G. T. Gentry

Sex Ratio of Bovine Embryos and Calves Originating from the Left and Right Ovaries

An asymmetric distribution of the sexes within the left and right uterine horns has been described in multiple species. A series of experiments were conducted to evaluate the sex ratio (% male) of calves gestated in the left and right uterine horns, as well as the sex ratio of embryos originating from the left and right ovaries of cattle. The sex ratio of calves gestated in the right uterine horn of naturally mated cows was significantly higher compared with the sex ratio of calves gestated in the left uterine horn. In addition, the sex ratio of the left and right uterine horns differed significantly from parity. The sex ratio of embryo transfer calves born following transfer to the left and right uterine horns was not significantly different. Additionally, the proportion of male embryos collected from the right uterine horns was significantly greater than from the left uterine horns of superovulated cows. The sex ratio of embryos collected from the left and right uterine horns of unilaterally ovariectomized cows was not significantly different. However, more female than male embryos were produced when left ovary oocytes fertilized in vitro. In conclusion, the results of these experiments demonstrate that a significantly greater proportion of males are gestated in the right uterine horn of cattle and a greater proportion of females in the left. Additionally, the data indicate that sex-specific selection pressure may be applied to embryos by ovarian factors rather than by the uterine environment.

Cryopreservation of Day 8 equine embryos after blastocyst micromanipulation and vitrification

Pregnancy rates after cryopreservation of large equine blastocyst stage embryos have remained lower than other domesticated livestock species. It is generally accepted that the embryonic capsule is the primary barrier to cryoprotectant entry into the embryo proper and techniques need to be developed to circumvent this obstacle. Therefore, the objective of this study was to develop an efficient Day 8 equine embryo cryopreservation protocol through blastocyst micromanipulation and vitrification. Grade 1 and 2 embryos recovered from mares (n = 15) 8 days after ovulation were used in these experiments. In experiment 1, the effect of either one- or two-puncture treatments before aspiration of blastocoel fluid and exposure to vitrification solutions was evaluated. No difference was detected in mean embryo volume across treatment groups after exposure to vitrification solutions or after 1, 24, 48, and 72 hours of culture. Percent of embryos re-expanding at 24 hours and percent of embryos showing diameter increase at 48 and 72 hours during in vitro culture were 100%, 83%, and 75% compared with 93%, 67%, and 50% for one- and two-puncture treatment groups, respectively. Capsule loss was 25% for one-puncture and 50% for two-puncture treatment groups. In experiment 2, no difference was detected in mean embryo volume for indirect introduction (aspiration of blastocoel fluid + equilibration) and direct introduction (injection of cryoprotectant into blastocoel cavity) treatment groups, after exposure to dilution solution or to culture medium. There was no difference in mean embryo volume for the indirect and direct introduction treatment groups after 1, 24, 48, and 72 hours of culture. Percent of embryos re-expanding at 24 hours and percent of embryos showing diameter increases at 48 and 72 hours during in vitro culture were 100%, 76.9%, and 69.2%, respectively, for both treatment groups. Those embryos subjected to the direct introduction treatment had a higher (P = 0.05) percent capsule loss (70%) compared with the indirect introduction treatment group (31%). The pregnancy rate after transfer of vitrified expanded Grade 1 blastocysts using the indirect introduction method was 83% (5/6). Three pregnancies were allowed to continue to term and resulted in the birth of three healthy foals. The vitrification protocol used in this study has the potential to become a key tool for the successful cryopreservation of equine expanded blastocysts.

The effect of exogenous follicle stimulating hormone (FSH) and endogenous plasma leptin concentrations on the pregnancy rate of beef heifers subjected to fixed-timed artificial insemination (FTAI)

In this study, 53 crossbred beef heifers were used to test the hypotheses that administration of exogenous FSH 2 days following CIDR insertion and administration of estradiol would increase the pregnancy rate in heifers synchronized for FTAI and that plasma leptin concentrations in beef heifers would be higher for heifers that became pregnant to FTAI. The heifers used in this study had a median age of 440 days, an average weight of 324 kg, an average body condition score of 5.1 and a mean reproductive tract score of 3.1. Heifers were stratified by weight and BCS into two groups and then treatments were randomly allotted to each group: (1) control (n=28) or (2) FSH (n=27). Both groups were administered 200mg estradiol benzoate (EB) and received an intravaginal controlled internal drug-releasing device (CIDR) on day 0. On day 2, females in the FSH treatment group were administered 20 mg of FSH, while the control group received 1 ml of saline. On day 7 all females were administered 25 μg PGF2α and the CIDR was removed. Then 24h following CIDR removal all females were administered 1mg EB and 24h later were subjected to FTAI. Pregnancy diagnosis was performed via transrectal ultrasonography 43 days following insemination. Blood samples were collected via jugular venipuncture on days 2, 6-10, 13 and 52 and plasma leptin concentrations were determined by radioimmunoassay. Pregnancy rates were higher (P=0.01) for FSH-treated females (60%) compared with females not receiving FSH (25%). Circulating plasma leptin concentrations were higher (P=0.0051) for pregnant females compared with females that did not become pregnant following FTAI during the experiment. Mean plasma leptin concentration was also higher (P=0.04) from day 2 to day 9 during the synchronization protocol in heifers that became pregnant compared with heifers that did not become pregnant from FTAI. There was no difference (P=0.38) in reproductive tract scores for heifers that became pregnant compared with heifers that did not become pregnant from FTAI. Circulating leptin concentrations were not different (P=0.11) for females receiving FSH compared with females in the non FSH-treated group. Circulating leptin concentrations were affected by sampling day (P<0.0001). However, there was no interaction between sampling day and pregnancy status (P=0.80), treatment and pregnancy status (P=0.14) or treatment and sampling day (P=0.12). These results indicate that the administration of FSH on day 2 of the synchronization protocol may increase pregnancy rates in beef heifers and that increased circulating concentrations of plasma leptin during the synchronization protocol may be indicative of subsequent pregnancy outcome.

The effect of equine chorionic gonadotropin (eCG) on pregnancy rates of white-tailed deer following fixed-timed artificial insemination.

Control of the white-tailed does reproductive cycle is not well documented. The objective was to determine the effects of giving equine chorionic gonadotropin (eCG) at progesterone device removal on fixed time artificial insemination (FTAI) pregnancy rates in white-tailed does. All does (n = 74) were synchronized with a vaginal progesterone implant (CIDR; 0.3 g progesterone), inserted on Day 0 (without regard to stage of estrous cycle), removed 14 days later, and subjected to FTAI, on average, 60 h post-CIDR removal. Of these, 34 were given 200 IU (im) of eCG at CIDR removal. Overall, FTAI pregnancy rate was 50% across 2 yrs (effect of year, P = 0.35). Administration of eCG at CIDR removal did not affect (P = 0.16) pregnancy rate (eCG = 59%; no eCG = 43%). Pregnancy rates were not affected by vulva score or doe disposition. Does that were ≤ 4 yrs old were more likely (P = 0.01) to become pregnant than does > 4 yrs of age. Does inseminated ≥ 60.5 h after CIDR removal were 22 times more likely (P = 0.002) to become pregnant to FTAI than does inseminated < 60.5 h. When frozen-thawed semen was deposited in the cervix or uterus, does were 17 times more likely (P = 0.005) to become pregnant compared with those receiving intravaginal insemination. Fecundity was not different (P = 0.73) across treatment groups (1.6 ± 0.11; no eCG vs. 1.7 ± 0.10; eCG). Furthermore, fecundity of does pregnant to FTAI was not different (P = 0.72) compared with does pregnant to clean-up bucks (1.7 ± 0.08; AI does vs. 1.7 ± 0.09; clean-up bucks). In summary, white-tailed does were successfully inseminated using a 14 days FTAI protocol, eCG may not be essential for acceptable pregnancy rates, and increased pregnancy rates may result when FTAI is done ≥ 60.5 h after progesterone device removal.

Pregnancy rates following low-temperature storage of large equine embryos prior to vitrification

&NA; Satisfactory pregnancy rates can now be achieved following the cryopreservation of large equine embryos. Nonetheless, its wide application might be limited by the fact that the cryopreservation of large equine embryos requires a specialized micromanipulation equipment and micromanipulation/vitrification skills. Alternatives should be developed to increase its utilization and widespread application in the commercial equine industry. To determine if large equine embryos are able to remain viable during transport from farms to specialized centers for embryo cryopreservation, we evaluated pregnancy rates following the low‐temperature storage of large equine embryos before vitrification. Grade 1 embryos (n = 37) were randomly assigned to six treatments consisting of day of collection (Day 7 or 8 after ovulation) and cooling for 0, 12, or 24 hours before vitrification in a factorial design. Pregnancy rates of Day 7 embryos cooled for 12 and 24 hours were 55.5% and 75%, respectively. Pregnancy rates of Day 8 embryos cooled for 12 and 24 hours were 0 and 16.6%, respectively. Day 7 cooled embryos resulted in higher pregnancy rate compared with Day 8 cooled embryos (64.7% and 7.7%, respectively; P < .05). Pregnancy rate comparison of cooled embryos grouped by diameter showed that embryos <550 &mgr;m resulted in a higher pregnancy rate compared with embryos >550 &mgr;m (71.4% and 12.5% respectively; P < .05). In conclusion, Day 7 equine embryos up to 550 &mgr;m can be cooled to temperatures of 9–12°C for 12 or 24 hours before vitrification and result in satisfactory pregnancy rates.

Impacts of incorporation of follicle stimulating hormone into an estrous synchronization protocol for timed artificial insemination of crossbred beef cattle

One-hundred-eighty crossbred beef cows and 66 crossbred beef heifers across three locations were stratified by body weight (BW), body condition score (BCS), and age (within location) to evaluate administration of follicle stimulating hormone (FSH) on Day 2 using a modified 7-day CO-Synch plus CIDR(®) protocol (Day 0=CIDR insertion) with timed-artificial insemination (TAI) at 72 h (cows) or 54 h (heifers) following CIDR removal. Estrous response following CIDR removal was determined using an Estrotect patch and TAI and final pregnancy rates were determined by transrectal ultrasonography 42-45 days following TAI and ≥ 45 days following removal of clean-up bulls. Estrous response rate, TAI and final pregnancy rates for cows were not affected (P ≥ 0.65) by treatment. Cows that exhibited estrus had greater (P<0.01) TAI pregnancy rate (66%) than cows not exhibiting estrus (38%). There was an estrous response by postpartum length interaction (P=0.02) where cows exhibiting estrus and ≥ 55 days postpartum had greater TAI pregnancy rates (75%) compared to cows not exhibiting estrus and < 55 days postpartum (39%) or ≥ 55 days postpartum (28%). For heifers, timed AI (P=0.46) and final pregnancy rates (P=0.45) were similar across treatments and estrous response had no effect (P=0.30) on TAI pregnancy rates. In conclusion, the addition of FSH to the CO-Synch plus CIDR estrous synchronization protocol did not increase TAI pregnancy rates in beef cows or heifers. However, a positive estrous response to the synchronization protocol was associated with increased TAI pregnancy rates in cows.