Previous studies at the Key Lake uranium mill (Saskatchewan, Canada) suggested the complex effluent discharged alters energetic stores of resident fish species. A second study at the same site demonstrated certain fish from lakes downstream of the mill produce larvae with elevated incidence of developmental deformities. The mechanisms by which energy homeostasis is affected in fish downstream of the Key Lake uranium mill are unknown, and the effects of deformities and altered metabolism on swimming ability have not been explored. Therefore, the overall objective of this thesis was to investigate whether effluent exposed fish exhibited differences in swimming performance and energy homeostasis. To achieve this objective two experiments were conducted. In the first experiment juvenile spottail shiner (Notropis hudsonius) were collected from a lake downstream of the Key Lake uranium mill, and compared to fish collected from a nearby reference lake. In the second experiment larvae were collected from laboratory raised fathead minnow (Pimephales promelas) exposed to 5% diluted uranium mill effluent or control (dechlorinated municipal) water, and reared in the same treatments to 60 days post hatch (dph). No gross deformities were observed in any fish, and only shiner collected from the exposure lake in the field experiment had enlarged heart ventricles relative to body size compared to fish from the reference lake. Swimming performance was similar between shiner from the exposure and reference lakes in the field study, but effluent exposure impaired swimming ability in 60 dph fathead minnow in the laboratory experiment compared to fish from the control water treatment. After swimming performance tests fish were considered fatigued and metabolic endpoints were compared to non-fatigued fish. In both non-fatigued and fatigued shiner, liver glycogen was significantly greater in fish collected from the exposure lake compared to the reference lake. There was no difference in liver triglycerides in non-fatigued shiner between lakes, but liver triglycerides decreased after swimming in the field study reference fish. Muscle energy stores were unaffected by site or swimming in the field experiment. Conversely, whole body triglycerides and glycogen were similar between treatments in non-fatigued fathead minnow in the laboratory experiment. Swimming significantly decreased whole body triglycerides in fathead minnow from both treatments, but whole body glycogen was unaffected. In the field experiment blood endpoints (hematocrit, plasma glucose, lactate) in fatigued and non-fatigued shiner from both lakes further supported the possibility of altered intermediary metabolism or blunted stress response in fish downstream of the Key Lake uranium mill. In the field study, shiner muscle citrate synthase activity (an indicator of tissue aerobic capacity) was similar between lakes, but muscle âhydroxyacyl coenzyme A dehydrogenase activity (an indicator of tissue lipolytic capacity) was elevated. In contrast, laboratory fathead minnow whole body âhydroxyacyl coenzyme A dehydrogenase activity was similar between treatments, but citrate synthase activity was significantly lower in fathead minnow from the 5% effluent treatment. In summary, shiner from the exposure lake in the field experiment had similar swimming endurance and greater energy stores compared to fish from the reference lake, despite metabolic alterations. Fathead minnow from the 5% effluent treatment in the laboratory experiment had reduced swimming endurance that was matched by reduced whole body citrate synthase activity, but no other metabolic alterations were observed. Therefore, effluent exposure caused metabolic alterations in both fathead minnow and shiner, but specific effects between experiments were inconsistent. Overall, the physiological significance of the metabolic and swimming effects of effluent exposure is unclear, but suggests discharged effluent has the potential to negatively affect wild fish survivability.

The overall objective of the research conducted and described in this thesis was to develop an environmentally relevant bioassay to assess the effects of complex effluents on a sentinel fish species. A short-term fathead minnow (FHM) reproductive bioassay was utilized to assess the effects of industrial effluents on multiple levels of biological organization (sub-organismal to population endpoints). The FHM bioassay was tested in both lab and on-site investigations using an artificial stream system. The incorporation of trophic-transfer into the bioassay was also developed to quantify the importance of contaminated food as a source of exposure. This work was conducted in two key phases. Phase I focused on testing and developing the FHM bioassay, in the lab and on-site with pulp mill effluent (PME), to firstly document response patterns and, secondly, to conduct an investigation of cause study. Phase II focused on developing the trophic-transfer system to document responses to metal mine effluent (MME) in the lab and on-site in an artificial stream system. Development of the trophic-transfer system was also conducted during this phase to compare responses to standard water-only exposures. <p>In Phase I, exposure to PME in both the lab and field studies resulted in disruptions in egg production and spawning events. By focusing on identifying response patterns I was able to determine that the effects observed were indicative of an estrogenic response. I was also able to identify a process stream that was the potential cause of responses observed after exposure to final effluent. Isolation of this process stream will assist the mill in developing approaches for future mitigation. The results from this research will also provide additional data for the environmental effects monitoring (EEM) program for pulp and paper and investigation of cause studies on a national basis. <p> In Phase II, in both the field and laboratory investigations, significant decreases in reproductive output (egg production and spawning events) were observed in the water-only system exposures. Significant decreases in hatching success and increases in deformities were observed in the trophic-transfer system only, suggesting that the combination of both food and water was important in assessing the effects on the F1 generation. Overall, the responses in the trophic-transfer system were not comparable between the lab and field studies. In the lab study, significant decreases in reproductive output occurred, compared to the field study where significant increases in egg production and spawning events occurred. In addition, the effects on the F1 generation in the field study were not as severe as those observed in the lab investigation. It was concluded that the presence of reference water and the environment within the trophic-transfer system were responsible for this reduction in toxicity. <p> Phases I and II of this research have made significant contributions to artificial stream development within Canada for the assessment of industrial effluents and their effects on aquatic biota. The results from these studies have also demonstrated that environmentally relevant testing is essential if we are to accurately assess effects on aquatic biota. Future development and application of this bioassay should be towards developing a standardized approach for not only assessing the effects of industrial effluents in a comparative manner, but also in investigation of cause studies.

Metal mines release treated effluents that contain a variety of metals, metalloids, and organics into the aquatic environment. A number of metal mine effluents (MMEs) have been found to contribute to adverse effects in fish and benthic invertebrates, such as decreased diversity and density, however the specific causal factors of toxic responses during chronic exposures to the MMEs are often unknown. Therefore, the overall objective of this dissertation was to explore causative and modifying factors of MME toxicity to a resident fish species, the fathead minnow (Pimephales promelas), during chronic, multi-trophic exposures. The representative MME used in this study was the process water effluent (PWE) of a Canadian metal mine, which is released into Junction Creek in Sudbury, Ontario, Canada. Chronic exposure to the MME has been a source of decreased reproductive output in fathead minnows in several previous studies, however, these same studies were not able to determine the potential causal factors of the reproductive impairment. In order to address the overall objective, several laboratory mesocosm studies were conducted, which consisted of three separate components. The first component included exploring several metals (Cu, Ni, and Se; alone and in mixture) that are consistently present in the MME and are known to cause toxicity at fairly low concentrations as potential causes for decreased egg production in fathead minnows. The second component included evaluating the role of decreased food availability (a possible indirect effect of MME in the receiving environment) as a potential cause of decreased egg production in fathead minnows. The third and final component included examining the role of water chemistry [(increased alkalinity and dissolved organic carbon (DOC)] as potential modifying factors of chronic MME toxicity to fathead minnows. In general, my results suggest that the metals present in the MME likely do not contribute directly to decreased reproductive performance in fathead minnows during chronic exposures, under the conditions examined. Instead, the MME appears to decrease food availability, therefore indirectly influence fathead minnow egg production. Furthermore, water chemistry modifications tested in this thesis were not able to entirely mitigate the reproductive effects in fish induced by the MME, although they did improve egg production relative to unmodified MME. Metal concentrations in fish tissues were not influenced by increases to alkalinity or DOC level in the exposure water, suggesting that bioavailability of metals during chronic exposure to metal-mixtures cannot be fully explained based on our understanding of metal complexation with abiotic ligands (inorganic and organic) during single metal or acute exposures. From a regulatory perspective, water chemistry modifications may somewhat improve fathead minnow reproductive performance during chronic exposure to the MME, however the MME would still not be entirely free of effects relative to the uncontaminated water. Future studies should focus on understanding the factors responsible for decreased food availability in MME-impacted aquatic ecosystems, and further explore potential approaches for ameliorating effluent quality.

Throughout their lives, prey organisms must balance the tradeoff between fitness-related activities and the risk of predation. To successfully mediate such tradeoffs, prey must have an accurate method to gauge current predation risk. For many aquatic organisms, the use of chemosensory information has been shown to be a ubiquitous and useful tool in mediating predation risk. The chemical cues to which aquatic organisms respond include the odour of known predators and the odour of a damaged conspecific or known or closely related heterospecific. In fishes, the response to damage-released cues from conspecifics or closely related heterospecifics has been shown to be innate, while the response to distantly related unknown heterospecific cues are likely learned. In a series of laboratory and field studies I examined the role of learning in the ability of fathead minnows to respond to damage-released cues of brook stickleback as an indication of predation risk. My results indicate that minnows from a population without stickleback do not recognize stickleback cues as dangerous. However, following the introduction of stickleback, minnows learn to recognize stickleback cues as dangerous. Further study indicated a low ratio of stickleback to minnows in a given population will decrease the likelihood of learning when compared with a similar sized population containing a higher ratio of stickleback to minnows. I also demonstrated that an increase in habitat complexity decreases the ability of minnows to learn to recognize stickleback cues. Studies have further demonstrated that in the face of predation (as indicated by chemical cues from minnows and stickleback) minnows will decrease their antipredator response when in the presence of a fish shoal, especially a shoal of conspecifics. Finally, an examination of the effects of a minnows length, body condition and breeding status indicate that morphological parameters can play a significant role in the intensity of response to heterospecific and conspecific damage-released cues. Previous works have demonstrated that behavioural responses to heterospecific damage-released cues increase the probability of the receiver surviving a predation event. This has important implications for predator/prey interactions and consequently factors affecting the learning and use of such cues deserve increasing attention from ecologists.

The overall objective of the research conducted and described in this thesis was to develop an environmentally relevant bioassay to assess the effects of complex effluents on a sentinel fish species. A short-term fathead minnow (FHM) reproductive bioassay was utilized to assess the effects of industrial effluents on multiple levels of biological organization (sub-organismal to population endpoints). The FHM bioassay was tested in both lab and on-site investigations using an artificial stream system. The incorporation of trophic-transfer into the bioassay was also developed to quantify the importance of contaminated food as a source of exposure. This work was conducted in two key phases. Phase I focused on testing and developing the FHM bioassay, in the lab and on-site with pulp mill effluent (PME), to firstly document response patterns and, secondly, to conduct an investigation of cause study. Phase II focused on developing the trophic-transfer system to document responses to metal mine effluent (MME) in the lab and on-site in an artificial stream system. Development of the trophic-transfer system was also conducted during this phase to compare responses to standard water-only exposures. <p>In Phase I, exposure to PME in both the lab and field studies resulted in disruptions in egg production and spawning events. By focusing on identifying response patterns I was able to determine that the effects observed were indicative of an estrogenic response. I was also able to identify a process stream that was the potential cause of responses observed after exposure to final effluent. Isolation of this process stream will assist the mill in developing approaches for future mitigation. The results from this research will also provide additional data for the environmental effects monitoring (EEM) program for pulp and paper and investigation of cause studies on a national basis. <p> In Phase II, in both the field and laboratory investigations, significant decreases in reproductive output (egg production and spawning events) were observed in the water-only system exposures. Significant decreases in hatching success and increases in deformities were observed in the trophic-transfer system only, suggesting that the combination of both food and water was important in assessing the effects on the F1 generation. Overall, the responses in the trophic-transfer system were not comparable between the lab and field studies. In the lab study, significant decreases in reproductive output occurred, compared to the field study where significant increases in egg production and spawning events occurred. In addition, the effects on the F1 generation in the field study were not as severe as those observed in the lab investigation. It was concluded that the presence of reference water and the environment within the trophic-transfer system were responsible for this reduction in toxicity. <p> Phases I and II of this research have made significant contributions to artificial stream development within Canada for the assessment of industrial effluents and their effects on aquatic biota. The results from these studies have also demonstrated that environmentally relevant testing is essential if we are to accurately assess effects on aquatic biota. Future development and application of this bioassay should be towards developing a standardized approach for not only assessing the effects of industrial effluents in a comparative manner, but also in investigation of cause studies.

The overall objective of the work described in this thesis was to examine the effects of both waterborne and dietary routes of exposure to fathead minnow (Pimephales promelas) when exposed to complex metal mining mixtures. This was conducted using a 21-day, multi-trophic, short-term fathead minnow (FHM) reproductive bioassay. The endpoints that were measured were used to assess the effects on multiple levels of biological organization (sub-organismal to population endpoints). The first phase of this research was conducted in situ using environmentally realistic concentrations of 3 separate metal mining effluents [20% surface water effluent (SWE), 30% mine water effluent (MWE), 45% process water effluent (PWE)] from Sudbury, Ontario, Canada. Metals were analyzed in several media (water, sediments) and tissues (biofilm, Chironomus dilutus, female fathead minnow carcass, ovaries, liver and gills). The incorporation of the biofilm (primary producers) into the bioassay also added another level of organization that was novel to this study. Significant increases in metal concentrations were observed in the water and biofilm tissues in all treatments [SWE, MWE, PWE], compared to reference. Cobalt and nickel increased significantly in C. dilutus tissues in SWE (1.4-fold and 1.5-fold respectively), and copper and selenium in PWE (5.2-fold and 3.3-fold respectively), however no significant increases occurred in MWE compared to reference. There were no significant increases in metal concentrations in female FHM tissues (carcass, liver, gonads, gills) in any of the treatments, suggesting that metal bioavailability was reduced. Cumulative number of eggs per female per day increased significantly (+127%) after exposure to SWE and decreased significantly (-33%) after exposure to PWE when compared to the reference fish. Mean total number of days to hatch was also reduced in PWE compared to reference. In order to gain a better understanding of the routes of exposure causing toxicity in FHM, the second phase of this research examined the effects of exposure through diet, through water or through both using a fully factorial food exposure design in a laboratory setting. In this experiment we pre-exposed C. dilutus to both 45% PWE and laboratory control water until they reached the 3rd-4th instar stage of development (approximately 21 days) where they were collected and frozen until the start of the FHM reproductive bioassay. We further examined the role of food quality on fish toxicity by assessing differences between multi trophic (where fish were fed both a live and frozen diet of C. dilutus) in the laboratory. This research was conducted at the Toxicology Centre in Saskatoon, Saskatchewan, Canada. The results showed that significant effects were observed when fish were fed a live diet versus a frozen diet. Condition factor and body weight increased, although inconsistent effects were observed for liver somatic index (LSI) in fathead minnows in both experiments when exposed to one or both routes of exposure. Cumulative total egg production and cumulative spawning events were both significantly affected by both waterborne and dietborne exposures with the greatest effects seen in the multi-trophic streams and particularly when fish were fed a live diet. This significance of this research has demonstrated the importance of including both routes of exposure when assessing effects of mine effluent. This research also shows that the artificial stream technology is a useful tool in isolating the effects of a particular point source input (metal mining mixtures) when a system is highly confounded. The results suggest that under environmentally relevant exposure conditions, trophic transfer and live diet may lead to greater reproductive effects and increased fish toxicity. This also suggests that trophic transfer is an important route of exposure that is virtually impossible to attain using typical laboratory bioassay techniques (food-borne study using artificial diets or waterborne exposures only).

Previous studies at the Key Lake uranium mill (Saskatchewan, Canada) suggested the complex effluent discharged alters energetic stores of resident fish species. A second study at the same site demonstrated certain fish from lakes downstream of the mill produce larvae with elevated incidence of developmental deformities. The mechanisms by which energy homeostasis is affected in fish downstream of the Key Lake uranium mill are unknown, and the effects of deformities and altered metabolism on swimming ability have not been explored. Therefore, the overall objective of this thesis was to investigate whether effluent exposed fish exhibited differences in swimming performance and energy homeostasis. To achieve this objective two experiments were conducted. In the first experiment juvenile spottail shiner (Notropis hudsonius) were collected from a lake downstream of the Key Lake uranium mill, and compared to fish collected from a nearby reference lake. In the second experiment larvae were collected from laboratory raised fathead minnow (Pimephales promelas) exposed to 5% diluted uranium mill effluent or control (dechlorinated municipal) water, and reared in the same treatments to 60 days post hatch (dph). No gross deformities were observed in any fish, and only shiner collected from the exposure lake in the field experiment had enlarged heart ventricles relative to body size compared to fish from the reference lake. Swimming performance was similar between shiner from the exposure and reference lakes in the field study, but effluent exposure impaired swimming ability in 60 dph fathead minnow in the laboratory experiment compared to fish from the control water treatment. After swimming performance tests fish were considered fatigued and metabolic endpoints were compared to non-fatigued fish. In both non-fatigued and fatigued shiner, liver glycogen was significantly greater in fish collected from the exposure lake compared to the reference lake. There was no difference in liver triglycerides in non-fatigued shiner between lakes, but liver triglycerides decreased after swimming in the field study reference fish. Muscle energy stores were unaffected by site or swimming in the field experiment. Conversely, whole body triglycerides and glycogen were similar between treatments in non-fatigued fathead minnow in the laboratory experiment. Swimming significantly decreased whole body triglycerides in fathead minnow from both treatments, but whole body glycogen was unaffected. In the field experiment blood endpoints (hematocrit, plasma glucose, lactate) in fatigued and non-fatigued shiner from both lakes further supported the possibility of altered intermediary metabolism or blunted stress response in fish downstream of the Key Lake uranium mill. In the field study, shiner muscle citrate synthase activity (an indicator of tissue aerobic capacity) was similar between lakes, but muscle âhydroxyacyl coenzyme A dehydrogenase activity (an indicator of tissue lipolytic capacity) was elevated. In contrast, laboratory fathead minnow whole body âhydroxyacyl coenzyme A dehydrogenase activity was similar between treatments, but citrate synthase activity was significantly lower in fathead minnow from the 5% effluent treatment. In summary, shiner from the exposure lake in the field experiment had similar swimming endurance and greater energy stores compared to fish from the reference lake, despite metabolic alterations. Fathead minnow from the 5% effluent treatment in the laboratory experiment had reduced swimming endurance that was matched by reduced whole body citrate synthase activity, but no other metabolic alterations were observed. Therefore, effluent exposure caused metabolic alterations in both fathead minnow and shiner, but specific effects between experiments were inconsistent. Overall, the physiological significance of the metabolic and swimming effects of effluent exposure is unclear, but suggests discharged effluent has the potential to negatively affect wild fish survivability.

As part of their reclamation plan, oil sands operators propose to transfer the mature fine tailings, which are a by-product of the oil sands extraction process, to open-pits and cap them with either a layer of surface water or oil sands process-affected waters (OSPW). These oil sands pit lakes are expected to develop habitats with productive capabilities comparable to natural lakes in the region. The studies presented in this thesis evaluate the potential impact of OSPW and its associated constituents [i.e. acid-extractable organics (e.g. naphthenic acids; NAs) and salts] on the reproductive physiology of adult fathead minnow (Pimephales promelas). Through 14-21 day fathead minnow reproduction assays it was demonstrated that aged OSPW can impair spawning, lower plasma sex steroid concentrations, and reduce male secondary sexual characteristics. The acid-extractable organics in OSPW were demonstrated to have an adverse effect on fathead minnow reproductive physiology. Other studies showed that the high salinity which characterizes OSPW also influences toxicity. When fathead minnows were exposed to the OSPW extract and 700 mg/l of NaHCO3, the NaHCO3 reduced the inhibitory effects of the extract on the numbers of reproductive tubercles and plasma testosterone levels by reducing the uptake of NAE to the fish. Embyro and larval bioassays also revealed that NaHCO3 reduces the acute toxic effects of the OSPW extract. An assessment of a wild population of fathead minnows inhabiting an OSPW pond determined that there were differences in the condition factor (CF), gonadosomatic indices (GSIs), liver somatic indices (LSIs), male secondary sexual characteristics, and 11-ketotestosterone concentrations in the fathead minnows from the OSPW pond relative to fish collected at reference sites. The opercula of fathead minnows from the OSPW pond also differed from those of reference fish and an examination of the gills revealed that were a number of proliferative and degenerative alterations relative to reference fish. Collectively, these studies demonstrate that aged OSPW has the potential to negatively affect the reproductive physiology of fathead minnows and suggest that aquatic habitats with high NAs concentrations (>10 mg/l) will have adverse effects on fish. / Canadian Natural Resources Ltd., Canadian Water Network, Canadian Oil Sands Network for Research and Development (CONRAD), Imperial Oil Ltd., NSERC, Shell Canada Energy, Suncor Energy Inc., Syncrude Canada Ltd., and Total E&P Canada

Site selection, and the means to access specific sites, is a keystone of parasitology. I evaluated migration and site selection behaviours of metacercariae of two congeneric species of strigeoid trematode throughout growth and encystment phases in the fathead minnow. Results showed that pre-encystment stages of Ornithodiplostomum ptychocheilus migrate along specific neural tracts to access sites in the optic lobes of the brain. Conversely, pre-encystment stages of Ornithodiplostomum sp. migrate via direct penetration, or via the vascular system to access visceral organs, especially the liver. Remarkably, both species have a bi-phasic pattern of development, with growth and encystment occurring in unique sites. Finally, I examined patterns of rodlet cell proliferation and maturation in response to growth and encystment phases of O. ptychocheilus. Cell densities were low (<11/mm2) in brain tissue adjacent to 1 and 2 week old metacercaria, but peaked to approximately 210/mm2 at 6 weeks. These results shed new light on the potential function of these enigmatic cells. / x, 125 leaves : ill. ; 29 cm.

No description available.