Putative epigenetic biomarkers of stress in the red blood cells of chickens reared across different biomes

Production animals are constantly subjected to adverse environmental conditions. These conditions influence the adult phenotype, mainly when they affect early life stages (pre and post-birth). These exposures will also produce epigenetic effects. DNA methylation of CpG dinucleotides in RedBloodCells(RBC) could be a useful epigenetic biomarker to identify animals subjected to chronic stress in the production environment. The present study compared the RBC methylome of chickens exposed to social isolation to non-exposed. This was performed in two experiments: one was run at LinköpingUniversity in Sweden and the other at Emprapa in Brazil. The aim was to identify stress associated with DNA methylation profiles in RBC across these populations, in spite of the variable conditions to which birds are exposed in each facility, and of chickens coming from independent lineages. Birds were exposed to social isolation and food and water deprivation at random periods of the day, from one to four hours from the first to the fourth week of their lives (BR, N=32 and SW, N=14). We collected RBC DNA and compared the reduced fraction of the RBC methylome between individuals of the socially isolated group and the non-isolated. We found 10 and 8 significant (p<0.0005) DifferentiallyMethylatedRegions(DMR) between treatments in the BR and SW populations, respectively. Most of these were located in intronic regions of Chr1, 2 and Z, but also in intergenic, up and downstream genes, and coding sequences. Interestingly, we found three significant (P <0.05) DMRs overlapping between the BR and SW lineages. The role of the genes associated to these DMRs (obtained through GO annotations and pathway analyses) are concordant with the function of RBC. We also found inter-treatment allele changes in animals before and after the experiment. Most of them overlapped with DMRs. Both, allele changes and DMRs were neighboring genes associated to cancer and coronary/heart diseases. This study is the first to track epigenetic biomarkers in production lineages, which, moreover, includes animals from different breeding programs and biomes. Our study represents the first approximation to develop an epigenetically based diagnostic tool for the detection of long-term stress in chickens, with possible application to other livestock species.