Comparison of Circulating Markers and Mucosal Immune Parameters from Skin and Distal Intestine of Atlantic Salmon in Two Models of Acute Stress.

Ensuring salmon health and welfare is crucial to maximize production in recirculation aquaculture systems. Healthy and robust mucosal surfaces of the skin and intestine are essential to achieve this goal because they are the first immunological defenses and are constantly exposed to multistressor conditions, such as infectious diseases, suboptimal nutrition, and environmental and handling stress. In this work, Atlantic salmon, split from a single cohort, were subjected to acute hypoxia stress or 15-min crowding stress and observed over a 24-h recovery period. Samples were collected from fish at 0, 1, 3, 6, 12 and 24 h post-stress to analyze plasma-circulating markers of endocrine function (cortisol), oxidative stress (glutathione peroxidase) and immune function (interleukin 10 (IL-10), annexin A1). In addition, mucosal barrier function parameters were measured in the skin mucus (Muc-like protein and lysozyme) and distal intestine (simple folds, goblet cell size and goblet cell area). The results showed that both acute stress models induced increases of circulating cortisol in plasma (1 h post-stress), which then returned to baseline values (initial control) at 24 h post-stress. Moreover, the hypoxia stress was mostly related to increased oxidative stress and IL-10 production, whereas the crowding stress was associated with a higher production of Muc-like protein and lysozyme in the skin mucus. Interestingly, in the distal intestine, smaller goblet cells were detected immediately and one hour after post-hypoxia stress, which could be related to rapid release of the cellular content to protect this organ. Finally, the correlation of different markers in the hypoxic stress model showed that the circulating levels of cortisol and IL-10 were directly proportional, while the availability of Muc-like proteins was inversely proportional to the size of the goblet cells. On the other hand, in the crowding stress model, a proportional relationship was established between plasma cortisol levels and skin mucus lysozyme. Our results suggest key differences in energy partitioning between the two acute stress models and support the need for further investigation into the interplay of multistressor conditions and strategies to modulate immunological aspects of mucosal surfaces.

Dietary soyasaponin supplementation to pea protein concentrate reveals nutrigenomic interactions underlying enteropathy in Atlantic salmon (Salmo salar).

BACKGROUND: Use of plant ingredients in aquaculture feeds is impeded by high contents of antinutritional factors such as saponins, which may cause various pharmacological and biological effects. In this study, transcriptome changes were analyzed using a 21 k oligonucleotide microarray and qPCR in the distal intestine of Atlantic salmon fed diets based on five plant protein sources combined with soybean saponins. RESULTS: Diets with corn gluten, sunflower, rapeseed or horsebean produced minor effects while the combination of saponins with pea protein concentrate caused enteritis and major transcriptome changes. Acute inflammation was characterised by up-regulation of cytokines, NFkB and TNFalpha related genes and regulators of T-cell function, while the IFN-axis was suppressed. Induction of lectins, complement, metalloproteinases and the respiratory burst complex parallelled a down-regulation of genes for free radical scavengers and iron binding proteins. Marked down-regulation of xenobiotic metabolism was also observed, possibly increasing vulnerability of the intestinal tissue. A hallmark of metabolic changes was dramatic down-regulation of lipid, bile and steroid metabolism. Impairment of digestion was further suggested by expression changes of nutrient transporters and regulators of water balance (e.g. aquaporin, guanylin). On the other hand, microarray profiling revealed activation of multiple mucosal defence processes. Annexin-1, with important anti-inflammatory and gastroprotective properties, was markedly up-regulated. Furthermore, augmented synthesis of polyamines needed for cellular proliferation (up-regulation of arginase and ornithine decarboxylase) and increased mucus production (down-regulation of glycan turnover and goblet cell hyperplasia) could participate in mucosal healing and restoration of normal tissue function. CONCLUSION: The current study promoted understanding of salmon intestinal pathology and establishment of a model for feed induced enteritis. Multiple gene expression profiling further characterised the inflammation and described the intestinal pathology at the molecular level.

Dietary Inclusion of Hydrolyzed Debaryomyces hansenii Yeasts Modulates Physiological Responses in Plasma and Immune Organs of Atlantic Salmon (Salmo salar) Parr Exposed to Acute Hypoxia Stress.

Stress related to salmon aquaculture practices (handling, sub-optimal nutrition, diseases, and environmental problems) may compromise fish welfare. This study describes the effects of two hydrolyzed Debaryomyces hansenii yeast-based products (LAN4 and LAN6) on physiological and immune responses of Atlantic salmon (Salmo salar) parr exposed to short hypoxia stress. A commercial-like diet (control diet: CD) and two experimental diets (CD supplemented with 0.1% of either component LAN4 or LAN6) were fed to fish for 8 weeks. At the end of the feeding experiment, fish were exposed to 1-min hypoxia and samples were collected at 0, 1, 3, 6, 12, and 24 h post-stress. Results showed that plasma cortisol reached a peak at 1 h post-stress in CD and LAN6 groups, whereas no significant increase in cortisol levels was detected in the LAN4 group. Moreover, the LAN6 group enhanced IL-10 responses to hypoxia, when compared to the control and LAN4 group. This suggests a regulation of immunosuppressive profiles in fish fed LAN4. Hypoxia stress increased TNFα in all groups, which indicates that fish may compensate for the short-term stress response, by modulating innate immune molecules. The apparent suppression of hypoxia responses in the LAN4 group coincided with the detection of differences in goblet cells size and Muc-like proteins production in DI; and upregulation (1 h post-stress) of pathways related to oxygen transport, hemoglobin complex, and glutathione transferase activity and the downregulation of fatty acid metabolism (6 h post-stress) in gills. To conclude, a 1-min hypoxia stress exposure affects the response to stress and immunity; and D. hansenii-based yeast products are promising components in functional aquafeeds for salmon due to their ability to counteract possible consequences of hypoxic stress.

Modulation of splenic immune responses to bacterial lipopolysaccharide in rainbow trout (Oncorhynchus mykiss) fed lentinan, a beta-glucan from mushroom Lentinula edodes.

Immunostimulants (IS) are considered a promising approach for improving resistance to pathogens in fish aquaculture. At present, development of IS are complicated due to limited knowledge on the mechanisms of their action. To assess the use of global gene expression analysis for screening of candidate IS we applied lentinan, a beta-glucan from the mushroom Lentinula edodes, as a model. After feeding rainbow trout (Oncorhynchus mykiss) with lentinan-supplemented (L) and control (C) diets for 37 days, fish were injected with bacterial lipopolysaccharide (LPS), a classical inducer of inflammation. Gene expression was analyzed in LPS-challenged compared to saline-injected fish using a salmonid 1.8k cDNA microarray (SFA2.0 immunochip) and real-time qPCR. Spleen was selected for data analyses due to highest magnitude of responses and its key role in the fish immune system. A group of genes implicated in acute inflammatory responses was higher induced in C versus L, including IFN-related and TNF-dependent genes (galectins and receptors, signal transducers and transcription factors), genes involved in MHC class I antigen presentation and leukocyte recruitment. A similar trend was observed in metabolism of iron and xenobiotics, markers of oxidative and cellular stress. Interestingly, differences between C and L were similar to those observed between salmon with low and high resistance to infectious salmon anemia virus. Genes with equal responses to LPS in L and C were related to cell communication (cytokines, chemokines and receptors), signal transduction, activation of immune cells, apoptosis, cellular maintenance and energy metabolism. In conclusion, lentinan decreased the expression of genes involved in acute inflammatory reactions to the inflammatory agent while major parts of the immune response remained unchanged. Such effects are expected for IS, which should modify immunity by enhancing beneficial and reducing detrimental responses.