A combined genotyping by sequencing (GBS) and methylated DNA immunoprecipitation (MeDIP) protocol was used to identify -in parallel- genetic variation (GWAS) and epigenetic differences in differentially methylated regions (DMR) in the genome of spermatozoa from the porcine animal model.

A combined genotyping by sequencing (GBS) and methylated DNA immunoprecipitation (MeDIP) protocol was used to identify -in parallel- genetic variation (GWAS) and epigenetic differences in differentially methylated regions (DMR) in the genome of spermatozoa from the porcine animal model. Breeding boars with good semen quality (n=11) and specific and well documented differ-ences in fertility (farrowing rate, FR) and prolificacy (litter size, LS) (n= 7) in artificial insemination programs were, using combined FR and LS, categorized as high-fertile (HF, n= 4) or low-fertile (LF, n=3), and results contrasted to boars with unknown fertility (UF, n=4). We identified 165,944 SNPs that explained 14-15% of variance among selection lines. Between HF and LF individuals (N=7, 4H and 3L), we identified 169 SNPs with P = 0.00015, which explained 58% of the variance among individuals. For the epigenetic analyses we considered fertility and period of ejaculate collection (late-summer and mid-autumn). Approximately 3-times more DMRs were observed in HF- than in LF-boars across these periods. Interestingly, UF-boars were clearly clustered with one of the other HF- or LF-groups. The highest differences in DMRs between HF and LF experimental groups across the pig genome were located in the chr 3, 9, 13 and 16, with most DMRs being hypermethylated in LF -boars. In both HF- and LF-boars DMRs are mostly hypermethylated in late-summer (LS) compared to mid-autumn (MA). Three overlaps were detected between SNPs (P=0.0005, n=1,318) and CpG sites within DMRs. In conclusion, fertility levels in breeding males including FR and LS can be discerned using methylome analyses, and the findings in this biomedical animal model ought to be applied besides sire selection, for andrological diagnosis of idiopathic sub/infertility.