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The impact of nutritional and environmental stressors on the immune response, oxidative stress and energy use of rainbow trout (Oncorhynchus mykiss).

Leonardo Magnoni1, 2*, Sara C. Novais3, Carla O. Silva3, Marco F. Lemos3, Rodrigo Ozorio1, 4, Inge Geurden5, Isabelle Leguen6, Patrick Prunet6, Ep Eding7 and Johan Schrama7
  • 1 CIIMAR, University of Porto, Portugal
  • 2 IIB-INTECH, Argentina
  • 3 MARE – Marine and Environmental Sciences Centre, ESTM, Instituto Politécnico de Leiria, Portugal
  • 4 ICBAS, University of Porto, Portugal
  • 5 NuMeA-INRA, France
  • 6 LPGP-INRA, Université Rennes, France
  • 7 WIAS, Wageningen University, Netherlands

Background: In aquaculture, raising fish in sub-optimal conditions may involve simultaneous exposure to multiple stressors, hampering health and growth. These stressors may act over a long period (chronic) or can be circumscribed to a short event (acute). To get a better picture of the limits between adaptation or stress responses, as well as the interaction between chronic and acute stressors, an approach using a multiple-parameter study is necessary. We have investigated the response of an isogenic heterozygous family of rainbow trout (Oncorhynchus mykiss) to a combination of nutritional and environmental challenges. We used an isogenic trout line (produced by GABI/La Peima, INRA, France) as experimental fish because they are genetically uniform, providing the best experimental conditions for measuring the genotype-environment interaction, allowing for the reproducibility of experimental outcomes. For this purpose, trout were fed for 7 weeks two diets differing in their cation-anion difference (CAD) and maintained at two different water dissolved oxygen (DO) levels (Normoxia or Hypoxia). At the end of the trial, trout were exposed to a standard acute confinement stress. We assessed a possible interactive effect between dietary CAD levels, DO levels and acute stress on several parameters indicating how energy allocation, oxidative status and health are modulated by concurrent conditions of stress. It is expected that in trout exposed to chronic stressors, such as high dietary CAD (acid-base imbalance) and low DO levels (limited energy use), the effects of acute stress will be enhanced. Materials and Methods: Rainbow trout approximately 115 g in weight (n=30) were randomly assigned to one of twelve tanks (200L). All tanks were connected to the same recirculating aquaculture system (RAS) and had identical inlet water quality. Water temperature was set at 14 ± 1°C. Photoperiod was maintained at 12: 12 (Light: Dark). Experimental tanks were distributed according to a 2 by 2 factorial design in triplicate tanks per experimental condition. One factor was CAD dietary content (200 or 700 mEqv/kg) and the other DO level (7 mg/L, Normoxia and 3.7 mg/L, Hypoxia). At sampling, fish from each experimental group were divided into Not Stressed (NS) or Stressed (S). In the stressed group, fish were netted and exposed to a standard acute confinement stress (2 minutes, density 200kg/m3). After the confinement, stressed fish were transferred into the original empty tank and sampled after 1 h. In all the sampling procedures, fish were netted and sedated/euthanized as quick and smoothly as possible. Plasma, liver and heart samples were obtained and frozen at -80oC for later analysis. Several parameters related to immune response were determined in plasma (alternative complement pathway (ACH50), lysozyme, and peroxidase). Enzyme activities as markers of oxidative stress were analysed in liver (lipid peroxidation, glutathione peroxidase (GPx), glutathione reductase (GR), total glutathione (TG), oxidized glutathione (GSSG), reduced glutathione (GSH)). Parameters related to energy metabolism were quantified in heart (cellular energy allocation (CEA), electron transport system (ETS) activity as measure of energy consumption, lactate dehydrogenase (LDH), and iso-citrate dehydrogenase (IDH)). Results and Discussion: Regarding the effects of diet (DI), DO level, acute stress (S) and their interactions on innate immune system, a significant effect of the diet was observed for ACH50 (P=0.001), but no interaction was observed between other stress factors. This difference was evident in trout subjected to hypoxia, where ACH50 activity was significantly higher in plasma of fish fed CAD 200 than in trout fed a CAD 700 diet when subjected to the acute stressor. Lysozyme activity in plasma of trout was differentially altered by acute stress condition (P<0.05), but no significant interactions were found with chronic stressors. Concerning markers of oxidative stress in liver of trout, this study showed that GPx activity was significantly affected by diet (P<0.034), displaying interactions with other stressors (DI/DO, DI/S, DI/DO/S). On the other hand, hepatic GR activity was significantly affected by diet (P<0.044), but no interaction was detected with the other factors studied. Additional markers analysed in relation to the glutathione system, indicate that diet (GSSH and TG, P<0.035 and <0.03, respectively) and acute stress (GSH, P<0.019) are the main factors affecting oxidative stress in this tissue. However, no significant differences in lipid peroxidation levels were detected between treatments, indicating that although diet (chronic) and acute stress affect redox balance, such changes occur without detectable oxidative damage. Energy metabolism markers analysed in trout´s heart showed a significant effect of acute stress in IDH activity (P<0.026), which activity was inhibited, indicating a less efficient generation of energy by aerobic metabolism in this group. An interaction between DO levels and acute stress was also detected for this enzyme activity as well as for LDH. Furthermore, the ratio LDH/IDH was significantly affected by diet (P<0.006), with an interaction with acute stress. Also, the ETS and CEA values in heart of trout showed interactions between DI/DO and DI/DO/S. In particular, total glycogen content in heart was modulated by the diet or acute stress (P<0.001 for both factors). All the parameters analysed in heart suggest that diet and acute stress are the main factors modulating energy use by the heart, although these factors could be altered by chronic hypoxic conditions to some extent. Conclusion: In general, we observed that the nutritional chronic stressor used in this study (CAD 700) was the main factor affecting innate immune response, oxidative stress responses and energy metabolism in trout, with a significant interaction with the acute stressor (netting and confinement). In the heart, this study showed that acute stress has a strong effect on how this organ alters the pattern of energy allocation and use in trout. Chronic hypoxia, below the incipient DO reported for trout (6 mg/L, Pedersen 1987), did not produce significant changes in most of the parameters studied, although it displayed interactive effects with either the diet or acute stressor.

Acknowledgements

This communication is part of a study funded within the EU-FP7 project AQUAEXCEL (262336). Additional funding was provided by Fundação para a Ciência e a Tecnologia (FCT) trough the Strategic Project UID/MAR/04292/2013 and the grants to Leonardo Magnoni (IF/01314/2014) and Sara C. Novais (SFRH/BPD/94500/2013). Leonargo Magnoni also wish to acknowledge WIAS Wageningen University for the research fellowship.

References

Pedersen CL (1987) Energy budgets for juvenile rainbow trout at various oxygen concentrations. Aquaculture, 62: 289-298. doi:10.1016/0044-8486(87)90171-2.

Keywords: Aquaculture, fish, Diet, Cation-anion difference, hypoxia, Oxidative Stress, immune response, Energy Metabolism

Conference: IMMR | International Meeting on Marine Research 2016, Peniche, Portugal, 14 Jul - 15 Jul, 2016.

Presentation Type: Poster presentation

Topic: Aquaculture

Citation: Magnoni L, Novais SC, Silva CO, Lemos MF, Ozorio R, Geurden I, Leguen I, Prunet P, Eding E and Schrama J (2016). The impact of nutritional and environmental stressors on the immune response, oxidative stress and energy use of rainbow trout (Oncorhynchus mykiss).. Front. Mar. Sci. Conference Abstract: IMMR | International Meeting on Marine Research 2016. doi: 10.3389/conf.FMARS.2016.04.00008

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Received: 14 May 2016; Published Online: 12 Jul 2016.

* Correspondence: Dr. Leonardo Magnoni, CIIMAR, University of Porto, Porto, Portugal, leonardo.magnoni@plantandfood.co.nz