Assessment of sperm longevity after diluting in different extenders

Differences in semen viability are still in the focus of clinicians working in artificial insemination (AI) centres. Clinical experience indicates that the quality of the produced fresh cooled and frozen semen is depending on individual stallion properties as well as the sensitivity to different extenders.

With the use of hormonal treatments (e.g. ovulation induction), pregnancy rates can be increased but the role of semen extenders is still a key factor in achieving good or excellent quality semen. Extenders provide the optimal environment for sperm cells during their journey and companies try to develop newer recipes; but over the last decades the base components of these liquids remained the same. When producing AI doses, the question arises: which extender will yield the best results for a particular sire with a specific sperm quality. In this study three different stallions with different sperm quality (excellent, good and poor) have been tested for viability with commercially available semen extenders offered for treating even lower quality semen. All stallions were kept at the same equine AI centre located in central Hungary under the same conditions. The animals have been clinically healthy and all of them were used as AI sires with good and acceptable pregnancy rates (65-100%). Fresh semen was collected, filtered and assessed at first for raw semen volume, total (TM%) and progressive motility (PM%), and concentration. Then the semen was diluted with 14 different commercially available extenders (INRA96, minitube EquiPlus, minitube Beyond, Hippex Prime, Hippex Plus, Hippex Equine Semen Extender, Spervital OVD white cap, EVD Plus red cap, EVD blue-old cap, EVD blue-new cap, EVD Special green cap, Nidacon BotuSemen, BotuSemen Gold, BotuTurbo, BotuSemen Special) according to the manufacturer's instructions. Diluted semen doses were cooled and stored at 4°Celsius and checked at 24 hours intervals for total and progressive motility at 12, 36, 60, 84, 108, 132 and 156 hours post collection. All data were inserted into Microsoft Excel and analysed for different parameters (sperm quality, stallion and extender as variables) relative to time (T) post collection. Significant differences were observed in TM% and PM% between stallions’ samples when applying different extenders. Best TM% and PM% could be seen at 12 hrs (37±9% mean±SD) and 36 hrs (23±8% mean±SD) motility assessment. Thereafter, the results slowly decreased until 60th hrs. Stallions with excellent or good initial motility had sperm with acceptable total and progressive motility for a longer time; four extenders-maintained sperm cells viable for more than 132 hours. Stallion-3 (low quality semen) produced poor results at the start, but interestingly, at 60 hrs, post-collection, both PM% and TM% increased for the next 24 hrs which may have clinical relevance. In conclusion, testing the extender before producing AI doses from a particular stallion is crucial for achieving the best results to provide our patients’ owners the best possible care.