research themes and future work are highlighted.

Urban Stormwater Characterization, Control, and Treatment.

A summary of 246 studies published in 2016 on topics related to the characterization and management of urban stormwater runoff is presented in the following review. The review is structured along three major topical areas: (1) general characterization of stormwater quantity and quality; (2) engineered systems for stormwater control and treatment, including erosion and sediment control practices, constructed stormwater ponds and wetlands, bioretention, permeable pavement, greenroofs, and rainwater harvesting and (3) watershedscale application of stormwater treatment and control practices. Common research themes and needs highlighted throughout this review include efforts to better understand stormwater transport and treatment mechanisms and their representation in models, advancements to optimize the design of stormwater control measures to meet specific hydrologic and/or water quality targets, and increasing understanding of the biophysical and social factors that influence watershed-scale implementation of low impact development and other stormwater control measures.

Urban Stormwater Characterization, Control, and Treatment.

The following review presents a synthesis of 181 journal articles published during 2015 that represent progress toward better characterizing, controlling, and treating urban stormwater runoff. The review is structured by general topical areas related to (1) stormwater quality and quantity characterization; (2) engineered stormwater control and treatment practices, including erosion and sediment control, stormwater ponds, constructed stormwater wetlands, bioretention, permeable pavement, greenroofs, and rainwater harvesting systems; and (3) watershed-scale modeling and optimization of stormwater control and treatment practices. Common research themes emerging from this collection of studies include potential to enhance hydrologic and pollutant treatment performance of stormwater practices via media amendments and the use of innovative outlet control structures, as well as development of a more mechanistic understanding of hydrologic and water-quality functions to inform modeling and performance predictions. These studies serve to expand the field's knowledge base and will inform future efforts to further improve stormwater control and treatment at various spatial and temporal scales.

Critical Review of Technical Questions Facing Low Impact Development and Green Infrastructure: A Perspective from the Great Plains.

Since its inception, Low Impact Development (LID) has become part of urban stormwater management across the United States, marking progress in the gradual transition from centralized to distributed runoff management infrastructure. The ultimate goal of LID is full, cost-effective implementation to maximize watershed-scale ecosystem services and enhance resilience. To reach that goal in the Great Plains, the multi-disciplinary author team presents this critical review based on thirteen technical questions within the context of regional climate and socioeconomics across increasing complexities in scale and function. Although some progress has been made, much remains to be done including continued basic and applied research, development of local LID design specifications, local demonstrations, and identifying funding mechanisms for these solutions. Within the Great Plains and beyond, by addressing these technical questions within a local context, the goal of widespread acceptance of LID can be achieved, resulting in more effective and resilient stormwater management.

Investigation of insecticide leaching from potted nursery stock and aquatic health benefits of bioretention cells receiving nursery runoff.

Tree nurseries and greenhouses within the USDA red imported fire ant (RIFA) quarantine zone are required to incorporate insecticides into their potting media to prevent artificial spread of RIFA. Bifenthrin and fipronil are two common insecticides that are incorporated into potting media. During irrigation and stormwater events, there is potential for insecticides to leach from nursery pots, resulting in the contamination of nearby surface waters. In this study, occurrences of insecticides in simulated nursery runoff were compared with two irrigation strategies and two types of containers in single pot leaching and field runoff simulations. In addition, toxicity of pot leachate to the aquatic invertebrate, Hyallela azteca, was measured, and removal efficiencies of insecticides from bioretention cell media were evaluated. Overhead irrigation resulted in significantly higher concentrations than drip irrigation, and RootMaker pots allowed more leaching as compared to standard slick-wall pots. However, in all tests, the average concentration of bifenthrin during 15 days of leaching in both pot and field simulations was greater than 200 ng/L; more than 100-fold greater than the LC50 for H. azteca. Toxicity studies confirmed this level of toxicity. Higher amounts of compost, 20 and 40%, in bioretention cell media resulted in greater percent reduction of both bifenthrin and fipronil. This study determined that management techniques may be able to limit the amount of insecticide that leaches from pots and runs off to receiving water bodies. Specifically, the selection of appropriate pot types, irrigation strategies, or filtering runoff through bioretention cells may reduce contamination loads. Thus, further best management strategies such as the use of bioretention cells are needed in nursery and greenhouse facilities to prevent surface water runoff from transporting toxic insecticides.

Using heat to characterize streambed water flux variability in four stream reaches.

Estimates of streambed water flux are needed for the interpretation of streambed chemistry and reactions. Continuous temperature and head monitoring in stream reaches within four agricultural watersheds (Leary Weber Ditch, IN; Maple Creek, NE; DR2 Drain, WA; and Merced River, CA) allowed heat to be used as a tracer to study the temporal and spatial variability of fluxes through the streambed. Synoptic methods (seepage meter and differential discharge measurements) were compared with estimates obtained by using heat as a tracer. Water flux was estimated by modeling one-dimensional vertical flow of water and heat using the model VS2DH. Flux was influenced by physical heterogeneity of the stream channel and temporal variability in stream and ground-water levels. During most of the study period (April-December 2004), flux was upward through the streambeds. At the IN, NE, and CA sites, high-stage events resulted in rapid reversal of flow direction inducing short-term surface-water flow into the streambed. During late summer at the IN site, regional ground-water levels dropped, leading to surface-water loss to ground water that resulted in drying of the ditch. Synoptic measurements of flux generally supported the model flux estimates. Water flow through the streambed was roughly an order of magnitude larger in the humid basins (IN and NE) than in the arid basins (WA and CA). Downward flux, in response to sudden high streamflows, and seasonal variability in flux was most pronounced in the humid basins and in high conductivity zones in the streambed.

Transport and fate of nitrate at the ground-water/surface-water interface.

Although numerous studies of hyporheic exchange and denitrification have been conducted in pristine, high-gradient streams, few studies of this type have been conducted in nutrient-rich, low-gradient streams. This is a particularly important subject given the interest in nitrogen (N) inputs to the Gulf of Mexico and other eutrophic aquatic systems. A combination of hydrologic, mineralogical, chemical, dissolved gas, and isotopic data were used to determine the processes controlling transport and fate of NO(3)(-) in streambeds at five sites across the USA. Water samples were collected from streambeds at depths ranging from 0.3 to 3 m at three to five points across the stream and in two to five separate transects. Residence times of water ranging from 0.28 to 34.7 d m(-1) in the streambeds of N-rich watersheds played an important role in allowing denitrification to decrease NO(3)(-) concentrations. Where potential electron donors were limited and residence times were short, denitrification was limited. Consequently, in spite of reducing conditions at some sites, NO(3)(-) was transported into the stream. At two of the five study sites, NO(3)(-) in surface water infiltrated the streambeds and concentrations decreased, supporting current models that NO(3)(-) would be retained in N-rich streams. At the other three study sites, hydrogeologic controls limited or prevented infiltration of surface water into the streambed, and ground-water discharge contributed to NO(3)(-) loads. Our results also show that in these low hydrologic-gradient systems, storm and other high-flow events can be important factors for increasing surface-water movement into streambeds.

Pesticide fate and transport throughout unsaturated zones in five agricultural settings, USA.

Pesticide transport through the unsaturated zone is a function of chemical and soil characteristics, application, and water recharge rate. The fate and transport of 82 pesticides and degradates were investigated at five different agricultural sites. Atrazine and metolachlor, as well as several of the degradates of atrazine, metolachlor, acetochlor, and alachlor, were frequently detected in soil water during the 2004 growing season, and degradates were generally more abundant than parent compounds. Metolachlor and atrazine were applied at a Nebraska site the same year as sampling, and focused recharge coupled with the short time since application resulted in their movement in the unsaturated zone 9 m below the surface. At other sites where the herbicides were applied 1 to 2 yr before sampling, only degradates were found in soil water. Transformations of herbicides were evident with depth and during the 4-mo sampling time and reflected the faster degradation of metolachlor oxanilic acid and persistence of metolachor ethanesulfonic acid. The fraction of metolachlor ethanesulfonic acid relative to metolachlor and metolachlor oxanilic acid increased from 0.3 to >0.9 at a site in Maryland where the unsaturated zone was 5 m deep and from 0.3 to 0.5 at the shallowest depth. The flux of pesticide degradates from the deepest sites to the shallow ground water was greatest (3.0-4.9 micromol m(-2) yr(-1)) where upland recharge or focused flow moved the most water through the unsaturated zone. Flux estimates based on estimated recharge rates and measured concentrations were in agreement with fluxes estimated using an unsaturated-zone computer model (LEACHM).

Pesticides in rain in four agricultural watersheds in the United States.

Rainfall samples were collected during the 2003 and 2004 growing seasons at four agricultural locales across the USA in Maryland, Indiana, Nebraska, and California. The samples were analyzed for 21 insecticides, 18 herbicides, three fungicides, and 40 pesticide degradates. Data from all sites combined show that 7 of the 10 most frequently detected pesticides were herbicides, with atrazine (70%) and metolachlor (83%) detected at every site. Dacthal, acetochlor, simazine, alachlor, and pendimethalin were detected in more than 50% of the samples. Chlorpyrifos, carbaryl, and diazinon were the only insecticides among the 10 most frequently detected compounds. Of the remaining pesticide parent compounds, 18 were detected in fewer than 30% of the samples, and 13 were not detected. The most frequently detected degradates were deethylatrazine; the oxygen analogs (OAs) of the organophosphorus insecticides chlorpyrifos, diazinon, and malathion; and 1-napthol (degradate of carbaryl). Deethylatrazine was detected in nearly 70% of the samples collected in Maryland, Indiana, and Nebraska but was detected only once in California. The OAs of chlorpyrifos and diazinon were detected primarily in California. Degradates of the acetanilide herbicides were rarely detected in rain, indicating that they are not formed in the atmosphere or readily volatilized from soils. Herbicides accounted for 91 to 98% of the total pesticide mass deposited by rain except in California, where insecticides accounted for 61% in 2004. The mass of pesticides deposited by rainfall was estimated to be less than 2% of the total applied in these agricultural areas.

Identifying fecal sources in a selected catchment reach using multiple source-tracking tools.

Given known limitations of current microbial source-tracking (MST) tools, emphasis on small, simple study areas may enhance interpretations of fecal contamination sources in streams. In this study, three MST tools-Escherichia coli repetitive element polymerase chain reaction (rep-PCR), coliphage typing, and Bacteroidales 16S rDNA host-associated markers-were evaluated in a selected reach of Plum Creek in south-central Nebraska. Water-quality samples were collected from six sites. One reach was selected for MST evaluation based on observed patterns of E. coli contamination. Despite high E. coli concentrations, coliphages were detected only once among water samples, precluding their use as a MST tool in this setting. Rep-PCR classification of E. coli isolates from both water and sediment samples supported the hypothesis that cattle and wildlife were dominant sources of fecal contamination, with minor contributions by horses and humans. Conversely, neither ruminant nor human sources were detected by Bacteroidales markers in most water samples. In bed sediment, ruminant- and human-associated Bacteroidales markers were detected throughout the interval from 0 to 0.3 m, with detections independent of E. coli concentrations in the sediment. Although results by E. coli-based and Bacteroidales-based MST methods led to similar interpretations, detection of Bacteroidales markers in sediment more commonly than in water indicates that different tools to track fecal contamination (in this case, tools based on Bacteroidales DNA and E. coli isolates) may have varying relevance to the more specific goal of tracking the sources of E. coli in watersheds. This is the first report of simultaneous, toolbox approach application of a library-based and marker-based MST analyses to flowing surface water.

Assessment of fecal pollution sources in a small northern-plains watershed using PCR and phylogenetic analyses of Bacteroidetes 16S rRNA gene.

We evaluated the efficacy, sensitivity, host-specificity, and spatial/temporal dynamics of human- and ruminant-specific 16S rRNA gene Bacteroidetes markers used to assess the sources of fecal pollution in a fecally impacted watershed. Phylogenetic analyses of 1271 fecal and environmental 16S rRNA gene clones were also performed to study the diversity of Bacteroidetes in this watershed. The host-specific assays indicated that ruminant feces were present in 28-54% of the water samples and in all sampling seasons, with increasing frequency in downstream sites. The human-targeted assays indicated that only 3-5% of the water samples were positive for human fecal signals, although a higher percentage of human-associated signals (19-24%) were detected in sediment samples. Phylogenetic analysis indicated that 57% of all water clones clustered with yet-to-be-cultured Bacteroidetes species associated with sequences obtained from ruminant feces, further supporting the prevalence of ruminant contamination in this watershed. However, since several clusters contained sequences from multiple sources, future studies need to consider the potential cosmopolitan nature of these bacterial populations when assessing fecal pollution sources using Bacteroidetes markers. Moreover, additional data is needed in order to understand the distribution of Bacteroidetes host-specific markers and their relationship to water quality regulatory standards.