122. PRODUCTION OF EMBRYOS IN SUPEROVULATED EWES USING FROZEN - THAWED, SEX-SORTED AND REFROZEN - THAWED SPERM

2010 ◽  
Vol 22 (9) ◽  
pp. 40
Author(s):  
K. H. Beilby ◽  
Y. B. Kaurivi ◽  
W. M. C. Maxwell ◽  
G. Evans ◽  
S. P. De Graaf ◽  
...  

Flow cytometric sex-sorting of sperm that has previously been cryopreserved allows sex-sorting technology to be applied more widely. While offspring have been produced following artificial insemination of synchronised ewes with frozen-thawed, sex-sorted and refrozen-thawed (FSF) sperm (1), the fertility of FSF-sperm in superovulated ewes has not been reported. The aim of this study was to determine the effect of cryopreservation prior to sperm sex-sorting and freezing on embryo production in superovulated ewes. Several ejaculates from 2 rams were either frozen-thawed, then sex-sorted and re-frozen (FSF X- and Y-chromosome enriched sperm), or immediately sex-sorted before freezing (SF X- and Y-chromosome enriched sperm). A portion of each ejaculate was also cryopreserved without sex-sorting (control). Thirty-one ewes were superovulated and inseminated (15 ×106 sperm per insemination dose) with either SF X, SF Y, FSF X, FSF Y or control sperm as previously described (2). Embryos were recovered 6 d after insemination and assessed. The superovulatory response (mean number of corpora lutea per ewe: 11.8 ± 1.3) and the embryo recovery rates (72.0 ± 5.9%) did not differ significantly among the groups. The fertilisation rates tended to differ (P=0.068) as a result of sperm treatment (control: 33%; SF: 54%; FSF: 18%) and were unaffected by sperm sex (X: 33%; Y: 37%). Of the embryos that were recovered, those derived from FSF-sperm were predominantly at the blastocyst stage (65%), whereas those derived from SF-sperm were evenly distributed among the blastocyst (30%), morula (38%) and arrested (32%) stages, suggesting that fertilisation lifespan of SF-sperm was greater than that of FSF-sperm. A greater proportion of embryos derived from Y-sperm were at the blastocyst stage compared with embryos derived from X-sperm (53% vs. 26%; P < 0.05). This study is the first to demonstrate that FSF-sperm is capable of fertilising oocytes of superovulated ewes. (1) de Graaf et al (2007) Theriogenology 67: 391–8.(2) de Graaf et al (2007) Theriogenology 67: 550–5.

2013 ◽  
Vol 25 (1) ◽  
pp. 268
Author(s):  
M. Kaymaz ◽  
A. R. Agaoglu ◽  
K. Karakas ◽  
I. Pir Yagci ◽  
O. Korkmaz Agaoglu ◽  
...  

The Angora, Kilis, Honamli, Hair, and Norduz are native goat breeds in Turkey and are currently in danger of extinction. This study aimed to assess the efficacy of the repeated administration of a superovulatory (SOV) protocol for in vivo embryo production in these breeds. A total of 14 Angora, 15 Kilis, 10 Honamli, 10 Hair, and 9 Norduz goats were used in this work. The synchronization procedure was started on Day 5 after visible oestrus by using controlled internal drug release dispensers (CIDR®) for 11 days. Administration of FSH (Folltropin®) began on Day 9 (twice daily) and continued for 3 days (total dose: 200 mg; 50 mg × 2.30 mg × 2.20 mg × 2). A dose of prostaglandin F2α (1.6 mg; Dalmazin®) was injected together with first FSH injection. Gonadotropin-releasing hormone (Receptal®: 100 µg) was injected 6 h before mating. All goats in oestrus were naturally mated twice a day for 3 days. Ovarian examination (number of corpora lutea) and embryo recovery were performed by laparotomy on Day 6 after CIDR® withdrawal. Each uterine horn was flushed, and the embryos were recovered and counted. To avoid intra-abdominal adhesions, a 2.5% heparin solution was used during flushing. The SOV procedure was repeated once per year during the breeding season (2009 to 2011). Fertilization and recovery rates were calculated. Differences in the SOV response and embryo yields were evaluated by Friedman’s test. In Hair goats, the number of corpora lutea decreased significantly (P < 0.05) during the third SOV cycle (12.7 ± 6.2, 14.0 ± 9.1, and 6.8 ± 5.6, respectively, for cycles 1, 2, and 3), whereas no effect of the cycle was observed in the remainder of breeds. The number of expanded blastocysts increased considerably during the third cycle in Angora (0, 0.2 ± 0.8, and 1.4 ± 2.9, respectively, for cycles 1, 2, and 3), Kilis (0.2 ± 0.4, 0.3 ± 1.3, and 4.2 ± 5.0), and Honamli (0, 1.3 ± 1.7, and 3.6 ± 4.5) goats, whereas a significant decrease was observed in Norduz goats (2.4 ± 5.0, 1.8 ± 2.0, and 0.1 ± 0.3; P < 0.05). The mean numbers of unfertilized oocytes were found to be significantly increased in Angora (0.4 ± 1.6, 0, and 2.1 ± 4.1, respectively, for cycles 1, 2, and 3), Kilis (0, 1.3 ± 3.9, and 3.1 ± 5.2), and Honamli (0, 4.9 ± 5.2, and 4.5 ± 7.8) goats (P < 0.05). As a result, fertilization rates (%) showed a decrease in Angora (50, 100, 24.5, respectively, for cycles 1, 2, and 3) and Honamli (100, 42.5, and 56.3) goats (P < 0.05), whereas recovery rates showed no difference among the different breeds. The methodology presented in this study was found to be an efficient technique for superovulation of the Angora, Kilis, and Honamli goats. Because Hair and Norduz are relatively small breeds, the dosage of FSH might have had a negative effect on the superovulation and embryo yield. Additionally, the use of intra-abdominal washing solutions for preventing adhesions as observed in previous works (data not shown) is believed to have a positive effect on achieving high levels of efficiency in in vivo embryo production.


Reproduction ◽  
2007 ◽  
Vol 134 (2) ◽  
pp. 233-239 ◽  
Author(s):  
Pritpal S Malhi ◽  
Gregg P Adams ◽  
Reuben J Mapletoft ◽  
Jaswant Singh

The study was designed to test the hypothesis that aging in cattle is associated with reduced developmental competence of oocytes. The hypothesis was tested by comparing embryo production and pregnancy rates between 13- to 16-year-old cows (n = 6 in Year 1 and n = 9 in Year 2) and their 3- to 6-year-old young daughters (n = 8 in Year 1 and n = 9 in Year 2) after superovulation and transfer of embryos into an unrelated group of young recipients. Embryos were transferred into 2- to 5-year-old recipient cows (n = 99) as singletons (n = 45) or in pairs (n = 54 pairs). Embryo survival in recipients was determined by ultrasonography and by the number of calves born. Between old versus young cows, the number of ovulations (31 ± 4 vs 38 ± 3; P = 0.2) and the number of corpora lutea (25 ± 3 vs 29 ± 2; P = 0.3) did not differ, but fewer (P = 0.04) embryos were recovered from old cows (6 ± 2) than their daughters (12 ± 2). A higher proportion (P < 0.0001) of unfertilized oocytes/uncleaved zygotes were recovered from old cows (222/312, 71%) than their daughters (119/316, 38%). Among the embryos recovered, the proportion of International Embryo Transfer Society Grades 1–2 embryos was similar (P = 0.9) between old (59/90, 66%) and young cows (130/194, 67%). The survival of embryos after transfer into recipients, and the proportion of calves born were also similar between old and young cows. In conclusion, recovery of fewer embryos and a greater proportion of unfertilized oocytes/uncleaved zygotes suggest reduced developmental competence of oocytes from old cows, but there was no difference between age groups in embryo survival after the morula/blastocyst stage.


2018 ◽  
Vol 30 (1) ◽  
pp. 238
Author(s):  
M. I. Cueto ◽  
J. Fernandez ◽  
M. M. Bruno-Galarraga ◽  
F. Pereyra-Bonnet ◽  
A. Gibbons

The Criolla breed is a local genetic resource, distributed in Patagonia, Argentina, whose primary production is meat. In the Criolla goat, efforts were made to locate productively superior males and to conserve their genetic material. Studies were carried on the feasibility of obtaining superior offspring through the implementation of embryo transfer programs. We assessed the fertilization rate and embryo production following AI with frozen semen or natural mating in Criolla goats subjected to a superovulation program. During the breeding season (May, 41° S), 26 Criolla goats were treated for oestrus with sponges (60 mg of medroxyprogesterone acetate, Progespon®, Syntex, Buenos Aires, Argentina) placed for 13 days. Goat donors were superovulated with a total of 80 mg of porcine (p)FSH (Folltropin V®, Bioniche, Ontario, Canada) every 12 h in 6 decreasing doses (18, 18, 14, 14, 8, and 8 mg) during the last 3 days of progestagen treatment. A dose of 125 μg of cloprostenol (Estrumate®, Schering-Plough, Quebec, Canada) was given in conjunction with the first dose of pFSH. Oestrus detection was performed every 12 h, starting at 24 h after sponge removal. Females were considered to be in oestrus if they passively accepted buck mounting. Goats were randomly assigned to the following treatments: (1) natural mating (NM, n = 12): donors detected in oestrus were individually mated with one proven fertile buck at oestrus and 12 h post-oestrus; does were remove from the male in between (Conventionally accepted treatment); (2) AI (n = 14): donors detected in oestrus were inseminated 12 ± 2 h after the onset of oestrus by laparoscopy with frozen-thawed semen (200 × 106 spermatozoa) from the same fertile buck. Embryo recovery was done by surgical prepubic laparotomy at Day 8 after sponge removal. Superovulation response was estimated by counting the number of corpora lutea (CL). Analysis of variance was used to compare fertilization rate (total number of embryos recovered for each animal, expressed as a proportion of the total number of embryos/oocytes recovered) and embryo production between treatments. Results were expressed as mean ± SEM. Statistical significance was accepted at P < 0.05. A total of 92.3% goats were recorded in oestrus (24/26) between 24 and 48 h after sponge removal (10 and 14 goats for NM and AI, respectively). An average of 16.6 ± 2.0 CL (range: 2–32) was observed in response to superovulation treatment. The recovery rate of embryos/oocytes was 60.0 ± 6.9%. No statistical difference was observed in the fertilization rate (52.1 ± 12.1 and 68.6 ± 12.1% for AI and NM, respectively) or the number of total (6.8 ± 2.0 and 4.8 ± 2.4 for AI and NM, respectively) and transferable embryos (5.6 ± 1.4 and 4.0 ± 1.7 for AI and NM, respectively) between treatments (P > 0.05). In conclusion, fertilization rates did not differ following laparoscopic insemination with frozen semen compared to natural mating in superovulated Criolla goats.


2002 ◽  
Vol 14 (4) ◽  
pp. 207 ◽  
Author(s):  
L. M. Mitchell ◽  
M. Silveira ◽  
M. J. A. Mylne ◽  
K. Matthews ◽  
W. S. Dingwall

The aim of this study was to determine whether previously observed seasonal differences in conceptus development in ewes are attributable to inherent differences in the oocyte and/or early embryo. Day 6 embryos were recovered from 50 ewes subjected to a standard oestrus synchronization, superovulation and laparoscopic artificial insemination protocol during October (peak breeding season) and April (transition to anoestrus). During the following October, 40 grade 1 and 2 embryos from each month, which had been cryopreserved at the late morula or unexpanded blastocyst stage, were thawed and transferred in singleton to synchronous recipients. Resulting pregnancies were monitored to term. For ewes receiving October- and April-produced embryos, overall mean SEM liveweight at the time of embryo transfer was 72 number of corpora lutea on the ovaries was 2.7 pregnant and their gestation lengths were 147 production on peripheral ovine pregnancy-associated glycoprotein concentrations during pregnancy or on fetal and placental characteristics at term, but, for each month, male lambs were heavier than females and were associated with larger placentae. Lamb birthweight was positively correlated with placental weight (r2 = 0.474, P<0.001) and the total weight of cotyledonary tissue (r2 = 0.429, P<0.001), but not to the number of cotyledons. Results demonstrate close relationships between fetal and placental weights at term, and that seasonal effects on conceptus development in ewes do not arise from inherent differences in the oocyte and/or early embryo.


2020 ◽  
Vol 60 (4) ◽  
pp. 497 ◽  
Author(s):  
Pedro Henrique Nicolau Pinto ◽  
Mario Felipe Alvarez Balaro ◽  
Helena Fabiana Reis de Almeida Saraiva ◽  
Viviane Lopes Brair ◽  
Vivian Angélico Pereira Alfradique ◽  
...  

Context In vivo embryo production, also called multiple ovulation and embryo transfer, can accelerate genetic gain, and thus improve animal production. However, there are issues limiting a wider use of this biotechnology in sheep livestock. Aims This study aimed to determine (1) whether a previous response to superovulation (SOV) can be used as a criterion to select ewes for in vivo embryo production, (2) whether the intensity of the SOV response (number of corpora lutea, CL) can affect the embryo recovery rate, and (3) whether the number of CL quantified by colour Doppler ultrasonography can be used to calculate the recovery rate. Methods Twenty-five Santa Inês ewes underwent SOV three times (SOV1, SOV2 and SOV3), with 200 mg FSH and natural mating. The number of CL after each SOV was determined by laparoscopy and by colour Doppler ultrasonography. Key results The number of CL significantly decreased (P &lt; 0.05) after SOV1 (7.5 ± 4.8) to 3.0 ± 5.0 at SOV 2 and 2.2 ± 3.5 at SOV3. Strong correlations were observed between SOV2 and SOV3 in terms of numbers of CL (r = 0.86, r2 = 0.74; P &lt; 0.0001) and viable embryos (r = 0.79, r2 = 0.63; P &lt; 00001). However, no correlations were observed between SOV1 and SOV2 or between SOV1 and SOV3. Recovery rate did not differ with the intensity of the SOV response (≤6, 7–10, &gt;10 CL) or between the methods used to quantify CL. Conclusions Ewes did not show the same pattern of response when submitted to successive FSH-based SOV. The intensity of the SOV response did not affect the recovery rate, and the number of CL estimated by colour Doppler ultrasonography can be used to calculate the recovery rate. Implications Selecting sheep embryo donors by a previous SOV response is not always feasible. The recovery rate is homogeneous and it is not affected by the intensity of the SOV response. A nonsurgical technique can be used to assess the recovery rate, improving animal welfare in MOET programs.


1983 ◽  
Vol 19 (1) ◽  
pp. 147 ◽  
Author(s):  
M.C. Schiewe ◽  
C.R. Looney ◽  
K.G. Hill ◽  
C.A. Johnson ◽  
R.A. Godke

Animals ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 216
Author(s):  
Hernan Baldassarre

The potential of laparoscopic ovum pick-up (LOPU) followed by in vitro embryo production (IVEP) as a tool for accelerated genetic programs in ruminants is reviewed in this article. In sheep and goats, the LOPU-IVEP platform offers the possibility of producing more offspring from elite females, as the procedure is minimally invasive and can be repeated more times and more frequently in the same animals compared with conventional surgical embryo recovery. On average, ~10 and ~14 viable oocytes are recovered by LOPU from sheep and goats, respectively, which results in 3–5 transferable embryos and >50% pregnancy rate after transfer. LOPU-IVEP has also been applied to prepubertal ruminants of 2–6 months of age, including bovine and buffalo calves. In dairy cattle, the technology has gained momentum in the past few years stemming from the development of genetic marker selection that has allowed predicting the production phenotype of dairy females from shortly after birth. In Holstein calves, we obtained an average of ~22 viable oocytes and ~20% transferable blastocyst rate, followed by >50% pregnancy rate after transfer, declaring the platform ready for commercial application. The present and future of this technology are discussed with a focus on improvements and research needed.


1979 ◽  
Vol 59 (4) ◽  
pp. 675-683 ◽  
Author(s):  
A. J. HACKETT ◽  
H. A. ROBERTSON ◽  
E. K. INSKEEP ◽  
J. N. B. SHRESTHA ◽  
M. S. WOLYNETZ

Synchronized estrus and ovulation were induced during the anestrous season (April–May 1974) in 373 ewes of three synthetic (one sire and two dam) strains and two unselected (Suffolk and Finnish Landrace) purebred strains by treatment with 30 mg fluorogestone acetate (FGA) impregnated in polyurethane intravaginal sponges for 12 days. Following sponge removal each ewe received 500 IU pregnant mares’ serum gonadotrophin (PMSG) IM. Of these, 167 were bred by artificial insemination (AI) at 48 and 60 h post sponge removal with 0.2 ml raw unextended semen collected by electroejaculation (EE). Five days after AI, ewes were exposed to a follow up ram for 16 days for subsequent mating if a second estrus occurred. The remaining 206 were exposed to rams for a period of 22 days for natural mating. Blood samples were collected from 69 ewes, 9, 19 and 27 days post sponge removal and analyzed for progesterone to ascertain if corpora lutea were formed and whether the ewes recycled. The age of ram by mating method interaction significantly affected both fertility and fecundity mainly because some of the younger rams lacked libido and experience for natural mating. There were no significant differences in prolificacy due to any of the main effects tested. Among the 69 ewes examined for progesterone levels, 93% had formed corpora lutea after hormone treatment and 16% recycled. Only 16 of the 255 ewes that did not conceive to the synchronized estrus lambed to the subsequent estrus.


2003 ◽  
Vol 2003 ◽  
pp. 82-82
Author(s):  
F. Forcada ◽  
J.A. Abecia ◽  
J.A. Valares

The efficacy of melatonin implants inserted around the spring equinox to improve fertility and ovulation rate or litter size in Mediterranean ewes has been previously reported (Chemineau et al., 1996; Forcada et al., 2002a.), indicating the ability of the hormone to regulate the hypothalamic activity (Viguié et al., 1995). Moreover, a direct effect of melatonin on corpora lutea and embryonic development has been also reported (Wallace et al., 1988; Abecia et al., 2002). The use of prolific Rasa Aragonesa (RA) ewes (a Mediterranean breed) before culling as embryo donors has been previously tested in the breeding season (Forcada et al., 2002b.). The aim of this experiment was to improve embryo production during the seasonal anoestrus period in selected superovulated RA ewes at the end of their reproductive lives through the use of melatonin.


2004 ◽  
Vol 16 (2) ◽  
pp. 207 ◽  
Author(s):  
A. Castro Neto ◽  
B.V. Sanches ◽  
S.H.V. Perri ◽  
M.M. Seneda ◽  
J.F. Garcia

The aim of the present study was to evaluate the effects of uterine double flushing on embryo recovery rates (total structures and viable embryos), after a resting period with the uterus filled with PBS. There were 210 embryo recovery procedures conducted using the uterine double flushing method, and the results were compaired with 432 conventional single flushing procedures. All procedures were conducted with Limousin (n=403) and Guzerá (n=239) cows, following the same superovulation protocol. Cyclic donors received a progestagen implant (CIDR) and 1mL of oestradiol benzoate (Estrogin®) at Day 0. Between Day 5 and Day 9 animals received doses ranging from 200 to 300UI (zebu cows) and 300 to 500UI (taurine cows) of FSH (Pluset® – Serono) in decreasing doses. Between Day 6 and Day 8 PGF2a (Ciosin®) was administered, followed by withdrawal of the progestagen after 24h. Artificial insemination was performed between 14 and 26h after the beginning of treatment. For the double flushing procedure, after the first uterine flushing of both horns with 1L DPBS (Nutricell), a Foley catheter was positioned in the uterine body in order to fill the uterus with the same solution (80 to 150mL). After this procedure the catheter was closed with a disposable 5-mL syringe gasket, and the animals were allowed to rest in the surroundings of the work place for 30min. After this period, a second flush was performed in order to recover the remaning liquid used during manipulation of the uterus. Animals from the control group (group A) were subjected to a single uterine flushing procedure. From 210 double flushing procedures (group B – test), 1409 viable embryos were produced, classified as grades I, II, III and IV (IETS), (average of 6.7 embryos per procedure), whereas, in the 432 single flushing procedure (group A – control), 1993 embryos were produced (average of 4.6). Statistical analysis showed the increase of viable embryo recovery rate. When consecutive double flushing was performed, the average of recovered embryos increased from 8.3 to 12.7 (P&lt;0.05) in Limousin cows, and from 7.9 to 11.5 (P&lt;0.05) in Guzerá cows. Comparing recovery after single flushing with that after double flushing, the mean number of viable embryos increased from 4.7 to 6.9 (P&lt;0.05) in Limousin cows and from 4.5 to 6.4 (P&lt;0.05) in Guzerá cows. In order to assure the nonexistence of negligence effects or operator influence on results, the mean values of total embryo recovery rate after single flushing (control group) was compared to the mean values of the same rate after the first uterine flushing on test group. Results indicated no difference between recovery rates. The present work showed the viability of using the uterine double flushing procedure for improvement of embryo recovery rates in cattle.


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