Using the fish plasma model to evaluate potential effects of pharmaceuticals in effluent from a large urban wastewater treatment plant.

Several pharmaceuticals and personal care products (PPCPs) were evaluated using the fish plasma model (FPM) for juvenile Chinook salmon exposed to effluent from a large urban wastewater treatment plant. The FPM compares fish plasma concentrations to therapeutic values determined in human plasma as an indication of potential adverse effects. We used human Cmax values, which are the maximum plasma concentration for a minimum therapeutic dose. Observed and predicted plasma concentrations from juvenile Chinook salmon exposed to a dilution series of whole wastewater effluent were compared to 1%Cmax values to determine Response Ratios (RR) ([plasma]/1%Cmax) for assessment of possible adverse effects. Several PPCPs were found to approach or exceed an RR of 1, indicating potential effects in fish. We also predicted plasma concentrations from measured water concentrations and determined that several of the values were close to or below the analytical reporting limit (RL) indicating potential plasma concentrations for a large number of PPCPs that were below detection. Additionally, the 1%Cmax was less than the RL for several analytes, which could impede predictions of possible effect concentrations. A comparison of observed and predicted plasma concentrations found that observed values were frequently much higher than values predicted with water concentrations, especially for low log10Dow compounds. The observed versus predicted values using the human volume of distribution (Vd), were generally much closer in agreement. These data appear to support the selection of whole-body concentrations to predict plasma values, which relies more on estimating simple partitioning within the fish instead of uptake via water. Overall, these observations highlight the frequently underestimated predicted plasma concentrations and potential to cause adverse effects in fish. Using measured plasma concentrations or predicted values from whole-body concentrations along with improved prediction models and reductions in analytical detection limits will foster more accurate risk assessments of pharmaceutical exposure for fish.

The screening and prioritization of contaminants of emerging concern in the marine environment based on multiple biological response measures.

There is ample evidence that a range of anthropogenic chemicals occur in the aquatic environment, some of which have the potential to cause harm. Contaminants of Emerging Concern (CECs) are a subset of anthropogenic compounds that are poorly characterized in terms of effects and occurrences, and that are generally unregulated. Due to the sheer number of chemicals used, it is necessary to identify and prioritize those that may cause biological impacts. A key challenge of doing so is the lack of traditional ecotoxicological information. The utilization of in vitro exposure-response studies or benchmarks based on in vivo data can provide a basis for developing threshold values for evaluation of potential impacts. There are challenges, including understanding the accuracy and range of application for modeled measures and translating in vitro response information from receptor models to apical endpoints. Despite this, the use of multiple lines of evidence increases the range of available information, and supports a weight-of-evidence approach to inform the screening and prioritization of CECs in the environment. The objective of this work is to perform an evaluation of CECs detected in an urban estuary, and to identify those that are most likely to elicit a biological response. Monitoring data from marine water, wastewater, and fish and shellfish tissue samples from 17 different campaigns combined with multiple biological response measures were compared with appropriate threshold values. CECs were categorized based on their potential to elicit a biological response; the uncertainty, based on consistency of lines of evidence, was also evaluated. Two-hundred-fifteen CECs were detected. Fifty-seven were identified as High Priority (likely to cause a biological effect), and 84 as Watch List (potential to cause biological effects). Due to the extent of the monitoring and range of the lines of evidence, this approach and results are applicable to other urbanized estuarine systems.

The influence of weevil herbivory on leaf litter chemistry in dioecious willows.

Leaf litter inputs can influence the structure and function of both terrestrial and adjacent aquatic ecosystems. Dioecy and herbivory are two factors that together have received little attention, yet have the potential to affect the quantity, quality, and timing of riparian litterfall, litter chemistry, and litter decomposition processes. Here, we explore litter chemistry differences for the dioecious Sitka willow (Salix sitchensis Sanson ex. Bong), which is establishing on primary successional habitats at Mount St. Helens (WA, USA) and is heavily infested with a stem-boring weevil (Cryptorhynchus lapathi). Weevil-attacked branches produced summer senesced litter that had significantly higher %N, lower C:N ratios, and lower condensed tannins than litter from branches that were unattacked by the weevil and senesced naturally in the autumn. Weevils more often attack female willows; however, these common litter chemicals did not significantly differ between males and females within the weevil-attacked and -unattacked groups. High-resolution mass spectrometry was used to isolate compounds in litter from 10 Sitka willow individuals with approximately 1500-1600 individual compounds isolated from each sample. There were differences between weevil-attacked litter and green leaf samples, but at this level, there was no clustering of male and female samples. However, further exploration of the isolated compounds determined a suite of compounds present only in either males or females. These findings suggest some variation in more complex litter chemistry between the sexes, and that significant differences in weevil-attacked litter chemistry, coupled with the shift in seasonality of litter inputs to streams, could significantly affect in-stream ecological processes, such as decomposition and detritivore activity.

Suspect and non-target screening of contaminants of emerging concern in streams in agricultural watersheds.

Surface water runoff is an important source of water contamination affecting nearby rivers and streams. Many rural creeks are documented habitats for important aquatic species and the focus of restoration activities. In this study, we collected creek water samples in watersheds with a range of commercial-to-agricultural land use during rain events, and applied suspect and non-target screening with high-resolution mass spectrometry (HRMS) to characterize the occurrence of contaminants of emerging concern (CECs). In total, 58 CECs were identified, and 36 of them were confirmed and semi-quantified with reference standards. Pesticides were detected in all land use, including urban/commercial areas. Some pesticides were observed at concentrations of >10,000 ng/L demonstrating the strong contamination input during rain events. Five pesticides (azoxystrobin, fludioxonil, 4-hydroxy-chlorothalonil, imidacloprid, 2-methyl-4-chlorophenoxyacetic acid) were prioritized based on their risk quotients. HRMS chemical profiles demonstrated the wide range of chemical exposures in a given stream system and that compounds associated with specific land uses occur across land uses. Temporal trends suggested that some CECs remain present in creek water for months, resulting in chronic exposures across the life stages of aquatic species. These findings highlight the potential for contamination from agricultural runoff and the associated ecological risk to aquatic species. SYNOPSIS: Suspect and non-target screening revealed the chronic occurrence of emerging contaminants in streams in agricultural catchments during rain events.

Evaluating exposures of bay mussels (Mytilus trossulus) to contaminants of emerging concern through environmental sampling and hydrodynamic modeling.

Bay mussels (Mytilus trossulus) were transplanted to 18 locations representing a range of potential exposures throughout Puget Sound, WA. Tissues were analyzed for over 200 organic contaminants. Results indicated the widespread exposure of marine organisms to trace levels of organic contaminants including the synthetic opioid oxycodone, present at three urban sites, and the chemotherapy drug melphalan, present at nine locations, at levels that may be of biological concern. Land-use and wastewater outfalls were evaluated as potential sources of CECs to the nearshore. Exposure to alkylphenol ethoxylates was associated with increased impervious surfaces in upland watersheds. A hydrodynamic simulation was performed using the Salish Sea Model to integrate inputs from 99 wastewater sources to Puget Sound. Predictions were consistent with concentrations of several wastewater-associated contaminants and δ15N enrichment. These results support the notion that Puget Sound nearshore biota suffer chronic exposures to a suite of contaminants from multiple sources and provide critical to focus future monitoring and management.

Suspect and Nontarget Screening for Contaminants of Emerging Concern in an Urban Estuary.

This study used suspect and nontarget screening with high-resolution mass spectrometry to characterize the occurrence of contaminants of emerging concern (CECs) in the nearshore marine environment of Puget Sound (WA). In total, 87 non-polymeric CECs were identified; those confirmed with reference standards (45) included pharmaceuticals, herbicides, vehicle-related compounds, plasticizers, and flame retardants. Eight polyfluoroalkyl substances were detected; perfluorooctanesulfonic acid (PFOS) concentrations were as high as 72-140 ng/L at one location. Low levels of methamphetamine were detected in 41% of the samples. Transformation products of pesticides were tentatively identified, including two novel transformation products of tebuthiuron. While a hydrodynamic simulation, analytical results, and dilution calculations demonstrated the prevalence of wastewater effluent to nearshore marine environments, the identity and abundance of selected CECs revealed the additional contributions from stormwater and localized urban and industrial sources. For the confirmed CECs, risk quotients were calculated based on concentrations and predicted toxicities, and eight CECs had risk quotients >1. Dilution in the marine estuarine environment lowered the risks of most wastewater-derived CECs, but dilution alone is insufficient to mitigate risks of localized inputs. These findings highlighted the necessity of suspect and nontarget screening and revealed the importance of localized contamination sources in urban marine environments.

Development of suspect and non-target screening methods for detection of organic contaminants in highway runoff and fish tissue with high-resolution time-of-flight mass spectrometry.

Untreated urban stormwater runoff contributes to poor water quality in receiving waters. The ability to identify toxicants and other bioactive molecules responsible for observed adverse effects in a complex mixture of contaminants is critical to effective protection of ecosystem and human health, yet this is a challenging analytical task. The objective of this study was to develop analytical methods using liquid chromatography coupled to high-resolution quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) to detect organic contaminants in highway runoff and in runoff-exposed fish (adult coho salmon, Oncorhynchus kisutch). Processing of paired water and tissue samples facilitated contaminant prioritization and aided investigation of chemical bioavailability and uptake processes. Simple, minimal processing effort solid phase extraction (SPE) and elution procedures were optimized for water samples, and selective pressurized liquid extraction (SPLE) procedures were optimized for fish tissues. Extraction methods were compared by detection of non-target features and target compounds (e.g., quantity and peak area), while minimizing matrix interferences. Suspect screening techniques utilized in-house and commercial databases to prioritize high-risk detections for subsequent MS/MS characterization and identification efforts. Presumptive annotations were also screened with an in-house linear regression (log Kowvs. retention time) to exclude isobaric compounds. Examples of confirmed identifications (via reference standard comparison) in highway runoff include ethoprophos, prometon, DEET, caffeine, cotinine, 4(or 5)-methyl-1H-methylbenzotriazole, and acetanilide. Acetanilide was also detected in runoff-exposed fish gill and liver samples. Further characterization of highway runoff and fish tissues (14 and 19 compounds, respectively with tentative identification by MS/MS data) suggests that many novel or poorly characterized organic contaminants exist in urban stormwater runoff and exposed biota.

A Field Trial of TCE Phytoremediation by Genetically Modified Poplars Expressing Cytochrome P450 2E1.

A controlled field study was performed to evaluate the effectiveness of transgenic poplars for phytoremediation. Three hydraulically contained test beds were planted with 12 transgenic poplars, 12 wild type (WT) poplars, or left unplanted, and dosed with equivalent concentrations of trichloroethylene (TCE). Removal of TCE was enhanced in the transgenic tree bed, but not to the extent of the enhanced removal observed in laboratory studies. Total chlorinated ethene removal was 87% in the CYP2E1 bed, 85% in the WT bed, and 34% in the unplanted bed in 2012. Evapotranspiration of TCE from transgenic leaves was reduced by 80% and diffusion of TCE from transgenic stems was reduced by 90% compared to WT. Cis-dichloroethene and vinyl chloride levels were reduced in the transgenic tree bed. Chloride ion accumulated in the planted beds corresponding to the TCE loss, suggesting that contaminant dehalogenation was the primary loss fate.

Evaluating Contaminants of Emerging Concern as tracers of wastewater from septic systems.

Bacterial and nutrient contamination from anthropogenic sources impacts fresh and marine waters, reducing water quality and restricting recreational and commercial activities. In many cases the source of this contamination is ambiguous, and a tracer or set of tracers linking contamination to source would be valuable. In this work, the effectiveness of utilizing a suite of Contaminants of Emerging Concern (CECs) as tracers of bacteria from human septic system effluent is investigated. Field sampling was performed at more than 20 locations over approximately 18 months and analyzed for a suite of CECs and fecal coliform bacteria. The sampling locations included seeps and small freshwater discharges to the shoreline. Sites were selected and grouped according to level of impact by septic systems as determined by previous field sampling programs. A subset of selected locations had been positively identified as being impacted by effluent from failing septic systems through dye testing. The CECs were selected based on their predominant use, their frequency of use, and putative fate and transport properties. In addition, two rounds of focused sampling were performed at selected sites to characterize short-term variations in CEC and fecal coliform concentrations, and to evaluate environmental persistence following source correction activities. The results indicate that a suite of common use compounds are suitable as generalized tracers of bacterial contamination from septic systems and that fate and transport properties are important in tracer selection. Highly recalcitrant or highly labile compounds likely follow different loss profiles in the subsurface compared to fecal bacteria and are not suitable tracers. The use of more than one tracer compound is recommended due to source variability of septic systems and to account for variations in the subsurface condition. In addition, concentrations of some CECs were measured in receiving waters at levels which suggested the potential for environmental harm, indicating that the possible risk presented from these sources warrants further investigation.

A methodology for evaluating and ranking water quantity indicators in support of ecosystem-based management.

Ecosystem-based Management (EBM) is an approach that includes different management priorities and requires a balance between anthropogenic and ecological resource demands. Indicators can be used to monitor ecosystem status and trends, and assess whether projects and/or programs are leading to the achievement of management goals. As such, the careful selection of a suite of indicators is a crucial exercise. In this paper we describe an indicator evaluation and selection process designed to support the EBM approach in Puget Sound. The first step in this process was the development of a general framework for selecting indicators. The framework, designed to transparently include both scientific and policy considerations into the selection and evaluation process, was developed and then utilized in the organization and determination of a preliminary set of indicators. Next, the indicators were assessed against a set of nineteen distinct criteria that describe the model characteristics of an indicator. A literature review was performed for each indicator to determine the extent to which it satisfied each of the evaluation criteria. The result of each literature review was summarized in a numerical matrix, allowing comparison, and demonstrating the extent of scientific reliability. Finally, an approach for ranking indicators was developed to explore the effects of intended purpose on indicator selection. We identified several sets of scientifically valid and policy-relevant indicators that included metrics such as annual-7 day low flow and water system reliability, which are supportive of the EBM approach in the Puget Sound.

Selecting indicator portfolios for marine species and food webs: a Puget sound case study.

Ecosystem-based management (EBM) has emerged as a promising approach for maintaining the benefits humans want and need from the ocean, yet concrete approaches for implementing EBM remain scarce. A key challenge lies in the development of indicators that can provide useful information on ecosystem status and trends, and assess progress towards management goals. In this paper, we describe a generalized framework for the methodical and transparent selection of ecosystem indicators. We apply the framework to the second largest estuary in the United States - Puget Sound, Washington - where one of the most advanced EBM processes is currently underway. Rather than introduce a new method, this paper integrates a variety of familiar approaches into one step-by-step approach that will lead to more consistent and reliable reporting on ecosystem condition. Importantly, we demonstrate how a framework linking indicators to policy goals, as well as a clearly defined indicator evaluation and scoring process, can result in a portfolio of useful and complementary indicators based on the needs of different users (e.g., policy makers and scientists). Although the set of indicators described in this paper is specific to marine species and food webs, we provide a general approach that could be applied to any set of management objectives or ecological system.

Phytoremediation of small organic contaminants using transgenic plants.

The efficacy of transgenic plants in the phytoremediation of small organic contaminants has been investigated. Two principal strategies have been pursued (1) the manipulation of phase I metabolic activity to enhance in planta degradation rates, or to impart novel metabolic activity, and (2) the enhanced secretion of reactive enzymes from roots leading to accelerated ex planta degradation of organic contaminants. A pair of dehalogenase genes from Xanthobacter autotrophicus was expressed in tobacco resulting in the dehalogenation of 1,2-dichloroethane, which was otherwise recalcitrant. A laccase gene from cotton was overexpressed in Arabidopsis thaliana resulting in increased secretory laccase activity and the enhanced resistance to trichlorophenol in soils. Although the results to date are promising, much of the work has been limited to laboratory settings; field demonstrations are needed.

A mass balance study of the phytoremediation of perchloroethylene-contaminated groundwater.

A mass balance study was performed under controlled field conditions to investigate the phytoremediation of perchloroethylene (PCE) by hybrid poplar trees. Water containing 7-14 mg L(-1) PCE was added to the test bed. Perchloroethylene, trichloroethylene, and cis-dichloroethylene were detected in the effluent at an average of 0.12 mg L(-1), 3.9 mg L(-1), and 1.9 mg L(-1), respectively. The total mass of chlorinated ethenes in the water was reduced by 99%. Over 95% of the recovered chlorine was as free chloride in the soil, indicating near-complete dehalogenation of the PCE. Transpiration, volatilization, and accumulation in the trees were all found to be minor loss mechanisms. In contrast, 98% of PCE applied to an unplanted soil chamber was recovered as PCE in the effluent water or volatilized into the air. These results suggest that phytoremediation can be an effective method for treating PCE-contaminated groundwater in field applications.

Degradation of low molecular weight volatile organic compounds by plants genetically modified with mammalian cytochrome P450 2E1.

Cytochrome P450 2E1 (CYP2E1) is a key enzyme in the mammalian metabolism of several low molecular weight volatile organic compounds (VOCs), such as trichloroethylene (TCE), vinyl chloride (VC), carbon tetrachloride (CT), benzene, chloroform, and bromodichloromethane (BDCM), which are all common environmental pollutants that pose risks to human health. We have developed a transgenic tobacco (Nicotiana tabacum cv. Xanthii) that expresses CYP2E1 with increased activity toward TCE and ethylene dibromide. In experiments with tobacco plant cuttings exposed to VOCs in small hydroponic vessels, the transgenic tobacco had greatly increased rates of removal of TCE, VC, CT, benzene, toluene, chloroform, and BDCM, compared to wild-type or vector control tobacco, but not of perchloroethylene or 1,1,1-trichloroethane.

Enhanced phytoremediation of volatile environmental pollutants with transgenic trees.

Small, volatile hydrocarbons, including trichloroethylene, vinyl chloride, carbon tetrachloride, benzene, and chloroform, are common environmental pollutants that pose serious health effects. We have developed transgenic poplar (Populus tremula x Populus alba) plants with greatly increased rates of metabolism and removal of these pollutants through the overexpression of cytochrome P450 2E1, a key enzyme in the metabolism of a variety of halogenated compounds. The transgenic poplar plants exhibited increased removal rates of these pollutants from hydroponic solution. When the plants were exposed to gaseous trichloroethylene, chloroform, and benzene, they also demonstrated superior removal of the pollutants from the air. In view of their large size and extensive root systems, these transgenic poplars may provide the means to effectively remediate sites contaminated with a variety of pollutants at much faster rates and at lower costs than can be achieved with current conventional techniques.

Microbial activity in soils following steam treatment.

Steam enhanced extraction (SEE) is an aquifer remediation technique that can be effective at removing the bulk of non-aqueous phase liquid (NAPL) contamination from the subsurface, particularly highly volatile contaminants. However, low volatility compounds such as polynuclear aromatic hydrocarbons (PAHs) are less efficiently removed by this process. This research evaluated the effects of steam injection on soil microbial activity, community structure, and the potential for biodegradation of contaminants following steam treatment. Three different soils were evaluated: a laboratory-prepared microbially-enriched soil, soil from a creosote contaminated field site, and soil from a chlorinated solvent and waste oil contaminated field site. Results from field-scale steaming are also presented. Microbial activity before and after steam treatment was evaluated using direct epifluorescent microscopy (DEM) using the respiratory activity dye 5-cyano-2,3, ditolyl tetrazolium chloride (CTC) in conjunction with the fluorochrome 5-(4,6-dichlorotriazinyl) aminofluorescein (DTAF) to yield a quantitative assessment of active and total microbial numbers. DEM results indicate that steamed soils that were analyzed while still hot exhibited microbial activity levels that were below detection. However, soil samples that were slowly cooled, more closely reflecting the conditions of applied SEE, exhibited microbial activity levels that were comparable to presteamed soils. Samples from a field-site where steam was applied continuously for 6 weeks also showed high levels of microbial activity following cooling. The metabolic capabilities of the steamed communities were investigated by measuring cell growth in enrichment cultures on various substrates. These studies provided evidence that organisms capable of biodegradation were among the mesophilic populations that survived steam treatment. Fluorescent in situ hybridization (FISH) analysis of the soils with domain-level rRNA probes suggest that both Archaea and Bacteria survived steam exposure.