Variation in oxytocin receptor DNA methylation in female chickens are associated with age, genetics, laying intensity, and housing conditions.

Animal welfare monitoring is rapidly becoming a critical concern, with an ongoing search for biomarkers. DNA methylation has the potential to serve as such a biomarker, as environment and genotype can interact to shape DNA methylation. We tested the hypothesis, whether genetically distinct laying hens display divergent methylation patterns at selected genes relevant for behavior in response to housing conditions. The aims of the present study were therefore to determine the effect of age (two-day-old chicks versus one-year-old laying hens), genetics (brown versus white laying hens), laying intensity (high versus low), and the effect of housing conditions (layer cage versus small group housing) on the methylation levels of selected genes implicated in behavior in chicken. DNA methylation at the OXT, OXTR, ESR1, HTR1A, HTR2A, and HTR3A gene were determined using direct bisulfite-PCR sequencing, with OXTR being the only gene displaying consistent effects. The greatest effect of age on methylation was evident for OXTR, with adult animals displaying significantly higher methylation than 2-day old chicks. While methylation levels for OXTR did not differ between chicks, significant differences in methylation levels were detected for OXTR when comparing samples from adult animals. For the majority of CpG sites, an interactive effect of laying intensity x housing and genetics x housing on OXTR DNA methylation was evident. When evaluating OXTR methylation status separated by housing condition, a clear difference between layer lines was revealed. While BLA (brown, high laying intensity) animals exhibited consistent methylation levels independent of the housing condition, WLA (white, high laying intensity), G11 (white, low laying intensity) and L68 (brown, low laying intensity) animals displayed differences in OXTR methylation depending on the housing condition. In summary, age, genetics, laying intensity, and housing conditions are associated with differences in DNA methylation of the oxytocin receptor in female chicken, suggesting that OXTR methylation may represent a candidate epigenetic biomarker linking genetics, environment, and laying performance. Future studies will have to determine its correlation with animal welfare status.