Infertility affects 10–15% of couples, and approximately 50% of cases are linked to male factor infertility. The purpose of this study was to evaluate the DNA methylation patterns in spermatozoa from males who are suffering from a reduction in fecundity. Thirty samples were subjected to 450K arrays as a screening study to evaluate the variation in sperm DNA methylation levels between cases and controls groups, and then four CpG sites (cg05799088, cg07227024, cg16338278, and cg08408433) underwent to deep bisulfite sequencing to validate the observed methylation differences in 111 samples (56 proven fertile males as ‘controls’ and 55 males suffering from a reduction in fecundity as ‘cases’). A significant difference in the mean methylation level was found between cases and controls in the CpGs of PRICKLE2 gene‐related amplicon (CpG1, p ≤ 0.002, and CpG2, p ≤ 0.004) and CpG of ALS2CR12 gene‐related amplicon (CpG1, p ≤ 0.015, and CpG2, p ≤ 0.009). Besides, a significant difference was found at seven from thirteen CpGs tested in the ALDH3B2 gene amplicon CpG2, CpG6, CpG9, CpG10, CpG11, CpG12, and CpG13 (p ≤ 0.005, p ≤ 0.004, p ≤ 0.012, p ≤ 0.028, p ≤ 0.012, p ≤ 0.009, and p ≤ 0.001, respectively). In addition, the results showed that nine CpGs out of the twenty‐six within the PTGIR gene‐related amplicon (CpG4, CpG6, CpG8, CpG9, CpG11, CpG15, CpG19, CpG23, and CpG26) had a significant difference in their mean methylation level (p ≤ 0.006, p ≤ 0.009, p ≤ 0.003, p ≤ 0.003, p ≤ 0.007, p ≤ 0.002, p ≤ 0.018, p ≤ 0.018, and p ≤ 0.040, respectively) in the case vs. control group. In conclusion, an alteration in the methylation levels of sperm DNA from males with reduced fecundity was observed. In addition, an association between changes in the methylation level for these CpGs and different semen parameters has been found.