Poster Presentation IXth International Conference on Boar Semen Preservation 2019

Update on the development of fresh and frozen sexed sorted boar sperm (#4.4)

Kilby L Willenburg 1 , Domingo Gomez-Lopez 1 , Paul Benesh 2 , Ofelia Lapacek 2 , Vish Vishwanath 3
  1. Fast Genetics, Middleton, WI, United States
  2. ST Genetics, Middleton, WI, United States
  3. ST Genetics, Navasota, TX, United States

The ability to gender separate boar sperm by DNA content with flow cytometry has been prevalent for several decades. The following data compared sorting protocols from ST-Fast Genetics (STF) and a published (P) sorting protocol with interest in sorting efficiencies and survivability for fresh and frozen boar sperm. Total and progressive motility, viability, and acrosome damage for fresh semen were analyzed at 0, 6, 24, 48, 72, 96 and 168h in addition to fragmented DNA at 0, 6, 24, 48 and 72h. In a separate experiment sorted semen was frozen-thawed with the STF sorting/freezing protocol and compared to a LEY freezing media with total and progressive motility, viability and acrosome damage analyzed at 0 and 2 h. The results show improvements in sorting rates (40%), orientation (24%) and PVR (peak to valley ratio, 103%) with the STF protocol compared to the P protocol (P<0.05). Immediately after sorting (0h) motilities were 76.6±2.8% for the P treatment, which was significantly lower (P<0.01) than STF (88.6 ±2.8) and a non-sorted control (92.6 ±2.8%). At 24h, motility dropped significantly (P<0.01) for P treatment (34.7 ±2.8%) compared to STF (90.2±2.8) and non-sorted cells (93.9 ±2.8%). The average motility for the next 120h was significantly different among all treatments (P<0.01) with non-sorted (91.0 ±2.8%) higher than STF (79.6 ±2.8%) and P (7.6 ±2.8%) treatments. Sperm viability followed a similar pattern with the P protocol significantly lower at 24h (68.3 ±1.8%) compared to STF (86.5 ±1.8%) and non-sorted control (92.2 ±1.8%, P<0.05). Furthermore, sorting did not increase the percentage of fragmented DNA as the non-sorted cells had a higher percentage of fragmented DNA over 72 h (2.2 ±0.1%) than P (0.78 ±0.1%) and STF protocols (0.4 ±0.1%, P<0.05). For frozen-thawed sorted sperm, STF had a significantly (P<0.05) higher motility at 0 (64.5 ±1.7%) and 2h (54.4 ±1.7%) compared to LEY treatment (50.0 ±1.7%, 41.8 ±1.7%) and significantly (P<0.05) lower acrosome damage (STF = 13.2 ±0.75 vs LEY = 22.7 ±0.75%). Interestingly, viability was lower in the STF method (42.3 ±1.25%) compared to LEY over 2h (46.5 ±1.25%, P<0.05). Overall, these data showed improvements in sorting rates, sorting efficiencies and sperm survival with the STF protocol compared to a published protocol. The STF protocol minimized the deleterious effects of sorting and maintained similar semen quality to non-sorted sperm up to 7 days in storage. Coupled with an AI strategy that includes synchronization, uterine semen deposition, and insemination with semen from boars of known fertility a breeding program with sexed sorted sperm is more feasible for commercial application.