268 TIMED INSEMINATION OF HEIFERS WITH SEXED SPERM

The objective of this field trial was to compare pregnancy rates in Angus heifers time-inseminated with sexed sperm 55–56 h or 67–68 h after CIDR removal and administration of 25 mg prostaglandin F2α (PGF2α). Timed insemination eliminates labor costs associated with visual detection of estrus. Anecdotally, use of sexed sperm in commercial timed insemination environments has resulted in reduced pregnancy rates compared to rates for visually observed estrus. However, the optimal fixed time of insemination for sexed sperm has not been studied previously. Sperm were stained at 200 � 106 sperm mL–1 with 49 µm Hoechst 33342 for 45 min at 34.5�C and then diluted to 100 � 106 sperm mL–1 for sorting. Sperm were sexed by flow cytometry/cell sorting at 40 psi using a quasi-cw (pulsed; Vanguard 350-HDM, Spectra-Physics, Mountain View, CA, USA) laser operating at 125 mW, resulting in >91% X-chromosome-bearing sperm. Sorted sperm(2 � 106 total) were packaged into 0.25-mL straws at 10 � 106 sperm mL–1 and cryopreserved. Straws contained >43% progressively motile sperm immediately after thawing for 30 s in a 37�C water bath, as determined by 'track' motility. Angus heifers (n = 67) were synchronized for estrus with IM administration of 100 µg gonadotropin-releasing hormone (GnRH) and a CIDR in place for 7 days, followed by 25 mg PGF2α IM. Heifers were inseminated in the uterine body either 55–56 h (the timing currently recommended for unsexed semen) or 67–68 h after CIDR removal, accompanied by 100 µg GnRH IM, balanced across two inseminators with sperm from two Angus bulls. Estrus was visually detected three times daily but not used as the basis for insemination. Two months post-insemination, pregnancywas determined using ultrasound. Datawere subjected to chi-square analysis. Pregnancy rates for heifers inseminated 55–56 h and 67–68 h after CIDR removal were 34% (n = 11/32) and 46% (n = 16/35), respectively (P > 0.1). Estrus was visually detected for 85% (n = 57/67) of the heifers. Four heifers not detected in estrus became pregnant (40%). Respective pregnancy rates resulting from inseminations 0–12 h and 18–24 h after observed onset of estrus were 25% (7/28) and 55% (16/29), respectively (P < 0.05). Based on the limited scope of this preliminary field trial, insemination of heifers with sexed sperm at 67–68 h post-CIDR removal may be preferable to insemination at 55–56 h. It appears to be best to wait 18 to 24 h post-onset of observed estrus to inseminate heifers with sexed sperm when not using fixed-time insemination procedures. A larger field trial is warranted to more accurately identify the optimal fixed time for insemination using sexed sperm.