β2-microglobulin (β2m) exists free of the major histocompatibility complex class I (MHC I) receptor in many bodily fluids. The amount of protein present in these fluids has been found to be a useful prognostic marker for various diseases but outside of its practical value not much is known about this form of β2m. In fish, soluble β2m has not been studied at all. Another unknown in fish is the effects that plants lectins might have on naturally carnivorous species in aquaculture. These plant proteins which bind to specific sugar groups found on cells have been shown to have a multitude of gastrointestinal and immune effects in mammals and can be found in the plant products being fed to carnivorous, cultured fish making them possible toxicants. The two studies of this thesis set out to pioneer knowledge on these subjects using rainbow trout as a model. The first investigation inspected the various body fluids of these fish for their free β2m content. Soluble β2m was found to be present in the plasma, the seminal fluid, ovarian fluid, and the mucus of the skin and intestines. This distribution shows that β2m could indeed make a good biomarker, not only for disease but also for pheromone release and alludes to some possible functions of soluble β2m while opening the way for future research on this form of the protein. The second study looked at the effects of lectins on the gut of rainbow trout by treating RTgutGC, an intestinal epithelial cell line derived from trout, with plant lectins from wheat (WGA) and soybean (SBA), among others. This study found WGA to be a potent inducer of morphological and cytotoxic effects in these cells while other lectins and plant factors were not. WGA was also observed to effect the expression of β2m and the α-chain of the MHC I receptor. This work suggests WGA ingested by trout through the wheat in their diet might be causing them harm and should be studied further. It is also interesting that both studies related β2m to the intestines of trout. This could allow soluble β2m to serves as a marker of WGA’s effect or for WGA to aid in the study of free β2m.

Proliferative Kidney Disease (PKD) caused by the Malacosporean parasite Tetracapsuloides bryosalmonae, is presently the most economically damaging disease of British rainbow trout farming, costing the industry in excess of £2.5 million per annum in the UK alone. With no vaccine or prophylactic treatment available, and only management techniques currently adopted to minimise the stress and mortality associated with the disease, alternative approaches must now be considered. This document investigates if selective breeding for PKD resistance is possible by assessing the level of additive genetic variation, and calculating the subsequent estimates of heritability, for commercial strains of rainbow trout. During a PKD outbreak on a commercial farm, 1500 communally reared juvenile rainbow trout from two strains (Houghton Spring and Isle of Man) were sampled on a single day, their body weight and fork length measured, and severity of kidney swelling scored according to the scale of Clifton-Hadley et al. (1987). Fish were assigned to individual families using microsatellite parentage assignment. Significant additive genetic variation was observed in the population, and families were ranked according to estimated breeding values. A combined estimate of heritability (h2 = 0.19 ± 0.08) for kidney score suggests the population will respond well to selective breeding for kidney score, which may be deemed a measure of resistance, whilst the favourable genetic correlations between kidney score and the production traits measured suggest simultaneous selection for kidney score and growth traits should also be effective. In order to support the findings of the initial research, controlled challenge experiments were conducted. Using the family EBV information on kidney score from the IoM strain (due to its certification as a disease-free site), four females, two with high and two with low response to PKD, were each crossed with a randomly selected neomale to produce twenty two families for PKD challenge experiments. The PKD experimental challenges showed evidence of additive genetic variation to kidney score over an eleven week period, supporting initial findings. A low score was deemed as evidence of greater resistance to the parasite in this study. Although female EBV was taken into consideration in the statistical model, there was found to be no significant difference in resistance according to family. Immunohistochemistry stained kidney sections from each individual involved in the challenges proved kidney score correlated significantly to the number of parasites in the kidney, suggesting that the scale of Clifton-Hadley et al. (1987) is a sufficient and accurate basis on which to describe the severity of PKD, and infection level in rainbow trout. Having discovered evidence that furunculosis, causative agent Aeromonas salmonicida, plays a major role in the mortality of fish suffering from PKD in the field, the bacterial disease was investigated to assess the resistance of the same families used in the PKD challenges. Twenty one of the families were used to discover that additive genetic variation for resistance to furunculosis is apparent when assessed as both a binary and longitudinal trait, suggesting significant genetic improvement can be made to increase resistance to furunculosis in the IoM stock. No significant correlation was observed between kidney score, EBV, and resistance to this bacterium, but there was a positive phenotypic correlation found between furunculosis resistance and size, suggesting simultaneous selection for performance and resistance is possible within this population.

<p>Diss. (sammanfattning) Umeå : Umeå universitet, 1995, härtill 5 uppsatser.</p> / digitalisering@umu

Η λεκτίνη ΜΒL συμμετέχει στην φυσική ανοσία, αφ΄ενός σαν ενεργοποιητής του συστήματος του συμπληρώματος και αφ΄ετέρου σαν οψωνίνη που προσδένεται σε συγκεκριμένες υδατανθρακικές δομές των μικροοργανισμών. Οι λεκτίνες τύπου C, είναι ασβέστιο εξαρτώμενες και φέρουν περιοχή δέσμευσης σε υδατάνθρακες (CRD). Προκειμενου να διασαφηνιστεί περαιτέρω η εξελικτική πορεία της λεκτινικής οδού του συμπληρώματος, απομονώθηκαν, κλωνοποιήθηκαν και χαρακτηρίστηκαν δύο ισομορφές της λεκτίνης ΜBL, οι ΜΒL1 και MBL2, στην ιριδίζουσα πέστροφα (Oncorhynchus mykiss). Οι συναγώμενες αμινοξικές αλληλουχίες των MBL1 και ΜΒL2 είναι 185 και 186 αμινοξέα, αντίστοιχα, παρουσιάζουν μεταξύ τους ταυτοσημία της τάξης του 83% ενώ εμφανίζουν το υψηλότερο σκορ ταυτοσημίας, 43% και 41% αντίστοιχα με τους τελεόστεους ιχθύες Αtlantic salmon και zebrafish. Το σκορ ταυτοσημίας των πρωτεϊνών της πέστροφας με τις αντίστοιχες πρωτεΐνες των πτηνών και των θηλαστικών κυμαίνεται μεταξύ 28 και 32%. Επιπλέον,οι περιοχές CRD των πρωτεϊνών αυτών χαρακτηρίζονται από την ύπαρξη του δομικού μοτίβου EPN που σχετίζεται με την εξειδίκευση της δέσμευσης ως προς την μαννόζη. Τα αποτελέσματα έδειξαν επίσης πως τα μόρια MBL1 και ΜBL2, εκφράζονται, σε επίπεδο mRNA, στο ήπαρ και στον σπλήνα της πέστροφας, αντίστοιχα. / Mannose-binding lectin(MBL) participates in the innate immune system as an activator of the complement system and as an opsonin after binding to certain carbohydrate structures on microorganisms. C-type lectins are all Ca2+ -dependent and they share a tightly folded carbohydrate recognition domain (CRD). In order to stydy the evolution of the complement lectin pathway, we report the isolation and characterization of two mannose-binding lectin isoforms, MBL1 and MBL2, in rainbow trout (Oncorhynchus mykiss). The deduced amino acid sequences of trout MBL1 and MBL2 are 185 and 186aa, respectively, presenting 83% identity to each other. These proteins exhibit the highest identity score 43 and 41% with the atlantic salmon and zebrafish counterparts. The identity with the bird and mammalian MBLs ranges from 28 to 32%. The trout MBL molecules contain in the CRD domain the EPN motif of mannose-binding C-type lectins, which is important for mannose specificity. Trout MBL1 and MBL2 are expressed exclusively in liver and spleen, respectively.

No description available.

The physiological responses to stressor exposure can be broadly grouped into the organismal and the cellular stress responses. The organismal stress response involves the release of hormones into general circulation, while the cellular stress response involves the synthesis of proteins, the most important being the heat shock proteins (HSPs), which play a role in maintaining protein homeostasis. Elevated HSP70 expression in response to stressors has been demonstrated in trout (Oncorhynchus mykiss) red blood cells (RBCs). The ease of repeated sampling of blood suggests the possibility of using this tissue as a non-lethal marker of cellular stress in fish. This study tested the hypothesis that stressor exposure will elevate HSP70 expression in trout RBCs and the role of stress hormones in mediating this response. Acute heat shock exposure (+12oC) significantly elevated plasma cortisol, glucose and lactate levels in heat shocked fish over 24 h. A tissue-specific response was seen in HSP70 expression in liver, brain, gill and RBCs. To enable measurement of RBC HSP70 concentrations, an enzyme-linked immunosorbent assay (ELISA) was developed using a commercially available rabbit anti-salmon HSP70 and a recombinant chinook salmon (Oncorhynchus tshawytscha) HSP70. To determine effects of chronic exposure, two studies were conducted exposing trout to either cadmium (0, 0.75 or 2.0 µg/L over 28 d) or municipal wastewater effluent (0, 20 or 90% over 14 d). However, neither exposure elicited a significant HSP70 response. Effects of stress hormones on RBC HSP70 levels were tested by exposing cells in vitro to either cortisol (10 and 100 ng/mL) or epinephrine (10 nM) with or without heat shock. Heat shock elevated HSP70 content in trout RBCs but no modulation by stress hormones was seen. It was shown for the first time that RBCs release HSP70 content into the medium in response to an acute heat shock and this release is attenuated by stress hormones. Overall, HSP70 levels in RBCs have the potential to be a reliable non-lethal marker of acute cellular stress effects in fish. The release of HSP70 from RBCs leads to the hypothesis that HSP70 may also have an extracellular role in fish, and warrants further study.

The purpose of this study was to investigate whether a cyclic feeding regime alters growth, lipid content and the expression of genes involved in appetite regulation and lipid metabolism in rainbow trout (Onchorynchus mykiss). Progeny from two lots of diallel crosses were raised under a daily fed control or a cyclic feeding regime containing alternating reduced and compensatory rations. Although the cyclic feeding regime did not improve overall growth, there was some degree of compensatory growth seen during re-alimentation, and the reduced ration led to sustained reductions in condition factor and muscle fat content. Appetite-regulating genes showed little or no differences in expression between feeding regimes. However, the lipid metabolism genes involved in fatty acid synthesis and muscle lipid uptake showed differences in expression during reduced and compensatory ration. This study helps to clarify optimal cyclic feeding regimes in order to enhance growth characteristics preferential to the fish culture industry. / NSERC Strategic Grant, OMAFRA

Approaches for the evaluation of plant protein ingredients for salmonid feeds were investigated in a series of four trials. The first trial compared the apparent digestibility coefficients (ADC) of crude protein (CP) and amino acids (AAs) of two novel products - Indian mustard protein concentrate (IMC, 62% CP) and Indian mustard protein meal (IMM, 42% CP), to a commercially available soy protein concentrate (SPC, 57% CP) for two salmonid species, rainbow trout and Atlantic salmon. The second trial involved assessment of relative bioavailability of arginine (Arg) from IMC, IMM and SPC compared to that of a crystalline Arg (L-Arg) in rainbow trout using slope-ratio assay. In the third trial, the effects of phytic acid (PA) and lignin on nutrient utilization and partitioning in rainbow trout were assessed. Finally, a series of experiments was conducted in the final trial to establish the evaluation criteria for pellet quality assessment. The ADC of CP and most AAs in IMC and IMM were high (>90%). Differences in the ADCs of some AAs can be attributed to the high PA intake by fish fed 30%-IMC diet. The significantly higher (P<0.05) bioavailability of Arg from IMC (123 to 187%) and IMM (116 to 211%) relative to that of L-Arg, as determined by various regression approaches, reaffirmed the findings of the first trial that these ingredients are of excellent protein quality and can readily be used in compounded fish feeds. It can be inferred from the lack of effects of PA, lignin or PA plus lignin on most indices of physiology, performance, and nutrient utilization in the pair-fed fish, that like any other animal, controls feed intake when in the presence of one or more dietary ANF. It was also shown in the pellet quality assessment trial that minor changes in dietary composition can significantly alter physical properties of aquaculture feed. This study highlighted the importance of a comprehensive assessment for the effective evaluation of the nutritive value of plant protein ingredients for use in aquaculture feeds. / Ontario Ministry of Natural Resources; BIOEXX Specialty Proteins Ltd.; Martin Mills Inc.; MITACS

The accumulation of persistent contaminants is a significant issue for the health of aquatic environments. This study aims to determine the effects of polycyclic aromatic hydrocarbons (PAH) on the stress response of fish by monitoring plasma cortisol levels and the expression of key hypothalamic-pituitary-interrenal (HPI) stress axis regulators. Injection of benzo(a)pyrene (BaP), a ubiquitous PAH, induced a differential dose- and time-dependentcortisol response in rainbow trout and brown bullhead. BaP exposure also elicited a species-specific transcriptional response at all levelsof the HPI axis.Similarly, the HPI axis response to a standardized emersionstressor revealed species-specific differences. In the field, exposure of different brown bullhead populations to sediment with complex PAH mixtures did not consistently affect cortisol levels and providedno evidence of genetic adaptation of the stress response. Thus, future studies are needed to bridge the gap in our understanding between the laboratory and field effects of PAHs on the stress response of fish.

Stocking trout to create successful sport fisheries is an irresistible lure to fisheries managers and sport anglers alike, but the implicit assumptions behind this simple process have seldom been questioned or assessed. Using common fisheries monitoring techniques, combined with social surveys, at nine Alberta lakes, I quantified three main assumptions behind put-and-take stocking. Surprisingly, 1) stocking high densities of Rainbow Trout created very low-density populations; 2) these populations supported mediocre fisheries; 3) these mediocre fisheries, if above a threshold catch rate, attracted very large numbers of satisfied anglers. Based on these findings, the stocking density and the direct cost of stocking were reduced by 80% at three experimental lakes. No major decreases in fishing quality, angler participation, or angler satisfaction were observed. I suggest refinements in the stocking process focus on determining how to provide adequate numbers of trout to create a basic level of satisfaction with the fishing experience.