Addendum to: The relationship between sperm nuclear DNA fragmentation, mitochondrial DNA fragmentation and copy number in normal and abnormal human ejaculates

Background While the kinetics of human sperm nuclear DNA fragmentation (SDF‐nDNA) following ejaculation have been described, the dynamics and relationships of mitochondrial DNA copy number per spermatozoon (mtDNAcn) and fragmentation (SDF‐mtDNA) remain unexplored. Objectives To compare post‐ejaculatory kinetics of mtDNAcn, SDF‐mtDNA and SDF‐nDNA, global, single‐strand DNA breaks (SDF‐SSBs) and double‐strand DNA breaks (SDF‐DSBs) in normozoospermic and non‐normozoospermic samples. Materials and methods 28 normozoospermic and 43 non‐normozoospermic ejaculates were evaluated at 0, 6, 24 and 48 h of incubation in vitro. SDF‐nDNA was determined by sperm chromatin dispersion (SCD) assays. mtDNAcn and SDF‐mtDNA were analysed by dPCR. Results SDF‐nDNA‐global values increased as a consequence of quadratic SDF‐SSBs and linear SDF‐DSBs kinetics. Non‐normozoospermic samples showed a slower SDF‐global rate between 6–24 h, due to lesser SSBs production. Regarding SDF‐DSBs, non‐normozoospermic samples exhibited a faster initial increase rate, followed by a slower final increment. The mtDNAcn median value decreased linearly, being 3.2× higher in non‐normozoospermics at all time points; mtDNAcn in both cohorts reduced to half of the baseline by 48 h. mtDNAcn was identified as a risk factor for discriminating non‐normozoospermia, a finding that was further strengthen when combined with SDF‐Global or SDF‐DSBs values. SDF‐mtDNA frequencies were identical, increasing over time correspondingly in both cohorts. The mtDNA fragmentation rate was initially fast, decreasing progressively with time for both cohorts; half of the initially unfragmented copies were fragmented after 48 h. Rates of mtDNAcn loss and SDF‐mtDNA increase were only marginally correlated with the rates of nuclear fragmentation. Conclusion mtDNA fragmentation and loss occur post ejaculation. Their dynamics are likely to be associated with different and/or uncoupled mechanisms to that which cause nuclear DNA fragmentation. Our results indicate that while mtDNA fragmentation is not influenced by the sperm quality, the number of copies of sperm mtDNAcn can potentially serve as a risk factor for predicting non‐normozoospermia.