Effect of a unilateral or bilateral twin embryo distribution on twinning and embryo survival rate in the cow

Uni- and bilateral twin embryo distributions were effected by the transfer of one embryo on Day 7 to the ipsi- or contralateral uterine horn of previously inseminated heifers (123, Exp. 1) or cows (95, Exp. 2). The embryo transfers were surgical in Exp. 1 and non-surgical in Exp. 2. Transferred and native embryos were distinguished by breed. Embryo survival rate was measured in a proportion (N = 40) of the heifers at Day 53 of gestation and in the remainder of the heifers and all of the cows at term. In the heifers (Exp. 1) overall pregnancy rates of 76% and 75% were recorded after uni- and bilateral twin embryo distributions respectively. Twinning rates of 55% and 60% at Day 53 of gestation and 60% and 60% at term were recorded for uni- and bilateral distributions respectively. In the cows (Exp. 2) calving rates of 61% and 63% and twinning rates of 33% and 38% were recorded following uni- and bilateral twin embryo distributions respectively. When the data from both experiments were combined, overall embryo survival rates were similar for both twin embryo distributions although the ipsilateral transfer of an embryo reduced the survival rate of the native embryo. It is concluded that the confinement of two embryos in one uterine horn on or after Day 7 does not depress pregnancy, twinning or overall survival rate to term.     

Embryonic and calving losses in bovine mixed-breed twins induced by transfer of in vitro-produced embryos to bred recipients

One or two in vitro-produced (IVP) Japanese Black (JB) cattle embryos at 8 days after in vitro fertilization were transferred to the contralateral uterine horn of previously bred Japanese Shorthorn (JSH) or JSH-JB cross recipients, and then the occurrence of early embryonic death, abortion during mid- and late gestation, and calving loss were recorded. The survival rate of embryos, including indigenous ones, was not affected by the number of embryos transferred, and a significantly higher twinning rate (68% of pregnant recipients at 80 days after transfer) was achieved when two IVP embryos were transferred, as compared with the rate when one IVP embryo was transferred (24%). In late ET (recipients at 8.5-9.0 days after the onset of oestrus), the embryo survival rate (22%) and the pregnancy rate (42%) at 80 days after ET were significantly lower than those rates in the synchronous ET (recipients at 8.0 days after the onset of oestrus; 47 and 79%, respectively). In the early ET (recipients at 6.0-7.5 days after the onset of oestrus), no significant differences from the synchronous ET were detected in these rates. Twenty-six percent of twin pregnant recipients were aborted during mid- or late-pregnancy, and 39% of twin calves were stillborn. The mean gestation length of the twin-bearing JSH dams (276 days) was 1 week shorter than that of the single-bearing JSH dams, and it was 2 weeks shorter than that of the JB dams bearing a single JB calf derived from the IVP embryos. The longer gestation length of single JB calves derived from IVP embryos resulted in a significantly higher mean birth weight than that of in vivo control calves with the standard length of gestation. In conclusion, the number of embryos to be transferred did not affect the embryo survival rate, and the transfer of two IVP embryos to previously inseminated recipients induced a significantly higher twinning rate during early pregnancy than that of one IVP embryo transfer. The incidence of embryonic losses during early pregnancy increased when Day 8 embryos were transferred to the recipients later in the oestrous cycle (>8.0 days). The results suggested that one cause of the high rate of abortions and stillbirths in twin-bearing dams is the difference in the mean gestation length between the native JSH and JB foetuses derived from transferred IVP embryos.     

Equine sperm-oocyte interaction: results after intraoviductal and intrauterine inseminations of recipients for oocyte transfer.

Insemination of recipients for oocyte transfer and gamete intrafallopian transfer (GIFT) in five experiments were reviewed, and factors that affected pregnancy rates were ascertained. Oocytes were transferred into recipients that were (1) cyclic and ovulated at the approximate time of oocyte transfer, (2) cyclic with aspiration of the preovulatory follicle, and (3) noncyclic and treated with hormones. Recipients were inseminated before, after, or before and after transfer. Intrauterine and intraoviductal inseminations were done.Pregnancy rates were not different between cyclic and noncyclic recipients (8/15, 53% and 37/93, 39%). The highest numerical pregnancy rates resulted when recipients were inseminated with fresh semen from fertile stallions before oocyte transfer or inseminated with cooled transported semen before and after oocyte transfer. Oxytocin was administered to recipients before oocyte transfer when fluid was imaged within the uterus. Administration of oxytocin to recipients at the time of oocyte transfer resulted in significantly higher pregnancy rates than when oxytocin was not administered (17/26, 65% and 28/86, 33%). Intraoviductal and intrauterine inseminations of recipients during oocyte transfer resulted in similar embryo development rates when fresh semen was used (12/22, 55% and 14/26, 55%). However, embryo development rates significantly reduced when frozen (1/21, 5%) versus fresh sperm were inseminated into the oviduct.Results suggest that insemination of a recipient before and after transfer could be beneficial when semen quality is not optimal; however, a single insemination before transfer was adequate when fresh semen from fertile stallions was used. Absence of a preovulatory follicle did not appear to affect pregnancy rates in the present experiments. The transfer of sperm and oocytes (GIFT) into the oviduct was successful and repeatable as an assisted reproductive technique in the equine.     

Development and survival after transfer of cow embryos cultured from 1-2-cells to morulae or blastocysts in rabbit oviducts or in a simple medium with bovine oviduct epithelial cells

This study compares development of bovine 1-2-cell embryos in bovine oviduct epithelial cell co-culture (Group EC) with a glucose- and serum-free simple medium (CZB), or after surgical transfer to ligated oviducts of rabbits (Group RO). Embryos were surgically collected from superovulated donor cows 40-48 h after the beginning of oestrus and randomly distributed between the two groups. Embryos were cultured or incubated for 5 days. In Exp. 1, embryo quality scores and total numbers of cells in the two groups were compared. In Exp. 2, pairs of similarly treated morulae were transferred to each of 10 or 12 recipients in the Groups RO and EC, respectively. Total cell counts per embryo in both groups averaged 52 (P greater than 0.05), and the in-vitro culture system was equivalent to the rabbit oviducts in promoting embryo development for all characteristics measured. Embryo survival, as determined by ultrasound between Days 39 and 43 after oestrus, in 13 ideal recipients was 57% for embryos in Group EC and 58% for embryos Group RO. None of the 9 less desirable recipients was pregnant for either group. These results establish that cattle zygotes can develop to morulae in culture with bovine oviduct epithelial cells in a simple medium and can produce normal pregnancy rates.     

Embryo and foetal loss in beef heifers between day 14 of gestation and full term

Following insemination, reproductive failure in cattle is largely manifested as embryo mortality and is a major source of financial loss to livestock producers. Ongoing studies at this laboratory into factors affecting embryo mortality have facilitated the collection of new data on the extent and timing of embryo and foetal mortality in cattle. Oestrus was synchronised in 158 beef cross heifers and following artificial insemination, embryo and foetal survival rates were determined on days 14 and 30 after insemination and subsequently at calving. Embryo survival rates measured on days 14, 30 and at full term were similar at 68%, 76% and 71.8%, respectively (P0.05). Based on morphological examination, all the 14-day-old embryos recovered were assessed as grade 1. These results provide new information indicating that most embryo losses in heifers have occurred before day 14 after insemination.     

Oocyte transfer in mares with intrauterine or intraoviductal insemination using fresh, cooled, and frozen stallion semen

The objectives were to compare embryo development rates after oocyte transfer with: (1) intrauterine or intraoviductal inseminations of fresh semen versus intraoviductal insemination of frozen semen; (2) intraoviductal versus intrauterine inseminations of cooled semen. In Experiment I, oocytes were transferred into the oviduct, and recipients were inseminated into the uterus with 1 x 10(9) fresh spermatozoa, or into the oviduct with 2 x 10(5) fresh or frozen-thawed spermatozoa. In Experiment II, semen was cooled to 5 degrees C before intrauterine insemination with 2 x 10(9) spermatozoa or intraoviductal inseminations of 2 x 10(5) spermatozoa (deposited with the oocytes). In Experiment I, embryo development rates were similar (P>0.05) for intrauterine versus intraoviductal inseminations when fresh semen was used (8/14, 57% and 9/11, 82%, respectively). However, embryo development rates were lower (P<0.05) when frozen spermatozoa were placed within the oviduct (1/12, 8%). In Experiment II, embryo development rates were higher (P<0.05) when cooled semen was used for intrauterine (19/23, 83%) versus intraoviductal (4/16, 25%) inseminations. We concluded that intraoviductal insemination can be successfully performed using fresh spermatozoa. However, the use of cooled and frozen spermatozoa for intraoviductal inseminations was less successful, and needs further investigation.     

The effect of timing of laparoscopic insemination in superovulated ewes, with or without sedation, on the recovery of embryos, their stage of development and subsequent viability

Twenty ewes were used as donors in a 2x2 factorial design experiment to investigate the effects of two different insemination times (48 vs 60 h after pessary withdrawal), with or without sedation, on the ovum recovery rate 5 d after insemination, the proportion of transferable embryos recovered, and the subsequent survival rate of embryos transferred to recipients. The ovum recovery rate following intauterine insemination at 48 h after progestagen pessary withdrawal was 63.8 and 53.4% for sedated and nonsedated control ewes, respectively. Following intrauterine insemination at 60 h the corresponding values for sedated and control ewes were 72.6 and 73.9%, respectively. The proportion of transferable quality embryos recovered was not affected by sedation but was improved by insemination at 48 h rather than 60 h after pessary withdrawal (100 vs 35.4%). Embryo survival following laparoscopic transfer to recipients from donor ewes inseminated at 48 h, with or without sedation was 38.8% (7 18 ) and 50% (7 14 ), respectively. Following intrauterine insemination of the donors at 60 h, the survival rate in recipients was reduced for embryos transferred from both the sedated and control ewes to 6.25% (1 16 ) and 36.3% (4 11 ). It is concluded that delaying the timing of intrauterine insemination relative to pessary withdrawal and the use of acepromazine maleate as a sedative at the time of insemination are deleterious to embryo development and subsequent viability.     

The usefulness of a single measurement of insulin-like growth factor-1 as a predictor of embryo yield and pregnancy rates in a bovine MOET program

The objective was to determine if a single measurement of plasma insulin-like growth factor-1 (IGF-1) could predict the number of viable embryos obtained from donors and the likelihood of pregnancy in recipients in multiple ovulation and embryo transfer (MOET) programs in cattle. The embryo yields from 101 embryo recoveries were examined in maiden Holstein heifers (n=75) and multiparous Holstein cows (lactating cows n=20, dry cows n=6). Donors were super stimulated with FSH and embryo recovery was done non-surgically 7 days after artificial insemination. Embryos were classified according to the IETS criteria. Pregnancy rates in 100 maiden Holstein heifer recipients were analysed. Recipients were on day 7+/-1 of the estrous cycle at transfer. Pregnancy diagnosis was carried out at day 30 (PD 30) and rechecked at day 60 (PD 60) after transfer. Blood samples from coccygeal vessels taken at the time of embryo recovery (donors) and transfer (recipients) were analysed for IGF-1, insulin, beta-hydroxybutyrate (beta-OHB), non-esterified fatty acids (NEFA), urea and cholesterol. There was a negative correlation between the number of viable embryos and insulin (r=-0.33, P=0.025) in donor heifers. In donor cows, the number of viable embryos was correlated with IGF-1 (r=0.43, P=0.028) and cholesterol (r=-0.43, P=0.027). In recipients, PD30 and PD 60 were not affected by any of the circulating parameters analysed. Insulin, IGF-1 and cholesterol only explained 8.9, 13.9 and 15.8% of the variation in the production of viable embryos, respectively. Several factors affect MOET programs and under the circumstances of the present study the usefulness of hormonal and metabolic profiles as predictors of the outcome of this biotechnology was limited.     

Laparoscopic transfer of ovine and cervine embryos using the transpic technique

A transpic technique was developed to transfer embryos to 352 sheep and 4 deer recipients using a laparoscope, a modified pair of Allis forceps and a modified Cassou aspic normally used for laparoscopic uterine insemination. The overall proportion of uncomplicated transfers in Experiment 1 in 216 recipient ewes was 90.7% (range between groups 80 to 100%), 3.7% of the transfers were presumed to be loss of embryos during expulsion from the transpic, and 5.6% were apparent transfers into the uterine wall. In Experiment 2,83% of transfers into 136 ewe recipients were uncomplicated, 5% were presumed to be loss of embryos during expulsion, 1% was apparent transfer into the uterine wall, and 11% involved 2 attempts at transfer. Only 34% of 116 recipients receiving low-quality frozen-thawed embryos were pregnant and 24% of the 226 embryos survived to term. In contrast, high pregnancy rates (>80%) and embryo survival rates (>70%) were achieved following uncomplicated and twice attempted transfers of fresh embryos. Pregnancy rates and embryo survival rates were low (<2%) following the presumed loss of embryos during expulsion and apparent transfers into the uterine wall. All 4 deer transfers were uncomplicated and 2 2 good-quality embryos survived to term compared with 0 2 low-medium quality embryos. The transpic technique is a moderately invasive technique which permits fast (15 to 20/h) and reliable transfer of embryos in small ruminants. With appropriate care, nearly all of the embryos can be correctly placed in the uterus, and high pregnancy rates and embryo survival rates can be achieved using this technique.     

Transfer of vitrified blastocysts from one or two superovulated Large White Hyperprolific donors to Meishan recipients: reproductive parameters at Day 30 of pregnancy

The present study was designed to determine the effect of pooling embryos from two donors on the reproductive success of transfer of vitrified/warmed porcine blastocysts. Intact blastocysts were collected from superovulated Large White Hyperprolific gilts (n = 24) on Days 5-5.5 after artificial insemination. Embryos were recovered by flushing the uterine horns, and unhatched blastocysts were selected. Vitrification and warming were performed as described by Berthelot et al. [Cryobiology 41(2000) 116]. To evaluate in vitro development, 37 vitrified/warmed blastocysts were cultured, non-vitrified embryos (n = 48) were used as controls. Embryo transfers were conducted in asynchronous (-24 h) Meishan gilts (n = 20). Twenty vitrified/warmed blastocysts were surgically transferred into one uterine horn. Ten recipients received embryos from one donor (Group 1) and the other 10 transfers were performed with mixed embryos from two donors (Group 2). Pregnancy was assessed ultrasonographically at Day 25 after estrus and recipients were slaughtered at Day 30 after transfer. In vitro survival rate of the vitrified/warmed blastocysts was lower (P < 0.01) than that from control embryos (73.0% versus 93.7%). The pregnancy rate for Group 1 (70%) was not different (P > 0.05) than that from Group 2 (90%). No significant differences were detected between Groups 1 and 2 for in vivo embryo development (number fetuses/transferred embryos in pregnant recipients) or in vivo embryo survival (number viable fetuses/transferred embryos in pregnant recipients). However, the in vivo efficiency (number viable fetuses/total transferred embryos) was higher (P < 0.05) when transfers were performed with embryos from two donors (19.5% versus 30.5%). These results indicate that pooling embryos from two donors increases the in vivo efficiency after transfer of vitrified/warmed porcine blastocysts.     

Successful embryo transfer in the silver fox (Vulpes vulpes)

Surgical embryo transfer in the silver fox was investigated as part of a larger project concerning the conservation of endangered canine species using modern artificial reproduction techniques with the farmed fox as a model. The animals were chosen on the basis of synchrony in natural oestrus. The timing of ovulation and artificial insemination was determined by measuring electrical resistance in the vagina. Twenty-nine embryos were flushed from eight humanely killed donor females and transferred surgically into the uteri of eight recipients. One recipient female gave birth to two male pups 47 days after the transfer of four expanded blastocysts and one embryo at the 16-cell stage derived from a donor female flushed 10 days after artificial insemination.     

Normal calves from transfer of biopsied, sexed and vitrified IVP bovine embryos

Data on biopsied, sexed and cryopreserved in vitro produced (IVP) bovine embryos, and their in vivo developmental competence are very limited. Two preliminary studies were conducted before the primary study. In Experiment 1, post-thaw in vitro developmental competence of biopsied and vitrified IVP embryos was evaluated using re-expansion as an endpoint. In Experiment 2, the pregnancy rates of biopsied fresh, frozen or vitrified embryos following single embryo transfer were compared. Since vitrified embryos resulted in a higher pregnancy rate than frozen-thawed embryos, in the primary study (Experiment 3), all IVP embryos were vitrified following biopsy and sexing (by DNA fingerprinting). In Experiment 3, we compared pregnancy initiation and calving results of heifers in the following treatments: 1) artificial insemination (AI); 2) AI plus contralateral transfer of a single embryo (AI + SET); 3) ipsilateral transfer of single embryo (SET); or 4) bilateral transfer of two embryos (DET). Birth weights, gestation lengths and dystocia scores were recorded. In Experiment 1, post-thaw re-expansion rate of biopsied and vitrified embryos was 85% (70/82). In Experiment 2, pregnancy rates (90 d) were 44% (7/16), 23% (3/13), and 50% (7/14) for vitrified, frozen and fresh embryos, respectively (P < 0.10). In Experiment 3, pregnancy rates of AI and SET were 65% (20/31) and 40% (16/40), respectively (P < 0.05). The pregnancy rate of AI + SET was 75% (27/36) with 11 carrying twins, and the pregnancy rate of DET was 72% (26/36) with 10 carrying twins. All AI fetuses were carried to term, but only half the SET fetuses were carried to term. Similar calving rates were observed in the AI + SET and DET groups, 76 and 70%, respectively, of those pregnant at Day 40. Mean birth weight, dystocia score and gestation length of AI calves were not different from those of SET calves. Mean birth weight and dystocia score of single-born calves were greater than those of twin born calves (P < 0.05). These data demonstrate that biopsied IVP bovine embryos can be successfully cryopreserved by vitrification and following post-thaw embryo transfer, acceptable rates of offspring with normal birth weights can be obtained without major calving difficulties.     

Embryo survival in pseudopregnant and in pregnant but genetically semi-sterile recipients after nonsurgical embryo transfer in the mouse

A new nonsurgical embryo transfer technique was used in the mouse that yielded survival rates of between 40 and 70% depending on embryo stage and, possibly, on the degree of synchrony between the embryo and recipient. Three variables were tested using this embryo transfer technique: a) pseudopregnant recipients vs pregnant but genetically semi-sterile recipients, b) embryos resulting from superovulation vs embryos from natural ovulation, and c) 12-hour vs 24-hour asynchrony between donors and recipients. None of these variables significantly affected the pregnancy rate or the percentage of transferred embryos developing to term. The pregnancy rates were between 77 and 90% in 6 experimental groups of 8 to 13 females. Survival rates were between 41 and 63% when all recipients were considered and between 53 and 68% when only the pregnant recipients were included. The embryo transfer procedure influenced litter size composition of the endogenous conceptuses of the semi-sterile recipients. Too many females were devoid of these. Recipients of expanded blastocysts had significantly better transfer results than recipients that also received morulae and early blastocysts. It was concluded that the transfer success rates were influenced by the recipients and possibly by their preparation for transfer.     

Donor and recipient ewe factors affecting in vitro development and post-transfer survival of cultured sheep embryos

Donor and recipient factors were assessed during development of embryos following superovulation, collection at the pronuclear and two-cell stage, culture in Synthetic Oviduct Fluid medium for 5 days and twin transfer into synchronised recipients to elucidate what factors affect embryo development and post-transfer survival. In particular, the administration of exogenous progesterone to recipients using an intravaginal CIDR^TM device immediately following embryo transfer was investigated.

From 138 embryos collected from 30 donor ewes, 75% (103) were of transferable quality following culture, of which 100 were transferred to 50 recipients. There was significant variation (P < 0.001) in embryo development to the blastocyst stage between different donor ewes, but this was not related to the donor ovulation rate. At ultrasound sonography (approximately Day 60 of pregnancy), 58% of recipients were pregnant and 42% embryos had survived. Donor ovulation rate was related to embryo survival (P < 0.05) after transfer; the survival rate of embryos from ewes with high ovulation rates was lower than that of embryos from ewes with low ovulation rates. Exogenous progesterone supplementation following transfer did not affect embryo survival, rate of embryo development or plasma progesterone levels. In general, the results from this study suggest that factors other than efficacy of embryo culture can affect the outcome of embryo survival following transfer and that, where possible, these factors should be considered and balanced in experimental designs.     

The effect of in vitro treatment of bovine embryos with IGF-1 on subsequent development in utero to Day 14 of gestation

Culture of bovine embryos with insulin-like growth factor-1 (IGF-1) can improve development to the blastocyst stage and embryo survival following transfer to heat-stressed, lactating dairy cows. Two experiments were conducted to determine whether IGF-1 could improve embryo survival and development at Day 14 after ovulation. In Experiment 1, non-lactating Holstein cows (n=58) were selected as recipients following synchronization for timed-embryo transfer. Embryos were produced in vitro and cultured with or without 100ng/mL IGF-1. At Day 7 after expected ovulation (Day 0), groups of 7-12 embryos were randomly transferred to each recipient. Embryos were recovered at Day 14. Embryo length and the presence or absence of an embryonic disc was recorded. Recovered embryos were cultured individually for 24h to determine interferon-tau (IFN-tau) secretion. There was no effect of IGF-1 on embryo recovery rate, embryo length or IFN-tau secretion. In Experiment 2, non-lactating (n=56) and lactating (n=35) Holstein cows were selected as recipients following synchronization for timed-embryo transfer. Embryos were produced as described in Experiment 1. At Day 7 after expected ovulation (Day 0), a single embryo was randomly transferred to each recipient. Embryos were recovered at Day 14. Embryo length and IFN-tau secretion were determined as in Experiment 1. Recovery rate at Day 14 tended (P=0.1) to be higher for recipients that received IGF-1 treated embryos compared to control embryos (43.2% versus 26.1%, respectively). There was no effect of IGF-1 on embryo length or IFN-tau secretion. In conclusion, results suggest that exposure to IGF-1 through Days 7-8 of development does not enhance capacity of embryos to prevent luteolysis. Results of the single embryo-transfer experiment suggested that IGF-1 treatment might affect embryo survival post-transfer as early as Day 14 after ovulation. Further experimentation is warranted to verify this finding.     

Embryos produced from fertilization with bovine viral diarrhea virus (BVDV)-infected semen and the risk of disease transmission to embryo transfer (ET) recipients and offspring

Bovine diarrhea virus (BVDV) causes a variety of economically important enteric and infertility problems in cattle. For that reason, several countries have eradicated the disease, and some others have schemes in progress to achieve freedom. Although there is a considerable amount of information about the risk of BVDV transmission through contaminated semen used for artificial insemination (AI), there is no evidence to indicate whether the resulting embryos, when used for embryo transfer, can lead to the transmission of BVDV to recipients or offspring. For this experiment, semen from a bull persistently infected with BVDV (10⁵ 50% tissue culture infective doses/mL NY strain) was used for insemination (two times at estrus) of BVDV-seronegative, superovulated cows (N = 35). Embryos were collected 7 days after insemination and subsequently were washed according to the International Embryo Transfer Society recommendations or left unwashed. Out of 302 collected oocytes and embryos, 173 (57%) were fertilized and the remaining 129 (43%) had degenerated. Infectious BVDV was detected in 24% (17/71) of unwashed and 10% (8/77) of washed embryos, and in all (N = 11) follicular fluid samples, oviductal epithelial cells, endometrium, and corpora lutea tissues as determined by the virus isolation test. After transfer of 39 washed embryos to 27 BVDV-seronegative recipients, 12 (44%) cows became pregnant and 17 calves free of BVDV and BVDV antibodies, including five sets of twins, were born. After embryo transfer, all pregnant and nonpregnant recipients remained free of BVDV and antibodies. In conclusion, results herein suggest that BVDV can be transmitted by AI resulting in the production of some proportion of contaminated embryos. However, it appears that such embryos, when washed according to International Embryo Transfer Society and the World Organization for Animal Health guidelines do not cause BVDV transmission to recipients or their offspring.     

Fertilization and ovum recovery rates in superovulated ewes following cervical insemination or laparoscopic intrauterine insemination at different times after progestagen withdrawal and in one or both uterine horns

In Exp. 1, 40 ewes were used in a 2 x 2 factorial design to investigate the effects of intrauterine versus cervical insemination and superovulation using pig FSH or PMSG and GnRH on egg recovery and fertilization rate. Cervical inseminations were carried out at 48 and 60 h (N = 20 ewes) and intrauterine insemination at 52 h (N = 20 ewes) after progestagen pessary withdrawal. Eggs were recovered on Day 3 of the oestrous cycle. Ovulation, egg recovery and fertilization rates were independent of the type of superovulatory hormone used. Fertilization rate was high irrespective of insemination site but intrauterine insemination at 52 h was associated with a significant (P less than 0.01) decrease in egg recovery of over 40% compared with cervically inseminated ewes. In Exp. 2 ewes were inseminated at 36 (N = 5), 48 (N = 6) or 60 (N = 6) h after pessary withdrawal to determine the optimum intrauterine insemination time to maximize both fertilization rate and egg recovery. Egg recovery per ewe flushed was 23, 59 and 67% after intrauterine insemination at 36, 48 and 60 h respectively. Correspondingly, 0, 85 and 100% of the eggs recovered were fertilized. The results of Exps 1 and 2 suggest that when intrauterine insemination occurs before or during ovulation it interferes with oocyte collection by the fimbria. In Exp. 3 egg recovery and fertilization rates were determined after cervical insemination at 48 and 60 h (N = 8) or intrauterine insemination at 48 (N = 9) or 60 (N = 8) h after progestagen withdrawal. Ewes in the last two groups were subdivided and inseminated unilaterally or bilaterally. Egg recovery was high after cervical insemination (95%) but only 36% of these eggs were fertilized. Unilateral intrauterine insemination was as effective as bilateral in ensuring high fertilization rates (100 versus 97%). Intrauterine insemination at 48 h compared with 60 h resulted in a significantly lower (P less than 0.05) percentage of eggs recovered (42 versus 90% respectively). However, reducing the degree of interference by adopting unilateral rather than bilateral insemination did not alleviate the detrimental effects of the 48-h insemination time on egg recovery. From these results we advocate the adoption of intrauterine insemination at 60 h after progestagen withdrawal to maximize fertilization rate and egg recovery in superovulated ewes.     

Bovine somatotropin increases embryonic development in superovulated cows and improves post-transfer pregnancy rates when given to lactating recipient cows

Previous studies indicated that the use of bovine somatotropin (bST) in concurrence with a timed artificial insemination (TAI) protocol increased pregnancy rates. However, the mechanisms for such a bST effect on fertility were not clear. Objectives of this study were to determine the effects of bST on fertilization and early embryonic development after cows received a superovulation treatment, test whether embryos recovered from bST-treated cows were more likely to survive after transfer to recipients, and evaluate whether treatment of recipient cows with bST affects pregnancy rates. Lactating (n = 8) and nonlactating (n = 4) Holstein donor cows were superovulated, inseminated at detected estrus and assigned to a nontreated control group or to a treatment group receiving a single injection of bST (500 mg, sc) at insemination. Embryos were nonsurgically flushed 7 days after AI and frozen in ethylene glycol for direct transfer. Embryos derived from bST-treated (bST-embryos) or control (control-embryos) donors were transferred to lactating Holstein recipient cows that received either bST treatment 1 day after estrus (500 mg, sc; bST-recipients) or were untreated controls (control-recipients). Thus, there were four treatment groups: control-embryos/control-recipients (n = 43), bST-embryos/control-recipients (n = 41), control-embryos/bST-recipients (n = 37), and bST-embryos/bST-recipients (n = 60). Pregnancy was determined by palpation per rectum 33-43 days after embryo transfer. Unfertilized ova per flush was less for bST than for control (1.0 +/- 0.9 < 3.7 +/- 0.9; P < 0.04). Percentage of transferable embryos was greater for bST than for control (77.2% > 56.4%; P < 0.01). Number of blastocysts per flush was greater for bST than for control (2.4 +/- 0.7 > 0.4 +/- 0.7; P < 0.04). Pregnancy rates following embryo transfer were 25.6% for control-recipient/control-embryo, 43.2% for bST-recipient/control-embryo, 56.1% for control-recipient/bST-embryo, and 43.3% for bST-recipient/bST-embryo. Transfer of bST-embryos increased pregnancy rates compared with transfer of control-embryos (P < 0.04). An interaction between embryo and recipient treatments (P < 0.05) indicated that treatment of recipient cows with bST increased pregnancy rates as compared to control-recipients that received a control-embryo. However, there was no additive effect when bST-recipients received a bST-embryo. Administration of bST at AI decreased the number of unfertilized ova, increased the percentage of transferable embryos, and stimulated embryonic development to the blastocyst stage. Moreover, bST affected both early embryonic development and recipient components to increase pregnancy rates following embryo transfer.     

The effect of recipient age on the success of embryo transfer in mice

Eight-hundred-sixteen morulae and blastocysts were transferred to the uteri of 102 recipients of various ages on Day 3 or 4 of pseudopregnancy (Day 1 is the day of the vaginal plug). Day 3 recipients had significantly higher pregnancy rates and embryo survival rates than Day 4 recipients. Recipient age had little effect on pregnancy rates, but had a significant effect on embryo survival in Day 3 recipients. Day 3 recipients of 11-13 weeks of age had the highest pregnancy rate (100%) and embryo survival (75%). The results suggest that recipient age should be considered an important factor in embryo transfer experiments.     

Effect of transfer of one or two in vitro-produced embryos and post-transfer administration of gonadotropin releasing hormone on pregnancy rates of heat-stressed dairy cattle

Pregnancy rates following transfer of an in vitro-produced (IVP) embryo are often lower than those obtained following transfer of an embryo produced by superovulation. The purpose of the current pair of experiments was to examine two strategies for increasing pregnancy rates in heat stressed, dairy recipients receiving an IVP embryo. One method was to transfer two embryos into the uterine horn ipsilateral to the CL, whereas the other method involved injection of GnRH at Day 11 after the anticipated day of ovulation. In Experiment 1, 32 virgin crossbred heifers and 26 lactating crossbred cows were prepared for timed embryo transfer by being subjected to a timed ovulation protocol. Those having a palpable CL were randomly selected to receive one (n = 31 recipients) or two (n = 27 recipients) embryos on Day 7 after anticipated ovulation. At Day 64 of gestation, the pregnancy rate tended to be higher (P = 0.07) for cows than for heifers. Heifers that received one embryo tended to have a higher pregnancy rate than those that received two embryos (41% versus 20%, respectively) while there was no difference in pregnancy rate for cows that received one or two embryos (57% versus 50%, respectively). Pregnancy loss between Day 64 and 127 only occurred for cows that received two embryos (pregnancy rate at Day 127=17%). Between Day 127 and term, one animal (a cow with a single embryo) lost its pregnancy. There was no difference in pregnancy rates at Day 127 or calving rates between cows and heifers, but females that received two embryos had lower Day-127 pregnancy rates and calving rates than females that received one embryo (P < 0.03). Of the females receiving two embryos that calved, 2 of 5 gave birth to twins. For Experiment 2, 87 multiparous, late lactation, nonpregnant Holstein cows were synchronized for timed embryo transfer as in Experiment 1. Cows received a single embryo in the uterine horn ipsilateral to the ovary containing the CL and received either 100 microg GnRH or vehicle at Day 11 after anticipated ovulation (i.e. 4 days after embryo transfer). There was no difference in pregnancy rate for cows that received the GnRH or vehicle treatment (18% versus 17%, respectively). In conclusion, neither unilateral transfer of two embryos nor administration of GnRH at Day 11 after anticipated ovulation improved pregnancy rates of dairy cattle exposed to heat stress.