Factors affecting the fertility of ewes after intrauterine insemination with frozen-thawed semen during the non-breeding season.

In this study, two successive field trials were conducted during the non-breeding season to investigate various factors affecting on fertility of Suffolk ewes after intrauterine insemination with frozen-thawed semen. In the first year (Experiment 1), three sperm numbers per insemination dose (0.25, 0.5 and 1 million sperm) and five sheep farms were used, and in the second year (Experiment 2), parity, age, body weight, body condition score (BCS) and postpartum days were investigated to compare pregnancy and lambing rates. High pregnancy and lambing rates (70.6 and 70.6%, respectively) were obtained with 0.25 million sperm per dose. There were no significant differences in the pregnancy and lambing rates among the five farms, but there was a tendency for one farm to have higher pregnancy (75.8%, P=0.065) and lambing (72.7%, P=0.077) rates than those (46.7-53.3% and 45.2-53.3% for the pregnancy and lambing rates, respectively) of the other farms. In Experiment 2, ewe age significantly affected both the pregnancy and lambing rates. Nulliparous ewes had a higher lambing rate (72.0%) than that (44.2%) of multiparous ewes, but a significant difference was not revealed. Regardless of body weight, BCS tended to be an important factor influencing on fertility of ewes. Body weight and the postpartum days did not affect the fertility of ewes. It was concluded from these results that the fertility of Suffolk ewes after intrauterine insemination with frozen semen was significantly influenced by sperm number per dose and ewe age. Nulliparous ewes at less than three years of age and with a BCS of more than 3.0 are expected to have higher fertility than other ewes.

Fertility after different artificial insemination methods using a synthetic semen extender in sheep.

The present study aimed to investigate the fertility of ewes artificially inseminated with three different methods using a synthetic semen extender, AndroMed. The three methods of artificial insemination (AI) were cervical AI with fresh-diluted or frozen-diluted semen at observed estrus, and an intrauterine AI with frozen-thawed semen. A total of 80 ewes were treated with a controlled internal drug release (CIDR) containing 0.3 g progesterone per device for 12 days. In Experiment 1 (26 Suffolk ewes), superovulation was induced with 20 mg follicle-stimulating hormone and 250 IU equine chorionic gonadotropin (eCG) two days and one day before CIDR removal, respectively, during the non-breeding season. In Experiment 2 (54 Suffolk and Suffolk crossbred ewes), an intramuscular injection of 500 IU eCG was administered one day before CIDR removal to synchronize estrus and ovulation during the breeding season. In Experiment 1, fresh-diluted or frozen-thawed semen was deposited into the cervical orifice after estrus detection, and an intrauterine AI with frozen-thawed semen was performed by laparoscopy at a fixed-time basis without estrus detection. Embryos were recovered by uterine flushing 6 days after AI, and the rates of recovered, fertilized (cleaved) ova and embryos at the morula or blastocyst stage were compared among the three AI methods. In Experiment 2, the pregnancy rates after the three AI methods were compared. In Experiment 1, the rates of recovered ova were not significantly different among the three AI methods (52.5-56.7%). The rate of fertilized ova (81.0%) by laparoscopic AI with frozen-thawed semen was significantly higher compared with cervical AI of fresh-diluted (25.5%) or frozen-thawed (3.5%) semen, but the rate of embryos at the morula or blastocyst stage (17.6%) was significantly lower than that of the cervical AI with fresh-diluted semen (69.2%). The rates of ewes yielding fertilized ova were not significantly different among the three groups (44.4, 11.1 and 62.5% for cervical AI with fresh-diluted and frozen-thawed semen and intrauterine AI with frozen-thawed semen). In Experiment 2, the pregnancy rate of ewes intrauterinally inseminated with frozen-thawed semen (72.2%) was significantly higher than those of ewes inseminated cervically with fresh-diluted (5.5%) or frozen-thawed (0.0%) semen. The present results showed that acceptable fertilization and pregnancy rates could be obtained by an intrauterine AI with frozen-thawed semen using a synthetic semen extender (AndroMed), but not sufficient by the cervical AI with either fresh or frozen semen.

Fertility after artificial insemination using a soybean-based semen extender in sheep.

The present study aimed to compare the fertility of ewes intrauterinally inseminated with frozen-thawed semen using a soybean-based semen extender (AndroMed) with those of ewes intrauterinally inseminated with frozen-thawed semen using a Tris-based extender containing either egg yolk or BSA. Suffolk ewes (n=104) were treated with an intravaginal sponge containing 40 mg fluoroprogesterone acetate (FGA) for 12 days and an intramuscular injection of 500 IU equine chorionic gonadotropin to induce estrus and ovulation during the non-breeding season (July, 2007). Intrauterine insemination was carried out 40-46 h after removal of the FGA sponge (n=90), regardless of the incidence of estrus. The pregnancy rates were not significantly different among the semen extenders containing egg yolk (64.5%) or BSA (58.6%) and AndroMed extender (56.7%). The lambing rates (64.5, 55.2 and 56.7% for the semen extenders containing egg yolk, BSA and AndroMed, respectively) and prolificacy (1.59 to 1.75) were also not significantly different. The present results indicate that an egg yolk-containing semen extender can be replaced with the non-animal derived extender AndroMed, which could be used for intrauterine insemination using frozen-thawed ram semen without reducing fertility.

Fertility of ewes inseminated intrauterinally with frozen semen using extender containing bovine serum albumin.

The present study was conducted to examine the fertility of ewes inseminated intrauterinally with frozen semen using semen extender containing either egg yolk or bovine serum albumin (BSA). Sixty Suffolk and cross-bred ewes were treated with controlled internal drug release (CIDR) devices during the non-breeding season (July 2006). A CIDR was inserted into the vagina for 12 days and an intramuscular injection of 500 IU equine chorionic gonadotropin was administered one day before its removal. Ejaculates from a suffolk ram were diluted with a Tris-based extender containing either 15% (v/v) egg yolk or 10% (w/v) BSA, and the diluted semen was frozen in 0.25 ml straws. A fixed-time intrauterine artificial insemination (AI) was performed 43-47 h after CIDR removal, regardless of incidence of estrus. There was no significant difference in pregnancy rates at 60 days after AI between the extenders containing egg yolk (66.7%, 20/30 animals) or BSA (65.5%, 19/29 animals). Furthermore, there were no significant difference in the lambing rates (66.7% and 62.1%) and prolificacy (1.25 and 1.56) between the two semen extenders. The present study indicates that a semi-defined semen extender containing 10% BSA produces fertility after intrauterine AI that is similar to that achieved with semen extender containing egg yolk.

Changes of fructose concentrations in seminal plasma and glucose and testosterone concentrations in blood plasma in rams over the course of a year.

The present study was performed to examine seasonal changes in the fructose concentrations of seminal plasma and glucose and testosterone concentrations of blood plasma over the course of a year (from November 2004 to November 2005) using 5 Suffolk rams. Osmolality of the seminal plasma was also measured. The fructose concentrations in the seminal plasma increased as the breeding season approached, with the maximum in October (179.8 mg/dl) and the minimum in May (6.9 mg/dl), although there were no significant differences during the year. Osmolality of the seminal plasma in February (304 mOsm) was significantly (P<0.05) lower than in January (325 mOsm), July (327 mOsm), and August (325 mOsm). It was also significantly (P<0.05) lower in November (308 mOsm) than in January and August. The blood plasma glucose concentration in October (79.3 mg/dl) was significantly (P<0.05) higher than in January and February (43.2 and 43.7 mg/dl, respectively). The blood plasma testosterone (T) concentrations were significantly (P<0.05) higher in September (8.5 ng/ml) and October (10.2 ng/ml) than in other months. The fructose concentrations in the seminal plasma appeared to be related to the glucose and T concentrations in the blood plasma. These results show that fructose concentrations in the seminal plasma and blood plasma glucose and T concentrations tended to increase during the breeding season, with the highest concentrations in October.

Effects of bovine serum albumin and trehalose in semen diluents for improvement of frozen-thawed ram spermatozoa.

In this study, two following experiments were performed to improve post-thaw motility and viability of frozen-thawed ram spermatozoa. We examined i) the effects of different concentrations of bovine serum albumin (0, 0.3, 1, 5, 10 and 15% BSA) in semen diluents lacking egg yolk and ii) the effects of four semen diluents, fructose (F: control) and trehalose (T) in semen diluents containing egg yolk, 15% BSA in semen diluents without egg yolk (BSA), and modified phosphate buffered saline (m-PBS). Frozen-thawed spermatozoa were examined for progressive sperm motility, viability, morphological abnormality, sperm tail swelling test, and sperm acrosome integrity. In Experiment 1, the rates of sperm motility immediately after thawing (0 h) were significantly (P<0.05) higher in the 10 and 15% BSA groups (55.0 +/- 2.9 and 58.3 +/- 6.7%, respectively) than in the positive control (F) group (41.7 +/- 4.4%). The rate of sperm viability in the negative control (0% BSA) group (80.2 +/- 3.3%) was significantly (P<0.05) lower than in the positive control (F) group (89.8 +/- 1.5%), but when compared with the F group, no significant differences were found among the 0.3, 1, 5, 10 and 15% BSA groups at 0 h. The rates of sperm morphological abnormality of the 10 and 15% BSA groups (6.5 +/- 1.3 and 6.3 +/- 1.1%, respectively) were significantly (P<0.05) lower at 0 h than that in the 1% BSA group (16.3 +/- 5.2%). In Experiment 2, T addition improved (P<0.05) the post-thaw motility compared with the F and BSA groups. Furthermore, at 3 and 6 h, the post-thaw motility of the T group (36.3 +/- 2.4 and 25.0 +/- 2.0%, respectively) was significantly (P<0.05) higher than in the BSA (26.3 +/- 2.4 and 18.8 +/- 1.3%, respectively) and F (28.8 +/- 3.8 and 18.8 +/- 2.4%, respectively) groups. The post-thaw sperm motility and viability in the m-PBS group were significantly (P<0.05) lower than those of the control (F), T, and BSA groups throughout all observation points. These results indicate that 10 and 15% BSA can be substituted for egg-yolk for ram semen diluent and that the addition of trehalose enhances motility and viability of ram spermatozoa after freezing and thawing.

Comparison of estrus induction and subsequent fertility with two different intravaginal devices in ewes during the non-breeding season.

Two experiments were conducted to compare the effect of estrus induction by controlled internal drug release (CIDR) and intravaginal cream containing 500 mg progesterone (P cream) in ewes during the non-breeding season. In the first experiment, twenty-four ewes were randomly grouped for two treatments with the different intravaginal devices for 12 days: Group A was the CIDR group and Group B was the P cream group. Blood was collected from all treated ewes, and progesterone (P(4)), estradiol 17-beta (E(2)) and luteinizing hormone (LH) concentrations were measured by enzyme immunoassay. In the second experiment, the conception rates from natural mating, estrus-detected AI (inseminated 12 h after estrus detection), or fixed-time AI (inseminated 42 h after removal of an intravaginal device) in 127 ewes treated with CIDR or P cream were compared. In Experiment 1, the rate of estrus induction and the time of estrus onset after device removal were 91.7% and 36.3 +/- 15.7 h in Group A, and 100% and 35.0 +/- 12.6 h in Group B, respectively. There were no significant differences between the devices. The mean plasma P(4) concentration in Group B was significantly (P < 0.01) lower than Group A between day -9 and day -1 (Day 0: the day of device removal). However, no significant differences were found in the mean E(2) concentrations of the two groups after treatment. The mean time of estrus onset in ewes with an observed LH surge and the time of LH surge after treatment were 23.3 +/- 8.7 h and 30.3 +/- 5.0 h for Group A and 27.6 +/- 6.5 and 26.3 +/- 8.0 h for Group B, respectively, and there were no significant differences. However, a significant difference (P < 0.05) was found in the mean time from the time of estrus onset to LH surge between Group A (6.4 +/- 6.7 h) and Group B (-1.3 +/- 4.1 h). In Experiment 2, the conception rates for natural mating, estrus-detected AI, and fixed-time AI were 55.0, 29.4, and 25.0% for Group A and 40.7, 25.0, and 42.1% for Group B, respectively, and there were no significant differences. These results suggest that the effect of induction of estrus and ovulation and the rate of conception after treatment were comparable to CIDR even though the plasma P(4) concentration of the P cream method tended to be low during the insertion period.

A comparative study of induction of estrus and ovulation by three different intravaginal devices in ewes during the non-breeding season.

The aim of the present study was to compare three methods of estrus synchronization in ewes during the non-breeding season. Forty-two ewes were randomly grouped for three treatments with different intravaginal devices for 12 days: Group A) CIDR, Group B) Self-made P sponge, Group C) MAP (medroxyprogesterone acetate) cream sponge. Furthermore, all groups were divided into two treatments with (R) or without ram presence to examine the "ram effect". Blood was collected from all treated ewes, and progesterone (P(4)), estradiol 17-beta (E(2)) and luteinizing hormone (LH) concentrations were measured by enzyme-immunoassay. All ewes showed estrus behavior between Day 0 to 3 after device removal, and the mean onset times of their estrus were 23.0, 33.0 and 21.0 h for Groups AR, BR and CR, respectively. On Day 5 as examined by laparoscopy, the ovulation rates (and number of ovulated ewes) were 1.45 (11/11), 1.25 (12/14) and 1.21 (14/14) for Groups A, B and C, respectively. In Group C, the time to LH surge was significantly (P<0.05) later (32.4 h) than those in Groups A (27.0 h) and B (25.5 h). Ram presence did not affect the number of ovulated ewes, ovulation rate or time to LH surge. The ram introduction group had significantly (P<0.05) lower E(2) concentrations during the period from 0 h to 36 h than the groups without ram presence. These results suggest that the self-made P sponge or MAP cream sponge was effective as well as CIDR, and ram introduction was not necessary, for induction of estrus and ovulation during the non-breeding season.