Abstract

Methionine presents a pivotal role in the regulation of many cellular events with crucial impact on the immune system, such as in processes involved in the control of inflammation and polyamines synthesis. Accordingly, the present study aimed to assess the modulatory effects of dietary methionine on the European seabass (Dicentrarchus labrax) immune status, inflammatory response and disease resistance to Photobacterium damselae subsp. piscicida (Phdp). For this purpose, fish were randomly distributed in three independent groups (three replicates per group) and each was fed the corresponding diet: a control diet (CTRL) formulated to meet the established amino acid requirements for the species; a diet supplemented with methionine at 0.5% of feed weight relative to the CTRL diet (8.2% of methionine concentration above CTRL); and one supplemented with methionine at 1% of feed weight to the CTRL diet (11.8% of methionine concentration above CTRL). To evaluate the immune status of fish fed with each of the diets before being submitted to bacterial infection fish were sampled from each group at 2 and 4 weeks after the beginning of feeding. Non-sampled fish were injected intraperitoneally with Phdp (5 × 103 cfu/fish) at 4 weeks after initiation of feeding and the inflammatory response (at 4, 24, and 48 h post-infection) and survival (lasting 21 days post-infection) evaluated. Fish hematological profile, peripheral cell dynamics, plasma humoral immune parameters, leucocyte migration to the inflammatory focus and head-kidney gene expression were evaluated. Results show that methionine dietary supplementation improves seabass cellular immune status without evidence of activation of pro-inflammatory mechanisms. Additionally, the observed enhanced immune status provided by methionine supplementation translated into an improved immune response to infection, as higher cellular differentiation/proliferation and recruitment to the inflammatory focus, improved plasma humoral immune parameters and modulation of key immune-related genes was observed. Lastly, after a bacterial challenge, higher survival was observed in fish fed supplemented diets, ultimately corroborating the positive effect of methionine administration for 4 weeks in the cellular immune status.

Highlights

  • A dependency of the immune system upon the availability of amino acids (AA) has been associated to their role as signaling molecules essential for cellular function as reviewed in [1,2,3,4], and as methyl group donors and precursors of physiological important molecules, such as hormones, bioactive amines, enzymes, neurotransmitters and nitric oxide

  • After 1 week, during which fish were all fed with the commercial diet with which they were being fed previously, the experiment was started by feeding of each group with the respective feed 3 times a day at an average ration of 2.5% biomass per day: (i) one group was fed a control diet (CTRL); (ii) another group was fed a diet supplemented with 0.5% methionine of feed weight to the CTRL diet (MET0.5); and (iii) another group was fed a diet supplemented with 1% methionine of feed weight to the CTRL diet (MET1)

  • The modulatory effect of dietary methionine supplementation on the European seabass immune status was here evaluated at two different levels and for two different feeding periods

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Summary

Introduction

A dependency of the immune system upon the availability of amino acids (AA) has been associated to their role as signaling molecules essential for cellular function as reviewed in [1,2,3,4], and as methyl group donors and precursors of physiological important molecules, such as hormones, bioactive amines, enzymes, neurotransmitters and nitric oxide. Methionine is precursor of cysteine, one of the three glutathione (GSH) elements, a molecule involved in scavenging free radicals, protecting cells from oxidative stress during inflammation [1]. Methionine plays a pivotal role in processes responsible for the control of inflammation and apoptosis, such as protein ubiquitination and autophagy [10]. Methionine and its downstream metabolite SAM are responsible for autophagy modulation [12]. Further in-depth studies are needed to understand the immune mechanisms that this particular AA is activating before and after infection episodes

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