Effluent Exposure Research Articles

Using Transcriptomic Tools to Evaluate Biological Effects Across Effluent Gradients at a Diverse Set of Study Sites in Minnesota, USA

The aim of this study was to explore the utility of "omics" approaches in monitoring aquatic environments where complex, often unknown stressors make chemical-specific risk assessment untenable. We examined changes in the fathead minnow (Pimephales promelas) ovarian transcriptome following 4-day exposures conducted at three sites in Minnesota (MN, USA). Within each site, fish were exposed to water from three locations along a spatial gradient relative to a wastewater treatment plant (WWTP) discharge. After exposure, site-specific impacts on gene expression in ovaries were assessed. Using an intragradient point of comparison, biological responses specifically associated with the WWTP effluent were identified using functional enrichment analyses. Fish exposed to water from locations downstream of the effluent discharges exhibited many transcriptomic responses in common with those exposed to the effluent, indicating that effects of the discharge do not fully dissipate downstream. Functional analyses showed a range of biological pathways impacted through effluent exposure at all three sites. Several of those impacted pathways at each site could be linked to potential adverse reproductive outcomes associated with the hypothalamic-pituitary-gonadal (HPG) axis in female fathead minnows, specifically signaling pathways associated with oocyte meiosis, TGF-beta signaling, gonadotropin-releasing hormone (GnRH) and epidermal growth factor receptor family (ErbB), and gene sets associated with cyclin B-1 and metalloproteinase. The utility of this approach comes from the ability to identify biological responses to pollutant exposure, particularly those that can be tied to adverse outcomes at the population level and those that identify molecular targets for future studies.

Metabolomics forin SituEnvironmental Monitoring of Surface Waters Impacted by Contaminants from Both Point and Nonpoint Sources

We investigated the efficacy of metabolomics for field-monitoring of fish exposed to wastewater treatment plant (WWTP) effluents and nonpoint sources of chemical contamination. Lab-reared male fathead minnows (Pimephales promelas, FHM) were held in mobile monitoring units and exposed on-location to surface waters upstream and downstream of the effluent point source, as well as to the actual effluent at three different WWTP sites in Minnesota. After four days of exposure, livers were collected, extracted, and analyzed by (1)H NMR spectroscopy and GC-MS to characterize responses of the hepatic metabolome. Multivariate statistical analysis revealed distinct metabolite profile changes in response to effluent exposure from each of the three WWTPs. Differences among locations (i.e., upstream, downstream, and effluent) within each of the three sites were also identified. These observed differences comport with land-use and WWTP characteristics at the study sites. For example, at one of the sites, the metabolomic analyses suggested a positive interactive response from exposure to WWTP effluent and nearby nonpoint (likely agricultural related) contamination. These findings demonstrate the utility of metabolomics as a field-based technique for monitoring the exposure of fish to impacted surface waters.

Development and evaluation of new behavioral indexes for a biological early warning system using <i>Daphnia magna</i>

Abstract. New behavioral indexes including combined index (CI), distribution index (DI), toxic index (TI), and altitude index (AI) for a biological early warning system (BEWS) were developed and evaluated using Daphnia magna in this study. The sensitivity and stability of each index were compared to evaluate the performance of the indexes through a real-time exposure test with a synthetic copper solution. The applicability of the CI to the field sample was evaluated through an effluent exposure test. The proportional relationship between toxicity level and magnitude of response was much lower in the effluent due to the complexity of water than in the copper solution. The results showed that the CI was most sensitive among the three indexes, while the DI was confirmed as the most useful index among the individual indexes. The combined index (CI) shows not only sensitivity but also stability in normal conditions below the statistically significant threshold (p < 0.01), whereas the individual indexes displayed unstable index values in normal conditions (p > 0.01). The CI improved performance of the BEWS in terms of sensitivity and stability, and it was confirmed as the higher correlation coefficient between the magnitude of the index and the toxicity level of the water sample.

Combined Amplicon Pyrosequencing Assays Reveal Presence of the Apicomplexan “type-N” (cf. Gemmocystis cylindrus) and Chromera velia on the Great Barrier Reef, Australia

BackgroundThe coral is predominantly composed of the metabolically dependent coral host and the photosynthetic dinoflagellate Symbiodinium sp. The system as a whole interacts with symbiotic eukaryotes, bacteria and viruses. Gemmocystiscylindrus (cf. “type-N” symbiont) belonging to the obligatory parasitic phylum Apicomplexa (Alveolata) is ubiquitous in the Caribbean coral, but its presence in the Great Barrier Reef coral has yet to be documented. Approaches allowing identification of the healthy community from the pathogenic or saprobic organisms are needed for sustainable coral reef monitoring.Methods & Principal FindingsWe investigated the diversity of eukaryotes associated with a common reef-building corals from the southern Great Barrier Reef. We used three tag encoded 454 amplicon pyrosequencing assays targeting eukaryote small-subunit rRNA gene to demonstrate the presence of the apicomplexan type-N and a photosynthetic sister species to Apicomplexa - Chromera velia . Amplicon pyrosequencing revealed presence of the small-subunit rRNA genes of known eukaryotic pathogens ( Cryptosporidium and Cryptococcus ). We therefore conducted bacterial tag encoded amplicon pyrosequencing assay for small-subunit rRNA gene to support effluent exposure of the coral. Bacteria of faecal origin (Enterobacteriales) formed 41% of total sequences in contrast to 0-2% of the coral-associated bacterial communities with and without C . velia , respectively.SignificanceThis is the first time apicomplexan type-N has been detected in the Great Barrier Reef. Eukaryote tag encoded amplicon pyrosequencing assays demonstrate presence of apicomplexan type-N and C. Velia in total coral DNA. The data highlight the need for combined approaches for eukaryotic diversity studies coupled with bacterial community assessment to achieve a more realistic goals of defining the holobiont community and assessing coral disease. With increasing evidence of Apicomplexa in coral reef environments, it is important not only to understand the evolution of these organisms but also identify their potential as pathogens.

Gene Expression of Fathead Minnows (Pimephales promelas) Exposed to Two Types of Treated Municipal Wastewater Effluents

Contaminants of emerging concern (CECs) in treated municipal effluents have the potential to adversely impact exposed organisms prompting elevated public concern. Using transcriptomic tools, we investigated changes in gene expression and cellular pathways in the liver of male fathead minnows (Pimephales promelas) exposed to 5% concentrations of full secondary-treated (HTP) or advanced primary-treated (PL) municipal wastewater effluents containing CECs. Gene expression changes were associated with apical end points (plasma vitellogenin and changes in secondary sexual characteristics). Of 32 effluent CECs analyzed, 28 were detected including pharmaceuticals, personal care products, hormones, and industrial compounds. Exposure to both effluents produced significantly higher levels of plasma VTG and changes in secondary sexual characteristics (e.g., ovipositor development). Transcript patterns differed between effluents, with <10% agreement in the detected response (e.g., altered production of transcripts involved in xenobiotic detoxification, oxidative stress, and apoptosis were observed following exposure to both effluents). Exposure to PL effluent caused changes in transcription of genes involved in metabolic pathways (e.g., lipid transport and steroid metabolism). Exposure to HTP effluent affected transcripts involved in signaling pathways (e.g., focal adhesion assembly and extracellular matrix). The results suggest a potential association between some transcriptomic changes and physiological responses following effluent exposure. This study identified responses in pathways not previously implicated in exposure to complex chemical mixtures containing CECs, which are consistent with effluent exposure (e.g., oxidative stress) in addition to other pathway responses specific to the effluent type.

Genomic and phenotypic response of hornyhead turbot exposed to municipal wastewater effluents

Laboratory tests with marine flatfish were conducted to investigate associations among gene expression, higher biological responses and wastewater effluent exposure. In the present study, male hornyhead turbot (Pleuronichthys verticalis) were exposed to environmentally realistic (0.5%) and higher (5%) concentrations of chemically enhanced advanced-primary (PL) and full-secondary treated (HTP) effluents from two southern California wastewater treatment plants (WWTP). Hepatic gene expression was examined using a custom low-density microarray. Alterations in gene expression (vs. controls) were observed in fish exposed to both effluent types. Fish exposed to 0.5% PL effluent showed changes in genes involved in the metabolism of xenobiotics, steroids, and lipids, among other processes. Fish exposed to 5% PL effluent showed expression changes in genes involved in carbohydrate metabolism, stress responses, xenobiotic metabolism, and steroid synthesis, among others. Exposure to 5% HTP effluent changed the expression of genes involved in lipid, glutathione and xenobiotic metabolism, as well as immune responses. Although no concentration-dependent patterns of response to effluent exposure were found, significant Spearman correlations were observed between the expression of 22 genes and molecular and/or higher biological responses. These results indicate that microarray gene expression data correspond to higher biological responses and should be incorporated in studies assessing fish health after exposure to complex environmental mixtures.

Biological response of high-back crucian carp (Carassius auratus) during different life stages to wastewater treatment plant effluent

This study presents the adverse effects of endocrine disrupting chemicals (EDCs) in effluent of wastewater treatment plants (WwTPs) on fish health. A study of chronic exposure to WwTPs effluent for 10 months was undertaken in high-back crucian carp (Carassius auratus) during different life stages, covering early-life-stage (ELS), prespawning period, and postspawning period. Condition factor (CF), gonadosomatic index (GSI), hepatosomatic index (HSI), and plasma vitellogenin (VTG) levels were employed as indicators to assess biological effects of effluent on this gynogenesis species. Meanwhile, some high-back crucian carp were caged in Demonstration Base of Biological Purification for Filter-feeding Fish (hereinafter, Demonstration Base), as WwTPs effluent exposure controls. In the meantime, a depuration study was carried out to determine whether or not the estrogenic effects caused by effluent exposure could be reduced after moving fish into EDCs-free water. CF, HSI, GSI, and plasma Vtg levels of high-back crucian carp caged in Demonstration Base were generally in accordance with seasonal change. Effluent exposure inhibited gonadal growth, reducing GSI in ELS while increasing it around spawning, sharpened liver burdens, increasing HSI, and induced abnormal Vtg expression in juvenile high-back crucian carp, augmenting Vtg concentrations in plasma. Around spawning period, Vtg in high-back crucian carp were mainly induced by endogenous estrogens, and EDCs in effluent had less influence on them. Staying in EDCs-free water for 30 days made high-back crucian carp recover from effects of previous effluent exposure, relieving inhibition of gonadal development and hypertrophy of liver as well as reducing Vtg induced by EDCs in effluent. The results revealed that high-back crucian carp in ELS are more sensitive to WwTPs effluent exposure. Additionally, the depuration study showed a clearance of the estrogenic effects caused by effluent.

Effect of battery-manufacturing effluent on endogenous antioxidant in freshwater fish

This study monitors and assesses the effect of battery-manufacturing effluent containing metals Pb, Zn and Cd on endogenous antioxidants. Malonaldehyde (MDA), reduced glutathione sulfyhydryl (GSH) and catalase (CAT) which are known biomarkers of effluent were exposed to 0%, 25%, 50%, 75% and 90% amendments for 74h on the gills, liver and kidneys of C. punctata. There was more metal Zn accumulation in the gills and GSH contents increased significantly in the gills (P<0.01), liver accumulation of Pb was found to be more (P<0.05), whereas lowest accumulation of Pb was found in kidneys and the highest accumulation of Cd (P<0.05). Over all amendments of the effluents, MDA contents were increased in the gills, liver and kidneys (P<0.01). GSH levels were decreased among the liver and kidneys compared to the gills (P<0.01) at 90% amendment. Effluent exposure caused a significant decrease in the activities of CAT in the gills, liver and kidneys (P<0.01, 0.05 and 0.05) of fish. Increased MDA activity was indicative of the formation of free radicals in the fish with exposure to amendments of battery manufacturing effluent, while increased levels of GSH pointed to the occurrence of a scavenging mechanism of free radicals.

STUDIES ON THE TOXICITY OF COMPOSITE TANNERY EFFLUENT USING GUPPY (Poecilia reticulata) AS A FISH MODEL

East Calcutta Wetland ecosystem is a Ramsar Site in India covering almost 51 km2 and is one of the largest of its type. This wetland receives a massive load of almost untreated industrial and domestic effluents every day, throughout the year, that has been effectively utilized in pisciculture for having a better yield (2.85 metric tons of fish per 0.01 km2 of water body per annum). The present investigation was carried out to study the toxicity of the untreated effluents discharged from tanneries located at the fringe of East Calcutta wetlands. Guppy (Poecilia reticulata) was chosen as a model fish and was exposed to three different sublethal concentrations (3%, 6% and 9%) of composite tannery effluents (TE) for different time periods (4 days, 7 days and 15 days). Exposed fish livers were analyzed for metallothionein (MT, a ubiquitous metal binding protein), cytochrome P4501A1+1A2 (CYP, a major family of xenobiotic metabolizing enzyme in fish biosystem) and nonspecific cytotoxic cells (NCCs, a major group of innate immune system in fish) expressions applying indirect enzyme-linked immunosorbent assay (ELISA) procedure. Results showed that expression of all the three chosen biomarkers were significantly dose and time dependent upon tannery effluent exposure. Major significances drawn from the present study was that fish species cultivated in east Calcutta wetland ecosystem are under potential threat of contamination and stress induced by composite tannery effluents that could lead to adverse physiological conditions. Moreover, these findings could be important in terms of designing biomarkers for early environmental warning system and also for monitoring fish health.

The stress response of three-spined sticklebacks is modified in proportion to effluent exposure downstream of wastewater treatment works

This study was conducted to investigate whether exposure to wastewater treatment works (WWTW) effluent affects the adaptive stress axis of fish resident within the receiving water. Three-spined sticklebacks (Gasterosteus aculeatus) were sampled from sites downstream of ten WWTWs in north-west England, selected to represent a range of human population equivalents between 1000 and 125,000. Following capture, indices of stress (whole-body cortisol and glucose concentrations) were measured both prior to, and following, the imposition of a standardised stressor to establish both baseline and stress-induced concentrations of cortisol and glucose. There was considerable between-site variation in size, and to a lesser extent condition, of the fish. Pre- and post-stress cortisol and glucose concentrations also varied significantly between-sites. A large proportion of the variation in both the somatic data and the stress response was explained by variation in the proportion of effluent contributing to total river flow at the study sites. Mass (r2=0.35, P<0.001) and length (r2=0.37, P<0.001) of the fish, and cortisol (r2=0.26, P<0.001) and glucose (r2=0.12, P<0.01) concentrations in unstressed sticklebacks, were positively related to the concentration of effluent across the sample sites. However, in stressed fish, cortisol (r2=0.32, P<0.001) and glucose (r2=0.14, P<0.001) concentrations exhibited a negative trend in relation to the effluent concentrations across sites. Individual variation in fish size did not account for the variation in either cortisol or glucose levels. These data provide the first indication that modulation of the stress axis in fish by anthropogenic factors might be widespread and of greater significance than hitherto assumed.

Expression profiling and gene ontology analysis in fathead minnow (Pimephales promelas) liver following exposure to pulp and paper mill effluents

Many studies link pulp and paper mill effluent (PPME) exposure to adverse effects in fish populations present in the mill receiving environments. These impacts are often characteristic of endocrine disruption and may include impaired reproduction, development and survival. While these physiological endpoints are well-characterized, the molecular mechanisms causing them are not yet understood. To investigate changes in gene transcription induced by exposure to a PPME at several stages of treatment, male and female fathead minnows (FHMs) were exposed for 6days to 25% (v/v) secondary (biologically) treated kraft effluent (TK) or 100% (v/v) combined mill outfall (CMO) from a mill producing both kraft pulp and newsprint. The gene expression changes in the livers of these fish were analyzed using a 22K oligonucleotide microarray. Exposure to TK or CMO resulted in significant changes in the expression levels of 105 and 238 targets in male FHMs and 296 and 133 targets in females, respectively. Targets were then functionally analyzed using gene ontology tools to identify the biological processes in fish hepatocytes that were affected by exposure to PPME after its secondary treatment. Proteolysis was affected in female FHMs exposed to both TK and CMO. In male FHMs, no processes were affected by TK exposure, while sterol, isoprenoid, steroid and cholesterol biosynthesis and electron transport were up-regulated by CMO exposure. The results presented in this study indicate that short-term exposure to PPMEs affects the expression of reproduction-related genes in the livers of both male and female FHMs, and that secondary treatment of PPMEs may not neutralize all of their metabolic effects in fish. Gene ontology analysis of microarray data may enable identification of biological processes altered by toxicant exposure and thus provide an additional tool for monitoring the impact of PPMEs on fish populations.

Temporal changes in stress and tissue-specific metabolic responses to municipal wastewater effluent exposure in rainbow trout

Sub-chronic exposure to municipal wastewater effluent (MWWE) in situ was recently shown to impact the acute response to a secondary stressor in rainbow trout (Oncorhynchus mykiss). However, little is known about whether MWWE exposure in itself is stressful to the animal. To address this, we carried out a laboratory study to examine the organismal and cellular stress responses and tissue-specific metabolic capacity in trout exposed to MWWE. Juvenile rainbow trout were exposed to 0, 20 and 90% MWWE (from a tertiary wastewater treatment plant), that was replenished every 2d, for 14d. Fish were sampled 2, 8 or 14d post-exposure. Plasma cortisol, glucose and lactate levels were measured as indicators of organismal stress response, while inducible heat shock protein 70 (hsp70), constitutive heat shock protein 70 (hsc70) and hsp90 expression in the liver were used as markers of cellular stress response. Impact of MWWE on cortisol signaling was ascertained by determining glucocorticoid receptor protein (GR) expression in the liver, brain and, heart, and metabolic capacity was evaluated by measuring liver glycogen content and tissue-specific activities of key enzymes in intermediary metabolism. Plasma glucose and lactate levels were unaffected by exposure to MWWEs, whereas cortisol showed a transient increase in the 20% group at 8d. Liver hsc70 and hsp90, but not hsp70 expression, were higher in the 90% MWWE group after 8d. There was a temporal change in GR expression in the liver and heart, but not brain of trout exposed to MWWE. Liver glycogen content and activities liver gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK), and alanine aminotransferase (AlaAT) were significantly affected by MWWE exposure. The glycolytic enzymes pyruvate kinase (PK) and hexokinase (HK) activities were significantly higher temporally by MWWE exposure in the gill and heart, but not in the liver and brain. Overall, a 14d exposure to MWWE evokes a cellular stress response and perturbs the cortisol stress response in rainbow trout. The tissue-specific temporal changes in the metabolic capacity suggest enhanced energy demand in fish exposed to MWWE, which may eventually lead to reduced fitness.

Swim performance and energy homeostasis in spottail shiner ( Notropis hudsonius) collected downstream of a uranium mill

The Key Lake uranium milling operation (Saskatchewan, Canada) releases complex effluent into the local watershed. The objective of the current study was to investigate whether fish from an effluent-receiving waterbody exhibited differences in swimming performance and energy homeostasis compared to fish from a local reference site. Juvenile spottail shiner ( Notropis hudsonius) were collected from a lake downstream of the uranium mill, and compared to fish collected from a nearby reference lake. Critical swimming speed ( U crit; fatigue velocity), tail beat frequency, and tail amplitude did not differ significantly when comparing fish collected from the exposure lake and reference lake. Captured shiner used in swim tests were considered fatigued, and metabolic endpoints were compared between this group and non-fatigued fish, which were treated similarly but not subjected to swim tests. In both non-fatigued and fatigued shiner, liver glycogen was significantly greater in fish collected from the exposure lake compared to the reference lake. However, it is unclear if this effect, and others related to condition, were the result of contaminant exposure or other environmental factors. While there were no differences in plasma lactate, hematocrit or liver triglycerides in non-fatigued fish between sites, only fatigued reference fish had increased lactate and hematocrit and decreased triglycerides. In non-fatigued fish, plasma glucose did not significantly differ between sites, but significantly decreased after swimming only in fish from the exposure lake. In summary, shiner from the exposure site demonstrated similar swim endurance and possessed greater energy stores despite metabolic alterations compared to shiner from the reference site. Therefore, because fish collected downstream of the uranium mill operation had similar swimming ability as fish from the reference lake, U crit test results presented here may not reflect or be indicative of metabolic effects of complex effluent exposure.

Tissue-specific metabolic changes in response to an acute handling disturbance in juvenile rainbow trout exposed to municipal wastewater effluent

The objective of this study was to evaluate the effects of municipal wastewater effluent (MWWE) exposure on aspects of both organismal and cellular stress response in rainbow trout ( Oncorhynchus mykiss). Juvenile rainbow trout were exposed for 14 d (2-d static renewal) to tertiary-treated MWWE at concentrations of 0%, 20% and 90%. Following the MWWE exposure, fish were subjected to an acute handling stress and sampled at 1, 4 and 24 h post-stressor, to evaluate the fish performance to additional stressors. Organismal stress response evaluation included measuring plasma cortisol, glucose and lactate concentrations, and tissue metabolic capacity, including gluconeogenic (liver) and glycolytic enzyme activities in the liver, brain, heart and gill. No significant differences between treatments were seen in plasma cortisol, glucose or lactate concentrations after 14 d exposure to MWWE. However, MWWE exposure significantly affected plasma cortisol and glucose response to the acute secondary stressor. Acute handling disturbance enhanced liver gluconeogenic capacity in the control group, but this response was altered in the MWWE exposed groups. MWWE exposure did not affect the acute stressor-mediated enhancement of brain or gill glycolytic capacity, but significantly reduced the glycolytic capacity of liver and heart in response to a secondary stressor compared to the control group. Altogether, chronic exposure to MWWE impacts the metabolic performances to a secondary stressor challenge and this includes disruptions in tissue-specific gluconeogenic and glycolytic capacities in rainbow trout.

Swimming performance and energy homeostasis in juvenile laboratory raised fathead minnow (Pimephales promelas) exposed to uranium mill effluent

Research at the Key Lake uranium mill (Saskatchewan, Canada) suggests effluent discharged from the mill affects energy stores of resident fish, but the mechanisms by which energy homeostasis is affected and the subsequent effects on swimming performance are unknown. In the present study larvae were collected from laboratory raised adult fathead minnow (Pimephales promelas) exposed to 5% diluted uranium mill effluent or control (dechlorinated municipal) water, and reared in the same treatments to 60days post hatch (dph). Critical swimming speed (Ucrit) was significantly lower in effluent exposed 60dph fish compared to control fish. Fish used in tests were considered fatigued and compared to fish without swim testing (non-fatigued). There were no differences in whole body glycogen or triglyceride concentrations between effluent exposed versus control fish. However, fatigued fish from both treatments had significantly lower triglycerides, but not glycogen, compared to non-fatigued fish from the same treatment. Whole body β–hydroxyacyl coenzymeA dehydrogenase activity was similar in fish from both treatments, but citrate synthase activity was significantly lower in effluent exposed fish. Our results suggest uranium mill effluent exposure in the laboratory affects aerobic energy metabolism and swimming performance in juvenile fathead minnow, which could affect wild fish survivability.

Short-term exposure to a treated sewage effluent alters reproductive behaviour in the three-spined stickleback ( Gasterosteus aculeatus)

Some UK sewage treatment work (STW) effluents have been found to contain high levels of anti-androgenic activity, but the biological significance of this activity to fish has not been determined. The aim of this study was to investigate the effects of exposure to a STW effluent with anti-androgenic activity on the reproductive physiology and behaviour of three-spined sticklebacks ( Gasterosteus aculeatus). Fish were exposed to a STW effluent (50 and 100%, v/v) with a strong anti-androgenic activity (328.56 ± 36.83 μg l −1 flutamide equivalent, as quantified in a recombinant yeast assay containing the human androgen receptor) and a low level of oestrogenic activity (3.32 ± 0.66 ng l −1 oestradiol equivalent, quantified in a recombinant yeast assay containing the human oestrogen receptor) for a period of 21 days in a flow-through system in the laboratory. Levels of spiggin, an androgen-regulated protein, were not affected by the STW effluent exposure, nor were levels of vitellogenin (a biomarker of oestrogen exposure), but the reproductive behaviour of the males was impacted. Males exposed to full strength STW effluent built fewer nests and there was a significant reduction in male courtship behaviour for exposures to both the 50 and 100% STW effluent treatments compared with controls. The effect seen on the reproduction of male sticklebacks may not necessarily have been as a consequence of the endocrine active chemicals present in the STW effluent alone, but could relate to other features of the effluent, such as turbidity that can impair visual signalling important for courtship interactions. Regardless the specific causation, the data presented show that effluents from STW have an impact on reproductive behaviour in male sticklebacks which in turn affects reproductive performance/outcome. The study further highlights the use of fish behaviour as a sensitive endpoint for assessing potential effects of contaminated water bodies on fish reproduction.

The use of field-based mesocosm systems to assess the effects of uranium milling effluent on fathead minnow (Pimephales promelas) reproduction

Northern Saskatchewan, Canada is home to a uranium milling operation that discharges a complex milling effluent containing nutrients, cations and anions, and many metals including selenium (Se). Se has the potential to accumulate in a system even when water concentrations are low. This study evaluated the effects of treated uranium milling effluent and contaminated sediment in combination and in isolation to determine the contribution and importance of each source to fathead minnow (Pimephales promelas) reproduction and survival. Trios of fathead minnows were allocated to one of four treatments for 21-days where the following were evaluated; survival (adult and 5 day larval), larval deformities, reproductive effects (egg production, spawning events) and metal tissue burdens (muscle, gonad, eggs and larvae). In addition Se speciation analysis was conducted on fish tissues. Effects were solely effluent-mediated with little contribution observed due to the presence of contaminated sediments. The contaminated sediments tested were taken from the actual receiving environment and represented the sediment composition found in greatest abundance. Results showed egg production significantly increased in the effluent treatments compared to the reference water treatments. Although egg production increased following effluent exposure, there was reduced hatching and larval survival and a significant increase in skeletal deformities in 5 day old larvae. Despite these effects on the offspring, when examined in an integrated manner relative to increased egg production, the mean number of normal larvae did not differ among treatments. Total selenium significantly increased in the effluent exposed, algae, female muscle, gonad, eggs and larvae in addition to other metals. A shift in the proportion of species of selenium was evident with changing exposure conditions. Biofilm/algae was key in the transfer of available Se into the food chain from the water and a source of direct dietary exposure in fish and possibly invertebrates.

Biological Degradation of Common Pharmaceuticals and Personal Care Products in Soils with High Water Content

Biological degradation rates of six pharmaceuticals and personal care products were examined in soil from a land application site and in adjacent soil with no prior history of effluent exposure. Microbial degradation rates were compared over 2 weeks under standing water or saturated conditions and draining conditions after having been saturated for 3 days. Biological degradation of 17β-estradiol exhibited rapid rates of biological degradation under both saturated and draining conditions. Half-lives for 17β-estradiol ranged from 1.5 to 4 days; 66–97% was lost from the soils. Estriol showed a pattern of biological degradation in both saturated and draining conditions though the half-lives were longer (8.7–25.9 days) than those observed for 17β-estradiol. Twenty-eight percent to 73% of estriol was lost over the 14 days treatment period. Estrone and 17α-ethinylestradiol exhibited slower rates of biological transformation under saturated and draining conditions. Half-lives for estrone ranged between 27.5 and 56.8 days with loss of at most 21%. 17α-ethinylestradiol exhibited half-lives of 22.6–207 days. Half-life data for ibuprofen ranged from 30.4 to 1,706.4 days in this experiment. Losses of up to 17% were observed in draining soils. Triclosan loss was at most 10%, and half-lives were 70.9–398.8 days. In all cases, soils that were draining from saturated conditions exhibited faster degradation rates than soils that remained saturated. Prior exposure of the soil to effluent did not always result in higher biological degradation rates.

River phosphorus cycling: Separating biotic and abiotic uptake during short-term changes in sewage effluent loading

Medium to small scale point sources continue to threaten river ecosystems through P loadings. The capacity and timescales of within-river processing and P retention are a major factor in how rivers respond to, and protect downstream ecosystems from, elevated concentrations of soluble reactive P (SRP). In this study, the bio-geochemical response of a small river (∼40 km 2 catchment area) was determined before, during and after exposure to a fourteen day pulse of treated sewage effluent using an upstream reach as a control. A wide array of approaches (batch and column simulations to in-situ whole stream metabolism) allowed independent comparison and quantification, of the relative contribution of abiotic and biotic processes in-river P cycling. This enabled, for the first time, separating the relative contributions of algae, bacteria and abiotic sorption without the use of labelled P (radioisotope). An SRP mass balance showed that the ecosystem switched from a P sink (during effluent inputs) to a P source (when effluent flow ceased). However, 65–70% of SRP was retained during the exposure time and remained sequestered two-weeks after-effluent flow ceased. Batch studies treated with biocide gave unrealistic results, but P uptake rates derived by other methods were highly comparable. Downstream of the effluent input, net P uptake by algae, bacteria and sediment (including the biofilm polysaccharide matrix) were 0.2 (±0.1), 0.4 (±0.3), and 1.0 (±0.9) mmol m −2 day −1 during effluent exposure. While autotrophic production did not respond to the effluent exposure, heterotrophic production increased by 67% relative to the control and this translated into a 50% increase in biological P uptake rate. Therefore, both biological and abiotic components of stream ecosystems uptake P during exposure to treated sewage effluent P inputs, and maintain a long ‘memory’ of this input in terms of P storage for considerable timescales after loading.

Recent Progress in Understanding the Causes of Endocrine Disruption Related to Pulp and Paper Mill Effluents

Abstract Reproductive impairment in fishes exposed to pulp and paper effluent has been observed in the wild for three decades. Despite intense research, identification of causative agents, if indeed the changes are strictly chemically mediated, has yet to be achieved. This review examines the body of research developed over the last six years that was directed at understanding the mechanisms of reproductive dysfunction in fishes associated with pulp mill effluent exposures. Research has continued to show diminishing effects of effluent exposure on fish reproductive physiology. Observations of exposure to androgens and estrogens continue to be observed and new evidence suggests that antiestrogens may be present. The presence of androgenic steroids remains a consistent observation, and those androgens appear to be native to wood. Recent studies have also suggested a number of alternate mechanisms beyond androgens or estrogens. One such possibility is that neuroactive substances are interfering with endocrine balance critical to reproduction. A second possibility is that some reproductive effects in the field may be due to nutritional factors, and thus reproductive impacts are caused by indirect effects of pulp mill effluent exposure. Ongoing mechanistic studies, particularly with paired field-lab components, are required to make further progress.