Internal nutrients dominate load and drive hypoxia in a eutrophic estuary.

The hypothesis that local hypoxia and chlorophyll concentration are spatially tethered to local, sediment-driven nutrient release was examined in a small, nutrient-impacted estuary in the Southern Gulf of St. Lawrence, Canada. Sediment reactor core samples were taken at 10 locations between 0.25 and 100% of the estuary area in spring and fall (2019) and used to estimate nitrogen and phosphate flux. Sediment organic matter, carbonate, percent nitrogen, percent carbon, δ13C, and δ15N were measured from the reactor core stations. Oxygen was recorded continually using oxygen loggers while chlorophyll and salinity were measured bi-weekly. A hydrodynamic model was used to determine water renewal time at each station. The most severe eutrophication effects were in the upper one-fifth of the estuary. There were strong local relationships between sediment biogeochemistry, hypoxia, and chlorophyll metrics but not with water renewal time. Internal nutrient loading represented 65% and 69% of total N loading, and 98% and 89% of total P loading to the estuary in June and September, respectively. Sediment nitrogen flux was highly predictable from a range of local sediment variables that reflect either nutrient content, or organic carbon enrichment in general. Percent nitrogen and percent carbon were highly correlated but sediment P flux was poorly predicted from sediment parameters examined. The highest correlations were with percent nitrogen and percent carbon. These results indicate that incorporating internal nutrient loading into nutrient monitoring programs is a critical next step to improve predictive capacity for eutrophication endpoints and to mitigate nutrient effects.

The Differential Effects of Salinity Level on Chlorpyrifos and Imidacloprid Toxicity to an Estuarine Amphipod.

Agricultural activity within coastal watersheds results in estuaries becoming the receiving environment for pesticide inputs. In estuaries, salinity can alter insecticide responses of exposed crustaceans. The acute toxicity of environmentally relevant doses of chlorpyrifos and imidacloprid were examined using the euryhaline amphipod Gammarus lawrencianus at 20 and 30 Practical Salinity Units (PSU). Responses were recorded every 24 h until an incipient (threshold) L(E)C50 was reached. For chlorpyrifos, LC50 ranged from 0.1 to 0.5 µg/L and was two-fold higher at 30 vs. 20 PSU at all time-points over the 96 h exposure. Imidacloprid immobility EC50 ranged from 4 to 40 µg/L over the 144 h exposure. An effect of salinity was only observed at 48 h and the EC50 values showed 1.4 times more potency at 20 PSU compared to 30 PSU. Measured concentrations of both compounds did not differ between salinities. Acetylcholinesterase activity in chlorpyrifos exposed amphipods showed no salinity effect at 96 h. We conclude that salinity level alters G. lawrencianus susceptibility to chlorpyrifos exposure, but not imidacloprid.

Assessing reproductive effects on fish populations: an evaluation of methods to predict the reproductive strategy of fishes.

Many environmental monitoring programs include an assessment of the health of fish populations using a sentinel species and include an indicator of reproductive potential. Knowledge of the reproductive strategy of the fish species is critical for data interpretation but is not always known. The reproductive strategy of a species can be determined from detailed histological analyses of ovaries throughout the reproductive cycle; however, these studies can be costly and can delay the implementation of a monitoring program. Three quick and cost-effective methods of predicting the reproductive strategy (annual single spawning or annual multiple spawning) are evaluated in this study using predicted probabilities from binary logistic regression models as a means of classifying the reproductive strategies of 18 different fish species in Atlantic Canada. The first method was based on the hypothesis that the variability in the ovary weight-body weight relationship in prespawning females is higher in multiple spawners. This method did not have a good classification rate due to some multiple spawners having low variability. The other two methods involved predictor variables representing the proportion of oocytes in different stages of development and predictor variables representing the distribution of oocyte sizes during the prespawning season for 111 fish (25 different samples for species). Predicted probabilities from these regression models could be used to correctly classify the reproductive strategies of all 25 samples (development stage model) and all but one sample (oocyte size distribution model). These models can be used to estimate the reproductive strategy of a species from a single sample of fish collected during the prespawning period to support species selection and data interpretation in environmental monitoring programs.

Oil Sands Derived Naphthenic Acids Are Oxidative Uncouplers and Impair Electron Transport in Isolated Mitochondria.

Naphthenic acids (NAs) are predominant compounds in oil sands influenced waters. These acids cause numerous acute and chronic effects in fishes. However, the mechanism of toxicity underlying these effects has not been fully elucidated. Due to their carboxylic acid moiety and the reported disruption of cellular bioenergetics by similar structures, we hypothesized that NAs would uncouple mitochondrial respiration with the resultant production of reactive oxygen species (ROS). Naphthenic acids were extracted and purified from 17-year-old oil sands tailings waters yielding an extract of 99% carboxylic acids with 90% fitting the classical O2-NA definition. Mitochondria were isolated from rainbow trout liver and exposed to different concentrations of NAs. Mitochondrial respiration, membrane potential, and ROS emission were measured using the Oroboros fluorespirometry system. Additionally, mitochondrial ROS emission and membrane potential were evaluated with real-time flow cytometry. Results showed NAs uncoupled oxidative phosphorylation, inhibited respiration, and increased ROS emission. The effective concentration (EC50) and inhibition concentration (IC50) values for the end points measured ranged from 21.0 to 157.8 mg/L, concentrations similar to tailings waters. For the same end points, EC10/IC10 values ranged from 11.8 to 66.7 mg/L, approaching concentrations found in the environment. These data unveil mechanisms underlying effects of NAs that may contribute to adverse effects on organisms in the environment.

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.

Monitoring stream sediment loads in response to agriculture in Prince Edward Island, Canada.

Increased agricultural land use leads to accelerated erosion and deposition of fine sediment in surface water. Monitoring of suspended sediment yields has proven challenging due to the spatial and temporal variability of sediment loading. Reliable sediment yield calculations depend on accurate monitoring of these highly episodic sediment loading events. This study aims to quantify precipitation-induced loading of suspended sediments on Prince Edward Island, Canada. Turbidity is considered to be a reasonably accurate proxy for suspended sediment data. In this study, turbidity was used to monitor suspended sediment concentration (SSC) and was measured for 2 years (December 2012-2014) in three subwatersheds with varying degrees of agricultural land use ranging from 10 to 69 %. Comparison of three turbidity meter calibration methods, two using suspended streambed sediment and one using automated sampling during rainfall events, revealed that the use of SSC samples constructed from streambed sediment was not an accurate replacement for water column sampling during rainfall events for calibration. Different particle size distributions in the three rivers produced significant impacts on the calibration methods demonstrating the need for river-specific calibration. Rainfall-induced sediment loading was significantly greater in the most agriculturally impacted site only when the load per rainfall event was corrected for runoff volume (total flow minus baseflow), flow increase intensity (the slope between the start of a runoff event and the peak of the hydrograph), and season. Monitoring turbidity, in combination with sediment modeling, may offer the best option for management purposes.

In vitro assessment of endocrine disrupting potential of naphthenic Acid fractions derived from oil sands-influenced water.

Oil sands-influenced process waters have been observed to cause reproductive effects and to induced CYP1A activity in fishes; however, little progress has been made in determining causative agents. Naphthenic acids (NAs) are the predominant organic compounds in process-affected waters, but due to the complexity of the mixture, it has been difficult to examine causal linkages in fishes. The aim of this study was to use in vitro assays specific to reproductive and CYP1A mechanisms to determine if specific acid extractable fractions of NAs obtained from oil sands-influenced water are active toward reproductive processes or interact with the Ah receptor responsible for CYP1A activity. NAs were extracted from aged oil sands-influenced waters by use of acid precipitation, and the mixture was fractionated into three acidic and one neutral fraction. The four fractions were examined for Ah receptor-mediated potency by use of the H4IIE-luc bioassay, effects on production of steroid hormones by use of the H295R steroidogenesis assay, and sex steroid receptor binding activity using the yeast estrogen screen and yeast androgen screen. The mixtures were characterized by high resolution mass spectrometry, (1)H nuclear magnetic resonance, and attenuated total reflectance infrared spectroscopy. The neutral fraction elicited Ah-receptor mediated activity after 24 h but not after 48 or 72 h. None of the fractions contained measurable levels of estrogen or androgen receptor agonists nor did they cause reductions in steroidogenesis. A number of fractions showed antiestrogenic or antiandrogenicity potency, with the neutral and main acidic fractions being the most potent. Neutral aromatic compounds are likely responsible for the CYP1A activity observed. Direct estrogenic, androgenic, or steroidogenic mechanisms are unlikely for NAs based on these results, but NAs act as potent antiandrogen or antiestrogens.

Reproductive strategies and seasonal changes in the somatic indices of seven small-bodied fishes in Atlantic Canada in relation to study design for environmental effects monitoring.

Small-bodied fishes are more commonly being used in environmental effects monitoring (EEM) studies. There is a lack of understanding of the biological characteristics of many small-bodied species, which hinders study designs for monitoring studies. For example, 72% of fish population surveys in Canada's EEM program for pulp and paper mills that used small-bodied fishes were conducted outside of the reproductive period of the species. This resulted in an inadequate assessment of the EEM program's primary effect endpoint (reproduction) for these studies. The present study examined seasonal changes in liver size, gonad size, and condition in seven freshwater and estuarine small-bodied fishes in Atlantic Canada. These data were used to examine differences in reproductive strategies and patterns of energy storage among species. Female gonadal recrudescence in all seven species began primarily in the 2-month period in the spring before spawning. Male gonadal development was concurrent with females in five species; however, gonadal recrudescence began in the fall in male three-spined stickleback (Gasterosteus aculeatus) and slimy sculpin (Cottus cognatus). The spawning period for each species was estimated from the decline in relative ovary size after its seasonal maximum value in spring. The duration of the spawning period reflected the reproductive strategy (single vs multiple spawning) of the species. Optimal sampling periods to assess reproductive impacts in each species were determined based on seasonal changes in ovary size and were identified to be during the prespawning period when gonads are developing and variability in relative gonad size is at a minimum.

A model naphthenic acid decouples oxidative phosphorylation through selective inhibition of mitochondrial complex activity.

The naphthenic acid fraction compound (NAFC), 3,5-dimethyladamantane-1-acetic acid, was tested for its ability to uncouple mitochondrial oxidative phosphorylation. Mitochondria isolated from rainbow trout (Oncorhynchus mykiss) liver were exposed to 3,5-dimethyladamantane-1-acetic acid in state 3 and 4 respiration, and mitochondrial membrane potential were quantified. Electron transport chain (ETC) protein complexes were isolated using pharmacological agents and inhibitors, and their activities measured. The NAFC compound completely inhibited states 3 and 4 respiration with an IC50 of 0.77 and 1.01 mM, respectively. The NAFC compound partially uncoupled mitochondrial membrane potential in state 3 and 4 respiration with an IC50 of 2.19 and 1.73 mM, respectively. The NAFC impaired the activities of ETC protein complexes with a 9.5-fold range in sensitivity. The relative inhibitory effect of the ETC protein complexes to NAFC was CIV≥CI>CIII>CII. The impairment of mitochondrial oxidative phosphorylation by adamantane 3,5-dimethyladamantane-1-acetic acid is mediated via its inhibition of ETC protein complexes.

A survey of Canadian mechanical pulp and paper mill effluents: insights concerning the potential to affect fish reproduction.

Building on breakthroughs recently made at kraft mills, a survey of mechanical pulp and paper mill effluents was undertaken to gain insights concerning potential effects on fish reproduction. Effluents from seven Canadian mills were characterized chemically for conventional parameters such as biochemical oxygen demand (BOD) and total suspended solids (TSS). Each sample was further subjected to solvent extraction followed by gas chromatographic separation for the determination of resin/fatty acids and for the estimation of a gas chromatography (GC) profile index. Each mill effluent was assessed for the potential to affect fish reproduction in the laboratory using a five day adult fathead minnow (Pimephales promelas) egg production bioassay with exposures to 100% effluent. The seven effluents were found to have substantial variation both in terms of chemical characterization and effects on fish reproduction. Temporal variations were also noted in effluent quality at mills sampled on different occasions. Similar to what has been observed for kraft mills, a general trend of greater reductions in egg production caused by effluents with greater BOD concentrations and GC profile indices was noted. Effluents with BOD > 25 mg/L and GC Profile indices >5.0 caused a complete cessation of egg production. At the same time, about half of the total effluents sampled had BOD < 25 mg/L and GC profile indices <5.0 and caused no significant effects on egg production, suggesting these values may be useful as effluent quality targets for mechanical pulp and paper mills. However, 3 out of 14 effluents sampled had BOD < 25 mg/L and GC profile indices <5.0 and caused significant reductions in egg production. The reason(s) for reproductive effects caused by such effluents is presently unclear. The effluent quality parameters considered in this study may require further refinement to address their utility in predicting the adverse reproductive effects induced by effluents from mechanical pulp and paper mills.

Immunological impacts of oil sands-affected waters on rainbow trout evaluated using an in situ exposure.

Rainbow trout were exposed in situ to oil sands-affected waters for 21 d, either with or without an immune stimulation using inactivated Aeromonas salmonicida. Three aquatic systems were utilized for the experiment: a pond containing oil sands tailings capped with approximately 3 m of natural surface water, a second pond where unextracted oil sands materials were deposited in the watershed, and a reservoir receiving Athabasca River water as a reference caging location. The three systems showed a gradient of oil sands-related compounds, most notably, total naphthenic acids were highest in the system containing tailings (13 mg/L), followed by the system influenced by unextracted oil sands (4 mg/L), followed by the reference cage location (1 mg/L). Biochemical and chemical measures of exposure in rainbow trout showed the same trend, with the tailings-influenced system having the highest hepatic EROD activity and elevated bile fluorescence measured at phenanthrene wavelengths. Trout caged in the tailings-influenced location had significantly fewer leukocytes and smaller spleens as compared to the reference fish, though liver size and condition factor were unaffected. Fish in the tailings-influenced waters also demonstrated increased fin erosion, indicative of opportunistic infection. The trout exposed to tailing-influenced waters also showed a significantly decreased ability to produce antibodies to the inactivated A. salmonicida. Given the complexity of the exposure conditions, exact causative agents could not be determined, however, naphthenic acids, polycyclic aromatic hydrocarbons and pH correlate with the immunotoxic effects while elevated salinity or metals seem unlikely causes.

Effluent monitoring at a bleached kraft mill: directions for best management practices for eliminating effects on fish reproduction.

A long-term monitoring study was conducted on effluents from a bleached kraft pulp and paper mill located in Eastern Canada. The study was designed to gain insights into temporal effluent variability with respect to fish reproduction as it related to production upsets, mill restarts and conditions affecting biological treatment performance. Final effluent quality was monitored between February 2007 and May 2009 using biochemical and chemical oxygen demand, total suspended solids, resin and fatty acids, a gas chromatographic profiling index, and the presence of methyl substituted 2-cyclopentenones. Selected effluent samples were evaluated for effects on fish reproduction (egg production) using a shortened version of the adult fathead minnow reproductive test. The events relating to negative effects on fish reproduction were upsets of the pulping liquor recovery system resulting in black liquor losses, operational upsets of the hardwood line resulting in the loss of oxygen delignification filtrates, and conditions that reduced the performance of biological treatment (e.g., mill shutdown and low ambient temperatures). The reductions in egg production observed in fathead minnow were associated with biochemical oxygen demand values > 20 mg/L, GC profiling indices > 1.2 and the presence of methyl-substituted 2-cyclopentenones at concentrations > 100 µg/L. This study demonstrated the importance of both in-plant measures for controlling the loss of organics as well as the optimum operation of biological effluent treatment for eliminating effluent-related effects on fish reproduction (egg production) in the laboratory.

Exposure of American lobster (Homarus americanus) to the pesticide chlorpyrifos results in changes in gene expression.

Chlorpyrifos is an organophosphate that is currently used to reduce arthropod pests for the protection of agricultural crops. Coastal marine ecosystems may be exposed to agricultural pesticides via runoff and pesticide exposure can impact the health and survival of non-target species such as the American lobster (Homarus americanus). In the current study, the gene expression changes of H. americanus stage IV larvae were evaluated to understand the physiological mechanisms affected by exposure to sublethal concentrations of chlorpyrifos. After 48 h chlorpyrifos exposure, surviving lobsters were processed for Illumina RNA sequencing (RNA-seq). Genes of interest that showed significant changes using RNA-seq were verified using reverse transcriptase quantitative polymerase chain reaction (RT-qPCR). Analysis of RNA-seq and the confirmation of gene expression patterns via RT-qPCR found altered expression in genes related to stress response (glutathione peroxidase 3 and heat shock protein 60), hypoxia response (hairy, astakine 2, hemocyanin), moulting (cytochrome P450 307a1 and chitinase), and immunity (astakine 2) pathways. Changes to gene expression were most notable in lobsters exposed to 0.57 µg/L chlorpyrifos.

Effects of Sublethal Chlorpyrifos Exposure on Postlarval American Lobster (Homarus americanus).

The organophosphate pesticide chlorpyrifos has been introduced to the marine environment via adsorption to agricultural soil runoff or as spray drift. Chlorpyrifos affects the survival of some larval decapod crustaceans, but no data exist on the impacts to the American lobster, Homarus americanus. The purpose of the present study was to assess the levels at which chlorpyrifos affects the survival of postlarval H. americanus. Using acute saltwater exposures, the 24- and 48-h median lethal concentrations were established for stage IV H. americanus (1.56 and 1.33 µg/L, respectively). Movement, acetylcholinesterase activity, intermoult period, specific growth rate, and moult increment were measured during exposure to sublethal concentrations. Movement patterns were assessed to establish a 48-h median inhibition concentration for cessation of normal movement (0.66 µg/L). Acetylcholinesterase activity was found to be inhibited immediately post-exposure to 0.50, 0.57, and 0.82 µg/L chlorpyrifos but could be recovered within a period (9-15 d) in clean seawater. Sublethal growth effects of increased intermoult period, decreased specific growth rate, and decreased moult increment were observed during exposure to an environmentally relevant concentration (0.82 µg/L). The present study suggests that H. americanus stage IV larvae were marginally less sensitive to chlorpyrifos compared with other decapods and that acute lethality of H. americanus postlarvae is not likely to occur with chlorpyrifos concentrations previously reported from aquatic environments. Environ Toxicol Chem 2019;38:1294-1301. © 2019 SETAC.

Distinct migratory and non-migratory ecotypes of an endemic New Zealand eleotrid (Gobiomorphus cotidianus) - implications for incipient speciation in island freshwater fish species.

BACKGROUND: Many postglacial lakes contain fish species with distinct ecomorphs. Similar evolutionary scenarios might be acting on evolutionarily young fish communities in lakes of remote islands. One process that drives diversification in island freshwater fish species is the colonization of depauperate freshwater environments by diadromous (migratory) taxa, which secondarily lose their migratory behaviour. The loss of migration limits dispersal and gene flow between distant populations, and, therefore, is expected to facilitate local morphological and genetic differentiation. To date, most studies have focused on interspecific relationships among migratory species and their non-migratory sister taxa. We hypothesize that the loss of migration facilitates intraspecific morphological, behavioural, and genetic differentiation between migratory and non-migratory populations of facultatively diadromous taxa, and, hence, incipient speciation of island freshwater fish species. RESULTS: Microchemical analyses of otolith isotopes (88Sr, 137Ba and 43Ca) differentiated migratory and non-migratory stocks of the New Zealand endemic Gobiomorphus cotidianus McDowall (Eleotridae). Samples were taken from two rivers, one lake and two geographically-separated outgroup locations. Meristic analyses of oculoscapular lateral line canals documented a gradual reduction of these structures in the non-migratory populations. Amplified fragment length polymorphism (AFLP) fingerprints revealed considerable genetic isolation between migratory and non-migratory populations. Temporal differences in reproductive timing (migratory = winter spawners, non-migratory = summer spawners; as inferred from gonadosomatic indices) provide a prezygotic reproductive isolation mechanism between the two ecotypes. CONCLUSION: This study provides a holistic look at the role of diadromy in incipient speciation of island freshwater fish species. All four analytical approaches (otolith microchemistry, morphology, spawning timing, population genetics) yield congruent results, and provide clear and independent evidence for the existence of distinct migratory and non-migratory ecotypes within a river in a geographically confined range. The morphological changes within the non-migratory populations parallel interspecific patterns observed in all non-migratory New Zealand endemic Gobiomorphus species and other derived gobiid taxa, a pattern suggesting parallel evolution. This study indicates, for the first time, that distinct ecotypes of island freshwater fish species may be formed as a consequence of loss of migration and subsequent diversification. Therefore, if reproductive isolation persists, these processes may provide a mechanism to facilitate speciation.

Carotenoid depletion and ovarian dysfunction in common bully exposed to a New Zealand pulp mill effluent.

In a previous cumulative impacts assessments of fish populations along the Waikato River, New Zealand, reduced ovary pigmentation was observed in the common bully (Gobiomorphus cotidianus) with an associated reduction in recruitment in the pulp mill-impacted Kopakorahi Arm of Lake Maraetai compared to an upstream reference site. Carotenoid pigments are typically responsible for the yellow to red colouration of fish eggs and a number of functions for these pigments such as pro-vitamins and antioxidants have been proposed. The purpose of the current study was first to quantify the observed pigment reduction and then to establish possible consequences of carotenoid deficiency. Significantly lower levels of all measured carotenoids were observed in the ovaries of fish from below the mill outfall compared to the reference site. Using in vitro incubations of ovarian follicles, it was also found that these fish had a significantly reduced ability to produce the sex steroids testosterone and estradiol. This study demonstrates changes in ovarian carotenoids associated with pulp mill effluent exposure. Significantly reduced biosynthetic capacity of ovarian follicles demonstrates reproductive endocrine disruption potential associated with exposure to this particular effluent. These results may provide an important link between reproductive dysfunction, oxidative stress and food chain modifications in pulp mill-impacted systems.

Altered reproduction in fish exposed to pulp and paper mill effluents: roles of individual compounds and mill operating conditions.

For the last 20 years, studies conducted in North America, Scandinavia, and New Zealand have shown that pulp and paper mill effluents affect fish reproduction. Despite the level of effort applied, few leads are available regarding the factors responsible. Effluents affect reproduction in multiple fish species, as evidenced by decreased gonad size, decreased circulating and gonadal production of reproductive steroids, altered expression of secondary sex characteristics, and decreased egg production. Several studies also have shown that effluent constituents are capable of accumulating in fish and binding to sex steroid receptors/ binding proteins. Studies aimed at isolating biologically active substances within the pulping and papermaking process have provided clues about their source, and work has progressed in identifying opportunities for in-mill treatment technologies. Following comparisons of manufacturing processes and fish responses before and after process changes, it can be concluded that effluent from all types of mill processes are capable of affecting fish reproduction and that any improvements could not be attributed to a specific process modification (because mills normally performed multiple modifications simultaneously). Improved reproductive performance in fish generally was associated with reduced use of molecular chlorine, improved condensate handling, and liquor spill control. Effluent biotreatment has been effective in reducing some effects, but biotreated effluents also have shown no difference or an exacerbation of effects. The role of biotreatment in relation to effects on fish reproduction remains unclear and needs to be resolved.

Opportunistic disease in yellow perch in response to decadal changes in the chemistry of oil sands-affected waters.

Oil sands-affected water from mining must eventually be incorporated into the reclaimed landscape or treated and released. However, this material contains petrogenic organic compounds, such as naphthenic acids and traces of polycyclic aromatic hydrocarbons. This has raised concerns for impacts of oil sands process-affected waters on the heath of wildlife and humans downstream of receiving environments. The objective of this study was to evaluate the temporal association of disease states in fish with water chemistry of oil sands-affected waters over more than a decade and determine the pathogens associated with disease pathologies. Yellow perch (Perca flavescens) captured from nearby lakes were stocked into two experimental ponds during 1995-1997 and 2008-2010. South Bison Pond is a drainage basin that has received unextracted oil sands-contaminated material. Demonstration Pond is a constructed pond containing mature fine tailings capped with fresh water. Two disease pathologies, fin erosion for which a suspected bacterial pathogen (Acinetobacter Iwoffi) is identified, and lymphocystis (confirmed using a real-time PCR) were associated with oil sands-affected water exposure. From 1995 to 1997 pathologies were most prevalent in the South Bison Pond; however, from 2008 to 2009, disease was more frequently observed in the Demonstration Pond. CYP1A activity was 3-16 fold higher in fish from experimental ponds as compared to reference populations and this pattern was consistent across all sampling years. Bile fluorescence displayed a gradient of exposure with experimental ponds being elevated over local perch populations. Naphthenic acids decreased in the Bison Pond from approximately 12 mg/L to <4 mg/L while naphthenic acids increased in the Demonstration Pond from 6 mg/L to 12 mg/L due to tailings densification. Temporal changes in naphthenic acid levels, CYP1A activity and bile fluorescent metabolites correlate positively with incidence of disease pathologies whereas all inorganic water quality changes (major ions, pH, metals) were not associated with disease responses.

An empirical model using dissolved oxygen as an indicator for eutrophication at a regional scale.

Simple empirical models can sometimes capture salient patterns without sacrificing predictive capacity when compared to more complex models. Herein we examine dissolved oxygen as an indicator of eutrophication status for shallow estuaries. Dissolved oxygen was measured hourly in the upper estuary of 15 watersheds along a nutrient-loading and geographic gradient. Metrics describing hypoxia and supersaturation were devised and then analyzed using multivariate statistics. Results revealed independent responses for hypoxia and supersaturation with hypoxia-related metrics correlating strongly with water residence. A metric integrating hypoxia and supersaturation effectively discriminated between seagrass and algae-dominated habitats and was significantly correlated with both water residence and nitrate-N loading. Chlorophyll, measured bi-weekly, was not correlated with our predictor variables likely because it does not account for benthic production. Over 70% of the variability in hypoxia was explained by water residence and nitrate-N loading indicating that this model can be of use for managers.

Transcriptional response in rainbow trout (Oncorhynchus mykiss) B cells and thrombocytes following in vivo exposure to benzo[a]pyrene.

Immune toxicity of polycyclic aromatic hydrocarbons (PAHs) in fishes has been frequently reported but the reasons for differential cell toxicity remains unclear. Rainbow trout were exposed in vivo with a single intraperitoneal injection of corn oil or 100mg/kg of the immunotoxic PAH benzo[a]pyrene (B[a]P) in corn oil. Leukocytes were harvested from head kidney, spleen and blood after 14days, the optimal time for B cell depletion found in a previous study. The mRNA expression of five cytochrome P450 (CYP) enzymes, the aryl hydrocarbon receptor (AhR), and an intrinsic pathway apoptosis checkpoint (p53) in B cells and thrombocytes were examined. Transcript levels were measured in immunomagnetically-isolated B cells and thrombocytes from those tissues as well as in liver as B cells had been previously shown to be responsive the BaP whereas thrombocytes were not. There was induction of CYP1A1 in liver, blood B cells, and blood and spleen thrombocytes; CYP1B1 in blood B cells, blood and spleen thrombocytes; CYP1A3 in liver, blood and spleen B cells, and blood thrombocytes; CYP1C1 in liver; and AhR in liver and spleen thrombocytes. There was no change in CYP1C2, or p53 mRNA levels across tissues or cell type. Induction in mRNA was observed 14 d after exposure, indicating a prolonged physiological effect of a single B[a]P injection. CYP1A1 and CYP1A3 were the most abundantly expressed CYP genes and CYP1B1 was generally least abundant. B[a]P-induced thrombocytes had a significantly different pattern of CYP expression than either liver or B cells. Given the importance of metabolites in the toxicity of PAHs, differences in CYP expression between tissues may explain differences in toxicity previously observed between B cells and thrombocytes.