Sources and Drivers of ARGs in Urban Streams in Atlanta, Georgia, USA.

The spread of antibiotic resistance genes (ARGs) in the aquatic environment is an emerging concern in the interest of protecting public health. Stemming the environmental dissemination of ARGs will require a better understanding of the sources and drivers of ARGs in the water environment. In this study, we used direct measurement of sewage-associated molecular markers, the class 1 integron gene, standard water quality parameters, and watershed characteristics to evaluate the sources and drivers of ARGs in an urban watershed impacted by a gradient of human activities. Quantitative polymerase chain reaction (qPCR) was used to quantify the abundance of the sewage-associated HF183, the E. coli fecal indicator, class 1 integron gene (int1), and the ARGs sulI, sulII, tetW, tetM, ampC, and blaSHV in stream water samples collected from the Proctor Creek watershed in Atlanta, Georgia. Our findings show that ARGs were widely distributed, with detection frequencies of 96% (sulI and sulII), 82% (tetW and tetM), and 49% (ampC and blaSHV). All the ARGs were positively and significantly correlated (r > 0.5) with the HF183 and E. coli markers. Non-linear machine learning models developed using generalized boosting show that more than 70% of the variation in ARG loads in the watershed could be explained by fecal source loading, with other factors such as class 1 integron, which is associated with acquired antibiotic resistance, and environmental factors contributing < 30% to ARG variation. These results suggest that input from fecal sources is a more critical driver of ARG dissemination than environmental stressors or horizontal gene transfer in aquatic environments highly impacted by anthropogenic pollution. Finally, our results provide local watershed managers and stakeholders with information to mitigate the burden of ARGs and fecal bacteria in urban streams.

Bacterial and viral fecal indicator predictive modeling at three Great Lakes recreational beach sites.

Coliphage are viruses that infect Escherichia coli (E. coli) and may indicate the presence of enteric viral pathogens in recreational waters. There is an increasing interest in using these viruses for water quality monitoring and forecasting; however, the ability to use statistical models to predict the concentrations of coliphage, as often done for cultured fecal indicator bacteria (FIB) such as enterococci and E. coli, has not been widely assessed. The same can be said for FIB genetic markers measured using quantitative polymerase chain reaction (qPCR) methods. Here we institute least-angle regression (LARS) modeling of previously published concentrations of cultured FIB (E. coli, enterococci) and coliphage (F+, somatic), along with newly reported genetic concentrations measured via qPCR for E. coli, enterococci, and general Bacteroidales. We develop site-specific models from measures taken at three beach sites on the Great Lakes (Grant Park, South Milwaukee, WI; Edgewater Beach, Cleveland, OH; Washington Park, Michigan City, IN) to investigate the efficacy of a statistical predictive modeling approach. Microbial indicator concentrations were measured in composite water samples collected five days per week over a beach season (∼15 weeks). Model predictive performance (cross-validated standardized root mean squared error of prediction [SRMSEP] and R2PRED) were examined for seven microbial indicators (using log10 concentrations) and water/beach parameters collected concurrently with water samples. Highest predictive performance was seen for qPCR-based enterococci and Bacteroidales models, with F+ coliphage consistently yielding poor performing models. Influential covariates varied by microbial indicator and site. Antecedent rainfall, bird abundance, wave height, and wind speed/direction were most influential across all models. Findings suggest that some fecal indicators may be more suitable for water quality forecasting than others at Great Lakes beaches.

Microbial source tracking (MST) in Chattahoochee River National Recreation Area: Seasonal and precipitation trends in MST marker concentrations, and associations with E. coli levels, pathogenic marker presence, and land use.

Escherichia coli levels in recreational waters are often used to predict when fecal-associated pathogen levels are a human health risk. The reach of the Chattahoochee River that flows through the Chattahoochee River National Recreation Area (CRNRA), located in the Atlanta-metropolitan area, is a popular recreation area that frequently exceeds the U.S. Environmental Protection Agency beach action value (BAV) for E. coli. A BacteriALERT program has been implemented to provide real-time E. coli estimates in the reach and notify the public of potentially harmful levels of fecal-associated pathogens as indicated by surrogate models based on real-time turbidity measurements from continuous water quality monitoring stations. However, E. coli does not provide information about the sources of fecal contamination and its accuracy as a human health indicator is questionable when sources of contamination are non-human. The objectives of our study were to investigate, within the Park and surrounding watersheds, seasonal and precipitation-related patterns in microbial source tracking marker concentrations of possible sources (human, dog, and ruminant), assess correlations between source contamination levels and culturable E. coli levels, determine which sources best explained model-based E. coli estimates above the BAV and detection of esp2 (a marker for the esp gene associated with pathogenic strains of Enterococcus faecium and Enterococcus faecalis), and investigate associations between source contamination levels and land use features. Three BacteriALERT sites on the Chattahoochee River were sampled six times per season in the winter and summer from December 2015 through September 2017, and 11 additional stream sites (synoptic sites) from the CRNRA watershed were sampled once per season. Samples were screened with microbial source tracking (MST) quantitative PCR (qPCR) markers for humans (HF183 Taqman), dogs (DogBact), and ruminants (Rum2Bac), the esp2 qPCR marker, and culturable E. coli. At the BacteriALERT sites, HF183 Taqman concentrations were higher under wet conditions DogBact concentrations were greater in the winter and under wet conditions, and Rum2Bac concentrations were comparatively low throughout the study with no difference across seasons or precipitation conditions. Concentrations of HF183 Taqman, DogBact, and Rum2Bac were positively correlated with culturable E. coli concentrations; however, DogBact had the largest R2 value among the three markers, and the forward stepwise regression indicated it was the best predictor of culturable E. coli concentrations at the BacteriALERT sites. Recursive partitioning indicated that BAV exceedances of model-based E. coli estimates were best explained by DogBact concentrations ≥3 gene copies per mL (CN/mL). Detections of esp2 at BacteriALERT sites were best explained by DogBact concentrations ≥11 CN/mL, while detections of esp2 at synoptic sites were best explained by HF183 Taqman ≥29 CN/mL. At the synoptic sites, HF183 Taqman levels were associated with wastewater treatment plant density. However, this relationship was driven primarily by a single site, suggesting possible conveyance issues in that catchment. esp2 detections at synoptic sites were positively associated with development within a 2-km radius and negatively associated with development within the catchment, suggesting multiple sources of esp2 in the watershed. DogBact and Rum2Bac were not associated with the land use features included in our analyses. Implications for Park management include: 1) fecal contamination levels were highest during wet conditions and in the off season when fewer visitors are expected to be participating in water-based recreation, 2) dogs are likely contributors to fecal contamination in the CRNRA and may be sources of pathogenic bacteria indicating further investigation of the origins of this contamination may be warranted as would be research to understand the human health risks from exposure to dog fecal contamination, and 3) high levels of the human marker at one site in the CRNRA watershed suggests more extensive monitoring in that catchment may locate the origin of human fecal contamination detected during this study.

PiSCES: Pi(scine) stream community estimation system.

The Piscine Stream Community Estimation System (PiSCES) provides users with a hypothesized fish community for any stream reach in the conterminous United States using information obtained from Nature Serve, the US Geological Survey (USGS), StreamCat, and the Peterson Field Guide to Freshwater Fishes of North America for over 1000 native and non-native freshwater fish species. PiSCES can filter HUC8-based fish assemblages based on species-specific occurrence models; create a community abundance/biomass distribution by relating relative abundance to mean body weight of each species; and allow users to query its database to see ancillary characteristics of each species (e.g., habitat preferences and maximum size). Future efforts will aim to improve the accuracy of the species distribution database and refine/augment increase the occurrence models. The PiSCES tool is accessible at the EPA's Quantitative Environmental Domain (QED) website at https://qed.epacdx.net/pisces/.

Determining global background soil PFAS loads and the fluorotelomer-based polymer degradation rates that can account for these loads.

In recent years, fluorotelomer-based polymers (FTPs) have been the dominant product of the fluorotelomer industry. For the last decade, whether FTPs degrade to toxic perfluorocarboxylates (PFCAs) has been vigorously contested, with early studies arguing that FTPs have half-lives >1000 years, and others concluding decadal half-lives. Given this FTP half-life discrepancy of 10- to >100-fold, here we investigate whether environmental loads of long-chain PFCAs might offer an independent approach to assess FTP half-lives. Specifically we: i) use surface soil-PFCA data to estimate terrestrial surface-soil background PFCA concentrations and loads; ii) extrapolate these data to generate global PFCA load estimates; iii) compare these estimates to published ocean-derived and industrial-emissions load estimates, finding agreement for perfluorooctanoate (C8), but an excess in longer-chain (C10,C12) PFCAs for ocean- and soil-derived loads relative to emissions; iv) model FTP degradation rates required to reconcile this discrepancy; and iv) compare our modeled estimates to existing experimental results. These findings show agreement for FTP half-lives at the decades-scale supporting existing laboratory studies that report decade-scale half-lives for FTPs. This suggests that global long-chain PFCA loads will increase for decades if legacy FTPs already manufactured are not contained upon disposal. These results suggest that FTPs comprised of novel poly- and perfluorinated alkyl substances (PFASs) now in production might constitute considerable sources to the environment of the new generation of PFASs.

Comparison of somatic and F+ coliphage enumeration methods with large volume surface water samples.

Coliphages are alternative fecal indicators that may be suitable surrogates for viral pathogens, but majority of standard detection methods utilize insufficient volumes for routine detection in environmental waters. We compared three somatic and F+ coliphage methods based on a paired measurement from 1 L samples collected from the Great Lakes (n = 74). Methods include: 1) dead-end hollow fiber ultrafilter with single agar layer (D-HFUF-SAL); 2) modified SAL (M-SAL); and 3) direct membrane filtration (DMF) technique. Overall, D-HFUF-SAL outperformed other methods as it yielded the lowest frequency of non-detects [(ND); 10.8%] and the highest average concentrations of recovered coliphage for positive samples (2.51 ± 1.02 [standard deviation, SD] log10 plaque forming unit/liter (PFU/L) and 0.79 ± 0.71 (SD) log10 PFU/L for somatic and F+, respectively). M-SAL yielded 29.7% ND and average concentrations of 2.26 ± 1.15 (SD) log10 PFU/L (somatic) and 0.59 ± 0.82 (SD) log10 PFU/L (F+). DMF performance was inferior to D-HFUF-SAL and M-SAL methods (ND of 65.6%; average somatic coliphage concentration 1.52 ± 1.32 [SD] log10 PFU/L, no F+ detected), indicating this procedure is unsuitable for 1 L surface water sample volumes. This study represents an important step toward the use of a coliphage method for recreational water quality criteria purposes.

Incidence of somatic and F+ coliphage in Great Lake Basin recreational waters.

There is a growing interest for the use of coliphage as an alternative indicator to assess fecal pollution in recreational waters. Coliphage are a group of viruses that infect Escherichia coli and are considered as potential surrogates to infer the likely presence of enteric viral pathogens. We report the use of a dead-end hollow fiber ultrafiltration single agar layer method to enumerate F+ and somatic coliphage from surface waters collected from three Great Lake areas. At each location, three sites (two beaches; one river) were sampled five days a week over the 2015 beach season (n = 609 total samples). In addition, culturable E. coli and enterococci concentrations, as well as 16 water quality and recreational area parameters were assessed such as rainfall, turbidity, dissolved oxygen, pH, and ultra violet absorbance. Overall, somatic coliphage levels ranged from non-detectable to 4.39 log10 plaque forming units per liter and were consistently higher compared to F+ (non-detectable to 3.15 log10 PFU/L), regardless of sampling site. Coliphage concentrations weakly correlated with cultivated fecal indicator bacteria levels (E. coli and enterococci) at 75% of beach sites tested in study (r = 0.28 to 0.40). In addition, ultraviolet light absorption and water temperature were closely associated with coliphage concentrations, but not fecal indicator bacteria levels suggesting different persistence trends in Great Lake waters between indicator types (bacteria versus virus). Finally, implications for coliphage water quality management and future research directions are discussed.

An Integrated Ecological Modeling System for Assessing Impacts of Multiple Stressors on Stream and Riverine Ecosystem Services within River Basins.

We demonstrate a novel, spatially explicit assessment of the current condition of aquatic ecosystem services, with limited sensitivity analysis for the atmospheric contaminant mercury. The Integrated Ecological Modeling System (IEMS) forecasts water quality and quantity, habitat suitability for aquatic biota, fish biomasses, population densities, productivities, and contamination by methylmercury across headwater watersheds. We applied this IEMS to the Coal River Basin (CRB), West Virginia (USA), an 8-digit hydrologic unit watershed, by simulating a network of 97 stream segments using the SWAT watershed model, a watershed mercury loading model, the WASP water quality model, the PiSCES fish community estimation model, a fish habitat suitability model, the BASS fish community and bioaccumulation model, and an ecoservices post-processer. Model application was facilitated by automated data retrieval and model setup and updated model wrappers and interfaces for data transfers between these models from a prior study. This companion study evaluates baseline predictions of ecoservices provided for 1990 - 2010 for the population of streams in the CRB and serves as a foundation for future model development.

Spatiotemporal characteristics of organic contaminant concentrations and ecological risk assessment in the Songhua River, China.

To control source pollution and improve water quality, an understanding of the spatiotemporal characteristics of organic contaminant concentrations in affected receiving waters is necessary. The Songhua River in northeast China is the country's third-largest domestic river and loadings of organic contaminants along an industrialized section have made it the focal point of a national pollution reduction plan. In addition to water quality issues, management of the Songhua River basin must also address local economic development, aquatic ecosystem sustainability and political relationships with Russia. In three periods spanning 2006 to 2010, eight polycyclic aromatic hydrocarbons (PAHs) and eight phenols were measured in surface waters at ten monitoring sites along the river. A generalized linear model (GLM) was used to characterize water quality at different sites and time periods. Chemical concentrations of the organic compounds showed significant sinusoidal seasonal patterns and the concentrations declined significantly from 2006 to 2010, possibly due to management practices designed to control water pollution. A critical body residue analysis showed that water concentrations measured during the winter of 2007 across all monitoring sites, but especially at S1-Shaokou and S2-Songhuajiangcun, presented a high risk for fish species. The spatiotemporal characteristics of water quality and estimated ecological risks shown here add to the body of knowledge to develop policies on industrial output and pollution management strategies for the Songhua River basin.

Pesticide Uptake Across the Amphibian Dermis Through Soil and Overspray Exposures.

For terrestrial amphibians, accumulation of pesticides through dermal contact is a primary route of exposure in agricultural landscapes and may be contributing to widespread amphibian declines. To show pesticide transfer across the amphibian dermis at permitted label application rates, our study was designed to measure pesticide body burdens after two simulated exposure scenarios. We compared direct exposures, where amphibians were present when spraying occurred, to indirect exposures, where amphibians were exposed to soils after pesticide application. During summer 2012, we reared barking (Hyla gratiosa) and green treefrogs (H. cinerea) through 60-90 days post-metamorphosis at a United States Environmental Protection Agency research laboratory. We tested exposure for 8 h to five pesticide active ingredients (imidacloprid, atrazine, triadimefon, fipronil, or pendimethalin) in glass aquaria lined with soil in the laboratory. We quantified total pesticide body burden and soil concentrations using liquid chromatography-mass spectrometry. All individuals in both treatments had measurable body burdens at the end of the study. A randomized block design analysis of variance (n = 18) showed that body burdens (p = 0.03) and bioconcentration factors (BCFs) (p = 0.01) were significantly greater in the direct overspray treatment relative to the indirect soil spray treatment for both species and tested pesticides. BCFs ranged from 0.1 to 1.16 and from 0.013 to 0.78 in the direct and indirect treatments, respectively. Our study shows dermal uptake for multiple pesticides from both direct spray and indirect soil exposures and provides empirical support for the degree to which terrestrial phase amphibians have higher body burdens after overspray pesticide exposure.

Estimating terrestrial amphibian pesticide body burden through dermal exposure.

Dermal exposure presents a potentially significant but understudied route for pesticide uptake in terrestrial amphibians. Our study measured dermal uptake of pesticides of varying hydrophobicity (logKow) in frogs. Amphibians were indirectly exposed to one of five pesticide active ingredients through contact with contaminated soil: imidacloprid (logKow = 0.57), atrazine (logKow = 2.5), triadimefon (logKow = 3.0), fipronil (logKow = 4.11) or pendimethalin (logKow = 5.18). All amphibians had measurable body burdens at the end of the exposure in concentrations ranging from 0.019 to 14.562 µg/g across the pesticides tested. Atrazine produced the greatest body burdens and bioconcentration factors, but fipronil was more permeable to amphibian skin when application rate was considered. Soil partition coefficient and water solubility were much better predictors of pesticide body burden, bioconcentration factor, and skin permeability than logKow. Dermal uptake data can be used to improve risk estimates of pesticide exposure among amphibians as non-target organisms.

Factors affecting the presence of human-associated and fecal indicator real-time quantitative PCR genetic markers in urban-impacted recreational beaches.

Urban runoff can carry a variety of pollutants into recreational beaches, often including bacterial pathogens and indicators of fecal contamination. To develop complete recreational criteria and risk assessments, it is necessary to understand conditions under which human contamination could be present at beaches solely impacted by urban runoff. Accurately estimating risk requires understanding sources, concentrations, and transport mechanisms of microbial contaminants in these environments. By applying microbial source tracking methods and empirical modeling, we assessed the presence and level of human contamination at urban runoff impacted recreational beaches. We also identified environmental parameters and pollution sources that can influence the concentration and transport of culturable and molecular fecal indicator bacteria (FIB) in systems impacted solely by urban runoff. Water samples and physico-chemical parameters were collected from shoreline locations from three South Carolina (SC) beaches (five locations per beach) and two Florida (FL) beaches (three locations per beach). Each SC beach was directly impacted by swashes or tidal creeks receiving stormwater runoff from the urbanized area and therefore were designated as swash drain associated (SDA) beaches, while FL beaches were designated as non-swash drain associated (NSDA). Sampling in swash drains (SD; three sites per SD) directly impacting each SC beach was also conducted. Results indicate that although culturable (enterococci) and real-time quantitative polymerase chain reaction (qPCR) (EC23S857, Entero1, and GenBac3) FIB concentrations were, on average, higher at SD locations, SDA beaches did not have consistently higher molecular FIB signals compared to NSDA beaches. Both human-associated markers (HF183 and HumM2) were concomitantly found only at SDA beaches. Bacteroidales species-specific qPCR markers (BsteriF1 and BuniF2) identified differences in the Bacteroidales community, depending on beach type. The marker for general Bacteroidales was most abundant at SD locations and exhibited a high correlation with both culturable and other molecular markers. Combining molecular information with predictive modeling allowed us to identify both alongshore movement of currents and SD outflow as significant influences on the concentration of molecular and culturable indicators in the bathing zone. Data also suggests that combining methodologies is a useful and cost effective approach to help understand transport dynamics of fecal contamination and identify potential sources of contamination at marine beaches.

Relation of fish and shellfish distributions to habitat and water quality in the Mobile Bay estuary, USA.

The Mobile Bay estuary in the northern Gulf of Mexico provides a rich habitat for many fish and shellfish, including those identified as economically and ecologically important. The National Estuary Program in Mobile Bay has focused on restoration of degraded estuarine habitat on which these species depend. To support this effort, we used statistical techniques of ordination, cluster analysis, and discriminant analysis to relate distributions of individual fish and shellfish species and species assemblages to two dozen water quality and habitat variables in a geo-referenced database. Species appeared to respond to dominant gradients of low to high salinity and upland to offshore habitat area; many of the 15 communities identified via cluster analysis showed aggregated spatial distributions that could be related to habitat characteristics. Species in the Mobile River Delta were distinct from those in other areas of the estuary. This analysis supports habitat management in the Mobile Bay estuary; however, due to mobility of organisms among sampling locations and the dynamic environmental conditions in estuaries, we conclude that the analyses presented here are most appropriate for an evaluation of the estuary as a whole.