The effect of interval between inseminations and semen handling on in-vivo and in-vitro fertility of frozen-thawed boar sperm

Cryopreserved gametes have played a substantial part in the advancement of genetic progress in the production livestock industries. However, the swine industry has not yet been able to capitalize on the potential of frozen-thawed boar sperm (FTS), due to the biological sensitivity of the sperm cells and the lower resulting pregnancy rates and litter sizes. This thesis will outline the current swine reproductive production practices, provide a brief history of boar sperm cryopreservation as well as the advancements in cryopreservation technology and evaluation. This will also report on the impact of artificial insemination in the swine industry with insight into the physiological response to AI, research using frozen-thawed boar sperm, and proper insemination timing relative to ovulation. Experiment one of this thesis determined the in vitro lifespan of frozen-thawed boar sperm to be used for artificial insemination. Many studies have been performed for determining the in vivo lifespan of frozen-thawed boar sperm, however very little is known about its in vitro lifespan. Samples (n = 20) were evaluated for motility and membrane integrity at the time of thawing and at 2, 6, 12 and 24 h intervals after thawing. Results indicated FTS can be diluted and stored at 17 or 26 °C for up to 2 h with no significant decrease in motility and membrane integrity (P > 0.10). Motility remained stable for only 2 h following thawing at all storage temperatures, however, storage at higher temperature showed a gradual decrease in post-thaw quality (P < 0.001). Additionally, it should be noted that viability was considerably less sensitive to storage temperature and duration, with samples held at 17 °C and 26 °C showing a marked reduction at 12 and 6 h, respectively. Storage of FTS at 37 °C, reduced motility and membrane integrity within 2 h, and samples were non-motile and non-viable by 24 hours. Experiment two was performed to determine the effect of interval between inseminations using FTS and its effect on pregnancy rate, litter size, and fetal paternity. We hypothesized that gilts inseminated at greater intervals would have increased pregnancy rates and litter sizes than those inseminated at a closer interval, accounting for variation in time of ovulation and the shortened lifespan of frozen-thawed boar sperm in the female reproductive tract. Gilts (n = 191) were assigned to one of the following treatments 1) AI at a 4 h interval (34 and 38 h); 2) AI at an 8 h interval (30 and 38 h); or 3) AI at a 16 h interval (22 and 38 h). First and second inseminations were from unique sires to allow for paternity identification. Interval between inseminations affected pregnancy rate (P 0.10). Gilts that ovulated by 36 and 48 h after the onset of estrus had greater pregnancy rates (P 0.10). Interval between inseminations had no effect on the proportion of fetuses sired by the second AI (P > 0.10); however, there was an interaction of interval between inseminations and estrus to ovulation interval (P < 0.05). The results of this experiment indicate that when FTS is used in a double AI system, highest fertility occurs when gilts are mated at 8 and 16 h intervals with estrus detection occurring at 12 hour intervals. The results of experiment one could have implications for handling and processing procedures for in vivo and in vitro fertility. Increased in vitro lifespan could allow for multiple dose preparation and storage prior to insemination. Additionally, the post-thaw quality results of samples held at 37 °C could be an indicator for in vivo post-thaw sperm survival patterns. With experiment two, very little information is available on the optimal insemination interval to account for the short lifespan of FTS, variation in time of ovulation, and an immune response due to artificial insemination. Knowledge from these studies may help improve timing for estrus detection and artificial insemination to optimize timing and increase fertility when using frozen-thawed boar sperm