77 PREGNANCY RATES AFTER VITRIFICATION OF FRESH AND COOLED EQUINE EMBRYOS USING THE CRYOTOP METHOD

2011 ◽  
Vol 23 (1) ◽  
pp. 144
Author(s):  
C. Baca Castex ◽  
G. Dalvit ◽  
M. Miragaya ◽  
A. Alonso ◽  
M. Pinto ◽  
...  

Cryopreservation of equine embryos is still not a routine procedure. Pregnancies have been obtained after transfer of vitrified embryos of less than 300 μm (Eldridge-Panuska et al. 2005). The aim of this study was to use the cryotop method (Kuwayama, 2007) to obtain pregnancies after transfer of vitrified thawed cooled and fresh embryos collected in our clinical embryo transfer programme. Embryos were assigned either to be vitrified within 3 h of collection or to be cooled for 18–24 h before vitrification. All embryos were vitrified and thawed by Cryotop Vitrification Kit® (Cryo Tech Laboratory®). Briefly, they were equilibrated in a solution containing ethylene glycol (EG), dimethylsulphoxide (DMSO) in TCM-199 for 10 to 25 min. Then they were moved to vitrification solution containing EG, DMSO, and sucrose in TCM-199 and loaded with a glass capillary onto the top of the film strip. After loading, almost all the solution was removed to leave only a thin layer covering the embryo, and the sample was quickly immersed into liquid nitrogen and covered with a protective cap. The time between entry to vitrification solution and nitrogen was from 1 to 3 min. At warming, the strip was immersed directly for 1 min into a 37°C medium containing sucrose in TCM-199. The embryo was incubated 3 min in a diluent solution, washed twice 5 min each in washing solution, and further cultured in DMEM F-12 with 10% FBS at 38.5°C 5% CO2 between 2 to 5 h. For transfer, the embryo was loaded in 0.5-mL straws. All recipient mares had ovulated 4 to 7 days before nonsurgical transfer. Pregnancies were detected 6 to 8 days later. A total of 15 embryos, grades 1 to 2, were obtained. Fresh embryos (n = 7) ranged between 250 and 800 μm, and refrigerated embryo (n = 8) diameter was between 130 and 550 μm. Pregnancy rates were 37.5% (3/8) for embryos cooled before vitrification and 28.6% (2/7) for embryos vitrified within 3 h. The overall pregnancy rate was 33.3% (5/15). Shipping cooled embryos allows maintaining a large number of recipients far away from donors, without decreasing pregnancy rate. It also makes it possible to send embryos to a specialised laboratory in order to be vitrified and preserved until recipients are available. Equine embryos collected 6 days after ovulation are generally smaller than 300 μm and have shown the highest survival rate after cryopreservation. However, the embryo recovery rate is higher when flushing is performed at Day 7 or 8. This cryopreservation protocol could provide a way to vitrify fresh and cooled embryos up to 550 μm, which would prevent the loss of valuable embryos collected in more advanced stages of development. In summary, pregnancies can be obtained after cooling for 18 to 24 h and vitrification of embryos collected 7 or 7.5 days after.

2007 ◽  
Vol 19 (1) ◽  
pp. 297
Author(s):  
S. Li ◽  
W. Yu ◽  
J. Fu ◽  
Y. Bai ◽  
F. Jin ◽  
...  

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.


2015 ◽  
Vol 27 (1) ◽  
pp. 168
Author(s):  
K. Karakas ◽  
H. Alkan ◽  
G. Onur ◽  
D. Ozen ◽  
M. Kaymaz ◽  
...  

The aims of this study were to compare the embryo recovery rate in Angora goats based on application timing; at the beginning (September – October; Group 1) and end (December; Group 2) of the breeding season and to evaluate the viability and survivability of fresh or vitrified-thawed embryos when transferred. For this purpose, nine Angora goats were used as donors and thirthy Angora goats were used as recipients. Donor goats were synchronized and superovulated with traditional protocol and were mated with fertile bucks. At the 156th hour of the mating, embryos were collected surgically and evaluated under a stereo microscope. In group 1, 103 embryos and in group 2, 63 embryos were collected from nine goats. Fresh or vitrified-thawed embryos were transferred surgically to synchronized recipients. In Group 1 fresh/thawed embryos were transferred to 15/15 goats and in group 2, fresh/thawed embryos were transferred to 8/8 goats, respectively. Each recipient received 1 or 2 embryos ipsilateral to the ovary containing one or more corpora lutea. On day 30 of the transfer, goats were examined by transrectal ultrasonography, pregnancy rates of fresh/thawed embryos were 66.6%/26.6% for group 1 and 62.5%/62.5% for group 2. On day 100 of the transfer, goats were examined again by ultrasonography, and pregnancy rates were 46.6%/0% for group 1 and 37.5%/0% for group 2, respectively. After about 50 days, goats were kidded. In group 1, 3 twins and 4 single kids were born; in group 2, 2 twins were born. The total number of collected embryos and pregnancy rates among the groups were analysed using SPSS® (version 14.01, Chicago, IL, USA) and for all comparisons, differences were considered with a minimum of 5% significance level. After statistical analyses, the numbers of collected embryos at the beginning and at the end of the breeding season were compared. There was no difference in freezable/transferable embryo quality. As a result, embryos could be collected after superovulation protocols in Angora goats both at beginning and end of the breeding season, however there might be a decrease in numbers of collected embryos and the reasons for this might not be only the seasonal factors but also the environment, care, nutrition and previous superovulation protocols. The pregnancy rate following transfer of fresh embryos was satisfactory but not all does confirmed pregnant kidded; hence, reducing the number of recipients kidding. The pregnancy rate following transfer of vitrified-thawed embryos was generally low and unsatisfactory. Further research is warrented in improving the cryopreservation techniques and thus the embryo survival rate of Angora goat embryos. This study was financed with the University of Ankara Grant.


2004 ◽  
Vol 16 (2) ◽  
pp. 207
Author(s):  
G. Bourgoin ◽  
H. Quinton ◽  
A. Rohou ◽  
P. Humblot ◽  
C. Ponsart

Success of embryo transfer (ET) is related to many well identified factors such as quality and stage of embryos, or parity of recipients. However, there are no data on the effects of factors associated with embryo recovery and ET policy (proportion of embryos transferred as fresh or frozen) on pregnancy rates (PR) after ET. A retrospective study was designed to study how these factors affect PR achieved with fresh embryos (FE). A total of 3380 embryo recovery sessions (RS) realized by Embryotop (France) between 1995 and 2002 were studied, corresponding to 14,755 transfers with FE. Forty-four RS were made with only frozen embryos and removed from data set. A total of 1716 RS involved transfer from both frozen and FE, whereas in 1620 RS all transfers were made with FE. The organization of RS was assessed by the number of operators (OP) per RS and per donor cow, the number of embryos collected per RS and per OP. The operator at RS and ET was also noted. Embryo characteristics such as stage and quality (according to IETS criteria), paternal origin, and breed of donor cow were recorded together with the breed, date of ET and parity of recipients. The association between each variable and PR was analyzed by Chi-square. Only the significant factors were considered for further analysis (P<0.05) and introduced with a stepwise procedure in a multivariate model of logistic regression to calculate adjusted odds ratio (OR). The number of RS was stable among years (463 to 493), whereas the number of embryos collected increased by 10% from 1995 to 2002. For 681 RS (20.4%), more than one donor cow were used for collection. Mean number of donor cows per RS and OP increased from 1.16 in 1995 to 1.34 in 2002. The mean number of transfers of FE per RS and OP was quite stable (3.82 in 1995 to 4.16 in 2002). This was explained by the increase in the number of frozen embryos per OP, averaging 1.99 in 1995 and 4.14 in 2002. The proportion of RS with only transfers of FE decreased by 20 points from 1995 (57%) to 2002 (36%). The number of transfers of FE per RS and OP was highly variable, ranging from 0 to 26. PR with FE averaged 52.1% and were significantly influenced by the number of donor cows per OP, operator of RS, embryo quality and paternal origin as well as recipient parity. Transfer of FE collected in RS with one donor cow resulted in higher PR than when more than one cow (53.0% v. 49.9%, OR=1.12, P=0.003). This could be partly explained by a higher mean number of transfers per RS and OP (1 donor cow=3.7±2.0 v.>1=5.3±3.1; P<0.05) as well as a higher proportion of RS with only FE transferred (1 donor cow=51.2% v.>1=38.5% ; P<0.05). Although the embryo quality was lower in RS combining ET of both frozen and FE than when only FE were transferred, these effects were not related to the lower quality of FE used in RS involving more than 1 donor cow (1 donor cow=1.74±0.50 v.>1=1.75±0.44; P>0.05), but may be explained by a longer time between embryo recovery and ET. After adjustment for the usually well identified sources of variation of PR, the conditions of RS as well as ET policy (leading to the selection of the best embryos for transfer as frozen and to influence on the quantity of work per OP) can significantly influence PR. These sources of variation should be taken into account when analyzing PR results.


2015 ◽  
Vol 27 (1) ◽  
pp. 163
Author(s):  
F. Du ◽  
X. Ma ◽  
B. Xu ◽  
Y. Wang ◽  
Y. Li ◽  
...  

The Japanese Black cattle (Wagyu) plays a significant part in the Japanese beef industry because it is numerically the largest breed group and comprises ~93% of the national purebred beef cow herd. Japanese Black cattle are genetically predisposed to intense marbling and to a high percentage of unsaturated fat resulting in a meat characterised by both high quality and price on the market. Like other breeds, genetic improvement and production traits in Wagyu can be fostered by the implementation of traditional reproductive strategies such as multiple-ovulation embryo transfer. A multiple-ovulation embryo transfer case study was performed in a leading production centre in Hainan island, South China. Donors (n = 40) were split into 2 groups receiving either a total dose of 400 mg of FSH Folltropin (FSH USA; Folltropin Bioniche Inc., USA; n = 24), or 10 mg of an FSH formulation produced from the Institute of Zoology of the Chinese Academy of Sciences (FSH CHN; n = 16). In both cases, dosages were equally distributed over a 4-day administration schedule. Both donors and recipients (198 heifers) were synchronized for fixed-time embryo transfer (ET) and AI, respectively, by adopting the Ovsynch protocol. Such protocol consists of GnRH administration at Day 0, followed by prostaglandin administration at Day 7 and a second administration of GnRH at Day 9. Artificial insemination was performed on donor animals at 12 and 24 h from the last GnRH administration, whereas for recipient synchronization receiving fresh embryos, the second GnRH administration was given at the time of second AI on donors, and ET was performed 7 days following the first AI. Final synchronization at the time of ET, judged by the ultrasonic presence of a functional corpus luteum, was 53% (105/198). The following parameters for FSH USA and FSH CHN were found to be not significantly different (Student's t; mean ± s.e.): i) ovulations (10.5 ± 1.2 v. 8.5 ± 1.2; P = 0.2); ii) embryos (6.3 ± 1.2 v. 5.1 ± 1.1; P = 0.5), and iii) embryos from ovulated donors (6.8 ± 1.3 v. 5.8 ± 1.1; P = 0.6). Recovered embryos from the 2 groups were also not different: i) degenerated embryos (1.0 ± 0.4 v. 0.8 ± 0.2; P = 0.6); ii) morula (4.0 ± 0.8 v. 2.6 ± 0.5; P = 0.1), and iii) early blastocysts (4.0 ± 1.2 v. 3.3 ± 0.5; P = 0.6). Blastocysts were recovered only from donors treated with FSH USA. Out of 233 recovered embryos, 34 were transferred as fresh and 71 as frozen/thawed. Pregnancy rate at 60 days following ET for fresh and frozen/thawed embryos was 47.1 and 35.2%, respectively (P = 0.2). Within frozen embryos, pregnancy rates derived from transferred morulas and blastocysts were 25 and 45.5% (P = 0.07). When considering the two sources of hormones, overall pregnancy rates were similar between the two groups (28/71, 39.4% v. 13/34, 38.2%; P = 0.9). Finally, pregnancy rates from the transfer of fresh embryos (9/21, 42.8% v. 7/13, 53.8%; P = 0.6) and frozen/thawed embryos (19/50, 38% v. 6/21, 23.8%; P = 0.6) were also not different. In conclusion, all parameters in this study did not differ between the 2 sources of FSH; however, a lower incidence of degenerated embryos and a higher pregnancy rate following transfer of frozen/thawed embryos occurred when FSH USA was used in donor animals.


2018 ◽  
Vol 52 (1-4) ◽  
pp. 22-30
Author(s):  
A K Paul ◽  
S M T Rahman

Hormonal treatment of cows at the coastal region of Barisal district of Bangladesh was performed to assess the improvement of pregnancy rate. A total of 100 cows and heifers with irregular history of cyclicity were selected randomly and divided into five treatment groups. The groups were A (treated with anthelmintic), B (treated with anthelmintic, vitamin ADE and multivitamin powder), C (treated with PGF2α), D (treated with GnRH) and E (treated with GnRH and PGF2α). Each group comprised of 20 animals. The age, breed and parity of experimental cows were considered during treatment. In the study, the cows treated with both GnRH and PGF2α (group E) showed significantly (p<0.05) higher estrus (80%) and pregnancy rate (60%) than that of group A, B, C and D. The overall estrus rates of local and crossbred cows were 64% and 70%, respectively and the pregnancy rates were 40 and 52%, respectively. The crossbred cows responded significantly (p<0.05) to hormonal treatment than that of local cows. Parity-2 cows showed higher estrus sign than that of other parities. However, the pregnancy rates were higher significantly (p<0.05) in parity-2 and parity ≥4 cows than that of parity-0, parity-1 and parity-3 cows. The pregnancy rate was also found higher in case of 4 to <5 years old cows than that of 2 to <3, 3 to <4, 5 to <6, and ≥6 years old. It may conclude that the hormonal regimen increases the pregnancy rate as well as decreases the undesired waiting of estrus and conception. Further study with more sample size will reveal the more effective treatment for cows at the coastal areas of Bangladesh.


2018 ◽  
Vol 52 (1-4) ◽  
pp. 22-30
Author(s):  
A K Paul ◽  
S M T Rahman

Hormonal treatment of cows at the coastal region of Barisal district of Bangladesh was performed to assess the improvement of pregnancy rate. A total of 100 cows and heifers with irregular history of cyclicity were selected randomly and divided into five treatment groups. The groups were A (treated with anthelmintic), B (treated with anthelmintic, vitamin ADE and multivitamin powder), C (treated with PGF2α), D (treated with GnRH) and E (treated with GnRH and PGF2α). Each group comprised of 20 animals. The age, breed and parity of experimental cows were considered during treatment. In the study, the cows treated with both GnRH and PGF2α (group E) showed significantly (p<0.05) higher estrus (80%) and pregnancy rate (60%) than that of group A, B, C and D. The overall estrus rates of local and crossbred cows were 64% and 70%, respectively and the pregnancy rates were 40 and 52%, respectively. The crossbred cows responded significantly (p<0.05) to hormonal treatment than that of local cows. Parity-2 cows showed higher estrus sign than that of other parities. However, the pregnancy rates were higher significantly (p<0.05) in parity-2 and parity ≥4 cows than that of parity-0, parity-1 and parity-3 cows. The pregnancy rate was also found higher in case of 4 to <5 years old cows than that of 2 to <3, 3 to <4, 5 to <6, and ≥6 years old. It may conclude that the hormonal regimen increases the pregnancy rate as well as decreases the undesired waiting of estrus and conception. Further study with more sample size will reveal the more effective treatment for cows at the coastal areas of Bangladesh.


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.


2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
S Watanabe ◽  
M Tomida ◽  
S Suzuki ◽  
Y Matsuda ◽  
K Yoshikai ◽  
...  

Abstract Study question When does blastomere exclusion from compaction increase and what effect does it have on the embryo? Summary answer More blastomere were excluded from compaction in embryos with irregular cleavage, resulting in lower blastocyst development rates, but no decrease in pregnancy rates at transfer. What is known already It has been reported that many of the chromosome analysis results of blastomere excluded from compaction were aneuploid, and pointed out that this exclusion may be related to the repair of blastocyst euploidy, but the effect of the number of excluded blastomere has not been reported. Study design, size, duration This is a retrospective study of 578 embryos that developed into morula with time-lapse monitoring by EmbryoScope (Vitrolife) in 2018–2019. Participants/materials, setting, methods The target embryos were classified into two groups: embryos with normal first and second cleavage (normal cleavage group) and embryos with irregular cleavage (dynamics of one cell dividing into three or more cells), called “direct cleavage”, at either cleavage (DC group), and the number of blastomere excluded from compaction during morula formation was recorded and compared. The blastocyst development rate and single blastocyst transfer pregnancy rates of the two groups were compared. Main results and the role of chance There are 286 in the normal cleavage group and 292 in the DC group. The mean number of excluded blastomere was 0.76 and 3.55, respectively, which was significantly higher in the DC group (P &lt; 0.01). Good blastocyst (Gardner classification 4 or higher) development rate was 84.5% (239/283) and 65.8% (181/275), respectively, and high grade blastocyst (Gardner classification BB or higher) development rate was 43.9% (105/239) and 14.9% (27/181) of them, both significantly higher in the normal cleavage group (P &lt; 0.01). The single blastocyst transfer pregnancy rates were 31.6% (25/79) and 32.4% (11/34), and the miscarriage rates were 24.0% (6/25) and 27.3% (3/11), respectively, neither was there a significant difference between the two groups. So, direct cleavage increased the number of blastomere excluded from compaction, decreased the rate of morula to good blastocyst development and reduced blastocyst grade, but did not affect blastocyst transfer pregnancy rate and miscarriage rate. Limitations, reasons for caution Please note that all target embryos must have developed into morula or larger (embryos that did not develop into morula will not be included in the study). Wider implications of the findings: Severe chromosomal aberrant blastomeres formed by direct cleavage were excluded from compaction, and the blastocyst development rate decreased due to a decrease in the amount of viable cells, but it is suggested that this blastomere exclusion mechanism is not related to euploidy after blastocyst development. Trial registration number Not applicable


2021 ◽  
Author(s):  
Kathryn Kost ◽  
◽  
Isaac Maddow-Zimet ◽  
Ashley C. Little

Key Points In almost all U.S. states, pregnancies reported as occurring at the right time or being wanted sooner than they occurred comprised the largest share of pregnancies in 2017, though proportions varied widely by state. The proportion of pregnancies that were wanted later or unwanted was higher in the South and Northeast than in other regions, and the proportion of pregnancies that occurred at the right time or were wanted sooner was higher in the West and Midwest. From 2012 to 2017, the wanted-later-or-unwanted pregnancy rate fell in the majority of states. However, no clear pattern emerged for any changes in the rate of pregnancies that were reported as wanted then or sooner or in the rate of those for which individuals expressed uncertainty.


2010 ◽  
Vol 39 (10) ◽  
pp. 2182-2186
Author(s):  
Cristiane Reinher ◽  
Júlio Otávio Jardim Barcellos ◽  
Vanessa Peripolli ◽  
Ênio Rosa Prates ◽  
Maria Eugênia Andrighetto Canozzi

It was evaluated in this work the effect of calving sub-season on the pregnancy rate of 7,726 multiparous Hereford beef cows grazing on natural pastures in southern Brazil, from 1994 to 2007. Calving sub-season periods were divided in 20-day intervals from August 12th to 31st; from September 1st to 20th; from September 21st to October 10th; from October 11th to 31st; from November 1st to 20th. Calving in each sub-season and pregnancy rate (PR) were analyzed by the Chi-square test. Overall means of each sub-season were 92.7% (from August 12th to 31st), 90.6% (from September 1st to 20th), 82.1% (from September 21st to October 10th), 77.7% (from October 11th to 31st), and 70.6% (from November 1st to 20th). Calving sub-season significantly influenced pregnancy rate. Cows which calved in the initial sub-season showed higher pregnancy rates than cows which calved from the fourth calving sub-season. However this difference did not occur in 1999 and in 2007 since calving rates did not statistically differ among the five sub-seasons. In 1998 and 2004, calving rates were lower and the effects of calving sub-season on pregnancy rates were higher, possibly due to climate variations. In general, pregnancy rate decreases as calving occurs later in the year, particularly in years when the weather adversely affects pasture growth.


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