Der Einsatz von Branntkalk war noch vor wenigen Jahren Bestandteil der »guten fachlichen Praxis« in der Karpfenteichwirtschaft. Im Hinblick auf den Schutz von Tier- und Pflanzenarten sowie geschützte Biotope wurde der Branntkalkeinsatz seit einigen Jahren vorsorglich eingeschränkt. Der Bericht dokumentiert die Ergebnisse einer Untersuchung zur Wirkung unterschiedlich hoher Branntkalkgaben auf die Wirbellosenfauna, Amphibien, Libellen und Makrophyten in Praxisteichen sowie in der Lehr- und Versuchsteichanlage Königswartha. In Laborversuchen wurden auch die Auswirkungen unterschiedlicher Branntkalkmengen auf Amphibienlaich und -larven untersucht. Die schädigende Wirkung von Branntkalk ist abhängig von der Dosierung. Während Einzelgaben unter 500 kg/ha praktisch ohne Auswirkungen bleiben, sind bei Desinfektionskalkungen auch Artenschutzbelange zu beachten.

The intestine and associated bacterial microbiota have a central role the physiology and homoeostasis of the host. The understanding of how farming conditions affect the intestine and associated microbiota of fish is the high importance to counteract the potential threats to health and welfare. Thus, this thesis aims to understand the role of stressful husbandry conditions on the intestine and associated microbiota of rainbow trout and Atlantic salmon. Within this context, the role of Pediococcus acidilactici as health promoter was also investigated Chapter 3 investigated the replacement of fishmeal by different plant protein ingredients in rainbow trout. The results of this chapter revealed that the effect of P. acidilactici on the microbiota of distal intestine in rainbow trout was dependent on the ingredients of the diet. The results also showed that the FM substitution induced major changes in the intestinal microbiota. Moreover, the modulation induced by plant-based diets on the microbiota varied according to the ingredients used. Chapter 4 studied the effect of dietary oxytetracycline in the distal intestinal microbiota of rainbow trout and the role of P. acidilactici to ameliorate the impact of antibiotic therapy. Experimental groups fed the diets with oxytetracycline had substantial changes in the distal intestinal microbiota including a decrease in the bacterial diversity. P. acidilactici did not ameliorate the effect of antibiotic therapy in the intestinal microbiota. Chapter 5 used Atlantic salmon during smoltification to study the changes in the microbiota of distal intestine and the role of P. acidilactici to promote intestinal health. The results showed that bacterial communities in the mucosa differed from the digesta. Seawater transfer and P. acidilactici had significant changes in the intestinal microbiota of both mucosa and digesta. However, the modulatory effect of both factors evaluated was larger in the mucosa-associated microbiota than in the digesta-associated microbiota. Furthermore, P. acidilactici induced a significant increase in antiviral-related genes. Chapter 6 investigated the replacement of fish oil by rapeseed oil alone or combined with P. acidilactici on the intestinal health and microbiota of two intestinal regions in Atlantic salmon. Replacement of fish oil by rapeseed oil alone or in combination with P. acidilactici supplementation did not induce major changes in the intestinal health and microbiota. The bacterial communities found were significantly different between the pyloric caeca and mid-intestine. In conclusion, this thesis contributes to new knowledge regarding the effect of dietary supplementation of P. acidilactici and the impact of different potential challenging factors in the health and intestinal microbiota of farmed salmonid species.

The study of the gut microbiota of fish began in the 1930’s and since that time a considerable amount of information has been collated on its composition and diversity. These studies have revealed that the microbial communities of the fish gastrointestinal tract are generally difficult to culture on bacteriological media and mainly consist of bacteria, archaea, viruses, yeasts and protists. The bacteria appear to be the most abundant of these microbial groups and their activity may have major implications for host health, development, immunity and nutrition. Therefore, much of the most recent published research has focused on developing improved methods of identifying the extent of the bacterial diversity within the fish gut and unravelling the potential influence of these microorganisms on the health of farmed fish species. However, whilst such studies have improved our knowledge of the dominant bacterial groups present in the rainbow trout gastrointestinal tract, the limited resolution capacity of many of the methods used has meant that our understanding of their baseline composition in healthy fish remains poorly understood. In this study, the bacterial communities that inhabit the intestine, now commonly referred to as the ‘microbiome’, of farmed Rainbow trout (Oncorhynchus mykiss) were characterized using a culture independent high-throughput molecular sequencing method. The microbiome of the intestinal lumen and mucosa was investigated to ascertain the true extent of the bacterial diversity present in this fish species prior to further experiments. It was found that the diversity of the intestinal microbiome was greater than previous studies had reported with a total of 90 and 159 bacterial genera being identified in both the lumen and mucosal regions respectively. The dominant bacterial phyla identified in both of the regions investigated were Proteobacteria, Firmicutes, Fusobacteria, Bacteroidetes and Actinobacteria. Furthermore, the data collected suggested that the intestinal microbiome may be similar in structure between individual fish, and illustrate the utility of next generation molecular methods in the investigation of the fish gut microbiome. A study was conducted to examine the effect of diet on the composition of the intestinal microbiome of rainbow trout. Two diets, one control and one treatment, were prepared which were identical apart from that the treatment diet contained a microalgal component at 5% of the total formulation. These diets were fed to rainbow trout for a total of 15 weeks. At the end of the trial period a total of 12 fish, three from each of four tanks, were sacrificed from each of the control and treatment groups and their intestinal tissue was sampled in order to compare the composition of the microbiome of both groups. The results revealed that both groups of fish shared similar microbiome compositions, with the Tenericutes being by far the most dominant phylum observed. The structure of the intestinal microbiome was not significantly different between both populations of trout tested. An increased level of bacterial diversity was noted in the treatment fish, however, this was not found to be statistically significant. A limited number of bacterial taxa were discriminatory between diets and were significantly elevated in the treatment group. These taxa were predominantly lactic acid bacteria of the genera Streptococcus, Leuconstoc, Lactobacillus, Lactococcus and Weissella. The results of this study suggested that the minor difference in the diets fed resulted in a correspondingly minor alteration in the intestinal microbiome of the tested rainbow trout. This may indicate that diet composition can modify the composition of the intestinal microbiome of these fish. A further study was conducted to investigate the structure of the intestinal microbiome from groups of fish reared in both freshwater cages and aquarium systems, in order to assess whether or not fish raised in different environments share similar microbiomes. This study also employed a novel computational tool, PICRUSt, to analyse the predicted functional capacity of the microbial communities of individual fish sampled from both environments. The data collected suggested that the structure of the intestinal microbiome was similar regardless of where the fish were raised, with the Tenericutes, Firmicutes, Proteobacteria, Spirochaetae and Bacteroidetes representing the dominant bacterial phyla recorded in the rainbow trout intestine. This suggests that the host may regulate the formation of the intestinal microbiome. A significant difference was however noted in community membership between the fish populations tested, which may point to an environmental influence on the intestinal microbiome. These data suggest that both deterministic host factors and stochastic environmental influences play important roles in shaping the composition of the bacterial communities in the intestine of these fish. The PICRUSt analysis revealed that gene pathways relating to metabolism, transport and cellular processes were enhanced in all of the fish studied, which may signal an involvement of these communities in the digestive processes of rainbow trout. In conclusion, this study used high-throughput sequencing methods in order to improve our understanding of the intestinal microbiome of farmed rainbow trout, and the effect of dietary and environmental factors on its composition. This research has generated scientific information relating to baseline bacterial community compositions in healthy fish, which may be used in future experiments including screening these baselines against the effects of novel aquafeed formulations, environmental perturbations or pathogenic challenges.

Over the past few years one of the major concerns in the Atlantic salmon (Salmo salar) farming industry has been the increasing incidence and severity of inflammatory viral diseases. Heart and skeletal muscle inflammation (HSMI) and cardiomyopathy syndrome (CMS) are currently two of the most prevalent viral diseases in commercial Atlantic salmon farms in Norway. Mortality levels in both diseases are generally low but morbidity can be very high with the associated chronic inflammatory response lasting for several months. The consequent reduced growth performance is causing considerable financial impact as HSMI has become increasingly widespread in recent years. The impact of CMS is further exacerbated as it generally affects large fish close to harvest. HSMI lesions occur in the atrium and ventricle in the heart including inflammation and necrosis in epi- endo- and myocardium along with myositis of red skeletal muscle. CMS lesions are commonly observed in the spongy myocardium in the atrium and ventricle of the heart with severe mononuclear inflammation and necrosis. Furthermore, circulatory disturbances associated with reduced cardiac function cause multifocal liver steatosis and necrosis in both diseases. Currently there are no vaccines or any other effective treatments for these diseases and so alternative therapies that could potentially modulate the intensity of the inflammatory response could be crucial to improve the clinical manifestation of the diseases. Therefore, the overall aim of the present study was to evaluate the concept of “clinical nutrition” to improve the clinical symptoms of both viral diseases, HSMI and CMS, through the use of functional feeds formulated with reduced lipid content and increased proportions of anti-inflammatory fatty acids to moderate the apparently uncontrolled inflammatory response in the heart tissue associated with both diseases and also alleviate the secondary hepatic lesions. The experimental work consisted of three major dietary trials in Atlantic salmon in seawater. Two large trials investigated the effects of functional feeds in Atlantic salmon challenged with Atlantic salmon piscine reovirus (ASRV) and piscine myocarditis virus (PMCV), the causal agents of HSMI and CMS, respectively. In both trials, heart transcriptome, heart and liver histopathology and tissue lipid and fatty acid compositions and metabolism were determined post-infection in fish fed with the functional feeds in comparison with fish fed with a standard commercial feed formulation considered as a reference diet. All the functional feeds were formulated to have reduced digestible energy through lower dietary lipid and higher protein contents, and increased levels and proportions of anti-inflammatory long-chain polyunsaturated fatty acids (LC-PUFA), particularly eicosapentaenoic acid (EPA) compared with the reference diets. Histopathology, fatty acid composition and gene expression of heart were assessed over a long time-period of 16 weeks and 14 weeks post-challenge with ASRV and PMCV, respectively. Viral load in heart tissue, hepatic histopathology and fatty acid composition of liver and head kidney along with expression of the genes involved in the eicosanoid and LC-PUFA and eicosanoid biosynthesis pathways were also determined in the HSMI trial. The third trial was a nutritional trial evaluating the effects of dietary digestible energy content on lipid and fatty acid metabolism in salmon fed diets containing graded amounts of lipid. Fatty acid composition of liver and heart were assessed over 12 weeks, along with the hepatic expression of genes of lipid and fatty acid metabolism. The results of this research are presented in four chapters (Chapters 2-5) as four paper manuscripts. The manuscripts/Papers are either published (Chapter 2), in review (Chapter 3 and 4) or drafted for submission (Chapter 5) in appropriate peer-reviewed international journals. Chapter 2 and 3 correspond to the HSMI trial, Chapter 4 to the nutritional trial, and Chapter 5 to the CMS trial. Chapter 2 showed that viral load and histopathology scores were lower in fish fed the functional feeds, especially diet FF1, which displayed better performance. Diet strongly influenced the expression of genes related with the immune and inflammatory responses, with delayed expression in fish fed the functional feeds. Up-regulation of pro-inflammatory genes was correlated with the higher viral load observed at early-mid stages of the disease in fish fed the reference diet (ST). Expression of genes related with the immune response at 16-weeks post challenge reflected the differences in immunomodulation between the functional feeds, with fish fed diet FF1 showing lower expression. Therefore, severity of the heart lesions was correlated with the intensity of the immune response and could be associated with tissue anti-inflammatory LC-PUFA levels. Chapter 3 was focused on liver histopathology, fatty acid composition and LC-PUFA biosynthesis, along with phospholipid fatty acid composition and eicosanoid production in head kidney and heart tissue at early and late stages of ASRV infection. Liver was severely affected by the virus at the beginning of the infection in fish fed the reference ST diet, but the level of lesions were similar in all dietary groups at the end of the trial. Hepatic expression of fatty acyl desaturases was significantly depressed in fish fed the ST diet compare with fish fed the functional feeds despite the lower levels of dietary LC-PUFA in that feed. Thus endogenous production and bioavailability of anti-inflammatory LC-PUFA was potentially enhanced in fish fed the functional feeds. Changes in tissue lipid content, mobilization of fatty acids involved in inflammatory responses and changes in expression of transcription factors and genes involved in eicosanoid biosynthesis were more prominent in head kidney, confirming the important role of this organ in dietary immunomodulation after viral infection. To a lesser extent similar changes were observed in heart tissue, suggesting in situ production of eicosanoids could also be important. The unexpected effects of diet on expression of genes of LC-PUFA biosynthesis were specifically investigated in the trial described in Chapter 4. One aim of this study was to clarify whether dietary lipid content or viral infection was the cause of altered expression of desaturase genes between the different diets. Hepatic expression of other genes of lipid and fatty acid metabolism were also determined to evaluate metabolic changes associated with dietary lipid/energy level. In general, reduction of dietary energy and lipid contents while maintaining similar proportions of dietary fatty acids, led to a general up-regulation of genes involved in lipid biosynthetic pathways. Thus salmon fed lower energy diet showed increased liver expression of fatty acyl desaturases in comparison with fish fed higher energy levels. Heart transcriptomic data in Chapter 5 showed a similar delay in the inflammatory response in fish fed the functional feeds after PCMV infection as observed in the HSMI study. Modulation of inflammatory responses, similar to that previously described after ASRV infection, was also observed in fish fed the functional feeds. However, the differences in the expression of immune related genes and the level of heart lesions were not as prominent at mid-late stages of the disease as in fish fed FF1 in the HSMI trial. The present study demonstrated the beneficial effects of a clinical nutrition approach via functional feeds in two viral inflammatory diseases, HSMI and CMS, currently affecting farmed Atlantic salmon.

Aquaculture is playing an important role in the development of fisheries in Vietnam, a role which has accelerated since 2000. Sustainability in aquaculture is receiving increasing attention, and this issue is not only the concern of government, but also stakeholders participating in the value chain. Therefore, this study aims to identify sustainability issues of farmed seafood by assessing the main sustainability issues raising concern. The Global Value Chain framework described by Gereffi et al. (2005) is applied for this study to explore the business relationships in supply chain and the perceptions of sustainability concerned by the value chain actors. A combination of qualitative and quantitative methods was used to collect data. An assessment of four species cultured on the Mekong Delta, the countries farmed seafood ‘hub’, found a clear distinction between species cultured with a local domestic market orientation (Giant Freshwater Prawn Macrobrachium rosenbergii; and Tilapia Oreochromis niloticus) and the two key export commodities - Striped catfish (Pangasianodon hypophthalmus) and Penaeid shrimp (Penaeus monodon & Litopenaeus vannamei). These orientations were based on a range of factors including the development of effective seed supplies and the cultural value of marketing in the live format. This study, conducted in ten provinces in the Mekong Delta from 2009 to 2013 had a focus on understanding the export-orientated commodities, striped catfish and shrimp through collection of baseline information on the value chain of farmed seafood, focusing on the farming sector, the actors and institutions involved and beneficiaries. Sustainability issues and perceptions of experts (top-down) and primary stakeholders (bottom-up) opinions were assessed through participatory workshops. Shrimp and striped catfish production are mainly farmed for export, with 83% and 95% of its production, respectively, leaving the country mainly after processing. Currently, mainly families operate small-/medium-scale farms; while large-farms are integrated within seafood processors. Production efficiency of large-farms tends to be better than small-/medium farms. Many striped catfish and shrimp farms are likely to reach several standard criteria such as economic feed conversion ratio (eFCR), stocking density, no banned chemical/drug and wild-seed use, and land property rights; however, there were still many standard criteria that existing farms could not meet such as effluent management, farm registration, fishmeal control, farm hygiene and record-keeping requirement. Hence, current farming practices, especially small-/medium farms have a long way to go to meet emergent international food standards. Recently, many small-/medium catfish farms faced problems with low fish prices, so they have had to cease catfish farming activities and temporarily stop farming; while some larger farms also had to temporarily stop farming. Therefore, fish price has tended to be a main driving force for catfish farm changes. In the shrimp industry, there were technical changes occurring in the high intensity level of shrimp farms (HiLI); whereas, the remaining shrimp farms had fewer changes in farm management. Most HiLI shrimp farms were affected by AHPNS disease, which was a main factor driving their farm changes. Many perceptions of sustainability were identified by stakeholder groups, however seven sustainability issues had a high level of agreement among stakeholders including input cost, capital & credit costs, unstable markets, government regulation & policy, disease, seed quality, water quality and water availability factors. Hatcheries, farmers and manager groups were more concerned about environmental issues; while for the input suppliers and processors, economics was the main issue. Farmers and processors were two main actors that played an important role in the production process of the value chain. Small-/medium farms dominated the number of farms overall and still played an important role in primary production. However, small-scale farms were considered as more vulnerable actors in the value chain, and they faced more difficulties in meeting increasing requirements on food quality/safety. To maintain the position in the value chain, the solutions could be horizontal and vertical coordination. Thus policy makers will need to find ways to include them in the planning processes. To reach sustainability will require the efforts of direct stakeholders, the role of the state agencies is essential in negotiation and diplomacy to create partnerships with the seafood importing countries. However, efforts to develop sustainable production become impossible without participation from importers, retailers and consumers.

The giant freshwater prawn, Macrobrachium rosenbergii (De Man, 1879), is gaining popularity as a key aquaculture species; global production currently exceed 220,000 tonnes, however, industry expansion is limited by high operational costs, with the feed accounting for between 40 to 60% of these. Attention, therefore, has been devoted to increasing the inclusion of plant proteins into the formulation of aqua feeds as a consequence of the limited, unpredictable supply and increasing price of fishmeal. The concomitant introductions of anti-nutritional factors (ANFs), such as phytic acid (PA) with the plant protein fraction, however, are major impediments in the efforts toward the increased use of plant protein ingredients in aqua feeds. Phytic acid is an anti-nutrient that can curtail the development of this as PA has been reported to suppress growth impairing proper nutrient intake, diminishing the availability of minerals, and causing damage to the body tissues and organs which can result in mortality. Although the anti-nutritive effects of PA have been studied extensively in terrestrial agriculture farm species, as well as in a variety of fish species, there is almost no information regarding the effects of PA in crustaceans, including the freshwater prawn, M. rosenbergii. The aims of this present thesis were, therefore, to gain a greater understanding of dietary PA and microbial phytase and their effect on growth performance, feed utilisation, nutrient utilisation and digestibility, mineral availability and whole body proximate composition in juvenile M. rosenbergii. Specifically, the first major experiment set out to investigate the effect of including increasing amounts of PA in the diets presented to M. rosenbergii on growth. The dose-response relationships between PA and growth performance, feed utilisation, nutrient digestibility and utilisation and whole proximate composition were investigated. Four replicate groups of M. rosenbergii with a mean initial carapace length of 6.03 ± 0.30 mm and mean initial weight of 0.29 ± 0.02 g were fed graded levels of PA for 140 days. The basal diet, to which different levels of PA were added to obtain 0.26 (control), 6.48, 11.28, 16.53, 21.45 and 26.16 g PA kg-1, contained fishmeal, soy protein concentrate, wheat meal and corn starch. The results indicated that growth performance, feed utilisation and survival did not differ significantly between the groups receiving the different inclusions of PA within their diets. The apparent protein, lipid and energy utilisations responded negatively, decreasing significantly (p<0.05) with an increasing inclusion of PA, particularly within the groups of prawns fed the diet with the highest inclusions of PA, i.e. the 21.45–26.16 g PA kg-1 diets. The digestibility of protein and lipid were also reduced as the inclusion of PA increased. The whole body composition of protein (p<0.04), lipid (p<0.01) and gross energy (p<0.05) decreased significantly with an increasing supplementation of PA, while the ash content significantly increased (p<0.01), most notably in the groups of prawns receiving the highest levels of dietary PA. The second major experimental trial investigated the effect of microbial phytase on the growth of juvenile M. rosenbergii, when fed diets supplemented with various doses for a period of 80 days. The study set out to improve the growth performance, feed utilisation, nutrient digestibility and utilisation and body composition of M. rosenbergii when fed diets high in plant protein ingredients. To investigate this, four plant protein based diets, which included soybean meal, wheat gluten and wheat meal, were formulated and supplemented with microbial phytase at levels of 0, 500, 1000 and 2000 FTU kg-1 (one phytase unit per kg) and fed to sixty juvenile M. rosenbergii (mean initial carapace length of 8.51 ± 0.52 mm; mean initial weight of 0.40 ± 0.07 g) for 80 days. High levels of plant protein in the diets supplemented with 0–2000 FTU kg-1 did not result in any negative effect on growth performance, feed utilisation nor on the survival of M. rosenbergii. Noticeable moderate growth improvements in line with increasing microbial phytase supplement levels were observed and the highest growth performance was seen in the group fed 2000 FTU kg-1. Supplementation of the diets with 500–2000 FTU kg-1 were found to affect the nutrient utilisation, resulting in a significant (p<0.05) increase in the protein and lipid utilisation when compared to the prawns analysed from the control group. In addition to this latter finding, an increasing supplement of phytase in the diet also resulted in an increase in the dry matter fraction as well as improvements in the digestibility of protein and lipid. There were, however, no significant differences in moisture, protein, lipid, gross energy and the ash content of the whole body of the M. rosenbergii among the groups. This thesis, in a third major trial, explored the impact of other potential ANFs associated to PA, the binding effect of PA with mineral. The effects of graded levels of PA (i.e. 0.26–control, 6.48, 11.28, 16.53, 21.45 and 26.16 g PA kg-1) on the moult frequency and mineral availability in juvenile M. rosenbergii fed over a period of 140 days were determined. The levels of PA assessed in this feed trial had no major detrimental effects on moult frequency. Negative effects (p<0.05) of high PA levels (i.e. 21.45–26.16 g PA kg-1), however, were found on the whole body P concentration. An inverse trend was recorded for the Ca content in the whole body (p<0.005) and carapace (p<0.004) with increasing PA inclusion. These results are consistent with findings for marine shrimp species such as Marsupenaeus japonicus and Litopenaeus vannamei. The graded inclusion of PA in the experimental diets also resulted in a significant reduced (p<0.03) P content in the carapace. Significant changes (p<0.05) were observed in the carapace Zn, Cu, K and Na compositions, particularly in the prawns fed the diet containing 11.28 g PA kg-1, which suggests that the specific minerals were either selectively utilised or retained in the carapace. Accordingly, this thesis investigates the potential of adding dietary supplements of microbial phytase in order to improve mineral availability as proven in several fish species. To explore this, four experimental diets were formulated - three incorporating different levels of microbial phytase (i.e. 0 FTU kg-1, 1000 FTU kg-1 and 2000 FTU kg-1) and a fourth, a control. An aliquot of 15 g PA kg-1 was also added to each treatment. The mineral premix was omitted from the test group diets except the control diet was identical to the 0 FTU kg-1 diet but included a mineral premix and an aliquot of 8 g kg-1 monosodium phosphate which replaced an equal amount of wheat meal fraction. The trial found no significant differences in growth, feed utilisation and moult frequency with the microbial phytase level within the diet, however, survival was compromised. The proximate composition of the prawn whole body was in most cases unaffected by the level of phytase. Supplementation of the diets with microbial phytase did, however, result in significant higher (p<0.05) concentrations of minerals including: 1) Ca, Mg, K and Na in the whole body; 2) Ca and Zn in muscle tissue; and, 3) Ca and Mg in the carapace. The Zn content of the carapace, however, was negatively affected by the inclusion of microbial phytase suggesting the necessity of this mineral within the diet of M. rosenbergii. This thesis contributes to current understanding surrounding the inclusion of dietary PA and the benefits of microbial phytase within the experimental diets consumed by juvenile M. rosenbergii. The knowledge gained from this work provides a means to optimise the use of plant protein ingredients and with the potential to decrease the dependability of fishmeal without compromising M. rosenbergii production and profitability, thus ultimately promoting the sustainable expansion of M. rosenbergii aquaculture.

This body of research explores the dietary application of a bioactive, solid-state fermentation (SSF) product in contemporary and lupin-containing diets of Nile tilapia (Oreochromis niltoticus) and rainbow trout (Oncorhynchus mykiss). Consequently, the work provides holistic assessment of the influences of SSF products on animal health and performance; depending upon rearing temperature, nutritional physiologies, feed formulations and extrusion conditions; alongside information on the performance of lupins in aquafeeds. The SSF product (at 0.1 % inclusion) improved growth performance of Nile tilapia fed diets containing lupins. Phosphorous retention appeared higher when the SSF product was included in a yellow lupin (Lupinus luteus) diet whilst Mg retention was significantly higher in fish fed narrow-leaf lupin (Lupinus angustifolius). The experimental ingredients did not appear to show any clear effects upon midgut macrostructure. At an ultrastructural level, the fish fed yellow lupin alone, displayed poorest brush border characteristics but those fed yellow lupin and the SSF product showed signs of amelioration since they did not differ significantly from those fed narrow-leaf lupin. Focus was then turned towards a yellow lupin-based diet in rainbow trout, with two inclusion levels of the SSF product. The SSF product significantly improved growth performance and feed efficiency at 0.5 %, with values closer to a fishmeal-based diet than the lupin control. The SSF product increased the digestibility of protein and energy and bioavailability of numerous elements. However, the digestibility and bioavailability of certain nutrients, e.g. fibre and Zn, were only increased with a 0.5 % inclusion. The SSF product influenced vertebral Ca:P ratio but no effect on vertebral morphology was identified. Fish fed yellow lupin kernel meal displayed high Mn concentrations throughout a number of tissues. The intestinal environment was explored in depth, revealing large differences dependent upon SSF product inclusion rate. Fish fed a 0.1 % inclusion exhibited deteriorated brush border characteristics and high diversity of microbes, including increased proportions of key salmonid pathogens. Those fed a 0.5 % inclusion displayed signs of increased surface area at an ultrastructural level, reduced goblet cell numbers and a low microbial diversity; with domination of one particular family, Enterobacteriaceae. Activities of alkaline phosphatase and leucine aminopeptidase within the anterior intestine also appeared to be influenced by SSF product inclusion. Variations in haemato-immunological parameters were also observed between the treatments. In the final experiment, the SSF product was applied, pre-extrusion, to a contemporary rainbow trout formulation, at 0, 0.5, 1.0 and 1.5 %. No significant improvements to performance were identified following SSF product inclusion. However, crude protein digestibility from SSF-supplemented diets was significantly higher than the control and tendencies towards elevated retention of Ca, P, and Mg were apparent. In vitro analysis of free-phosphate release was conducted upon pre- and post-extruded diets, at varying temperatures. This indicated that neither extrusion conditions (105 °C) nor an ambient temperature of 10 °C were sufficient to cease P-liberating capabilities by the SSF product; suggesting that the two conditions combined limited the product’s efficacy in practice. This research evidenced that SSF products can be an effective means of improving the available nutrient profiles of compound diets for both omnivorous, warmwater and carnivorous, temperate finfish. Lupins are a promising alternative protein source but their nutritional value can be substantially improved by SSF product application. Exploration of the effects of SSF products on the intestinal environment revealed that both negative and positive effects on intestinal health can occur, which is highly dependent upon product inclusion rate. The holistic approaches adopted within this series of studies have seldom been performed on monogastric animals and thus provide valuable, transferable information for advancing knowledge in the application of SSF products, exogenous enzymes and lupins to farm animal feeds, in general.

Atlantic salmon (Salmo salar L.) has been domesticated since the 1960s and has undergone over 10 generations of artificial selection for economically important traits. As a result, domesticated salmon have diverged with respect to a number of phenotypic, genotypic and behavioural traits from their wild counterparts. Since the selection pressures that are present in the wild differ greatly from the ones that shape salmon under culture conditions, domesticated salmon stocks are considered to be maladapted to natural conditions. Despite strict regulations, insoluble issues pertaining to large-scale cage rearing of farmed fish mean that there is a continuous presence of farm escapees in the wild. Gene flow from escapees has been perceived as a factor in the decline of wild populations, suggested to occur through disruption of local adaptation. This study aims to improve understanding of the genetic differences between wild and domesticated stocks by comparing the transcriptomes of Figgjo (wild) and Mowi (domesticated) strains. A series of common garden experiments have been performed, utilizing pure and reciprocal hybrid crosses of the wild and domesticated stocks, reared under two different conditions and sampled at four time points and three distinct life stages (embryo, sac-fry and feeding fry). Microarray interrogations were performed employing a 44K custom microarray design to identify genes and gene pathways that are differentially expressed between the stocks. KEGG-based functional analyses have been implemented using different gene set enrichment packages, and dominance and additive parameters were calculated from normalized expression values to predict the mode of heritability of the genes identified as differentially expressed between stocks. Most biological functions represented in wild and domesticated crosses were consistent across life stages and environments. The transcriptomic differences detected between stocks in multiple developmental stages likely reflected adaptations to selection pressures differing between natural and aquaculture environments. Down-regulated environmental information processing and immune and nervous system functions in domesticated vs. wild fish may be due to local adaptation to captivity. These included reduced information acquisition and processing systems, altered stress responsiveness and changes in feeding behaviour. In line with the resource allocation theory of production trait animals, reduced immune function was coupled with increased expression of growth and development related pathways in domesticated salmon, compared to wild counterparts. Although there is support for this trade-off in all life-stages, resource allocation showed a shift over time; possibly reflecting variation in the utilization of energy sources during the transition from endogenous to exogenous feeding. Differences in cell communication and signalling pathways between wild and domesticated stocks, associated with organogenesis during the embryo stage, reflect sampling time and are indicative of altered organ development in response to domestication. Stress responses common across stocks included the down-regulation of cellular processes, including cell cycle and meiosis, and genetic information processing, such as replication and repair, transcription and translation pathways, probably reflecting the reallocation of energy resources away from growth and towards the restoration of homeostasis. Moreover, the mobilization of energy to cover the increased demands of maintaining homeostasis was indicated by the up-regulation of some metabolic pathways, mostly involved in energy, lipid and carbohydrate metabolism in response to stress. The analysis also revealed cross-specific stress responses, including indicators of a non-additive stress response in hybrid crosses. Most differentially expressed transcripts exhibited additive (31-59%) or maternal dominant (19-33%) inheritance patterns, although maternal over-dominance (23-26%) was also significant in the embryo stage. The mode of heritability of some immune transcripts was suggestive of maternal environmental influence having been affected by aquaculture. This study has demonstrated that biological functions affected by domestication include those associated with allocation of resources, involve reduction of information acquisition and processing systems and may lead to loss of local adaptation to wild conditions. Since such changes may affect key systems, such as immunity and responsiveness to stress, they can potentially have serious negative consequences under natural conditions. Transcriptomic differences observed between wild and domesticated stocks primarily exhibited additive and maternal dominant inheritance modes. Since gene-flow from farmed fish can be frequent and primarily concerns farmed females, this suggests that introgression due to repeated large scale escape events has the capacity to significantly erode local adaptation.

The increasing incidence of infections due to antibiotic resistant bacteria has led to renewed interest in bacteriophages (= phages) and phage therapy. Although phage therapy has been applied to control bacterial diseases in plants, poultry, livestock and humans, its application in aquaculture is still relatively limited. The emergence of phage-resistant bacterial mutants has been considered to be one of the major limitations of phage therapy. This study aimed to (i) isolate and characterise phages; (ii) select phages and their bacterial hosts to set up in vivo phage therapy models with aquaculture animals, and estimate the efficiency of phage therapy; (iii) investigate the generation and characteristics of phage-resistant mutants, and thus estimate the consequence of applying phage therapy when phage-resistant mutants emerge; and (iv) discuss the prospects for application of phages in aquaculture. Two Vibrio isolates and their phages were isolated from a Scottish marine fish farm. Based on the results of conventional phenotype testing and 16S rRNA gene sequencing analysis, the two vibrios, V9 and V13, were identified as Vibrio splendidus and Vibrio cyclitrophicus, respectively. The bacterial characteristics including morphology, temperature and salinity range of growth, production of extracellular enzymes, and the possession of virulence genes were examined. According to the morphological characteristics observed using transmission electron microscopy by negative staining, phage PVS9 of V. splendidus V9 was identified as a myophage, while phage PVC13 of V. cyclitrophicus V13 was identified as a siphophage. The phages could only lyse one bacterial host strain and their genomic DNA was double stranded with a size of ~46 kb. The two Vibrio isolates were found to be non- or of low virulence to rainbow trout, goldsinny wrasse and Artemia in pathogenicity experiments. Thus an in vivo phage therapy model could not be set up using these Vibrio isolates and their phages. Two phages pAS-3 and pAS-6 were isolated using the Aeromonas salmonicida subsp. salmonicida Hooke strain as the host. Phages pAS-3 and pAS-6 had a similar genome size of ~50 kb, and the same relatively narrow host range within A. salmonicida subsp. salmonicida strains. The siphophage pAS-3 formed clear plaques and inhibited A. salmonicida Hooke growth in vitro completely for at least 18 hours when using MOI = 1,000, whereas the podophage pAS-6 formed turbid plaques and weakly inhibited Hooke growth. Rainbow trout exposed by intraperitoneal injection with 0.1 mL of the raw phage preparations at a concentration of 108 PUF mL-1 showed no adverse effects over 14 days. In the phage therapy trial, fish were firstly injected with 1 x 102 CFU fish-1 of A. salmonicida Hooke, then immediately injected with phage preparations of pAS-3 and pAS-6, respectively, using MOI = 10,000. Compared with the control group (which did not receive phage treatment), phage treated groups showed a delay in the time to death, and lower mortalities. However, the mortalities and time to death between phage treated and non-treated groups were not significantly different. Phage-resistant mutants of pathogenic A. salmonicida strain Hooke were induced by repeatedly challenging with phage pAS-3. One of the mutants, termed HM, was chosen to compare the characteristics with the parental wild-type strain Hooke. Test results including the formation of ‘smooth’ colonies on TSA, autoagglutination negative, the formation of creamy colonies on Coomassie Brilliant Blue agar, and the degradation of a thick/furry layered structure on the cell surface indicated a deficiency of the A-layer in the phage-resistant mutant HM. Therefore, it was deduced that the A-layer either directly acted as the receptor of A. salmonicida phage pAS-3, or was affected indirectly by the change of an unknown phage receptor. The greater wax moth larvae model was used to compare the virulence of the phage-resistant mutant HM and the parental wild-type strain Hooke, as it is an ethically acceptable animal model, which has the advantages of being low cost and convenient for injection, and is also a recognised alternative model for bacterial pathogens of fish. The results showed that virulence of the phage-resistant mutant HM did not decline in the greater wax moth larvae model compared with that of the parental wild-type strain Hooke. In conclusion, different approaches were used to isolate and characterise phages from different aquaculture environments for potential use in phage therapy. A rainbow trout model was set up using pathogenic A. salmonicida strain Hooke and two A. salmonicida phages pAS-3 and pAS-6. The use of phage treatment led to lower cumulative mortalities and delay to the time of death, although the differences between the groups were not significant, futher work is required to determine if these phages have potential in phage therapy. The consequence of applying phage therapy when phage-resistant mutants emerge was estimated based on their characteristics and virulence, and no decline in virulence of the phage-resistant mutant from this study indicates the importance of fully testing the virulence of phage-resistant mutants before carrying out large scale field trials of phage therapy. It appears feasible to use phage therapy as an alternative approach to control bacterial infections in aquaculture, but further studies are required to focus on improving effectiveness, and also to overcome the concrete limitations and hurdles in application and commercialisation. Moreover, a broader range of applications of phages in aquaculture should be explored.

Dans le contexte du développement spectaculaire de l’aquaculture, les farines et huiles de poissons, qui étaient les composantes principales des aliments aquacoles, ont progressivement été remplacées par des mélanges de produits végétaux. Cependant, le remplacement total n’est pas encore possible sans dégrader les performances des poissons (croissance et survie). Pour continuer son développement durable, l’aquaculture doit davantage s'affranchir de ces matières premières. Une voie de progrès possible est l'utilisation de la variabilité génétique existante dans les populations de truites pour la capacité à survivre et grandir avec des aliments à base de végétaux (Aliments V). Cependant très peu d'études se sont intéressées aux mécanismes mis en place par certains génotypes pour utiliser plus efficacement ces aliments. L'objectif de la thèse était d'étudier les caractéristiques à l'origine d'une meilleure utilisation des aliments V chez différents génotypes de truite arc-en-ciel (trois lignées isogéniques et lignée sélectionnée). L'originalité de ce travail repose sur l'étude de la réponse de différentes lignées à un aliment 100 % V, en combinant des approches de physiologie nutritionnelle et de biologie moléculaire sans a priori ou ciblées. Ainsi les transcriptomes de trois lignées isogéniques, ayant des performances contrastées lorsqu'elles sont nourries avec des aliments V, ont été comparés à court et long terme. Puis des caractères liés à la nutrition, ainsi que l'expression de certains gènes, identifiés grâce à l'étude transcriptomique, ont été étudiés chez une lignée sélectionnée pour sa capacité à survivre et grandir avec un aliment V. Les résultats obtenus ont permis de conclure que l'acceptation de l'aliment V joue un rôle clef, notamment lors des stades précoces. Les résultats obtenus mettent également en avant le rôle prépondérant de la composante immunitaire qui est impactée différemment en fonction des génotypes. Finalement, après 6 mois d'alimentation, alors que le métabolisme des lipides participe à expliquer les différences d’utilisation de l’aliment V chez les lignées isogéniques, il ne parait pas déterminant chez la lignée sélectionnée. Globalement, nos résultats suggèrent l'existence d'une grande diversité de mécanismes dans les populations de truites leur permettant de survivre et grandir avec un aliment à base de végétaux dépourvu de farine et huiles de poisson. / In the context of the exponential development of aquaculture, fish meal and oils, previously the main components of aquaculture feed, have been increasingly replaced by a blend of plant products. However, total replacement is not yet possible without impairing fish performance (survival and growth). To continue its sustainable development, aquaculture must further decrease its reliance on both fish oil and fish meal. In addition to the changes in diet composition, a possible way forward is the use of the genetic variability for the ability to survive and grow with plant-based diets (V diets) existing in trout populations. However, very few studies have investigated the mechanisms implemented by some genotypes to use these diets more efficiently. The objective of this thesis was to study the characteristics that led to a better utilisation of V diet in different rainbow trout genotypes. The originality of this PhD work relies on the genetic rainbow trout lines whose responses to a 100% V diet were studied by combining nutritional physiology and molecular biology with or without a priori approaches. Thus, the transcriptomes of three isogenic lines, which have contrasting performances when fed V diets, were compared at short and long term. Then nutrition-related traits, as well as the expression of some genes identified through the transcriptomics analysis, were studied in a line selected for its ability to survive and grow with a V diet. The results obtained led to the conclusion that the acceptance of V diets plays a key role, particularly in the early stages. The results also highlight the essential role of the immune system, which is impacted differently according to genotypes. Finally, after 6 months of feeding, while lipid metabolism seems to play a role in the differences between genotypes for their ability to efficiently utilize the V diet, this pathway does not appear to play a determinant role in the selected line. Overall, our results suggest that there are a wide variety of mechanisms allowing trout to survive and grow with a total plant-based diet, devoid of both fish meal and oils.