249 EFFECT OF REFREEZING BULL SEMEN ON IVF SUCCESS RATE
P. McCue A , J. Kelly B , S. Ashworth C , D. Kleemann B and S. Walker BA Animal Reproduction and Biotechnology Laboratory, Colorado State University, For Collins, CO 80523, USA
B South Australia Research and Development Institute, Turretfield Research Centre, Rosedale, SA, Australia
C Total Livestock Genetics, Camperdown, Victoria, Australia. Email: pmccue@colostate.edu
Reproduction, Fertility and Development 17(2) 275-275 https://doi.org/10.1071/RDv17n2Ab249
Submitted: 1 August 2004 Accepted: 1 October 2004 Published: 1 January 2005
Abstract
Cryopreserved semen from valuable sires may be available in limited quantities in some situations. A large percentage of the spermatozoa in a thawed straw is potentially wasted since a relatively small number of spermatozoa are required for most assisted reproduction techniques. The goal of this study was to determine the effect of dilution and refreezing of bull semen on fertilization and blastocyst development rates following in vitro fertilization. The hypothesis was that frozen bull semen that was thawed, diluted, and refrozen could be used successfully for IVF. Oocytes were harvested from cow ovaries collected from a local abattoir and matured in vitro for 24 hours. Ova were subsequently assigned to one of four in vitro fertilization treatment groups. Group 1 ova (n = 158) were fertilized with bull semen frozen at a concentration of 20 × 106 spermatozoa per 0.25 mL straw. Group 2 ova (n = 157) were fertilized with semen frozen at an initial concentration of 2 × 106 spermatozoa. Group 3 ova (n = 157) were fertilized with semen that had been thawed and refrozen at a concentration of 20 × 106 spermatozoa. Group 4 ova (n = 150) were fertilized with semen that had initially been frozen at a concentration of 20 × 106 spermatozoa and then thawed, diluted to a concentration of 2 × 106 spermatozoa, and refrozen. IVF was performed in a medium volume of 100 μL using 1 × 106 spermatozoa/mL. Cleavage and blastocyst rates were determined 2 days and 7 days, respectively after IVF. Cleavage rates following IVF was highest with semen frozen at 20 × 106 spermatozoa (89.9%), intermediate with semen frozen at 2 × 106 spermatozoa or refrozen at 20 × 106 spermatozoa (71.3% and 73.9%, respectively), and lowest with semen refrozen at 2 × 106 spermatozoa (38.7%) (P < 0.05). Blastocyst development rate was not significantly different (P > 0.05) among treatment groups. This study confirmed the hypothesis that refrozen bovine semen can be used successfully for in vitro fertilization. Although the overall IVF efficiency was lower using diluted refrozen semen, multiple IVF procedures could theoretically be performed over time from one initial straw. Consequently, if a limited amount of frozen semen is available, thawing of a single straw followed by dilution, re-allocation into multiple straws, and refreezing should be considered to facilitate the more efficient use of semen in future assisted reproduction endeavors.
This study was supported by the PEG Program, Colorado State University.
