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The skin barrier consists of mucus, primarily comprising highly glycosylated mucins, and the epithelium. Host mucin glycosylation governs interactions with pathogens and stress is associated with impaired epithelial barrier function. We characterized Atlantic salmon skin barrier function during chronic stress (high density) and mucin O -glycosylation changes in response to acute and chronic stress. Fish held at low (LD: 14-30 kg/m 3 ) and high densities (HD: 50-80 kg/m 3 ) were subjected to acute stress 24 h before sampling at 17 and 21 weeks after start of the experiment. Blood parameters indicated primary and secondary stress responses at both sampling points. At the second sampling, skin barrier function towards molecules was reduced in the HD compared to the LD group (P app mannitol; p < 0.01). Liquid chromatography-mass spectrometry revealed 81 O -glycan structures from the skin. Fish subjected to both chronic and acute stress had an increased proportion of large O -glycan structures. Overall, four of the O -glycan changes have potential as indicators of stress, especially for the combined chronic and acute stress. Stress thus impairs skin barrier function and induces glycosylation changes, which have potential to both affect interactions with pathogens and serve as stress indicators.

We have developed a mathematical model which estimates the growth performance of Atlantic salmon in aquaculture production units. The model consists of sub-models estimating the behaviour and energetics of the fish, the distribution of feed pellets, and the abiotic conditions in the water column. A field experiment where three full-scale cages stocked with 120,000 salmon each (initial mean weight 72.1  ± SD 2.8 g) were monitored over six months was used to validate the model. The model was set up to simulate fish growth for all the three cages using the feeding regimes and observed environmental data as input, and simulation results were compared with the experimental data. Experimental fish achieved end weights of 878, 849 and 739 g in the three cages respectively. However, the fish contracted Pancreas Disease (PD) midway through the experiment, a factor which is expected to impair growth and increase mortality rate. The model was found able to predict growth rates for the initial period when the fish appeared to be healthy. Since the effects of PD on fish performance are not modelled, growth rates were overestimated during the most severe disease period. This work illustrates how models can be powerful tools for predicting the performance of salmon in commercial production, and also imply their potential for predicting differences between commercial scale and smaller experimental scales. Furthermore, such models could be tools for early detection of disease outbreaks, as seen in the deviations between model and observations caused by the PD outbreak. A model could potentially also give indications on how the growth performance of the fish will suffer during such outbreaks., Statement of Relevance: We believe that our manuscript is relevant for the aquaculture industry as it examines the growth performance of salmon in a fish farm in detail at a scale, both in terms of number of fish and in terms of duration, that is higher than usual for such studies. In addition, the fish contracted a disease (PD) midway through the experiment, thus resulting in a detailed dataset containing information on how PD affects salmon growth, which can serve as a foundation to understanding disease effects better. Furthermore, the manuscript describes an integrated mathematical model that is able to predict fish behaviour, growth and energetics of salmon in response to commercial production conditions, including a dynamic model of the distribution of feed pellets in the production volume. To our knowledge, there exist no models aspiring to estimate such a broad spectre of the dynamics in commercial aquaculture production cages. We believe this model could serve as a future tool to predict the dynamics in commercial aquaculture net pens, and that it could represent a building block that can be utilised in a future development of knowledge-driven decision-support tools for the salmon industry.

The objective of this study was to determine the underlying physiological and molecular responses to long-term sublethal ammonia exposure in Atlantic salmon (Salmo salar) parr. Previous studies have predominately focused on mechanisms during acute, short-term exposure. For that purpose Atlantic salmon parr were exposed to four ammonia concentrations between 4 and 1800 μmol l(-1) total ammonia nitrogen (TAN), and subjected to two feeding regimes for 15 weeks. Elevated environmental ammonia and full feeding strength caused an initial increase in plasma ammonia levels ([T(amm)]) after 22 days of exposure, which thereafter declined and remained similar to the control animals towards the end of the study. On the other hand, a progressive decrease in plasma urea levels was evident throughout the entire exposure period and depended on the concentration of environmental ammonia, with the largest decrease in urea levels observed at the highest ammonia concentrations (1700 and 1800 μmol l(-1) TAN). We hypothesized that the successful adaptation to long-term elevated ammonia levels would involve an increased capacity for carrier-facilitated branchial excretion. This hypothesis was strengthened by the first evidence of an up-regulation of branchial transcription of the genes encoding the Rhesus (Rh) glycoproteins, Rhcg1 and Rhcg2, urea transporter (UT) and aquaporin 3a (Aqp3a), during long-term exposure. Of the Rhesus glycoprotein (Rh) mRNAs, Rhcg1 was up-regulated at all tested ammonia levels, while Rhcg2 showed a concentration-sensitive increase. Increased transcription levels of V-type H(+)-ATPase (H(+)-ATPase) were observed at the highest ammonia concentrations (1700 and 1800 μmol l(-1) TAN) and coincided with an up-regulation of Rhcg2 at these concentrations. Transcription of UT and Aqp3a was increased after 15 weeks of exposure to low ammonia levels (470 and 480 μmol l(-1) TAN). A significant increase in brain glutamine (Gln) concentration was observed for full fed Atlantic salmon after 22 days and in fish with restricted feeding after 105 days of exposure to 1800 and 1700 μmol l(-1) TAN, respectively, without any concomitant decrease in brain glutamate (Glu) concentrations. These results suggest that Gln synthesis is an ammonia detoxifying strategy employed in the brain of Atlantic salmon parr during long-term sublethal ammonia exposure. Full feed strength had an additive effect on plasma [T(amm)], while the restricted feeding regime postponed the majority of the observed physiological and molecular responses. In conclusion, Atlantic salmon parr adapts to the long-term sublethal ammonia concentrations with increased branchial transcription levels of ammonia and urea transporting proteins and ammonia detoxification in the brain., (Copyright © 2012 Elsevier B.V. All rights reserved.)

We have previously documented increased survival by feeding tetradecylthioacetic acid (TTA) during a natural outbreak of infectious pancreatic necrosis in post-smolt S1 Atlantic salmon. The aim of the present study was to test the effects of dietary TTA in S0 smolt at a location where fish often experience natural outbreaks of heart and skeletal muscle inflammation (HSMI) during their first spring at sea. The experimental groups were fed a diet supplemented with 0.25% TTA for a 6-week period prior to a natural outbreak of HSMI in May 2007. Relative percent survival for the groups fed TTA was 45% compared with control diets, reducing mortality from 4.7% to 2.5%. Expression of genes related to lipid oxidation was higher in cardiac ventricles from salmon fed TTA compared with controls. In addition, salmon fed TTA had periodically reduced levels of plasma urea, and increased cardiosomatic index and growth. Reduced mortality and increased growth after administration of TTA may be related to a combination of anti-inflammatory effects, and an altered metabolic balance with better protein conservation because of increased lipid degradation.

The peroxisomal enzyme urate oxidase plays a pivotal role in the degradation of purines in both prokaryotes and eukaryotes. However, knowledge about the purine-induced expression of the encoding gene is lacking in vertebrates. These are the first published sequences of fish urate oxidase, which were predicted from PCR amplified liver cDNAs of Atlantic salmon (Salmo salar), Atlantic cod (Gadus morhua), Atlantic halibut (Hippoglossus hippoglossus) and African lungfish (Protopterus annectens). Sequence alignment of different vertebrate urate oxidases revealed amino acid substitutions of putative functional importance in the enzyme of chicken and lungfish. In the adult salmon, expression of urate oxidase mRNA predominated in liver, but was also identified in several nonhepatic organs including brain, but not in skeletal muscle and kidney. Juvenile salmon fed diets containing bacterial protein meal (BPM) rich in nucleic acids showed a significant increase in liver urate oxidase enzyme activity, and urea concentrations in plasma, muscle and liver were elevated. Whereas salmon fed the 18% BPM diet showed a nonsignificant increase in liver mRNA levels of urate oxidase compared with the 0% BPM-fed fish, no further increase in mRNA levels was found in fish receiving 36% BPM. The discrepancy between urate oxidase mRNA and enzyme activity was explained by rapid mRNA degradation or alternatively, post-translational control of the activity. Although variable plasma and liver levels of urate were detected, the substrate increased only slightly in 36% BPM-fed fish, indicating that the uricolytic pathway of Atlantic salmon is intimately regulated to handle high dietary purine levels.

The premise that a dietary dipeptide approach will improve the understanding of amino acid utilization in the fastest-growing vertebrate, the teleost fish, was tested by examining the muscle free amino acid (FAA) pool and enzyme activities, in concert with growth response, when dietary amino acids were provided in free, dipeptide or protein molecular forms. We present the first evidence in fish that, in response to a synthetic dipeptide diet, muscle FAA varies as a result of both growth rate and amino acid availability of specific peptides. We demonstrate significantly diminished muscle indispensable FAA (3-10-fold) in rainbow trout alevins fed a dipeptide-based diet compared with a protein-based diet. The dipeptide-based diet did not contain proline, resulting in 10-27-fold less muscle free proline and hydroxyproline in alevins. The response of alevins fed FAA-based or peptide-based diets can be indicative of collagen turnover (Hyp/Pro ratio) and showed significant differences between dietary treatments. Pyrroline-5-carboxylate (P5C) reductase activity was detected, suggesting that P5C may ameliorate proline deficiency, but synthesis from glutamate could not maintain free proline levels in muscle. This finding will provide an impetus to test whether proline is conditionally indispensable in young fish, as in mammals and birds. This study shows that amino acids given entirely as dipeptides can sustain fish growth, result in muscle FAA and enzyme responses in line with dietary levels and identify growth-limiting amino acids. The understanding of these factors necessitates a diet formulation that will improve the accuracy of determining amino acid requirements in the early life stages of vertebrates.

Most studies on the antioxidants, lipoic acid (LA) and ascorbic acid (AA), focused on species that, unlike teleost fish, are not scurvy-prone, and are able to synthesize AA. The antioxidant properties of LA may make it useful in aquaculture nutrition, but several effects must first be investigated, and we address here plasma free amino acids (FAA). In mammals, LA and AA in high doses were claimed to alter plasma FAA profile; to our knowledge, however, no data are available in fish. We therefore studied the effects of dietary LA and AA on plasma FAA in the South American teleost fish pacu, which is being used increasingly in aquaculture. LA treatment decreased concentrations of 18 of 23 individual FAA; specifically, dispensable and total FAA were significantly affected. Ornithine was elevated (+26%) in LA-treated fish and significantly decreased ratios of plasma [Arg]/[Orn] and other individual [FAA]/[Orn] were observed. LA and AA both affected sulfur FAA concentrations. Plasma cystine levels were significantly increased in the LA-supplemented groups. AA had little effect on most amino acids, and no interaction with LA was detected. AA supplementation did, however, significantly lower taurine (-42%) and cystathionine (-31%) levels in plasma. No effect on the branched chain:aromatic amino acid ratios was observed. The data indicate that at the dietary level studied, LA and AA independently affect selected plasma FAA in pacu, and suggest that any use of LA in particular as a dietary supplement should take into account an altered plasma FAA profile.

Considering that amino acids constitute an important energy fuel during early life of the Atlantic halibut (Hippoglossus hippoglossus L.), it is of interest to understand how the nitrogenous end products are handled. In this study we focused on the kinetics and fates of ammonia, urea and uric acid. The results showed that ammonia (T(Amm): NH(3)+NH(4)(+)), and urea-N contents increased during final oocyte maturation. Urea-N excretion dominated the total nitrogenous end product formation in early embryos. Later, yolk T(Amm) levels increased in embryos and ammonia excretion was low. In the last part of the embryonic stage T(Amm) accumulation dominated, and was apparently due to yolk storage. Around hatching, the larval body tissues (larva with yolk-sac removed) accounted for 68% of whole animal urea-N accumulation, while T(Amm) levels increased predominately by yolk accumulation. Afterwards, ammonia excretion dominated and uric acid accumulation accounted for less than 1%. Urea, synthesised either through the ornithine-urea cycle, argininolysis or uricolysis, accounted for approximately 8% of total nitrogenous end product formation in yolk-sac larvae. The results suggested that a sequence occurred regarding which nitrogenous end products dominated and how they were handled. Urea excretion dominated in early embryos (<7 dPF), followed by yolk ammonia accumulation (7-12 dPF), and finally, ammonia excretion dominated in later embryonic and yolk-sac larval stages (>12 dPF).

Embryos and larvae of the African catfish Clarias gariepinus excrete significant quantities of urea. The present study focused on the potential urea-generating pathways during early development of this teleost; uricolysis, argininolysis and the ornithine-urea cycle (OUC). Uricase, allantoinase, allantoicase and ureidoglycollate lyase of the uricolytic pathway were expressed in all early life stages and in adult liver of C. gariepinus. Uricase activity increased in starved larvae compared with yolk-sac larvae. The key regulatory enzyme of the teleost OUC, carbamoyl phosphate synthetase III (CPSase III), was expressed predominantly in muscle of developing C. gariepinus larvae and showed negligible activity in the absence of its allosteric effector N-acetyl-L-glutamate. CPSase III and ornithine carbamoyl transferase activities increased in fed larvae compared with starved larvae. In contrast to the early developmental stages, adult C. gariepinus expressed only low and variable levels of CPSase III, suggesting that, under the experimental conditions employed, OUC expression is influenced by developmental stage in this species. The data indicate that early C. gariepinus life stages express the enzymes necessary for urea production by uricolysis, argininolysis and the OUC, and this may explain why urea tissue levels and urea excretion rates are substantial during the early development of this air-breathing teleost.

The presence of carbamoyl phosphate synthetase III (CPSase III), catalyzing the first step of the urea cycle in fish, in Atlantic halibut (Hippoglossus hippoglossus L.) yolk-sac larvae and adult white muscle has been established using gel filtration chromatography to separate the CPSase III from the pyrimidine-pathway related CPSase II. The results are consistent with the hypothesis that teleostean fish express urea cycle enzymes during early development and with recent observations of low levels of CPSase III in muscle tissue. The presence of CPSase III in crude extracts could not be established using sensitive assay conditions to discriminate between CPSase III and CPSase II. However, kinetic characterization after chromatographic separation identified each as typical CPSase II and CPSase III activities, respectively. The CPSase III was less sensitive to activation by N-acetyl-L-glutamate and had a higher Km for ammonia than CPSase III found in other species. These results suggest that precise quantitation of low levels of CPSase III in the presence of CPSase II by assaying crude extracts may be difficult unless the enzymes are first separated and the kinetic properties of CPSase III are determined; the results indicate that assaying larval extracts of Atlantic halibut in the presence of uridine triphosphate results in CPSase activity that reflects mostly CPSase III and can, therefore, be used to measure changes in CPSase III activity.