Effects of metformin on wild fathead minnows (Pimephales promelas) using in-lake mesocosms in a boreal lake ecosystem.

Due to its widespread use for the treatment of Type-2 diabetes, metformin is routinely detected in surface waters globally. Laboratory studies have shown that environmentally relevant concentrations of metformin can adversely affect the health of adult fish, with effects observed more frequently in males. However, the potential risk to wild fish populations has yet to be fully elucidated and remains a topic of debate. To explore whether environmentally relevant metformin exposure poses a risk to wild fish populations, the present study exposed wild fathead minnows (Pimephales promelas) to 5 or 50 µg/L metformin via 2 m diameter in-lake mesocosms deployed in a natural boreal lake in Northern Ontario at the International Institute for Sustainable Development - Experimental Lakes Area (IISD-ELA). Environmental monitoring was performed at regular intervals for 8-weeks, with fish length, weight (body, liver and gonad), condition factor, gonadosomatic index, liver-somatic index, body composition (water and biomolecules) and hematocrit levels evaluated at test termination. Metabolic endpoints were also evaluated using liver, brain and muscle tissue, and gonads were evaluated histologically. Results indicate that current environmental exposure scenarios may be sufficient to adversely impact the health of wild fish populations. Adult male fish exposed to metformin had significantly reduced whole body weight and condition factor and several male fish from the high-dose metformin had oocytes in their testes. Metformin-exposed fish had altered moisture and lipid (decrease) content in their tissues. Further, brain (increase) and liver (decrease) glycogen were altered in fish exposed to high-dose metformin. To our knowledge, this study constitutes the first effort to understand metformin's effects on a wild small-bodied fish population under environmentally relevant field exposure conditions.

Intersex manifestation in the rainbow darter (Etheostoma caeruleum): Are adult male fish susceptible to developing and recovering from intersex after exposure to endocrine active compounds?

For over a decade, intersex has been observed in rainbow darter (RD) (Etheostoma caeruleum) populations living downstream wastewater treatment plants (WWTPs) in the Grand River, Ontario, Canada. To further our understanding of intersex development in adult male fish, the current study addressed three objectives: i) can intersex be induced in adult male fish, ii) is there a specific window of exposure when adult male fish are more susceptible to developing intersex, and iii) can pre-exposed adult male fish recover from intersex? To assess intersex induction in adult male fish, wild male RD were exposed in the laboratory for 22 weeks (during periods of spawning, gonadal regression, and gonadal recrudescence) to environmentally relevant concentrations of 17α-ethinylestradiol (EE2) including nominal 0, 1, and 10 ng/L. Intersex rates and severity at 10 ng/L EE2 were similar to those observed historically in adult male populations living downstream WWTPs in the Grand River and confirmed previous predictions that 1-10 ng/L EE2 would cause these adverse effects. To assess a window of sensitivity in developing intersex, male RD were exposed to nominal 0, 1 or 10 ng/L EE2 for 4 weeks during three different periods of gonadal development, including (i) spawning, (ii) early recrudescence and (iii) late recrudescence. These short-term exposures revealed that intersex incidence and severity were greater when RD were exposed while gonads were fully developed (during spawning) compared to periods of recrudescence. To assess if RD recover from intersex, wild fish were collected downstream WWTPs in the Grand River and assessed for intersex both before and after a 22-week recovery period in clean water that included gonadal regression and recrudescence. Results showed that fish did not recover from intersex, with intersex rates and severity similar to those both before and after the transition to clean water. This study further advances our knowledge on intersex manifestation in adult male fish including their sensitivity to endocrine active compounds during different periods of their annual reproductive cycle and their limited ability to recover from intersex after onset of the condition.

Wild fish responses to wastewater treatment plant upgrades in the Grand River, Ontario.

Municipal wastewater treatment plant (WWTP) effluent is one of several point sources of contaminants (nutrients, pharmaceuticals, estrogens, etc.) which can lead to adverse responses in aquatic life. Studies of WWTP effluent impacts on rainbow darter (Etheostoma caeruleum) collected downstream of WWTPs in the Grand River, Ontario have reported disruption at multiple levels of biological organization, including altered vitellogenin gene expression, lower levels of in vitro steroid production, and high frequency of intersex. However, major upgrades have occurred at treatment plants in the central Grand River over the last decade. Treatment upgrades to the Waterloo WWTP were initiated in 2009 but due to construction delays, the upgrades came fully on-line in 2017/2018. Responses in rainbow darter have been followed at sites associated with the outfall consistently over this entire time period. The treatment plant upgrade resulted in nitrification of effluent, and once complete there was a major reduction in effluent ammonia, selected pharmaceuticals, and estrogenicity. This study compared several key responses in rainbow darter associated with the Waterloo WWTP outfall prior to and post upgrades. Stable isotopes signatures in fish were used to track exposure to effluent and changed dramatically over time, corresponding to the effluent quality. Disruptions in in vitro steroid production and intersex in the darters that had been identified prior to the upgrades were no longer statistically different from the upstream reference sites after the upgrades. Although annual variations in water temperature and flow can potentially mask or exacerbate the effects of the WWTP effluent, major capital investments in wastewater treatment targeted at improving effluent quality have corresponded with the reduction of adverse responses in fish in the receiving environment.

Comparative Effects of Embryonic Metformin Exposure on Wild and Laboratory-Spawned Fathead Minnow (Pimephales promelas) Populations.

Metformin is routinely detected in aquatic ecosystems because of its widespread use as a treatment for Type 2 diabetes. Laboratory studies have shown that exposure to environmentally relevant concentrations of metformin can alter metabolic pathways and impact the growth of early life stage (ELS) fish; however, it is unknown whether these effects occur in wild populations. Herein, we evaluate whether findings from laboratory studies are representative and describe the relative sensitivities of both populations. Duplicate exposures (0, 5, or 50 µg/L metformin) were conducted using wild- and lab-spawned fathead minnow (Pimephales promelas) embryos. Apart from the water source, exposure conditions remained constant. Wild embryos were exposed to previously dosed lake water to account for changes in bioavailability, while reconstituted freshwater was used for the laboratory study. Developmental metformin exposure differentially impacted the growth and morphology of both cohorts, with energy dyshomeostasis and visual effects indicated. The fitness of wild-spawned larvae was impacted to a greater extent relative to lab-spawned fish. Moreover, baseline data reveal important morphological differences between wild- and lab-spawned ELS fatheads that may diminish representativeness of lab studies. Findings also confirm the bioavailability of metformin in naturally occurring systems and suggest current exposure scenarios may be sufficient to negatively impact developing fish.

A 30-Year Study of Impacts, Recovery, and Development of Critical Effect Sizes for Endocrine Disruption in White Sucker (Catostomus commersonii) Exposed to Bleached-Kraft Pulp Mill Effluent at Jackfish Bay, Ontario, Canada.

Jackfish Bay is an isolated bay on the north shore of Lake Superior, Canada that has received effluent from a large bleached-kraft pulp mill since the 1940s. Studies conducted in the late 1980s found evidence of reductions in sex steroid hormone levels in multiple fish species living in the Bay, and increased growth, condition and relative liver weights, with a reduction in internal fat storage, reduced gonadal sizes, delayed sexual maturation, and altered levels of circulating sex steroid hormones in white sucker (Catostomus commersonii). These early studies provided some of the first pieces of evidence of endocrine disruption in wild animals. Studies on white sucker have continued at Jackfish Bay, monitoring fish health after the installation of secondary waste treatment (1989), changes in the pulp bleaching process (1990s), during facility maintenance shutdowns and during a series of facility closures associated with changing ownership (2000s), and were carried through to 2019 resulting in a 30-year study of fish health impacts, endocrine disruption, chemical exposure, and ecosystem recovery. The objective of the present study was to summarize and understand more than 75 physiological, endocrine, chemical and whole organism endpoints that have been studied providing important context for the complexity of endocrine responses, species differences, and challenges with extrapolation. Differences in body size, liver size, gonad size and condition persist, although changes in liver and gonad indices are much smaller than in the early years. Population modeling of the initial reproductive alterations predicted a 30% reduction in the population size, however with improvements over the last couple of decades those population impacts improved considerably. Reflection on these 30 years of detailed studies, on environmental conditions, physiological, and whole organism endpoints, gives insight into the complexity of endocrine responses to environmental change and mitigation.

Endocrine Disruptor Impacts on Fish From Chile: The Influence of Wastewaters.

Industrial wastewaters and urban discharges contain complex mixtures of chemicals capable of impacting reproductive performance in freshwater fish, called endocrine-disrupting compounds (EDCs). In Chile, the issue was highlighted by our group beginning over 15 years ago, by analyzing the impacts of pulp and paper mill effluents (PPME) in the Biobio, Itata, and Cruces River basins. All of the rivers studied are important freshwater ecosystems located in the Mediterranean region of Central Chile, each with a unique fish biodiversity. Sequentially, we developed a strategy based on laboratory assays, semicontrolled-field experiments (e.g., caging) and wild fish population assessments to explore the issue of reproductive impacts on both introduced and native fish in Chile. The integration of watershed, field, and laboratory studies was effective at understanding the endocrine responses in Chilean freshwater systems. The studies demonstrated that regardless of the type of treatment, pulp mill effluents can contain compounds capable of impacting endocrine systems. Urban wastewater treatment plant effluents (WWTP) were also investigated using the same integrated strategy. Although not directly compared, PPME and WWTP effluent seem to cause similar estrogenic effects in fish after waterborne exposure, with differing intensities. This body of work underscores the urgent need for further studies on the basic biology of Chilean native fish species, and an improved understanding on reproductive development and variability across Chilean ecosystems. The lack of knowledge of the ontogeny of Chilean fish, especially maturation and sexual development, with an emphasis on associated habitats and landscapes, are impediment factors for their conservation and protection against the threat of EDCs. The assessment of effects on native species in the receiving environment is critical for supporting and designing protective regulations and remediation strategies, and for conserving the unique Chilean fish biodiversity.

Polycyclic aromatic compounds (PACs) in the Canadian environment: A review of sampling techniques, strategies and instrumentation.

A wide variety of sampling techniques and strategies are needed to analyze polycyclic aromatic compounds (PACs) and interpret their distributions in various environmental media (i.e., air, water, snow, soils, sediments, peat and biological material). In this review, we provide a summary of commonly employed sampling methods and strategies, as well as a discussion of routine and innovative approaches used to quantify and characterize PACs in frequently targeted environmental samples, with specific examples and applications in Canadian investigations. The pros and cons of different analytical techniques, including gas chromatography - flame ionization detection (GC-FID), GC low-resolution mass spectrometry (GC-LRMS), high performance liquid chromatography (HPLC) with ultraviolet, fluorescence or MS detection, GC high-resolution MS (GC-HRMS) and compound-specific stable (δ13C, δ2H) and radiocarbon (Δ14C) isotope analysis are considered. Using as an example research carried out in Canada's Athabasca oil sands region (AOSR), where alkylated polycyclic aromatic hydrocarbons and sulfur-containing dibenzothiophenes are frequently targeted, the need to move beyond the standard list of sixteen EPA priority PAHs and for adoption of an AOSR bitumen PAC reference standard are highlighted.

Development of a thin-film solid-phase microextraction (TF-SPME) method coupled to liquid chromatography and tandem mass spectrometry for high-throughput determination of steroid hormones in white sucker fish plasma.

Steroid hormones (SH) play a number of important physiological roles in vertebrates including fish. Changes in SH concentration significantly affect reproduction, differentiation, development, or metabolism. The objective of this study was to develop an in vitro high-throughput thin-film solid-phase microextraction (TF-SPME)-liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for targeted analysis of endogenous SH (cortisol, testosterone, progesterone, estrone (E1), 17ß-estradiol (E2), and 17α-ethinylestradiol (EE2)) in wild white sucker fish plasma where the concentrations of the analytes are substantially low. A simple TF-SPME method enabled the simultaneous determination of free and total SH concentrations. The use of biocompatible coating allowed direct extraction of these hormones from complex biological samples without prior preparation. The carryover was less than 3%, thereby ensuring reusability of the devices and reproducibility. The results showed that TF-SPME was suitable for the analysis of compounds in the polarity range between 1.28 and 4.31 such as SH at different physicochemical properties. The proposed method was validated according to bioanalytical method validation guidelines. The limit of detection (LOD) and limit of quantification(LOQ) for cortisol, testosterone, progesterone, E1, E2, and EE2 were from 0.006 to 0.150 ng/mL and from 0.020 to 0.500 ng/mL, respectively. The recovery for the method was about 85%, and the accuracy and precision of the method for cortisol, testosterone, and progesterone were ≤ 6.0% and ≤ 11.2%, respectively, whereas those for E1, E2, and EE2 were ≤ 15.0% and ≤ 10.2%, respectively. On the basis of this study, TF-SPME demonstrated several important advantages such as simplicity, sensitivity, and robustness under laboratory conditions. Graphical abstract.

Fish Performance Indicators Adjacent to Oil Sands Activity: Response in Performance Indicators of Slimy Sculpin in the Steepbank River, Alberta, Adjacent to Oil Sands Mining Activity.

Since 2009, the Canadian and Alberta governments have been developing monitoring plans for surface water quality and quantity of the lower Athabasca River and its tributaries (2010-2013). The objectives of the present study to the fish monitoring program were to 1) assess the current status of fish in a tributary of the lower Athabasca River, 2) identify existing differences between upstream reference and within the oil sands deposit exposure sites, and 3) identify trends/changes in fish performance indicators relative to historical studies. The present study examines the fish performance indicators in slimy sculpin (Cottus cognatus) in the Steepbank River, Alberta, in terms of growth, gonad size, condition, and hepatic 7-ethoxyresorufin-O-deethylase (EROD) activity as an indicator of exposure to oil-sands-related compounds. The sampling program followed historical sampling methods (1999-2000) to provide comparable data over time with an additional upstream site (n = 2) added as development progressed. Consistent changes were documented in sculpin collected from downstream sections of the Steepbank River within the oil sands deposit (n = 2) in 2010 through 2013. Sculpin demonstrated increased liver size with corresponding induction of EROD activity consistent with historical data and reductions in energy investment relative to reproductive development and gonadal steroid production capacity. There was no consistent evidence of changes in fish performance indicators with increased surface mining development, particularly adjacent to the Steepbank River Mid site. Although physical development in the Steepbank watershed has increased over the last 15 yr, these results are consistent with historical data suggesting that the magnitude of the response in the aquatic environment adjacent to the development has not changed. Environ Toxicol Chem 2020;39:396-409. © 2019 SETAC.

Development of environmental effects monitoring protocol in Brazil: a fish guide study of three river estuaries.

In Brazil, there are no unified and effective environmental monitoring models for bodies of water. Thus, several methodologies are used that result in information that is often difficult to compare, especially for stakeholders involved in regional water management. Studies in some countries such as Australia, Chile, the USA, and Sweden use the monitoring model implemented in Canada that was developed in the early 1990s. This model was designed to evaluate whether the current environmental regulations are sufficiently protective for pulp and paper effluents and for metal mining effluents. In this study, the Canadian Environmental Effects Monitoring methodologies were applied to three different Brazilian river basins, with the goal of constructing a framework for monitoring environmental effects. Pilot studies were carried out in the estuarine regions of the Benevente, Jucu, and Santa Maria da Vitória river basins, which are important rivers in the state of Espírito Santo. Evaluations included fish health, bioaccumulation studies, benthic invertebrate survey, and physical-chemical analyses of water and sediment. The quality of the environments was evaluated by means of seasonal samplings and comparisons between discharge, upstream, and downstream areas. This study made it possible to identify appropriate fish species to be used in environmental effects monitoring and the environmental quality of the rivers themselves as well as knowledge and policy gaps to implement such monitoring programs in Brazil. The study raises questions about the adequacy of Brazilian environmental legislation concerning tidal rivers.

Developing Triggers for Environmental Effects Monitoring Programs for Trout-Perch in the Lower Athabasca River (Canada).

Baseline variations in trout-perch energy use (growth, gonad size) and energy storage (condition, liver size) were characterized between 2009 and 2015 in 8 reaches of the Athabasca River (AB, Canada), including 2 reaches upstream of the city of Fort McMurray (AB, Canada) and 6 reaches downstream of Fort McMurray among existing oil sands operations. Generalized linear models, used to account for background variation, indicated that fork length, gonad size, and liver size decreased, whereas body weight increased, in relation to river discharge, for both male and female trout-perch. Air temperature was positively correlated with liver size and negatively correlated with gonad size and body weight for females, but only positively correlated with gonad weight for males. These linear models explained approximately 20 to 25% of the variation in adjusted body size, and upward of 80% of the variation in adjusted body weight, liver weight, and gonad weight. Residuals from linear models were used to estimate normal ranges of variation for each of the fish population performance measures. Combined, the models and normal ranges can be used to assess subsequent monitoring data, providing potential triggers for follow-up monitoring activities. Environ Toxicol Chem 2019;38:0-0. Environ Toxicol Chem 2019;38:1890-1901. © 2019 SETAC.

In vivo solid-phase microextraction sampling combined with metabolomics and toxicological studies for the non-lethal monitoring of the exposome in fish tissue.

Various environmental studies have employed the biomonitoring of fish in their aquatic ecosystems in order to identify potential metabolic responses to the exposome. In this study, we applied in vivo solid-phase microextraction (SPME) to perform non-lethal sampling on the muscle tissue of living fish to extract toxicants and various endogenous metabolites. Sixty white suckers (Catastomus commersonii) were sampled from sites upstream, adjacent, and downstream from the oil sands development region of the Athabasca River (Alberta, Canada) in order to track their biochemical responses to potential contaminants. In vivo SPME sampling facilitated the extraction of a wide range of endogenous metabolites, mainly related to lipid metabolism. The obtained results revealed significant changes in the levels of numerous metabolites, including eicosanoids, linoleic acids, and fat-soluble vitamins, in fish sampled in different areas of the river, thus demonstrating SPME's applicability for the direct monitoring of exposure to different environmental toxicants. In addition, several classes of toxins, including petroleum-related compounds, that can cause serious physiological impairment were tentatively identified in the extracts. In vivo SPME, combined with the analysis of contaminants and endogenous metabolites, provided important information about the exposome; as such, this approach represents a potentially powerful and non-lethal tool for identifying the mechanisms that produce altered metabolic pathways in response to the mixtures of different environmental pollutants.

Assessing wild fish exposure to ligands for sex steroid receptors from pulp and paper mill effluents in the Biobio River Basin, Central Chile.

Bioactive substances in the Biobio River Basin in Chile were examined by deploying Semi Permeable Membrane Devices (SPMDs) upstream and downstream of 4 pulp mill effluent discharges. Androgenic and estrogenic activity of SPMD extracts were then evaluated using in vitro fish sex steroid receptor binding assays. The results indicated the occurrence of estrogenic type compounds associated with one of the mill discharges. A significant correlation among the presence of these compounds, an increase in gonadosomatic index GSI and induction of hepatic EROD activity of two native fish species was observed. However, no significant presence of mature oocytes in female gonads was detected. Although EROD induction was observed in sites impacted by mill effluents, an increase of its activity occurred towards the downstream areas, suggesting other non-mill sources. More research is needed to understand the environmental changes in context of the new technological improvements in treatment systems to MBBR (Moving Bed Biofilm Reactor) recently implemented by the pulp mill industries.

An International Perspective on the Tools and Concepts for Effluent Toxicity Assessments in the Context of Animal Alternatives: Reduction in Vertebrate Use.

Since the 1940s, effluent toxicity testing has been used to assess potential ecological impacts of effluents and help determine necessary treatment options for environmental protection prior to release. Strategic combinations of toxicity tests, analytical tools, and biological monitoring have been developed. Because the number of vertebrates utilized in effluent testing is thought to be much greater than that used for individual chemical testing, there is a new need to develop strategies to reduce the numbers of vertebrates (i.e., fish) used. This need will become more critical as developing nations begin to use vertebrates in toxicity tests to assess effluent quality. A workshop was held to 1) assess the state of science in effluent toxicity testing globally; 2) determine current practices of regulators, industry, private laboratories, and academia; and 3) explore alternatives to vertebrate (fish) testing options and the inclusion of modified/new methods and approaches in the regulatory environment. No single approach was identified, because of a range of factors including regulatory concerns, validity criteria, and wider acceptability of alternatives. However, a suite of strategies in a weight-of-evidence approach would provide the flexibility to meet the needs of the environment, regulators, and the regulated community; and this "toolbox" approach would also support reduced reliance on in vivo fish tests. The present Focus article provides a brief overview of wastewater regulation and effluent testing approaches. Alternative methodologies under development and some of the limitations and barriers to regulatory approaches that can be selected to suit individual country and regional requirements are described and discussed. Environ Toxicol Chem 2018;37:2745-2757. © 2018 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals, Inc. on behalf of SETAC.

Spatial trends in a biomagnifying contaminant: Application of amino acid compound-specific stable nitrogen isotope analysis to the interpretation of bird mercury levels.

Levels of biomagnifying contaminants are greatest in high-trophic level biota (e.g., predatory birds such as gulls). Gull eggs have been used to assess contaminant spatial patterns and sources, but such assessments must consider how organism trophic position may influence spatial inferences. Stable nitrogen isotopes (δ15 N) in bulk tissue are routinely used in this context. However, bulk δ15 N values are only useful if spatial differences in baseline δ15 N values are considered. Amino acid compound-specific stable nitrogen isotope analysis can generate estimates of baseline δ15 N values and trophic position from the same sample. In the present study, eggs (n = 428) of California (Larus californicus), herring (Larus argentatus smithsonianus), and ring-billed (Larus delawarensis) gulls were used to assess spatial patterns in mercury (Hg) availability in 12 western Canadian lakes located over 14 degrees of latitude, with amino acid compound-specific stable isotope analysis adjustment of egg Hg levels for trophic position. Mean trophic position-adjusted egg Hg levels (micrograms per gram, dry wt) were greatest at sites in receiving waters of the Athabasca River (X¯ = 0.70) compared to southern (X¯ = 0.39) and northern (X¯ = 0.50) regions. Research is required to investigate factors (e.g., local Hg released as a result of human activities, processes influencing Hg methylation) which may be responsible for greater Hg availability in the lower Athabasca River basin. However, it is clear that amino acid compound-specific stable isotope analysis is a valuable tool for assessing contaminant spatial patterns. Environ Toxicol Chem 2018;37:1466-1475. © 2018 SETAC.

Assessing recovery of in vitro steroid production in male rainbow darter (Etheostoma caeruleum) in response to municipal wastewater treatment plant infrastructure changes.

The present study examined in vitro 11-ketotestosterone and testosterone production by the testes of rainbow darter (Etheostoma caeruleum) collected from selected reference sites and downstream of 2 municipal wastewater treatment plants (MWWTPs; Waterloo and Kitchener) on the central Grand River (Ontario, Canada), over a 6-yr period (2011-2016). The main objective was to investigate if infrastructure upgrades at the Kitchener MWWTP in 2012 resulted in a recovery of this response in the post-upgrade period (2013-2016). Two supporting studies showed that the fall season is appropriate for measuring in vitro sex steroid production because it provides stable detection of steroid patterns, and that the sample handling practiced in the present study did not introduce a bias. Infrastructure upgrades of the Kitchener MWWTP resulted in significant reductions in ammonia and estrogenicity. After the upgrades, 11-ketotestosterone production by MWWTP-exposed fish increased in 2013 and it continued to recover throughout the study period of 2014 through 2016, returning to levels measured in reference fish. Testosterone production was less sensitive and it lacked consistency. The Waterloo MWWTP underwent some minor upgrades but the level of ammonia and estrogenicity remained variable over time. The production of 11-ketotestosterone and testosterone in rainbow darter below the Waterloo MWWTP was variable and without a clear recovery pattern over the course of the present study. The results of the present study demonstrated that measuring production of sex steroids (especially 11-ketotestosterone) over multiple years can be relevant for assessing responses in fish to environmental changes such as those resulting from major infrastructure upgrades. Environ Toxicol Chem 2018;37:501-514. © 2017 SETAC.

In vivo microsampling to capture the elusive exposome.

Loss and/or degradation of small molecules during sampling, sample transportation and storage can adversely impact biological interpretation of metabolomics data. In this study, we performed in vivo sampling using solid-phase microextraction (SPME) in combination with non-targeted liquid chromatography and high-resolution tandem mass spectrometry (LC-MS/MS) to capture the fish tissue exposome using molecular networking analysis, and the results were contrasted with molecular differences obtained with ex vivo SPME sampling. Based on 494 MS/MS spectra comparisons, we demonstrated that in vivo SPME sampling provided better extraction and stabilization of highly reactive molecules, such as 1-oleoyl-sn-glycero-3-phosphocholine and 1-palmitoleoyl-glycero-3-phosphocholine, from fish tissue samples. This sampling approach, that minimizes sample handling and preparation, offers the opportunity to perform longitudinal monitoring of the exposome in biological systems and improve the reliability of exposure-measurement in exposome-wide association studies.

Halogenated phenolic compounds in wild fish from Canadian Areas of Concern.

Concentrations of halogenated phenolic compounds were measured in the plasma of brown bullhead (Ameiurus nebulosus) from 4 Canadian Areas of Concern (AOCs), to assess exposure to suspected thyroid-disrupting chemicals. Hydroxylated polychlorinated biphenyls (OH-PCBs) were detected in every sample collected in 3 of the AOCs; the detection frequency was lower in samples from the Detroit River AOC. The OH-PCBs most frequently detected were pentachloro, hexachloro, and heptachloro congeners, which are structurally similar to thyroid hormones. Pentachlorophenol (PCP) was detected at highest concentrations (1.8 ng/g) in fish from Prince Edward Bay, the Bay of Quinte Lake reference site, and Hillman Marsh (the Wheatley Harbour reference site), suggesting local sources of contamination. Elevated PCP concentrations were also detected in the plasma of brown bullhead from exposed sites in the Toronto and Region AOC (0.4-0.6 ng/g). Triclosan was consistently detected in the Toronto and Region AOC (0.05-0.9 ng/g), consistent with wastewater emission. Greater concentrations were occasionally detected in the plasma of brown bullhead from the Bay of Quinte AOC. Concentrations of polybrominated diphenyl ethers were highest in the Toronto and Region AOC, and at 2 of the Bay of Quinte AOC exposed sites near Trenton and Belleville. Distribution patterns reflected the properties and usage of the compounds under investigation and the characteristics of each AOC. Environ Toxicol Chem 2017;36:2266-2273. © 2017 SETAC.

The Relationship between Organic Loading and Effects on Fish Reproduction for Pulp Mill Effluents across Canada.

This study builds upon the work of a multiagency consortium tasked with determining cost-effective solutions for the effects of pulp mill effluents on fish reproduction. A laboratory fathead minnow egg production test and chemical characterization tools were used to benchmark 81 effluents from 20 mills across Canada, representing the major pulping, bleaching, and effluent treatment technologies. For Kraft and mechanical pulp mills, effluents containing less than 20 mg/L BOD5 were found to have the greatest probability of having no effects. Organic loading, expressed as the total detected solvent-extractable components by gas chromatography/mass spectrometry (GC/MS), also correlated with decreased egg laying. Exceptions were found for specific Kraft, mechanical, and sulfite mills, suggesting yet unidentified causative agents are involved. Recycled fiber mill effluents, tested for the first time, were found to have little potential for reproductive effects despite large variations in BOD5 and GC/MS profiles. Effluent treatment systems across all production types were generally efficient, achieving a combined 82-98% BOD5 removal. Further reductions of final effluent organic loadings toward the target of less than 20 mg/L are recommended and can be realized through biotreatment optimization, the reduction of organic losses associated with production upsets and selecting best available technologies that reduce organic loadings to biotreatment.

Returning to normal? Assessing transcriptome recovery over time in male rainbow darter (Etheostoma caeruleum) liver in response to wastewater-treatment plant upgrades.

The present study measured hepatic transcriptome responses in male rainbow darter (Etheostoma caeruleum) exposed to 2 municipal wastewater-treatment plants (MWWTPs; Kitchener and Waterloo) over 4 fall seasons (2011-2014) in the Grand River (Ontario, Canada). The overall goal was to determine if upgrades at the Kitchener MWWTP (in 2012) resulted in transcriptome responses indicative of improved effluent quality. The number of differentially expressed probes in fish downstream of the Kitchener outfall (904-1223) remained comparable to that downstream of Waterloo (767-3867). Noteworthy was that year and the interaction of year and site explained variability in more than twice the number of transcripts than site alone, suggesting that year and the interaction of year and site had a greater effect on the transcriptome than site alone. Gene set enrichment analysis revealed a gradual reduction in the number of gene ontologies over time at exposure sites, which corresponded with lower contaminant load. Subnetwork enrichment analysis revealed that there were noticeable shifts in the cell pathways differently expressed in the liver preupgrade and postupgrade. The dominant pathways altered preupgrade were related to genetic modifications and cell division, whereas postupgrade they were associated with the immune system, reproduction, and biochemical responses. Molecular pathways were dynamic over time, and following the upgrades, there was little evidence that gene expression profiles in fish collected from high-impact sites postupgrade were more similar to those in fish collected from reference site. Environ Toxicol Chem 2017;36:2108-2122. © 2017 SETAC.