10 CASA EVALUATION OF SEXED AND NON-SEXED FROZEN BULL SEMEN

Abstract

Flow citometry cell sorting has been proven successfully to separate X and Y sperm; however, the technology is still too stressfull for the viability of the sorted semen. The objective of this study was to evaluate nonsexed and sexed frozen sperm motility characteristics using a CASA technology. Ejaculates from 4 different bulls (3 Holstein and 1 Angus) were collected, and processed as split non-sexed and sexed semen samples using Tris egg yolk extenders. X and Y sperm were separated using a high-speed sorter (SX Moflo). Cryopreservation was done at the same time under appropiate conditions using a programmed cryochamber. Thawing procedure was done at 37°C for 30 s and a sample of each straw was placed in the evaluation chamber. The experiment was repeated twice and two chambers with 30 observations each were analyzed each time. Mean and standard deviation of each characteristic were calculated, compared and analyzed statistically. The sperm concentration was determined by means of a burker counting chamber. Sperm quality was determined at 0 h after thawing, and later at 1 h, 2 h and 3 h after incubation in a glass tube at 30°C. The following sperm motility parameters were determined with the Hamilton Thorne (HTM-ceros 12.1) on at least 1000 spermatozoa: velocity average path (VAP), velocity straight line (VSL), amplitude lateral head (ALH), beat cross frequency (BCF), straightness (STR), linearity (LIN), and percentage of progressively motile spermatozoa (PMS). Linearity of nonsexed spermatozoa was 53 ± 3.5, 47 ± 0.8, 43 ± 7.8 and 42 ± 4.5 for the 0 h and the 3 test incubation times and 49.5 ± 3.7, 51.2 ± 3.7, 43.3 ± 7.8 and 44.5 ± 7.6, respectively, for sexed semen. There were no significant differences (P > 0.05) in the progressive velocity, track speed and linearity between sexed and nonsexed semen. The percentage of static cells was 33%, 30%, 47% and 50% for the 0 h and the 3 test incubation periods; however, the percentage of static cells for the sexed semen was 53%, 71%, 77% and 82%, respectively. Results from the analysis indicate a significant increase (P < 0.01) in the number and the percentage of static cells with time. The lateral amplitude of sperm motility for nonsexed semen was 5.9 ± 0.5, 6.8 ± 0.8, 6.0 ± 0.4 and 5.1 ± 0.7, and for sexed semen 6.6 ± 0.7, 6.8 ± 0.4, 6.4 ± 0.4 and 5.5 ± 1.7, respectively. The percentage of progressively motile sperm was significantly different at 0 time 49.7 ± 4.9 and 23.1 ± 4.9 for nonsexed and sexed semen respectively. After 3 hours of incubation the percentage of progressively motile sperm was 38.7 ± 10.2 and 3.7 ± 3.2 for nonsexed and sexed semen, respectively. In conclusion, sexed frozen semen seems to have characteristics similar to those of normal nonsexed semen. However, a significant decrease in the percentage of progressively motile cells could affect pregnancy rates. More research needs to be done to detect differences between bulls and cryoprotectans.Research supported by Centro Genetico Bovino de EOLIA sa Argentina.