Studies in Europe, Japan, and North America have reported that wild fish exposed to treated sewage effluents can exhibit significant physiological and reproductive abnormalities consistent with exposure to hormonally active chemicals. The main objective of this research project was to examine the estrogenic and androgenic activity in treated sewage to determine the risk associated with treated sewage discharges in Australia and New Zealand. Several bioassays, including a sheep estrogen receptor and a rainbow trout androgen receptor binding assay, were set up and validated with model compounds. The assays were then used to measure the estrogenic and androgenic activity in sewage samples from 15 municipal sewage treatment plants (STP) utilizing a variety of treatment technologies. Raw sewage samples contained high levels of both estrogenic and androgenic activity, up to 185 ng/L estradiol equivalents (EEq) and up to 9330 ng/L testosterone equivalents (TEq), respectively. Secondary treatment processes such as activated sludge had the greatest impact on removal of biological activity from the wastewater. The estrogenic and androgenic activity in final treated effluents were <1 to 4.2 ng/L EEq and <6.5 to 736 ng/L TEq, respectively. Based on lowest observable effective concentrations reported in the literature, these levels are unlikely to induce biological effects in exposed fish in the short term. To examine potential long-term effects, resident mosquitofish chronically exposed to undiluted treated sewage were sampled. Several morphological biomarkers indicative of endocrine disruption were measured and compared with mosquitofish captured at a reference site. Mosquitofish captured in a constructed wetland for tertiary treatment of secondary treated sewage exhibited morphological differences such as elongated anal fins consistent with exposure to androgenic chemicals, although this effect was not measurable in fish collected at sites further downstream or at any of the other sites. Based on these results, it is unlikely that mosquitofish populations would be significantly affected by exposure to final treated sewage. A reverse transcription real-time polymerase chain reaction (RT-PCR) method to measure the production of a female-specific protein (vitellogenin) mRNA in adult male mosquitofish was developed, and this could be used as a rapid test to detect early changes in individuals exposed to estrogenic activity.

The aim of this thesis was to examine the influence of nitrite on fish. Influence of nitrite was evaluated on the basis of acute and sub-chronic toxicity tests results on rainbow trout (Oncorhynchus mykiss). The effects of nitrite were also observed in wels catfish {--} albino (Silurus glanis). On the basis of the results of acute toxicity tests, values of lethal concentration of nitrite for rainbow trout (Oncorhynchus mykiss) (96hLC50 = 11.2 mg.l-1 NO2-) and for wels catfish {--} albino (Silurus glanis) (96hLC50 = 15.8 mg.l-1 NO2-) were calculated. Using results of acute toxicity test for rainbow trout (Oncorhynchus mykiss) and with respect to legislative requirements, concentrations of nitrite for sub-chronic toxicity test were selected. The results of sub-chronic toxicity test showed that nitrite concentration of 3 mg.l-1 NO2- during 28-day exposition caused 65 % fish mortality. This concentration also caused growth rate decrease compared to control. Growth rate among fish exposed to concentrations lower then 3 mg.l-1 NO2- was comparable to control. Among fish exposed to nitrite concentration of 0.6 mg.l-1 NO2- and higher nitrite accumulation in muscle and in blood plasma was observed. Haematological examination showed statistically significant decrease of haematocrit value and concentration of haemoglobin and increase of the number of leukocytes in experimental fish. Other measured haematological parameters (Ery, MCV, MCH and MCHC) were comparable with control.

A indoleamina 2,3-dioxigenase (IDO) é uma enzima que cataboliza o aminoácido triptofano, levando à inibição da proliferação de linfócitos T, seja pela exaustão desse aminoácido no ambiente, ou pela indução via catabólitos induzindo-os a apoptose. Em mamíferos, esta enzima atua em diversas condições do organismo como a gestação, infecções, inflamações crônicas, transplantes e tumores, sendo reconhecida como uma autêntica reguladora imunológica. Estudos recentes identificaram a presença de moléculas homólogas a IDO em espécies filogeneticamente mais antigas que os mamíferos placentados, sendo que o papel da IDO primitiva não está totalmente elucidada. Sabe-se que em fungos e bactérias a IDO limita-se ao metabolismo do triptofano para obtenção de energia e que em vertebrados inferiores sua função ainda não é bem esclarecida. Desta forma, este estudo teve por objetivo averiguar a presença da IDO em células sanguíneas e órgãos hematopoiéticos de trutas arco-íris, colaborando para o estudo filogenético do sistema imune nos vertebrados, particularmente dos peixes. A expressão de IDO foi observada em todos os órgãos hematopoiéticos estudados incluindo os leucócitos e também foi observada marcação positiva para a enzima em pequenos vasos sanguíneos do rim cefálico, baço e fígado. Os resultados obtidos, apresentando marcações positivas da IDO restritas em leucócitos e tecidos hematopoiéticos de trutas arco-íris, poderiam significar a primeira evidência de que a IDO, à semelhança do que ocorre nos mamíferos, possa estar relacionada ao sistema imunológico nesta espécie. / The indoleamine 2,3-dioxygenase (IDO) is an enzyme that catabolizes the amino acid tryptophan, leading to inhibition of T lymphocyte proliferation by exhaustion of this amino acid in the environment, or by induction via the catabolites inducing apoptosis. In mammals, this enzyme acts on various conditions such as pregnancy, infections, chronic inflammation, transplantation and tumors, being recognized as an authentic immune regulator. Recent studies have identified the presence of homologous IDO in older phylogenetically related species molecules, the placental mammals, and the role of early IDO is not fully elucidated, it is known that in fungi and bacteria the role of IDO is limited to tryptophan metabolism for energy, and is function in lower vertebrates is still not well understood. This study aims to investigate the presence of IDO in peripheral blood cells and hematopoietic organs of rainbow trout, thereby contributing to the phylogenetic study of the immune system in vertebrates, particularly in fish. The expression of IDO was observed in the inner region of small blood vessels in the head kidney, spleen, liver and peripheral blood cells, mainly in monocytes and lymphocytes. The restricted localized expression of IDO in these tissues and cells, as occurred in mammals, may suggest a first evidence for a putative role of this enzyme in the immune system in rainbow trout.

Ce travail de thèse avait pour objectif d’étudier la régulation de l’expression des gènes du métabolisme intermédiaire par les acides aminés alimentaires dans le foie de truite arc-en-ciel. Ces études ont permis de caractériser, pour la première fois dans le foie de truite, les principaux acteurs de la voie de signalisation Akt/TOR et leurs régulations. Nos résultats in vitro montrent qu’un mélange d’acides aminés, seul ou avec l’insuline, est capable de réguler l’expression de nombreux gènes impliqués dans la lipogenèse, la néoglucogenèse et la glycolyse. Les régulations observées en présence conjointe d’un mélange d’acides aminés et d’insuline semblent être, pour la plupart, dépendantes de la voie TOR. Par la suite, nous avons étudié l’effet de certains acides aminés comme la leucine (connue pour son effet « signal ») ainsi que la lysine et la méthionine (souvent ajoutées dans les aliments piscicoles riches en matières premières végétales afin d’atteindre l’équilibre en acides aminés). En présence d’insuline, la leucine, contrairement à la lysine et la méthionine, active la voie de signalisation TOR et régule l’expression de certains gènes (néoglucogenèse et lipogenèse) de façon similaire à un mélange d’acides aminés. Parallèlement, in vivo, nous avons étudié la régulation de l’expression des gènes du métabolisme intermédiaire lors d’un remplacement partiel ou total des huiles et farines de poisson par des produits végétaux dans l’aliment piscicole. Cette expérimentation a montré que, ni les voies de signalisation Akt/TOR, ni l’expression des gènes cibles ne sont affectés par ces nouveaux aliments. En conclusion, ces travaux ont montré que les acides aminés semblent jouer un rôle important dans la régulation de l’expression des gènes hépatiques du métabolisme intermédiaire chez la truite arc-en-ciel. / The objective of my PhD was to characterize the regulation of the intermediary metabolism related gene expression by dietary amino acids in the liver of rainbow trout. This work allowed us to characterize, for the first time in the liver of trout, the main proteins of the Akt/TOR signalling pathway and their regulations. In vitro results showed that a mixture of amino acids, in the presence or absence of insulin, is able to regulate the expression of numerous genes involved in lipolysis, gluconeogenesis and glycolysis. Such regulations induced by an amino acid mix together with insulin appear to be, at least partly, TOR-dependent. Afterwards, I studied the effect of specific amino acids known to be a signalling molecule (leucine) or having potential application as supplements to reach essential amino acid balance in plant ingredients-rich diet (lysine and methionine). In the presence of insulin, leucine, in contrast to lysine and methionine, is able to activate the TOR signalling pathway and regulate the expression of several genes involved in gluconeogenesis and lipogenesis in the same way as a mixture of amino acids. Furthermore, we studied in vivo, the effect of partial or total replacement of fish oil and fish meal by plant products in fish feed on .the regulation of intermediary metabolism related gene expression. This study showed that neither Akt/TOR signalling pathway nor the expression of the target genes were affected by such diets. In conclusion, these studies showed that amino acids seem to play an important role in the hepatic regulation of intermediary metabolism gene expression in the rainbow trout.

The construction of the Katse dam was completed in 1996. The dam started filling in 1995 and reached full capacity in early 1998. It forms part of Phase 1A of the Lesotho Highlands Water Project (LHWP). The main aim of the project is to provide revenue to Lesotho, by transferring water from the catchment of the Senqu (Orange) River in Lesotho to South Africa’s major industrial and population centres. During the implementation of the project, an estimated 130 people were displaced. However, the most serious impact has been the loss of the traditional way of life in the form of arable and grazing land as a consequence of inundation. One of the obligations of the project is to ensure that the standard of living of those affected by the project is not impaired. The Lesotho Highlands Development Authority identified a number of rural development projects, which included fisheries development. This study is an integral part of fisheries development in Lesotho. The aim of this study was to gain an understanding of the biology and the demographics of the fish species in the lake. This information would serve as the basis for the development of a management plan for the sustainable utilisation of the fisheries resources. To achieve this aim, the following specific objectives were addressed: 1. Investigation of the biology of the three principal species. 2. Description of the key population parameters (growth, mortality & recruitment). 3. Investigation of the distribution and relative abundance of the three species. The three principal species in the Katse dam are rainbow trout, Oncorhynchus mykiss, and the two cyprinids, Barbus aeneus and Labeo capensis. Sectioned otoliths were used to age O. mykiss while both otoliths and scales were used to age B. aeneus and L. capensis. Marginal zone analysis revealed that slow growth was experienced in winter for the three species. The maximum-recorded age for both B. aeneus and L. capensis was 12 years while O. mykiss reached 4 years. There was no significant difference in growth rates of the different sexes and growth was best described by the 3 parameter Von-Bertalanfy growth model as Lt = 603[1-e⁻°·¹⁵⁽t ⁺ °·°³⁾] for B. aeneus, Lt = 526[1-e⁻°·²¹⁽t ⁺ °·⁹⁾] for O. mykiss and Lt = 521[1-e⁻°·¹⁷⁽t ⁺ °·²¹⁾] for L.capensis. Male and female O. mykiss attained 50% sexual maturity (Lm₅₀) at 235 and 275mm FL, respectively. There was no difference in Lm₅₀ for male and female B. aeneus and L. capensis. B. aeneus reached sexual maturity at 285mm FL while L. capensis reached sexual maturity at 244mm. Both cyprinid species spawned in summer (November to January) while O. mykiss spawned in winter (May to August). The mean total mortality rate (Z) estimated from catch curves and Butterworth et al (1989) equation was 0.72 yr⁻¹ for B. aeneus, 0.62 yr⁻¹ for L. capensis and 1.32 yr⁻¹ for O. mykiss. The estimate of natural mortality was 0.41 yr⁻¹ for both B. aeneus and L. capensis and 0.81yr⁻¹ for O. mykiss.

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.

The growth of vertebrates is primarily regulated by the growth hormone-insulin-like growth factor (GH-IGF) system, but not in isolation. The central question of this dissertation was how do other hormones peripheral to the GH-IGF system interact with the system, including feedbacks by GH and IGF themselves on various tissues in rainbow trout (Oncorhynchus mykiss)? The representative hormones selected were thyroxine, cortisol, and the sex steroids testosterone and estrogen, along with GH and IGF. These hormones were chosen because they are known to affect overall growth and development during specific life events, but exactly what target genes and what mechanisms are involved are only at the early stages of being delineated in fish. Liver and gill tissues were selected as representative tissues to assess the in vitro effects on growth-related genes of the GH-IGF system. A total of more than thirty experiments were conducted, including time- and concentration-response, inhibitory studies, hormone combination studies, and radio-receptor binding assays. Hormones were applied to whole tissue cultures and real-time quantitative-PCR was used to measure hormonal effects on GHR, IGF, and IGFR1 genes. Microsomal preparations were treated with selected hormones and radio-labeled GH or IGF. A gamma counter was used to measure receptor-ligand activity. GH and IGF were found to possess autocrine and/or paracrine actions in self-regulating target growth genes. Thyroxine had no direct effects on targeted growth genes but may interact with other molecules or hormones to elicit its effects on growth and development. Cortisol directly influenced target growth genes in a tissue-specific and isoform-specific manner. Finally, sex steroids differentially regulated the growth genes: estradiol inhibited growth genes while testosterone directly stimulated growth genes. These findings contribute to understanding how hormones peripheral to the GH-IGF system interact with the growth system. / National Science Foundation grant IOS 0920116 to Dr. Mark Sheridan

Examination of variation at polymorphic microsatellite loci is a widely accepted method for determining parentage and examining genetic diversity within rainbow trout (Oncorhynchus mykiss) breeding programs. Genotyping costs are considerable; therefore, we developed a single-step method of co-amplifying twelve microsatellite loci in two hexaplex reactions. The protocol is explicitly described to ensure reproducible results. I applied the protocol to samples previously analyzed at the National Center for Cool and Coldwater Aquaculture (NCCCWA) with previously reported marker sets for a comparison of results. Each marker within the multiplex system was evaluated for duplication, null alleles, physical linkage, and probability of genotyping errors. Data from four of the 12 markers were excluded from parental analysis based on these criteria. Parental assignments were compared to those of a previous study that used five independently amplified microsatellites. Percentages of progeny assigned to parents were higher using the subset of eight markers from the multiplex system than with five markers used in the previous study (98% vs. 92%). Through multiplexing, use of additional markers improved parental allocation while also improving efficiency by reducing the number of PCR reactions and genotyping runs required. I evaluated the methods further through estimation of F-statistics, pairwise genetic distances, and cluster analysis among brood-years at the NCCCWA facility. These estimates were compared to those from nine independently amplified microsatellites used in a previous study. Fst metrics calculated between brood-years showed similar values of genetic differentiation using both marker sets. Estimates of individual pairwise genetic distances were used for constructing neighbor-joining trees. Both marker-sets yielded trees that showed similar subpopulation structuring and agreed with results from a model-based cluster analysis and available pedigree information. These approaches for detecting population substructure and admixture portions within individuals are particularly useful for new breeding programs where the founders' relatedness is unknown. The 2005 NCCCWA brood-year (75 full-sib families) was challenged with Flavobacterium psychrophilum, the causative agent of bacterial coldwater disease (BCWD). The overall mortality rate was 70%, with large variation among families. Resistance to the disease was assessed by monitoring post-challenge days-to-death. Phenotypic variation and additive genetic variation were estimated using mixed models of survival analysis. The microsatellite markers used were previously isolated from BAC clones that harbor genes of interest and mapped onto the rainbow trout genetic linkage map. A general relationship between UBA gene sequence types and MH-IA-linked microsatellite alleles indicated that microsatellites mapped near or within specific major histocompatibility (MH) loci reliably mark sequence variation at MH genes. The parents and grandparents of the 2005 brood-year families were genotyped with markers linked to the four MH genomic regions (MH-IA, MH-IB, TAP1, and MH-II) to assess linkage disequilibrium (LD) between those genomic regions and resistance to BCWD. Family analysis suggested that MH-IB and MH-II markers are linked to BCWD survivability. Tests for disease association at the population level substantiated the involvement of MH-IB with disease resistance. The impact of MH sequence variation on selective breeding for disease resistance is discussed in the context of aquaculture production. / Master of Science

Tesis por compendio / En la presente Tesis Doctoral se utilizaron cebada y remolacha como fuentes alternativas de carbohidratos en dietas para trucha arcoíris con el objetivo de evaluar su efecto sobre parámetros productivos, histología hepática e intestinal, estrés y calidad de la carne. Los marcadores estudiados fueron los mismo tanto para los experimentos de cebada como de remolacha. En las pruebas de la cebada se utilizaron concentraciones crecientes de este ingrediente (0-32%) en la dieta, se inició con un peso medio de 127.72 ± 5.65 g y se finalizó cuando alcanzaron el peso comercial al cabo de 84 días. En las pruebas de remolacha se utilizaron dos niveles de remolacha (14% y 28%) y dos de betaína (0.9% y 1.63%), se inició con un peso medio de 69 ± 2.2 g y se finalizó cuando los peces alcanzaron el peso comercial al cabo de 105 días. La inclusión de cebada en dietas de trucha arcoíris no mostró diferencias significativas en los parámetros de crecimiento ni biométricos. La inclusión de 14% remolacha y 0.9% betaína no afectó al crecimiento, parámetros nutritivos, biométricos y retenciones de nutrientes en comparación con el control, concentraciones mayores de remolacha y betaína tuvieron un efecto negativo. Cuando se estudiaron los parámetros histológicos y morfométricos en hígado e intestino, los resultados mostraron que los peces alimentados con concentraciones crecientes de cebada mostraron hepatocitos más pequeños, mientras que los peces alimentados con remolacha y betaína presentaban hepatocitos más grandes, en ambos casos el hígado mostró un menor nivel de vacuolización. La concentración de cebada de un 8% produjo un efecto negativo a nivel morfológico del intestino, sin embargo, la inclusión de remolacha no produjo ningún efecto. Los peces fueron sometidos a estrés por ausencia de oxígeno y aumento de densidad durante 10 minutos. Cuando el estrés fue analizado a nivel bioquímico los resultados mostraron que a niveles basales, la inclusión de cebada y remolacha no causó ningún cambio en los niveles de cortisol, glucosa y MDA, aunque los valores basales de lactato fueron significativamente más bajos en los peces alimentados con remolacha y betaína. Treinta minutos después del estrés la concentración de cortisol, glucosa y lactato aumentó significativamente en los peces de todos los grupos y la concentración de MDA disminuyó. La inclusión de cebada mostró valores más bajos de cortisol, glucosa y lactato que la dieta control. Cuando los peces alcanzaron el peso comercial se analizó la calidad de los filetes y las propiedades antioxidantes. En ambas pruebas los peces alimentados con los ingredientes objeto de estudio mostraron valores de actividad de agua más bajos y una mejora en la textura y el color de los filetes comparado con los peces alimentados con la dieta control. La inclusión de remolacha y betaína no tuvo ningún efecto sobre los parámetros oxidativos del filete, mientras que la inclusión de cebada inhibió su oxidación lipídica. Concentraciones de cebada por encima del 8% mejoraron la actividad antioxidante de los filetes y aumentaron el contenido en alpha-tocoferol, sin embargo, los peces alimentados con remolacha y betaína mostraron un mayor contenido en flavonoides y fenólico pero no se observó ningún efecto sobre las propiedades antioxidantes del filete. Por último, cuando se llevó a cabo el análisis sensorial se observó que los peces alimentados con dietas con más de un 8% de cebada mostraron unas agallas más rojas y mejor textura, además de un color rojo del filete más intenso, parámetros que se correlacionan con una mejora de la frescura del pescado. Sin embargo, la inclusión de cebada o remolacha en dietas de trucha arco iris no tuvo ningún efecto sobre la aceptabilidad de los filetes. / The current Doctoral Thesis used barley and red beet as alternative carbohydrate sources in rainbow trout diets. The aim was to evaluate their effect on productive, histological and morphometric parameters, their effect on biochemical indexes after an acute stress and their effect on the quality of rainbow trout (Oncorhynchus mykiss). The different markers studied were the same in the barley and red beet experiments. In the barley experiment, increasing concentrations of barley (0-32%) were used in the diet, starting with an average weight of 127.72 ± 5.65 g and ending when they reached commercial weight after 84 days. In the red beet experiment, two red beet levels (14% and 28%) and two betaine levels (0.9% and 1.63%) were used, starting with an average weight of 69 ± 2.2 g and finishing when they reached commercial weight at the end of 105 days. The inclusion of barley in rainbow trout diets did not show significant differences in growth and biometric parameters. While the inclusion of 14% of red beet and 0.9% betaine did not affect growth, nutritive or biometric parameters and nutrient retentions compared to control, higher red beet and betaine concentrations showed a negative effect. When the histological and morphometric parameters were studied in liver and intestine, the results showed that fish fed at high barley concentrations showed smaller hepatocytes than control, while hepatocytes were larger in fish fed with red beet and betaine than control, however, in both cases, the liver had a lower level of vacuolization. Barley inclusion at 8% produced a negative effect at intestine morphological level; however, no effects were observed with the inclusion of red beet. Fish were submitted to stress, a lack of oxygen and increased of fish density, during 10 minutes. The results showed that at basal levels, the inclusion of barley and red beet did not cause any change in cortisol, glucose and MDA levels, although basal lactate values were significantly lower in the fish fed with red beet and betaine. Thirty minutes after stress the concentration of cortisol, glucose and lactate increased significantly in the fish of all groups and the concentration of MDA decreased. The inclusion of barley showed lower cortisol, glucose and lactate values than control. When fish reached commercial weight, the quality of the fillets and antioxidant properties were analysed. In both trials, fish fed with the experimental ingredients showed lower water activity values and textural and colour properties were enhanced. Red beet and betaine inclusion did not show any effect on the oxidative parameters of the fillets, while the inclusion of barley showed an inhibitory effect on fillets lipid oxidation. Concentrations of barley above 8% improved the antioxidant activity of the fillets and increased the content of alpha-tocopherol, however, fish fed with red beet and betaine showed a higher content of flavonoids and phenolics but no effect on the antioxidant properties of the fillet. Finally, when the sensory analysis was carried out, it was observed that fish fed diets with more than 8% barley showed redder gills and better texture than control, in addition to a more intense red colour of the fillet, these characteristics are correlated with an improvement of fish freshness. However, the inclusion of barley or red beet in rainbow trout diets had no effect on the acceptability of fillets. / En la present tesi doctoral es van utilitzar ordi i remolatxa com a fonts alternatives de carbohidrats en dietes per a truita amb l'objectiu d'avaluar el seu efecte sobre paràmetres productius, histologia hepàtica i intestinal, estrès i qualitat de la carn. Els marcadors estudiants fòren els mateixos en els experiments d`ordi I remolatxa. En les proves de l'ordi es van utilitzar concentracions creixents d'aquest ingredient (0-32%), es va iniciar amb un pes mitjà de 127.72 ± 5.65 g i es va finalitzar quan van arribar al pes comercial al cap de 84 dies. En les proves de remolatxa es van utilitzar dos nivells de remolatxa (14% i 28%) i dos de betaïna (0.9% i 1.63%), es va iniciar amb un pes mitjà de 69 ± 2.2 g i es va finalitzar quan els peixos van aconseguir el pes comercial al cap de 105 dies. La inclusió d'ordi en dietes de truita no va mostrar diferències significatives en els paràmetres de creixement i biomètrics. La inclusió de 14% remolatxa i 0,9% betaïna no va afectar el creixement, paràmetres nutritius, biomètrics i retencions de nutrients en comparació amb el control, concentracions majors de remolatxa i betaïna van tenir un efecte negatiu. Quan es van estudiar els paràmetres histològics i morfomètrics en fetge i intestí, els resultats van mostrar que els peixos alimentats amb concentracions creixents d'ordi tenien hepatòcits més petits, mentres que els peixos alimentats amb remolatxa i betaïna presentaven hepatòcits més grans, i en ambdós casos el fetge va mostrar un menor nivell de vacuolització. La concentracion d'ordi d'un 8% va produir un efecte negatiu a nivell morfològic de l'intestí, mentres que la inclusió de remolatxa no va produir cap efecte a nivell d'intestí. Els peixos van ser sotmesos a estrès per absència d'oxigen i augment de densitat durant 10 minuts. Quan l'estrès va ser analitzat a nivell bioquímic els resultats van mostrar que a nivells basals, la inclusió d'ordi i remolatxa no va causar cap canvi en els nivells de cortisol, glucosa i MDA, encara que els valors basals de lactat van ser significativament més baixos en els peixos alimentats amb remolatxa i betaïna. 30 minuts després de l'estrès la concentració de cortisol, glucosa i lactat va augmentar significativament en els peixos de tots els grups i la concentració de MDA va disminuir. La inclusió d`ordi va mostrar valors més baixos de cortisol, glucosa i lactat que la dieta control. Quan els peixos van aconseguir el pes comercial es va analitzar la qualitat dels filets i propietats antioxidants. En ambdues proves els peixos alimentats amb els ingredients objecte d'estudi van mostrar valors d'activitat d'aigua més baixos i una millora en la textura i el color dels filets comparat amb els peixos alimentats amb la dieta control. La inclusió de remolatxa i betaïna no va tenir cap efecte sobre els paràmetres oxidatius del filet, mentre que la inclusió d'ordi va inhibir l'oxidació lipídica dels filets. Concentracions d'ordi per sobre del 8% van millorar l'activitat antioxidant dels filets i van augmentar el contingut en alpha-tocoferol, però, els peixos alimentats amb remolatxa i betaïna van mostrar un major contingut en flavonoides i fenòlic però no es va observar cap efecte sobre les propietats antioxidants del filet. Finalment, quan es va dur a terme l'anàlisi sensorial es va observar que els peixos alimentats amb dietes amb més d'un 8% d'ordi van mostrar unes ganyes més vermelles i millor textura, a més d'un color vermell del filet més intens, paràmetres que es correlacionen amb una millora de la frescor del peix. No obstant això, la inclusió d'ordi o remolatxa en dietes de truita no va tenir cap efecte sobre l'acceptabilitat dels filets. / Pinedo Gil, J. (2018). Study of novel carbohydrate sources on rainbow trout (Oncorhynchus mykiss) diets [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/107364 / Compendio

Current literature suggests that little, if any, research has been conducted in South Africa to determine the impact of alien invasive trout on indigenous amphibian biodiversity. The primary aim of the research project was to establish whether waterfalls are seasonally important in protecting the indigenous Natal Cascade Frog Hadromophryne natalensis tadpole populations from predation by alien rainbow trout Oncorhynchus mykiss and brown trout Salmo trutta at two sampling areas located at Injesuthi and Monk‟s Cowl Nature Reserve within the uKhahlamba Drakensberg Park World Heritage Site (UDPWHS). The dissertation argues in favour of a biotic rationale, namely trout predation, as being the primary cause for the decline of H. natalensis tadpoles below the waterfalls and systematically negates the influence of geo-physical (abiotic) environmental factors on tadpole abundance. Habitat isolation and fragmentation is identified as a latent threat to the continued persistence of H. natalensis populations in the UDPWHS. / Environmental Sciences / M. Sc. (Environmental Science)