363 FACTORS INFLUENCING PREGNANCY RATES FOLLOWING TRANSFER OF BOVINE IN VIVO EMBRYOS BIOPSIED FOR SEX DETERMINATION

2007 ◽  
Vol 19 (1) ◽  
pp. 297
Author(s):  
S. Li ◽  
W. Yu ◽  
J. Fu ◽  
Y. Bai ◽  
F. Jin ◽  
...  
Keyword(s):  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Pregnancy Rates ◽  
Chi Square ◽  
Embryo Survival ◽  
Chi Square Test ◽  
First Time ◽  
Fresh Embryos

Data collected from commercial embryo transfer programs in 63 farms in China during June 2002 to December 2005 was analyzed to examine the effects of various factors (biopsy, freezing, sample size, embryo development and quality, in vitro culture, and recipient quality) on pregnancy rates of in vivo-biopsied embryos. Embryos were flushed from superovulated dairy cattle and subjected to a biopsy for sexing determination using protocols and sexing kits supplied by AB Technology Ltd. Fresh embryos were implanted on the same day or frozen with AG freeze medium (AB Technology Ltd., Pullman, WA, USA) for later transfer. Recipients were synchronized with CIDA + PG protocols. Embryos were cultured in 6-well dishes containing 1.3 mL of holding medium (AB Technology Ltd.) in each well at room temperature (20–25�C) for examination of embryo survival in vitro. The chi-square test was used in statistic analysis. The implantation of fresh embryos after biopsy did not affect pregnancy rates (49.6%, 257/518) compared to that of non-biopsied fresh and frozen–thawed embryo groups (52.9%, 47/140 and 46.6%, 177/380, respectively). However, for biopsied embryos subjected to frozen and thawed procedures before implantation, particularly for those subjected to the removal of a larger biopsy, a reduced pregnancy rate was observed (41.8%, 297/710; P < 0.01). Pregnancy rates among biopsied embryos at 3 different development stages (morula-early blastocyst, blastocyst, and expanded blastocyst) were not different. Similar results were found between embryo groups of grade 1 and 2. A significant decrease in pregnancy rate (0/10) was observed with embryos held in vitro for a longer period of time (>5 h), suggesting detrimental effects of in vitro conditions on embryo survival. The highest pregnancy rate (68.0%) was observed in recipients synchronized for the first time before being implanted with biopsied embryos. Significant decreases in such rates were found in recipients synchronized for the second or third times or those with an abortion history at the first or second synchronization-implantation treatment (P < 0.01). Better pregnancy rates (45.6%, 41/90; 46.1%, 76/165; and 45.5%, 5/11) were obtained for recipients implanted with biopsied embryos at Days 7.5, 8.0, and 8.5 post-heat detection, respectively, compared to 16% at Day 7 (3/18, P < 0.05). It is concluded that mechanical treatment (cutting) does not reduce the survival of biopsied embryos; however, cryopreservation reduces their ability to survive in vivo. The analyses also suggest that holding embryos in vitro should not be longer than 5 h unless more favorable in vitro conditions can be provided. To achieve better results of implantation of biopsied embryos, embryo transfer should be performed during 7.5–8.5 days post-estrus, and the healthy recipients synchronized for the first time should be used.

289 EFFECTS OF YEAR PERIOD, TECHNICAL TEAM, AND BREED ON THE PREGNANCY RATES AFTER TRANSFER OF IVF BOVINE EMBRYOS IN THE SOUTHERN REGION OF RIO DE JANEIRO

2010 ◽  
Vol 22 (1) ◽  
pp. 301
Author(s):  
B. G. Moura ◽  
J. Almeida ◽  
F. L. Lima ◽  
G. Balbi ◽  
R. Calmerani ◽  
...  
Keyword(s):  
Beef Cattle ◽  
Dairy Cattle ◽  
Embryo Transfer ◽  
Pregnancy Rates ◽  
Chi Square ◽  
Chi Square Test ◽  
Few Data ◽  
Rectal Palpation ◽  
Fresh Embryos

The aim of the work was to study the effects of year period, technical team, breed, beef cattle and dairy cattle on the pregnancy rates in fresh embryos used in bovine transfer of IVF programs. The study was carried out at the fertilization laboratory In Vitro Nyltta Britto de Carvalho, in partnership with In Vitro Brazil, located at the Boa Vista farm, Barra do Pirai, during August 2007 to September 2008, seeking subsidies to improve the use of the technique in the field. During that period, aspirations and inovulations in 3 different periods I (August to December), II (January to April), and III (May to September) were carried out. The jobs were accomplished by 9 technical teams (A, B, C, D, E, F, G, H, and I) rendering services to the laboratory, by working with 2 beef breeds (Brahman and Nelore) and 3 dairy breeds (Gir, Girolando, and Holstein). The different breed receivers were synchronized, and in general, from 6 to 8 days after heat, they received embryo transfer, the cervical way, under low epidural anesthesia, where each female received 1 fresh embryo of IVF. All cows were submitted to gestation diagnosis by rectal palpation and ultrasonography, in general, 42 days after embryo transfer. The numbers of embryo transferred and pregnancy rates were submitted to the chi-square test, which presented significant differences (P < 0.05). There were pregnancy rates of 36.25%a (n = 960), 39.83%a (n = 1180), and 32.59%b (n = 919) in the I, II, and III periods, respectively. Among the 9 technical teams, there were verified pregnancy rates (%) of 33.51d (n = 1313), 30.30d (n = 330), 35.00cd (n = 405), 39.24cd (n = 1060), 59.25a (n = 7), 33.33d (n = 24), 53.57bc (n = 28), 43.31c (n = 157), and 58.33ab (n = 12) for A, B, C, D, E, F, G, H, and I teams, respectively. Among breeds there were rates (%) of 36.89ab (n = 412), 34.68b (n = 1286), 35.13ab (n = 74), 38.94a (n = 1140), and 37.80ab (n = 82) for Brahman, Nelore, Gir, Girolando, and Holstein, respectively. In the study, pregnancy rates (%) of 35.21b (n = 1698) in beef cattle and 38.65a (n = 1296) in dairy cattle were observed. The differences in pregnancy rates with respect to the evaluated factors, may be explained by individual, breed, and nutritional variations of the animals. There are few data in the literature with results on the embryo transfer use of IVF bovine under field conditions.

188 EFFECT OF TRANSFER OF FROZEN-THAWED IVF EMBRYOS ON PREGNANCY IN REPEAT-BREEDER INSEMINATED OR NON-INSEMINATED HOLSTEIN CATTLE

2006 ◽  
Vol 18 (2) ◽  
pp. 202 ◽  
Author(s):  
O. Dochi ◽  
M. Tanisawa ◽  
S. Goda ◽  
H. Koyama
Keyword(s):  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Calf Serum ◽  
Pregnancy Rates ◽  
Holstein Cattle ◽  
Chi Square ◽  
Vitro Fertilization ◽  
Chi Square Test ◽  
Repeat Breeder

Repeat-breeding is one of the important factors that affect dairy management. The objective of this study was to investigate the effect of transfer of frozen–thawed IVF embryos on pregnancy in repeat-breeder Holstein cattle. Cumulus–oocyte complexes (COCs) were collected by aspiration of 2–1-mm follicles from ovaries obtained at a local abattoir. COCs were matured for 20 h in TCM-199 supplemented with 5% calf serum (CS) and 0.02 mg/mL of FSH at 38.5°C under a 5% CO2 atmosphere in air. Matured oocytes were inseminated with spermatozoa of 5 × 106/mL in BO solution (Brackett and Oliphant 1975 Biol. Reprod. 12, 260–274) containing 10 mM hypotaurine and 4 units/mL heparin. After 18 h of gamete co-culture, presumptive zygotes were cultured in CR1aa (Rosenkrans et al. 1991 Theriogenology 35, 266) supplemented with 5% CS for 8 days at 38.5°C under 5% CO2, 5% O2, 90% N2 atmosphere in air. After in vitro fertilization, Day 7 and Day 8 blastocysts were frozen in 1.5 M ethylene glycol (EG) in Dulbecco's PBS (DPBS) supplemented with 0.1 M sucrose and 20% CS. Embryos were transferred into a freezing medium, loaded into 0.25-mL straws, and allowed to stand for 15–20 min for equilibration. The straws were then plunged into a −7°C methanol bath of a programmable freezer for 1 min, seeded at −7°C, maintained at −7°C for 15 min, cooled to −30°C at the rate of −0.3°C/min, and then plunged into liquid nitrogen. Recipient animals (43 heifers, 131 cows) included those that did not conceive after being artificially inseminated (AI) 3 to 15 times. The frozen–thawed IVF embryos were directly transferred to the recipient animals 7 days after estrus or AI. Pregnancy rates were analyzed by chi-square test. The results are presented in Table 1. There were no significant differences in the pregnancy rates between treatments. However, a slightly higher pregnancy rate was achieved by embryo transfer after AI. These results suggest that embryo transfer may increase the pregnancy rate in repeat-breeder Holstein cattle. Table 1. Pregnancy rates after transfer of IVF frozen–thawed embryos in repeat-breeder Holstein cattle

106 PREGNANCY RATE AFTER EMBRYO TRANSFER OF IN VIVO-PRODUCED OVINE EMBRYOS CRYOPRESERVED BY SLOW FREEZING OR VITRIFICATION

2010 ◽  
Vol 22 (1) ◽  
pp. 212
Author(s):  
N. Mucci ◽  
F. Hozbor ◽  
G. G. Kaiser ◽  
E. Sanchez ◽  
R. H. Alberio
Keyword(s):  
Ethylene Glycol ◽  
Pregnancy Rate ◽  
Liquid Nitrogen ◽  
Embryo Transfer ◽  
Slow Freezing ◽  
Embryo Cryopreservation ◽  
Chi Square ◽  
Chi Square Test

Although slow freezing is the method of choice to cryopreserve in vivo-produced ovine embryos, vitrification has became an alternative procedure mostly developed for in vitro-produced bovine embryos. The aim of this work was to compare pregnancy rates after cryopreservation of in vivo-produced ovine embryos with slow freezing or open pulled straw (OPS) vitrification method. Ewes were synchronized using intravaginal sponges containing 60 mg of medroxyprogesterone acetate for 14 d. Superovulation was performed using a total dose of 176 IU of ovine FSH (Ovagen), in 6 decreasing doses (i.m.) from Day 12 to 14 of treatment (Day 0 = sponge placing). Ewes were hand mated with 2 rams of proven fertility. Embryos were recovered 6 days after estrous detection by surgical procedure, evaluated under stereomicroscope, and randomly assigned to the cryopreservation treatments. Slow freezing was performed in D-PBS supplemented with 1.78 M ethylene glycol, 0.1 M sucrose, 4 mg mL-1 of BSA, and 20% serum. Embryos were loaded into 0.25-mL plastic straws and placed into a -7°C methanol bath chamber. After seeding embryos were cooled to -35°C at a rate of 0.5°C/min and then stored in liquid nitrogen. Thawing was performed by placing the straws in a 30°C water bath for 30 sec. Vitrification was performed by using the OPS method (Vajta et al. 1998) with minor modifications. Embryos were incubated in D-PBS supplemented with 1.78 M ethylene glycol, 1.3 M DMSO for 3 min and then transferred for 25 s in vitrification solution of D-PBS with 3.56 M ethylene glycol, 2.6 M DMSO, and 0.5 M sucrose, loaded in a 1 mL drop in the OPS, and immediately submerged into and stored in liquid nitrogen. Warming was performed in D-PBS plus 0.25 M sucrose for 5 min and then into D-PBS plus 0.15 M sucrose for another 5 min. Before embryo transfer, the presence of corpus luteum (CL) was detected by laparoscopic examination. One embryo per recipient was surgically transferred in the apical extreme of the uterine horn ipsilateral to the CL. Pregnancies were determined by ultrasonography 41 days after embryo transfer. Data were analyzed using the chi-square test. We found 47.8% pregnancy rate using slow freezing (11/23) and 43.5% pregnancy rate using OPS vitrification (10/23). Statistical differences were not detected (P = 0.09). We conclude that vitrification by OPS system, with minor modifications, is a suitable procedure for in vivo-produced ovine embryo cryopreservation.

scholarly journals 183PREGNANCY RATE FOLLOWING TRANSFER OF IN VITRO- AND IN VIVO-PRODUCED BOVINE EMBRYOS TO LH-TREATED RECIPIENTS

2004 ◽  
Vol 16 (2) ◽  
pp. 213 ◽  
Author(s):  
J. Small ◽  
M. Colazo ◽  
D. Ambrose ◽  
R. Mapletoft ◽  
J. Reeb ◽  
...  
Keyword(s):  
Pregnancy Rate ◽  
Animal Health ◽  
Beef Cows ◽  
Water Bath ◽  
Bovine Embryos ◽  
Chi Square ◽  
Embryo Survival ◽  
Frozen Embryos

The objective was to evaluate the effect of pLH treatment on pregnancy rates in recipients receiving in vivo- or in vitro-produced bovine embryos. Heifers (n=37) and lactating (n=28) and non-lactating (n=150) beef cows were treated at random stages of the cycle with 100μg GnRH i.m. (Cystorelin, Merial Canada Inc., Victoriaville, Quebec, Canada) on Day −9, 500μg cloprostenol i.m. (PGF; Estrumate, Schering Plough Animal Health, Pointe-Claire, Quebec, Canada) on Day —2 and GnRH on Day 0 (66h post-PGF; without estrus detection). Cattle were placed at random, by class, into three groups: no further treatment (Control; n=71), or 12.5mg pLH (Lutropin-V, Bioniche Animal Health, Belleville, Ontario, Canada) on Day 5 (n=72) or on Day 7 (n=72) after the second GnRH. On Day 7, cattle with a CL &gt;10mm in diameter (determined ultrasonically) received in vivo-produced, fresh (Simmental) or frozen (Holstein), or in vitro-produced frozen (Holstein) embryos (embryo type balanced among groups). Embryos were cryopreserved in 10% ethylene glycol; in vivo-produced frozen embryos were thawed 5 to 10s in air, 15s in a water-bath at 30°C and then “direct-transferred” nonsurgically. In vitro-produced frozen embryos (donated by IND Lifetech Inc., Delta, British Columbia, Canada) were thawed in a water-bath at 27°C for 10s and placed in ViGro Holding Plus medium (AB Technology, Pullman, WA, USA) at room temperature, evaluated and then transferred nonsurgically. Pregnancy was determined by ultrasonography on Day 35. Data were analyzed with CATMOD, chi-square and GLM procedures (SAS Institute, Cary, NC, USA.). Twenty cattle (9.3%) did not receive embryos; five heifers had cervical problems, and five heifers and 10 cows did not have a CL &gt;10mm. Overall, 7.1% of the recipients had two CL on the day of embryo transfer. There was no effect (P&gt;0.05) of treatment, embryo type (or interaction) or class of recipient on pregnancy rate (overall, 44.1%, 86/195; Table 1). Similarly, mean (±SD) CL diameter and luteal area did not differ (P&gt;0.05) among groups or between pregnant and open recipients (overall, 22.0±3.4mm and 352.0±108.7mm, respectively). However, recipients with a CL diameter ≥18mm tended (P&lt;0.1) to have a higher pregnancy rate (45.8 vs 25.0%). In a subset of 40 recipients examined ultrasonically on Day 12, 50% of those treated on Day 5 and 70% of those treated with pLH on Day 7 had two CL. In summary, overall pregnancy rate in GnRH-synchronized recipients receiving in vitro- or in vivo-produced embryos by nonsurgical transfer was 44.1%. Embryo survival to Day 35 was not affected by type of embryo or treatment with pLH 5 or 7 days after ovulation. Table 1 Pregnancy rate in recipients on Day 35 based on pLH treatment and embryo-type

204 RELATIONSHIP BETWEEN RESPIRATORY ACTIVITY AND THE PREGNANCY RATE OF BISECTED BOVINE EMBRYOS IN VIVO

2007 ◽  
Vol 19 (1) ◽  
pp. 219 ◽  
Author(s):  
S. Moriyasu ◽  
H. Hirayama ◽  
K. Sawai ◽  
S. Kageyama ◽  
S. Aoyagi ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Respiratory Activity ◽  
Pregnancy Rates ◽  
Bovine Embryos ◽  
Chi Square ◽  
Important Indicator ◽  
Consumption Rates

Oxygen consumption is an important indicator of the metabolic activity of living cells, which may provide valuable information for evaluating embryo quality. We have found that the bovine embryos with high oxygen consumption possess stronger potential for further development. However, the relationship between respiratory activity and the pregnancy rate of embryos is still unclear. In this study, we investigated the respiration rates of bisected bovine embryos and the pregnancy rates of demi-embryos after embryo transfer. Compact morula-stage embryos were bisected evenly by micro glass needle. One hundred bisected embryos were incubated for 24 h in embryo culture medium (IVD101; Research Institute for the Functional Peptides, Yamagata, Japan) at 39�C under 5% CO2, 5% O2, 90% N2. After the incubation, demi-embryos were classified into 2 groups: blastocoel-formed (BC) and blastocoel-not-formed (CM) embryos. Oxygen consumption rates of demi-embryos were measured by scanning electrochemical microscopy (SECM; Hokuto Denko Corporation, Tokyo, Japan). Within 3 h after the measurement, 80 demi-embryos were transferred into recipient cows (one demi-embryo/one recipient) at 7–8 days after estrus. Recipient cows were diagnosed for pregnancy by ultrasonography approximately 40 days after estrus. Statistical difference was analyzed by Tukey's post-hoc test and chi-square test. A total of 27 recipient cows became pregnant; the pregnancy rates for cows with CM and BC demi-embryos were 40.6% (13/32) and 29.2% (14/48), respectively. Mean oxygen consumption rates (� 10-14 mol s-1) in pregnant and non-pregnant cows were 0.47 and 0.39 for CM demi-embryos and 0.63 and 0.52 for BC demi-embryos, respectively. Retrospective analysis showed that the respiratory activity of demi-embryos in the pregnant group was higher than those in the non-pregnant group. In particular, the pregnancy rates for demi-embryos with respiratory activity higher than 0.35 in CM and 0.40 in BC groups were 52.0% (13/25) and 35.9% (14/39), respectively. On the other hand, cows with demi-embryos having an oxygen consumption rate under 0.35 in CM (n = 7) and 0.40 in BC (n = 9) groups did not become pregnant. These results demonstrated that bovine demi-embryos with higher respiratory activity showed a high pregnancy rate after embryo transfer. It is generally known that the pregnancy rate after the transfer of bisected embryos is lower than that of whole embryos. The measurement of oxygen consumption by SECM procedures is a useful tool to assess the quality of pre-implantation embryos and may contribute to the improvement of the success rate for bisected embryo transfer.

110 First ovum pickup-in vitro-produced lidia breed calves using Lidia breed recipients: influence of age and state of the recipients and in vitro-produced embryos on pregnancy rates

2019 ◽  
Vol 31 (1) ◽  
pp. 181
Author(s):  
G. Gamarra Lazo ◽  
D. Di Scala ◽  
S. Maunas ◽  
R. Chaubet ◽  
S. Lacaze
Keyword(s):  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Development Stage ◽  
Pregnancy Rates ◽  
Chi Square ◽  
Related Factors ◽  
Single Operator ◽  
Pregnancy Status ◽  
Chi Square Analysis

We previously demonstrated the success of in vitro embryo production (IVP) in Lidia breed cattle (Gamarra Lazo et al. 2017 Reprod. Fertil. Dev. 30, 187). As in other species, the success of IVP is linked to the birth of calves from this technique. In the Lidia breed, an important factor to consider is the use of Lidia recipients in order to keep the temperament characteristic of this breed to next generations. The aim of the study was to produce ovum pickup (OPU)-IVP calves in the Lidia breed and to assess the effects of recipient and embryo related factors (status of the recipients; development stage of IVF embryos) on pregnancy rate following embryo transfer. Ovum pickup-IVP embryos from Lidia breeds were produced by a standard protocol (Gamarra Lazo et al. 2017 Reprod. Fertil. Dev. 30, 187). Numbers of blastocysts and expanded blastocysts were recorded on Day 7. A total of 27 blastocysts (B) and 34 expanded blastocysts (EB) of excellent quality (grade 1 according to IETS classification) were selected for fresh transfer. All embryos were transferred to Lidia breed recipients (heifers or cows) by a single operator under similar environmental and field conditions. Recipients were synchronized by subcutaneous insertion of an ear implant of 3.3mg of Norgestomet (Crestar®, MSD, Courbevoie, France) for 9 days. Two days before implant withdrawal, 0.5mg of Cloprostenol (Estrumate®, MSD) was injected. No oestrous detection was performed and synchronized females were selected as recipients when they presented a well developed corpus luteum at Day 9 after implant withdrawal (Day 6 to 7 after the expected oestrus). Blood samples were collected from recipients to determine pregnancy status using the bovine pregnancy associated glycoprotein (Idexx, Westbrook, ME, USA) 50-60 days after transfer. Pregnancy rates were analysed by chi-square analysis to compare results between heifers and cows and between B and EB embryo stages. The overall pregnancy rate after transfer of IVP fresh embryos from Lidia breed averaged 41.0% (n=25). A higher pregnancy rate was achieved in cows compared to heifers [51.2% (21/41) v. 20.0% (4/20) respectively, P&lt;0.05]. There was no difference in pregnancy rate between grade 1B [37% (10/27)] and EB [44.1% (15/34)] embryos (P&gt;0.05). Surprisingly, these results suggest that Lidia breed cows are the best recipients for OPU-IVP embryos. This may be related to the limited feasibility of manipulating the uterine horn during the embryo transfer in Lidia breed heifers, which have a low weight (less than 280kg) and present a narrow rectum diameter. It has been also observed that the cervix is very thin and difficult to cross, thus increasing the stress and potentially inflammatory and immune products secretion. Development stage of embryos did not affect pregnancy rate. To our knowledge, no OPU-IVP Lidia breed calves have been reported previously following transfer into Lidia breed recipients. In the current work, 13 OPU-IVP Lidia breed calves were born. Therefore, we confirmed the possibility of applying OPU-IVP and embryo transfer techniques in this breed within a genetic program.

150 EFFECT OF TIMELY FLUNIXIN MEGLUMINE TREATMENT ON PREGNANCY RATES IN HOLSTEIN HEIFERS

2006 ◽  
Vol 18 (2) ◽  
pp. 183
Author(s):  
A. Guzeloglu ◽  
H. Erdem ◽  
M. K. Saribay ◽  
W. W. Thatcher ◽  
T. Tekeli
Keyword(s):  
Pregnancy Rate ◽  
Inhibitory Effect ◽  
Early Embryo ◽  
Pregnancy Rates ◽  
Control Group ◽  
Chi Square ◽  
Embryo Survival ◽  
Flunixin Meglumine ◽  
Chi Square Test ◽  
Pregnancy Recognition

The objective was to determine if administration of flunixin meglumine (FM) to heifers following insemination would increase pregnancy rate due to its inhibitory effect on prostaglandin F2� (PGF2�) synthesis. Fifty-two 15-month-old Holstein heifers were synchronized with single or double injections of PGF2� followed by an injection of gonadotropin-releasing hormone (GnRH) 48 h later and a timed artificial insemination at 12-14 h after injection of GnRH (Day 0). Heifers randomly assigned to the treatment group (FMG) were injected twice with FM (1.1 mg/kg BW; i.m.), given 12 h apart on the evening of Day 15 and the morning of Day 16. The control group (CG) was not treated. Pregnancy rates were defined as the percentage of heifers diagnosed pregnant by ultrasound at Days 29 and 65 after AI. Effects of treatment on pregnancy rates and pregnancy losses were analyzed by chi-square test and logistic regression analysis. Pregnancy rates in the heifers treated with FM were higher at Day 29 (76.9%; 20/26 [FMG] vs. 50%; 13/26 [CG]; P < 0.04) and tended to be higher at Day 65 (69.2%; 18/26 [FMG] vs. 46.2%; 12/26 [CG]; P < 0.09). Administration of FM two times at a critical stage leading up to pregnancy recognition, associated with corpus luteum maintenance, increased both early embryo survival and pregnancy rate via an additive antiluteolytic effect due to a delay in the luteolytic secretion of PGF2�; this provided conceptuses extra time to develop the capability to inhibit the luteolytic process.

229 OOCYTE RECOVERY, IN VITRO FERTILIZATION AND EMBRYO TRANSFER IN THE SERVAL (LEPTAILURUS SERVAL)

2006 ◽  
Vol 18 (2) ◽  
pp. 223 ◽  
Author(s):  
C. E. Pope ◽  
M. C. Gómez ◽  
A. Cole ◽  
C. Dumas ◽  
B. L. Dresser
Keyword(s):  
Embryo Transfer ◽  
Oocyte Retrieval ◽  
Sub Saharan Africa ◽  
Medium Size ◽  
Embryo Survival ◽  
Frozen Semen ◽  
Old Donor ◽  
Fresh Embryos

Servals are medium size (9 to 18 kg) spotted cats found in sub-Saharan Africa that are protected by CITES under Appendix II regulations. There are at least six sub-species, one of which is listed as Endangered by the U.S. Endangered Species Act. In vitro-derived embryos have been produced in at least one-half of the 36 species of nondomestic cats, and kittens have been born after embryo transfer in six species. In the present study we evaluated (1) ovarian response of servals to repeated exogenous gonadotropin stimulation, and (2) in vitro and in vivo developmental ability of in vitro-derived embryos. One two-year-old and one five-year-old female were treated six and three times, respectively, over a 3.5-year period, with a total of 20 or 25 IU of porcine FSH (i.m.; Sioux Biochem, Sioux City, IA, USA) administered daily over four days during interestrus. On Day 5, 15 IU of porcine LH (i.m.; Sioux Biochem) was given, and laparoscopic oocyte retrieval was performed 24 h later. A total of 234 preovulatory oocytes were recovered: 182 (mean = 30.3) from the two-year-old and 52 (mean = 17.3) from the five-year-old female. A total of 91 and 91 oocytes were recovered at retrievals 1 through 3 and 4 through 6, respectively, from the two-year-old donor. Eighty oocytes from the two-year-old donor were inseminated with cooled (24 h, 4°C) semen. Frozen semen from the same male was used to inseminate 102 oocytes from the two-year-old female and 52 oocytes from the five-year-old female. Overall, 136 embryos (58% cleavage frequency) were produced: 119 (65% cleavage frequency) from the two-year-old and 17 (33% cleavage frequency) from the five-year-old female. Cleavage frequency of oocytes from the two-year-old female inseminated with cooled or frozen semen was similar, 68% (54/80) and 64% (65/102), respectively. Embryos were cultured for 5 or 6 days before controlled rate cryopreservation or uterine transfer (Gómez et al. 2003 Theriogenology 60, 239–251). On Day 5, 66 early to mid-stage morulae were cryopreserved at a slow controlled rate. Sixty Day 5 and 18 Day 6 embryos were auto-transferred to a recipient (8 to 26/transfer) in a total of six surgical procedures, of which five were with fresh embryos (n = 70) and one was with cryopreserved embryos (n = 8). The sixth embryo transfer procedure (26 fresh embryos) resulted in the unassisted birth of a live male kitten on Day 77 of gestation. We have shown that in vitro-derived embryos can be generated in the serval and that oocyte retrieval rates and cleavage frequencies are comparable to those reported for other species of mid-sized nondomestic cats. The nominal incidence of pregnancy and frequency of embryo survival may be improved by transferring early cleavage staged embryos into the oviduct, as demonstrated in the African wildcat (Felis silvestris lybica; Gómez et al. 2004 Cloning and Stem Cells 6, 247–258). This work was partially funded by the Dan Heard Conservation Challenge Grant.

135 IN VITRO DEVELOPMENT OF IN VIVO PRODUCED EMBRYOS FROZEN AT MORULA OR BLASTOCYST STAGES

2008 ◽  
Vol 20 (1) ◽  
pp. 148
Author(s):  
R. Sartori ◽  
G. M. Machado ◽  
M. M. Guardieiro ◽  
M. R. Bastos ◽  
L. Leme ◽  
...  
Keyword(s):  
Ethylene Glycol ◽  
Liquid Nitrogen ◽  
Zebu Cattle ◽  
Hatching Rate ◽  
Chi Square ◽  
Fluid Medium ◽  
Chi Square Test ◽  
Fresh Embryos

This study was designed to compare cryotolerance between morulae and blastocysts collected from superovulated heifers. Twenty pubertal beef heifers (10 Nelore and 10 crossbred Nelore � Simmental) were superovulated with 100 mg of FSHp (Folltropin-V, Bioniche, Ontario, Canada), and embryos were collected and evaluated 7 days after estrus. Grades 1 and 2 embryos (IETS) were divided into four groups: morulae cryopreserved (MC) in liquid nitrogen (n = 24); blastocysts cryopreserved (BC; n = 19); morulae fresh (MF; n = 23); and blastocysts fresh (BF; n = 18). For freezing, embryos were immersed in ethylene glycol (Ethylene Glycol Freeze Plus with 0.1 m sucrose, Bioniche, Pullman, WA, USA), and a standard protocol (cooling rate of –0.5�C/min) was used. Prior to in vitro culture, embryos were removed from nitrogen, kept at room temperature for 5 s, and put in a water bath at 30�C for 20 s. Within 5 h after recovery, thawed and fresh embryos were washed five times in holding solution (Holding Plus, Bioniche), transferred to synthetic oviduct fluid medium (SOF, Nutricell, Campinas, SP, Brazil), and cultured for 72 h. Embryos were evaluated at 48 and 72 h of culture. After the last evaluation, degenerate and non-hatched embryos were removed from culture, and the remaining embryos were measured by a graduated ocular coupled to the Motic Images Plus 2.0 program. Hatched blastocysts were kept in culture for an additional 48 h for post-hatching development assessment. For post-hatching culture PHD medium (Brand�o DO et al. 2005 Biol. Reprod. 71, 2048–2055) was added into each well, to have a final composition of 50% SOF and 50% SOF PHD. At 120 h of culture (48 h of PHD culture) only morphologically normal blastocysts were measured. Comparison among groups was performed by ANOVA or chi-square test. Data are presented as mean � SEM. After 48 h of culture, hatching rate (%) was significantly lower in cryopreserved (MC = 8.3 and BC = 21.5) than in fresh (MF = 56.5 and BF = 77.8) embryos (P < 0.05). However at 72 h, hatching rate was similar among BC (75.9), MF (78.3), and BF (88.9), being MC (41.7) still lower (P < 0.05). The diameter (µm) of hatched embryos after 72 h of culture was 272.8 � 27.1a (n = 8), 320.6 � 18.6ab (n = 14), 385.3 � 14.2c (n = 17), and 378.0 � 22.0bc (n = 16) for MC, BC, MF, and BF, respectively (a–cP < 0.05). After 120 h of culture, the diameter of MC (379.0 � 39.9; n = 8), although similar to BC (495.4 � 59.6; n = 10), was smaller than MF (509.1 � 36.5; n = 11) and BF (511.8 � 41.2; n = 14). The results of this study with zebu cattle suggest that morulae are less resistant to cryopreservation in liquid nitrogen than blastocysts. Moreover, frozen/thawed embryos, when put in culture, present a slower development compared with fresh embryos. Financial support from CNPq and FAPESP from Brazil.

100 FACTORS AFFECTING PREGNANCY RATES IN THE TRANSFER OF IN VITRO-PRODUCED JAPANESE BLACK CATTLE EMBRYOS

2017 ◽  
Vol 29 (1) ◽  
pp. 157
Author(s):  
T. Nishisouzu ◽  
A. Abe ◽  
S. Matoba ◽  
O. Dochi ◽  
K. Okamura
Keyword(s):  
Pregnancy Rate ◽  
Embryo Transfer ◽  
Pregnancy Rates ◽  
Oestrous Cycle ◽  
Rapid Expansion ◽  
Japanese Black Cattle ◽  
Factors Affecting ◽  
Increased Activity

Despite the rapid expansion of in vitro embryo production (IVP) technology for genetic improvement in the cattle industry in the last decades, pregnancy rates by the transfer of IVP embryos are still lower than those derived from in vivo-produced embryos. The objective of this study was to analyse factors affecting pregnancy rates after the transfer of IVP Japanese Black cattle embryos under farm conditions. Holstein heifers (n = 4,475) and cows (n = 8,541) were selected as recipients. Most cows (80%) were managed in tie-stall barns and most heifers (80%) were managed in pens. Embryo transfers were performed for 9 years, from 2004 to 2012. The embryos were produced from oocytes derived from a local abattoir and semen from 14 proven bulls by the Livestock Improvement Association of Japan (Hamano and Kuwayama 1993 Theriogelogy 39, 703–712). The fresh IVP embryos (quality; IETS code 1) that reached the blastocyst stage after 7 to 8 days (insemination = Day 0) were transported by an airplane (2 h) and subsequently by a car (1.5 h). Embryos were non-surgically transferred to each recipient on Day 7 to 9 of their natural oestrous cycle on farms. Pregnancy was diagnosed on Day 40 to 60 after oestrus. Pregnancy results were statistically analysed using the GLM procedures of SAS. The following variables were included in the model: recipient parity (0, 1, 2, or 3), day (7, 8, or 9) of the oestrous cycle at the time of embryo transfer, oestrus behaviour (increased activity observed by farmers), presence of mucus at oestrus, presence of blood after oestrus, and year (1, 2, 3, 4, 5, 6, 7, 8, or 9) and season (April–June as spring, July–September as summer, October–December as fall, or January–March as winter) of embryo transfer. The Bonferroni correction was used to counteract the problem of multiple comparisons. Heifers had significantly higher pregnancy rates than cows (51.0% v. 37.9%, respectively; P < 0.01), and first parity cows had higher pregnancy rates than third parity cows (42.9% v. 35.7%, respectively; P < 0.01). Pregnancy rates were significantly higher in recipients that received an embryo transfer on Day 8 of their oestrous cycle, than on Day 7 (46.6% v. 42.4%, respectively; P < 0.01). Recipients without oestrus behaviour had higher pregnancy rates than those with oestrus behaviour (46.3% v. 43.4%, respectively; P < 0.01). The presence of mucus and/or blood after oestrus and the season of transfer were not found to significantly affect pregnancy rates. The results of this study indicated that performing IVP embryo transfers on Day 8 of a recipient’s oestrous cycle will improve the pregnancy rate, season does not have an effect on pregnancy rate, and the detection of oestrus by monitoring increased activity is not always reliable and instead should be determined by multiple symptoms on farm conditions.

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