Abstract
BackgroundIn fish, minimally invasive blood sampling is widely used to monitor physiological stress with blood plasma biomarkers. As fish blood cells are nucleated, they might be a source a potential new markers derived from ‘omics technologies. We modified the epiGBS (epiGenotyping By Sequencing) technique to explore changes in genome-wide cytosine methylation in the red blood cells (RBCs) of challenged European sea bass (Dicentrarchus labrax), a species widely studied in both natural and farmed environments.ResultsWe retrieved 501,108,033 sequencing reads after trimming, with a mean mapping efficiency of 73.0% (unique best hits). Minor changes in RBC methylome appeared to manifest after the challenge test and a family-effect was detected. Only fifty-seven differentially methylated cytosines (DMCs) close to 51 distinct genes distributed on 17 of 24 linkage groups (LGs) were detected between RBCs of pre- and post-challenge individuals. Thirty-seven of these genes were previously reported as differentially expressed in the brain of zebrafish, most of them involved in stress coping differences. While further investigation remains necessary, few DMC-related genes associated to the Brain Derived Neurotrophic Factor, a protein that favors stress adaptation and fear memory, appear relevant to integrate a centrally produced stress response in RBCs.ConclusionOur modified epiGBS protocol was powerful to analyze patterns of cytosine methylation in RBCs of D. labrax and to evaluate the impact of a challenge using minimally invasive blood samples. This study is the first approximation to identify epigenetic biomarkers of exposure to stress in fish.
Highlights
In fish, minimally invasive blood sampling is widely used to monitor physiological stress with blood plasma biomarkers
We showed that a modified epiGenotyping By Sequencing (epiGBS) protocol originally proposed by Van Gurp et al [34] was applicable to further analyze patterns of cytosine methylation in Red blood cell (RBC) of D. labrax
The European sea bass has become one of the most studied species in fish epigenetics [20,21,22,23,24,25,26,27], and for this species or for other cultured fish species, our modified version of the original epiGBS protocol seems to be a powerful and affordable method to screen a significant number of individuals with sufficient depth and coverage to reach meaningful conclusions
Summary
Minimally invasive blood sampling is widely used to monitor physiological stress with blood plasma biomarkers. Because samples are easy to obtain, poorly invasive, and can be stored in large collections that may reflect variation in many parameters at both the individual and the population levels, blood is certainly the most commonly used tissue to check for and to monitor the response of cells, organs, or whole organism to environmental perturbations, to assess health status of organisms, and to diagnose metabolic impairments and dysfunctions in vertebrates. Because fish blood cells are nucleated and, apart from blood plasma in which cortisol, glucose, lactate and other metabolites are measured, mobilized as part of the stress response in fish [12, 13], it is appealing to investigate if components of their genomic machinery may respond to environmental stressors and broaden the panel for poorly invasive stress monitoring. The use of red blood cells (RBCs) in ‘omics fish studies has received little attention [14,15,16], and a single study investigated RBC epigenome in steelhead (Onchorhynchus mykiss) [17]
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