Reproduction in the ewe
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The literature pertaining to the reproduction in the ewe was reviewed. Where the available literature on the ewe was deficient, references were drawn from other species and extrapolated to the ewe. Details are given of the events occurring in the ovary in preparation for the release of the gametes, their transport, fertilization, early embryonic development and uterine capacity. Also reviewed are the hormones, progesterone and the gonadotropins, their measurement in the systemic circulation and their use in the manipulation of the oestrous cycle of the ewe. 2. Materials and methods used throughout this study were discussed in Chapter 2. The reader is referred to this chapter in all the subsequent chapters. 3. In an attempt to establish the most efficient type of teaser, two vasectomised rams, two testosterone-treated ewes and two testosteronetreated wethers were compared with two rams on their mating behaviour and their ability to detect ewes in oestrus. No significant difference was observed between them on their ability as teasers although during a ten minutes test period, testosterone-treated ewes stayed longer with and mounted the oestrous ewes more times (P < 0.01) than the others. It was concluded that testosterone-treated ewes were the best type of teasers because of thei.r availability and ease of handling. 4. Four experiments were carried out to establish the best superovulatory and oestrus synchronization regimen for donor Merino ewes. In the first experiment ewes were superovulated with 1000 IU, 1500 IU or 2000 IU of pregnant mare serum gonadotropin (PMSG). The ovulation rates, the embryo recovery and fertilization rates were similar in the three groups. However, ewes treated with 2000 IU had significantly more unovulated follicles than the others (P < 0.05). The second experiment was of a factorial design, involving treatment with three dosages of PMSG for superovulation and either the prostaglandin F2a analogue cloprostenol, or progestagen sponges for oestrus synchronization. There was no difference in ovulation and ova recovery rates between treatments. However, at all PMSG dosages, a significantly higher fertility rate (P < 0.01) was observed in the ova collected after progestagen synchronization than after prostaglandin synchronization. The time to subsequent oestrus was significantly longer (P < 0.05) in the prostaglandin synchronized ewes than the others, but the pattern of plasma progesterone levels was the same in the two groups. In the second part of this experiment repeated superovulation for three times, at six week intervals, was carried out in 12 ewes. Although extensive post-surgical adhesions occurred after three operations, the ovarian response was not affected. In the third experiment, ewes were superovulated with 1000 IU PMSG at different days of the luteal phase of their oestrous cycle, followed two days later by an intramuscular cloprostenol injection. Ovarian activity and ova recovery rate increased with the advancing luteal phase, but the percentage of ova fertilized was reduced during the similar period. In the fourth experiment, ewes were superovulated during the middle of their luteal phase (days 7 to 11) of the oestrous cycle) and their oestrus synchronized using either progestagen sponges or cloprostenol. Ovarian response was similar in all the groups, but the two progestagen synchronized groups had better oestrus synchronization and a significantly higher (P < 0.05) percentage of ova fertilized than the others. It was concluded from this chapter that in Merino ewes:- a) 1000 IU PMSG is the most efficient dosage for superovulation compared with higher dosages. b) Although superovulation is equally effective on any day of the luteal phase of the oestrous cycle, it should be carried out preferably late than early in the cycle. c) Progestagen synchronization is preferable to cloprostenol synchronization in superovulated ewes. 5. Analysis of the results obtained from 80 Merino ewes superovulated with 1000 IU PMSG each, showed that 74 ewes ovulated with a mean of 5.9 ovulations. Ova recovery rate was 68.6% and 52.1% of the ova recovered were fertilized. There was no difference in the recovery rates when embryos were collected on either days 4, 6 or 8 of the oestrous cycle. Of the ewes that ovulated, 54% had all the ova fertilized, 34% had all the ova unfertilized and 12% had some ova fertilized and others unfertilized. 6. An experiment to establish what hormone parameters in the systemic circulation may be monitored to help predict superovulatory response in donor ewes showed that the rate of progesterone increase from day a to day 9 of the oestrous cycle and the magnitude and length of time from luteinizing hormone (LH) peak to basal levels may be the best criteria. In the second experiment, a study of gonadotropin receptor levels in unilaterally ovariectomised ewes, which were subsequently superovulated, showed that the differences in superovulatory response at the same PMSG dosages may be due to differences in FSH receptor concentrations. 7. Attempts to develop alternative methods to the standard surgical procedure for embryo collection from sheep failed. Indwelling uterine balloon catheters failed because their presence blocked ovulation by preventing the LH surge and also elicited an immunological reaction in the uterus. Attempts to fertilize follicular oocytes in the oviducts of catheterized ewes failed and the oocyte recovery rate was reduced considerably. 8. Ewes responded to PMSG superovulation during the post partum period, but although ova were recovered, none of them was fertilized. Reduction of daylight length during the non-breeding season did not change the oestrus incidence in Merino ewes, but superovulatory response was significantly improved (P < 0.01) and more embryos were collected in ewes with reduced daylight length compared with the controls. Superovulation of the neonatal and prepubertal lambs showed that response could only be achieved after 4 weeks of age and fertilization occurred when these ewe lambs were inseminated. Due to the immature reproductive tracts, it was not possible to flush the uteri and the oviducts of these lambs for embryo collection in situ. Successful in vitro fertilization was achieved using follicular oocytes and Dulbecco's phosphate buffer as the oocyte collection and incubation medium, but development of the fertilized ova did not proceed beyond the first cleavage stage. It was concluded that the reduction of daylight length in summer was the ideal method of improving superovulatory response in Merino ewes and that there is optimism for successful in vitro fertilization. However, further experimentation is necessary. 9. Uterine capacity was studied in Merino ewes by introducing one, two, three and five embryos into either the ipsilateral or contralateral uterine horns. Pre-natal losses were highest the first 20 days after both the contralateral and ipsilateral transfers. Fifty-five lambs (42%) were born after 131 embryos were transferred into ipsilateral uterine horns compared with 11 lambs (12%) after 89 embryos were transferred into the contralateral uterine horns. The highest percentage of lambs born (83%) occurred after single embryos were transferred and the lowest (28%) after five embryos were transferred per recipient ewe. Superovulation of the recipient ewes before embryo transfer did not improve the conception rates. In triplet transfers conception rate was significantly improved when two embryos were deposited in one uterine horn and one in the other, than when all the three were deposited in either the contralateral horn (P < 0.01) or ipsilateral horn (P < 0.05). It was concluded that although the number of lambs born per ewe can be increased through embryo transfer, four lambs seemed to be the maximum that the uterus would accommodate pt a time in anyone ewe of this breed. Plasma progesterone levels were measured on day 8 of the oestrous cycle in all the recipient ewes. One group of recipient ewes received embryos after superovulation to increase the number of corpora Zutea and therefore the luteal support. The plasma progesterone levels on day 8, were similar between and within the groups in all recipient ewes. 10. A technique was developed for transferring sheep embryos through a laparoscope introduced at the paralumbar fossa of the recipient ewe. This was achieved with ewes in a standing position under mild sedation and local anaesthesia. Two of the three ewes used in this procedure became pregnant. In another experiment, day 4 embryos were transferred to recipient ewes through cannulated oviducts. Three of the five recipient ewes became pregnant and two of them subsequently lambed. One of these ewes lambed the following year after another transfer through the same cannulae. It was concluded however that further investigation is needed to perfect these techniques for practical application.