The estuarine sheepshead minnow (Cyprinodon variegatus) is the most eurythermic fish species, with a thermal tolerance window between 0.6°C and 45.1°C. However, little is known about the physiological mechanisms that allow this species to survive this temperature range. In order to understand how sheepshead minnow physiology is affected by temperature acclimation and acute changes in temperature, I conducted research on this species using a multi-level approach. I began at the organismal level, and examined the effects of these temperature changes on the sheepshead minnow's metabolic rate and swimming performance. The next chapter investigated the effects of changing temperatures on cardiac function (i.e., tissue/organ specific effects). In the final chapter, I conducted research at the sub-cellular level, and determined how mitochondrial bioenergetics / function is impacted by changing temperatures. This research shows that while sheepshead minnows are able to sustain heart function and mitochondrial respiration over a broad range of temperatures; they also display a plastic temperature response which is associated with the downregulation of standard metabolic rate and cardiac remodeling to maintain force generation. Collectively, these physiological responses may contribute to the sheepshead minnow's ability to maintain physiological and organismal function across a large temperature range.

In many rivers, dams have interrupted the connectivity of migration routes for fish. While fishways can provide access between downstream and upstream habitats, it is important that passage can occur with minimal delay, energy expenditure, and physiological stress. The research presented here is based on investigations into fishway attraction and passage for the Gates Creek sockeye salmon (Oncorhynchus nerka, Walbaum) stock at the Seton River dam in British Columbia. The first part of this thesis examined the effect of changes in water discharge from the dam on the relationship between the physiological condition of sockeye and their behaviour in approaching the fishway entrance. Fish were caught and non-lethally biopsied under three normal operating discharge conditions at Seton River dam, and subsets of sampled fish were implanted with radio transmitters and released downstream of the dam. Indices of physiological stress and exhaustive exercise (e.g. plasma cortisol, glucose, lactate, osmolality and hematocrit) did not differ among the water discharge levels that were examined. Fish delayed in the tailrace below the fishway entrance significantly longer under intermediate discharge (19.9 h @ 12.7 m³s‾¹) than either the high discharge (9.3 h @ 15.8 m³s‾¹), or the low discharge (7.0 h @11.0 m³s‾¹;) conditions (P = 0.022, and P = 0.015, respectively). Delay time was similar under high and low discharge conditions (P = 0.617), and passage success was found to be independent of discharge (P = 0.356). The second part of this thesis investigated how prior physiological condition and subsequent swimming energetics and behaviours effected fishway passage success. Fish were captured and biopsied, before being implanted with electromyogram (EMG) transmitters and released near the downstream entrance of the fishway. Very few differences existed between successful and unsuccessful fish in body size, initial plasma physiology and energy state, and mean swim speed and energy use during passage. However, plasma Na+ concentration was significantly lower in unsuccessful fish (P = 0.022), which is suggestive of a depressed ionic state for unsuccessful fish. Generally, fish did not employ burst swimming during successful or failed attempts at passage, indicating that failure was probably not related to metabolic acidosis.

In many rivers, dams have interrupted the connectivity of migration routes for fish. While fishways can provide access between downstream and upstream habitats, it is important that passage can occur with minimal delay, energy expenditure, and physiological stress. The research presented here is based on investigations into fishway attraction and passage for the Gates Creek sockeye salmon (Oncorhynchus nerka, Walbaum) stock at the Seton River dam in British Columbia. The first part of this thesis examined the effect of changes in water discharge from the dam on the relationship between the physiological condition of sockeye and their behaviour in approaching the fishway entrance. Fish were caught and non-lethally biopsied under three normal operating discharge conditions at Seton River dam, and subsets of sampled fish were implanted with radio transmitters and released downstream of the dam. Indices of physiological stress and exhaustive exercise (e.g. plasma cortisol, glucose, lactate, osmolality and hematocrit) did not differ among the water discharge levels that were examined. Fish delayed in the tailrace below the fishway entrance significantly longer under intermediate discharge (19.9 h @ 12.7 m³s‾¹) than either the high discharge (9.3 h @ 15.8 m³s‾¹), or the low discharge (7.0 h @11.0 m³s‾¹;) conditions (P = 0.022, and P = 0.015, respectively). Delay time was similar under high and low discharge conditions (P = 0.617), and passage success was found to be independent of discharge (P = 0.356). The second part of this thesis investigated how prior physiological condition and subsequent swimming energetics and behaviours effected fishway passage success. Fish were captured and biopsied, before being implanted with electromyogram (EMG) transmitters and released near the downstream entrance of the fishway. Very few differences existed between successful and unsuccessful fish in body size, initial plasma physiology and energy state, and mean swim speed and energy use during passage. However, plasma Na+ concentration was significantly lower in unsuccessful fish (P = 0.022), which is suggestive of a depressed ionic state for unsuccessful fish. Generally, fish did not employ burst swimming during successful or failed attempts at passage, indicating that failure was probably not related to metabolic acidosis.

In many rivers, dams have interrupted the connectivity of migration routes for fish. While fishways can provide access between downstream and upstream habitats, it is important that passage can occur with minimal delay, energy expenditure, and physiological stress. The research presented here is based on investigations into fishway attraction and passage for the Gates Creek sockeye salmon (Oncorhynchus nerka, Walbaum) stock at the Seton River dam in British Columbia. The first part of this thesis examined the effect of changes in water discharge from the dam on the relationship between the physiological condition of sockeye and their behaviour in approaching the fishway entrance. Fish were caught and non-lethally biopsied under three normal operating discharge conditions at Seton River dam, and subsets of sampled fish were implanted with radio transmitters and released downstream of the dam. Indices of physiological stress and exhaustive exercise (e.g. plasma cortisol, glucose, lactate, osmolality and hematocrit) did not differ among the water discharge levels that were examined. Fish delayed in the tailrace below the fishway entrance significantly longer under intermediate discharge (19.9 h @ 12.7 m³s‾¹) than either the high discharge (9.3 h @ 15.8 m³s‾¹), or the low discharge (7.0 h @11.0 m³s‾¹;) conditions (P = 0.022, and P = 0.015, respectively). Delay time was similar under high and low discharge conditions (P = 0.617), and passage success was found to be independent of discharge (P = 0.356). The second part of this thesis investigated how prior physiological condition and subsequent swimming energetics and behaviours effected fishway passage success. Fish were captured and biopsied, before being implanted with electromyogram (EMG) transmitters and released near the downstream entrance of the fishway. Very few differences existed between successful and unsuccessful fish in body size, initial plasma physiology and energy state, and mean swim speed and energy use during passage. However, plasma Na+ concentration was significantly lower in unsuccessful fish (P = 0.022), which is suggestive of a depressed ionic state for unsuccessful fish. Generally, fish did not employ burst swimming during successful or failed attempts at passage, indicating that failure was probably not related to metabolic acidosis. / Forestry, Faculty of / Graduate

Die Bedeutung ökologischer Faktoren bei der Entstehung phänotypischer sowie genetischer Vielfalt durch natürliche Selektion, besonders bei sympatrischer Artbildung, ist derzeit ein Fokus der Evolutionsforschung. Gemeinsam vorkommende und nah verwandte Arten werden daher als Modellorganismen verwendet, um die Ursachen und Mechanismen ökologischer und evolutionärer Diversifizierung zu untersuchen. Ein sympatrisches Fisch-Artenpaar, für das eine sympatrische Artbildung auf Basis genetischer Analysen vermutet wird, existiert im norddeutschen Stechlinsee. Die zwei Maränenarten sind morphologisch kaum zu unterscheiden, zeigen eine ähnliche Nahrungszusammensetzung und kommen gemeinsam im Freiwasser, allerdings in leicht unterschiedlichen Wassertiefen, vor. Die Hypothese meiner Arbeit war, dass sich die Physiologie bzw. das Verhalten der Arten in Bezug auf die wichtigsten Umweltfaktoren ihres Lebensraumes, Futterdichte, Lichtintensität und Wassertemperatur unterscheiden. Daher haben wir Fraßeffizienz, Stoffwechselraten und Temperaturpräferenzen in Abhängigkeit dieser Faktoren bei beiden Arten untersucht, nachdem diese zuvor unter identischen Laborbedingungen herangezogen wurden. Wir fanden keinen Unterschied in der Fraßeffizienz, allerdings zeigten beide Arten je nach Temperatur unterschiedliche Stoffwechselraten sowie entsprechende Unterschiede in der Temperaturpräferenz. Die Ergebnisse legen nahe, dass temperaturbedingte physiologische Anpassungen sowie die Nutzung unterschiedlicher thermischer Mikrohabitate die zwischenartliche Konkurrenz verringern und eine gemeinsame Existenz ermöglichen. Um die mögliche Bedeutung einer solchen öko-physiologischen Spezialisierung für die Artbildung innerhalb des Sees einschätzen zu können, haben wir basierend auf den Freiland- und Labordaten ein mathematisches Evolutionsmodell entwickelt. Demnach ist ein Aufspalten einer Ausgangspopulation in zwei Populationen mit unterschiedlichen Temperaturoptima wahrscheinlich. Eine ökologische und evolutionäre Diversifizierung entlang des Temperaturgradienten ist somit ein empirisch und theoretisch plausibles Szenario für die sympatrische Artbildung der Stechlinsee-Maränen. / Abstract The role of ecological factors in generating phenotypic and genetic diversity through natural selection has received increasing attention in evolutionary biology during the last decade, especially with respect to diversification in sympatry. Sympatrically occurring and closely related species are used as model systems to study the causes and mechanisms of ecological and evolutionary diversification. A sympatric species pair of coregonid fish, for which a speciation in sympatry has been suggested based on genetic analyses, coexists in the German Lake Stechlin. The two species are morphologically similar planktivores with weak divergence in diet composition that co-occur within the pelagic area of the lake at slightly different water depths. Accordingly, it was hypothesized that the species would differ in physiology and behaviour with respect to the most important environmental factors of their natural habitat, food density, light intensity and water temperature. We studied the feeding efficiencies, metabolic rates, and temperature preferences of both species previously hatched and raised under identical laboratory conditions. We found no divergence in feeding efficiency, but significant differences in temperature-related metabolic costs as well as a corresponding difference in thermal preference. These results suggest that temperature-related physiological adaptations and the associated use of slightly different thermal microhabitats reduces exploitative competition between the species and facilitates their coexistence. To evaluate a potential role of this eco-physiological specialization for a speciation in sympatry, we developed a mathematical evolutionary model, based on our field observations and laboratory experiments. The model showed that an evolutionary splitting of an ancestral into two coexisting populations with different temperature optima is likely in this system. In conclusion, an eco-physiological and evolutionary diversification along the temperature-depth gradient of the lake is an empirically and theoretically plausible scenario for the sympatric speciation of the coregonids.

Stressors in the aquaculture environment can lead to negative impacts on growth and immune health, resulting in susceptibility to infectious diseases. These stressors are expected to increase as the growth of aquaculture continues to rise to meet demands for quality fish protein. Isoflavones, as a crude extract or as a pure isolate, may be effective in modulating the stress response, promoting growth and immunity. The objective of these studies was to examine the effect of various pure isoflavone isolates and crude isoflavone extracts on stress, growth, and immunity. Nile tilapia (Oreochromis niloticus) were stressed by adding hydrocortisone to the feed. In a 7-week study, pure isoflavone isolates of genistein and puerarin were evaluated to determine their respective effects on stress, growth, and immunity. A separate 10-day physiological and 6-week growth study focused on crude isoflavone extracts from kudzu (Pueraria lobata), red clover (Trifolium pratense), and soybean (Glycine max) was performed to determine their respective effects on stress, growth, and immunity. Numerous physiological parameters of the fish were measured (serum cortisol concentration, blood glucose concentration, hematocrit, hepatosomatic index, plasma protein concentration, lysozyme activity, and spleensomatic<br>index) to determine the effects of these pure isoflavone and crude isoflavone extracts on the modulation of stress and immunity. Many growth parameters were examined (length, weight, condition factor, weight gain, specific growth rate, feed intake, feed conversion ratio, and protein efficiency ratio) as well to determine the effects of these pure isoflavones and isoflavone extracts on growth. The addition of isoflavone and crude isoflavone extracts to the diet of Nile tilapia ameliorated some of the negative consequences of stress. Compared to stressed fish fed commercial feed, genistein and puerarin added to the diet appeared to improve serum cortisol concentrations, which resulted in increased plasma protein, albeit at different durations of stress. Puerarin, as well as all three crude isoflavone extracts, significantly increased spleen-somatic index compared to non-supplemented stressed fish, although the crude isoflavone extracts did not appear to improve serum cortisol concentrations. Crude isoflavone extracts also showed overall increases in lysozyme activity compared to non-supplemented stressed fish, although this was not significant. Genistein, puerarin, and red clover showed increased growth rates, feed conversion ratio, and protein efficiency. Overall, pure isolates of isoflavone appear to be more effective in modulating stress, immunity, and growth than the crude isoflavone extracts, although red clover extract showed promises in the ability to modulate the stress response and improve growth and immunity. There are likely substantial interactions between the isoflavones in the crude extracts that cannot be fully understood by measuring the effects of single isoflavones. Regardless, isoflavone supplementation (pure or crude) appeared to generally have an overall positive impact on stressed Nile tilapia, requiring more research to better understand the effects and mechanisms behind these isoflavones.

Made available in DSpace on 2017-07-10T14:38:30Z (GMT). No. of bitstreams: 1 Dissertacao - adriane (pdf).pdf: 931863 bytes, checksum: 9362d32483efab607a95a16a4db0a7fd (MD5) Previous issue date: 2016-02-19 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Stress is an adaptive component that allows the fish to deal with various stressful events that occur during the life cycle of fish, such as the reproductive migration that demands physical efforts. The aim was to assess whether the steps for implementation to fish in the reservoir dam is promotes stress in curimba (Prochilodus lineatus). Adult fish species were sampled in March 2015 at three sites in the Hydroelectric Powerplant of Porto Primavera, Upper Paraná River, Brazil: downstream, the fish ladder and upstream of the dam, totaling 48 individual. Variations in body weight, standard length, and determination of plasma concentrations of cortisol, glucose, lactate, choloride and hematocrit they were analyzed in relation to gender and local sampled. The body weight and standard length presented comparable values for each location between genders, but the averages of females were significantly higher. Significant increase in plasma levels of glucose and lactate were observed but without significant changes in cortisol, chloride levels, as well as in hematocrit. The intense exercise of the fish by the transposition of fish ladder represented moderate stressor, without causing harmful effects to these animals, since the recovery of fish at the upstream from the dam. These results are deepening about the physiology used as a tool for conservation by providing knowledge and subsidies for the conservation and management of migratory species. / Resumo O estresse é um componente adaptativo que permite ao peixe lidar com eventos estressores que ocorrem durante o ciclo de vida dos peixes, como, por exemplo, a migração reprodutiva que exige esforço fisiológico dos animais. O objetivo do trabalho foi avaliar, por meio de indicadores fisiológicos, se a transposição de escada para peixes em barragem de reservatório representa um evento estressor para o curimba (Prochilodus lineatus). Peixes adultos da espécie foram amostrados em março de 2015 em três locais na Usina Hidrelétrica de Porto Primavera, Alto Rio Paraná, Brasil: a jusante, na escada para peixes e a montante da barragem, totalizando 48 indivíduos. Variações do peso corporal, comprimento padrão e das concentrações plasmáticas de cortisol, glicose, lactato, cloreto e hematócrito foram analisadas em relação ao gênero e local amostrado. O peso corporal e o comprimento padrão apresentou valores similares em todos os locais de coleta, nos dois gêneros, porém as fêmeas foram significativamente maiores a montante. Aumento significativo nos níveis plasmáticos de glicose e lactato foram observados, mas sem alterações significativas das concentrações circulantes de cortisol, cloreto e do hematócrito. O exercício intenso dos peixes pela transposição da escada para peixes representou uma condição moderadamente estressante, sem causar prejuízo biológico, visto a recuperação observada a montante. Estes resultados são aprofundamentos sob a fisiologia utilizada como ferramenta para conservação fornecendo conhecimento e subsídios para a conservação e manejo de espécies migratórias.

Animals rely on lipids as a major fuel for endurance exercise because they pack more joules per gram than any other fuel. However, in contrast to mammals, information on how the mobilization of lipids from endogenous stores is managed to meet the needs of energy metabolism in swimming fish is sparse. Information on in vivo rates of lipid mobilization in swimming fish has been limited to relatively low exercise intensities and has only been investigated in a single species. Therefore, the goal of my thesis was to address this paucity of information by quantifying lipolytic rate in rainbow trout during graded exercise and fatty acid mobilization in Atlantic salmon during prolonged endurance exercise. In the first part of my work, I hypothesized that like mammals, rainbow trout stimulate lipolysis above resting levels to a peak with increasing work intensity, but subsequently lower its rate at high intensities when ATP production from carbohydrates becomes dominant. To test this hypothesis, I measured the rate of appearance of glycerol (Ra glycerol) in the blood (resulting from the breakdown of triacylglycerol (TAG)) of trout at rest (control) and during graded exercise from rest to Ucrit. Results showed that Ra glycerol in trout averaged 1.24 ± 0.10 µmol kg -1 min-1 and that this rate was unaffected by exercise of any intensity. These experiments revealed that rainbow trout do not modulate lipolysis during exercise. Furthermore, I calculated that baseline lipolytic rate was much higher in trout than in mammals and that this rate is in constant excess of the requirements of energy metabolism. My second investigation focused on measuring fatty acid mobilization in Atlantic salmon. To date, the majority of studies on energy metabolism in salmonids have used rainbow trout as the ubiquitous model for salmonids. I postulated that domesticated rainbow trout may be far less impressive athletes than their wild anadromous form and other salmonids. In this regard, I proposed that studying energy metabolism in Atlantic salmon (even those from aquaculture) may help to deepen our understanding of the physiology of true long-distance migrant fish. To study the effects of prolonged endurance exercise on the mobilization of fatty acids from endogenous stores in these fish, I monitored the rate of appearance of fatty acids (Ra NEFA calculated from Ra Palmitate) in the blood during 72 hours of sustained swimming. I found that contrary to what has been previously described in rainbow trout, Ra Palmitate (and by proxy, Ra NEFA) is reduced by approximately 64% (from 0.75 ± 0.12 µmol kg-1min-1 to 0.27 ± 0.06 µmol kg-1min-1 and from 19.3 ± 7.8 µmol kg-1min-1 to 6.9 ± 2.0 µmol kg-1min-1 for Ra Palmitate and Ra NEFA, respectively) during prolonged endurance exercise in Atlantic salmon. However, like in trout, even this reduced rate of fatty acid mobilization exceeds the requirements of energy metabolism at rest and during swimming. While further experiments will be necessary, I speculated that this reduction in Ra NEFA may be caused by a partial inhibition of lipolysis to reduce the energetic cost of TAG:FA cycling and optimize fuel budgets during prolonged endurance exercise. This thesis provides the first in vivo measurements of lipolysis during graded exercise in salmonids and the first in vivo measurements of fatty acid mobilization in Atlantic salmon. From the results mentioned above, I concluded that salmonids mobilize lipids in constant excess of the requirements for energy metabolism, possibly to allow for rapid reorganization of membrane phospholipids in response to changing environmental conditions. However, more anadromous and migratory phenotypes may rely on a tighter control of lipolysis to minimize the costs of substrate cycling and conserve energy on limited fuel stores.

Sperm competition is a widely-recognized and powerful selective force. Male accessory glands are organs found across animal taxa that can influence sperm performance, and thus may be selected for in competitive contexts. In fishes, these organs are in fact rare, but display great diversity in form and function across species. Although the accessory gland is known to play a role in mate attraction, parental care, fertilization, or post-copulatory competition in a few select species, the role of this organ remains a mystery in most species. Many fishes with accessory glands also exhibit alternative reproductive tactics (ARTs), which add an extra layer of complexity to how species respond to sperm competition. Because males of different ARTs typically experience different levels of sperm competition risk, it’s possible they may differentially invest in accessory glands to overcome this competition. In this thesis, I used the plainfin midshipman fish (Porichthys notatus), a species with both ARTs and an accessory gland, to experimentally investigate the role of the accessory gland in sperm competition and uncover how this organ may differ between ARTs. Over a two-year period, I studied tactic-specific investment in the accessory gland in fish from the beaches of British Columbia. I also examined the effects of seminal fluid, produced in part by the accessory gland, on sperm performance and morphology. I found that males adopting the ‘guarder’ male tactic invested more in one region (the lobules) of this organ, while males adopting the smaller ‘sneaker’ male tactic invested more the other region of the gland (the nodes). Using data collected over five years, I found that guarder males also invested more in their whole accessory glands. Additionally, I report that sperm swam faster in the presence of seminal fluid, and seminal fluid increased sperm head size in both male tactics and increased midpiece size in guarder males. These results suggest that the plainfin midshipman accessory gland may have dual functions, one of which may be to aid sperm competitive ability through enhancements in swimming speed and potentially more successful sperm morphology. Taken together, the results of my thesis improve our knowledge of the role of non-sperm components like seminal fluid and the accessory gland in sperm competition, and demonstrate how species with ARTs can have varying physiological responses to such competition. Only a handful of studies have considered the effects of seminal fluid on sperm performance. By examining sperm competition in a more biologically relevant way and incorporating the effects of a little-studied organ that impacts sperm competition, we should be able to more generally and accurately appreciate the dynamics of post-copulatory competition and fertilization. / Thesis / Master of Science (MSc)

Steelhead and rainbow trout are common names for marine-migratory (anadromous) and freshwater-resident forms of Oncorhynchus mykiss, a partially migratory salmonid fish. Anadromous and resident forms are sympatric and can produce offspring with a life history different from their own (i.e., steelhead parents can produce rainbow trout offspring and vice versa). The expression of these alternative life histories is a plastic response to individual patterns of energy acquisition, assimilation, and allocation during juvenile life stages. Individual performance during early stream life is of particular interest because of potential carry-over effects on subsequent growth and developmental trajectories. In a series of experiments in laboratory streams, I determined the influence of individual variation in energy metabolism on behavior, growth, and life-history expression in O. mykiss. Individual variation in energy metabolism was a strong predictor of feeding territory acquisition by juvenile fish during the transition from dependence on maternal provisioning of egg yolk reserves to independent feeding. Feeding territory acquisition was positively associated with standard metabolic rate (SMR) under conditions of an abundant and predictable food supply. When the density of intraspecific competitors was manipulated, the association between SMR and territory acquisition was strongest at intermediate stocking densities, moderate at the highest stocking densities, and weakest at the lowest stocking densities. However, reducing the spatial predictability of food resources within streams reversed the influence of SMR on competitive outcomes. These experiments determined that variation in ecological conditions during early life stages imposes different selection regimes on juvenile O. mykiss and results in physiological divergence among cohorts. Subsequent rearing experiments determined that behavioral dominance influences rates of anadromy and freshwater maturation, most likely through the association between SMR and territory acquisition. In addition to the effects of behavioral dominance, I observed a significant influence of sex, rearing temperature, and individual growth trajectories on the expression of anadromy and freshwater maturation. Partially migratory populations of O. mykiss maintain an exceptionally diverse portfolio of life-history strategies. Results from this work lend insight into a suite of behavioral and physiological processes influencing individual life histories. / Graduation date: 2013