scholarly journals Establishing an in vitro production program for buffalo embryos (Bubalus bubalis) in Colombia

2015 ◽  
pp. 4495-4504
Author(s):  
Felipe Gamarra P ◽  
Viviana Rendón V ◽  
Aldemar Chávez R ◽  
Leonardo Perez S ◽  
Walter Cardona-Maya ◽  
...  
Keyword(s):  
Bubalus Bubalis ◽  
Fertilization In Vitro ◽  
Natural Cycle ◽  
Ultrasound Guided ◽  
In Vitro Production ◽  
Production Program ◽  
Frozen Semen ◽  
Sexed Semen ◽  
Vitro Production

Objective. Evaluate the results of the standardization of the in vitro production program of buffalo embryos, using oocytes obtained by ultrasound guided oocyte puncture during the 2012 breeding season in Colombia. Materials and methods. Fifty seven buffalo females were selected for ultrasound guided transvaginal aspiration of follicles, oocytes were identified within follicular fluid, classified and transported to the laboratory and matured in vitro for 18 to 20 hours. Frozen semen of seven Mediterranean bulls were used, motile sperm was obtained using the Percoll technique and oocytes were inseminated with 2 million sperm/ml. Presumptive zygotes were cultured for 6 days, grade 1 embryos obtained were frozen using ethylene glycol. Embryos were transferred to females on day 5 during natural cycle. Results. 97 aspirations were performed on the 57 animals, in 8.2% of the aspirations no oocytes were found. 8 oocytes/aspiration were obtained. Of the 783 oocytes, 92% were classified as viable (721/783) and were fertilized. The cleavage and blastocyst rate were 23% and 19% respectively. 37 embryos were transferred and 11 pregnancies were obtained, confirmed by rectal palpation 60 days after transfer, with a pregnancy rate of 29.7%. Conclusions. The results presented here are comparable with others in literature and shows the feasibility of producing in vitro embryos and pregnancies after the standardization of current protocols, with normal and sexed semen and transfer during natural cycle in buffalo.Key words: Buffaloes, embryo, fertilization in vitro, reproduction (Source: MeSH).

scholarly journals Differences in Parameters of an Embryo In Vitro Production Program between Cattle (Bos Indicus) and Buffaloes (Bubalus bubalis)

2020 ◽  
Vol 9 ◽  
pp. 29-37
Author(s):  
Jesús Alfredo Berdugo ◽  
Ariel Marcel Tarazona-Morales ◽  
José Julian Echevererry ◽  
Jose Luis Konrad ◽  
Gustavo Angel Crudeli ◽  
...  
Keyword(s):  
Bos Indicus ◽  
Bubalus Bubalis ◽  
In Vitro Production ◽  
Production Program ◽  
Vitro Production

scholarly journals Differences in Parameters of an Embryo In Vitro Production Program between Cattle (Bos Indicus) and Buffaloes (Bubalus bubalis)

2020 ◽  
Vol 9 ◽  
pp. 29-37
Author(s):  
Jesús Alfredo Berdugo ◽  
Ariel Marcel Tarazona-Morales ◽  
José Julian Echevererry ◽  
Jose Luis Konrad ◽  
Gustavo Angel Crudeli ◽  
...  
Keyword(s):  
Bos Indicus ◽  
Bubalus Bubalis ◽  
In Vitro Production ◽  
Production Program ◽  
Vitro Production

292 EXPRESSION PROFILES OF STRESS AND METABOLIC MARKER GENES DURING IN VITRO PRODUCTION OF BUFFALO (BUBALUS BUBALIS) EMBRYOS

2006 ◽  
Vol 18 (2) ◽  
pp. 253
Author(s):  
R. Rajhans ◽  
G. S. Kumar ◽  
G. T. Sharma
Keyword(s):  
Bubalus Bubalis ◽  
Marker Genes ◽  
Hsp 70 ◽  
In Vitro Production ◽  
Glucose Transporter 1 ◽  
Total Rna ◽  
Gene Transcripts ◽  
Vitro Production ◽  
Glut1 Gene

An increased understanding of the pre-implantation embryo developmental stage, with respect to physiological interaction of embryo with its micromilieu both in vivo and in vitro, is imperative to comprehend the events of pre-implantation development.The objective of the present study was to examine the temporal expression of heat shock protein 70 (Hsp-70) and glucose transporter 1 (Glut1) genes in pre-implantation-stage buffalo embryos produced under the standard in vitro production (IVP) system. Embryos were produced from slaughterhouse ovaries employing standard in vitro embryo production protocol, and presumptive zygotes produced following IVM/IVF were cultured in vitro in mSOF under mineral oil; FCS (10%) was added at 48 h post-insemination (hpi).The time series of development at stages being zygote (18-20 hpi), 2-1 cell (48 hpi), 8-16 cell (94-96 hpi), morula (120-144 hpi), and blastocyst (168-192 hpi), pre-implantation embryos conforming to the above developmental pattern were considered as 'fast-cleaving embryos', and all the embryos that did not conform to the above developmental timing were regarded as 'slow-cleaving embryos'. Pools of immature oocytes (IM, 120), Matured oocytes (MO, 120), 8-16 cell stages (8-16, 70), morula (M, 28), and blastocyst (B, 9) were collected and prepared for total RNA isolation and RT-PCR for the specific transcripts, with �-actin as loading control. The total RNA content ranged from 2.5 to 5.0 ng per oocyte/embryo. Presence of Hsp 70 and Glut1 gene transcripts was assessed in different stages of buffalo pre-implantation embryos using primers designed from bovine Hsp 70 and Glut1 by using the OLIGO program. RT was standardized using the embryo equivalent of 1-10 oocytes/embryo as the template as described by Arcellana-Panlilio and Schultz 1993 (Methods Enzymol. 225, 303-328) with PCR conditions being 59�C and 62�C for 45 s with 39 cycles for Glut1 and Hsp 70 gene transcripts, respectively. Amplicons were subjected to restriction digestion and sequencing (Acc. No. AJ812563, AJ812564). The expression of Hsp 70 throughout pre-implantation development in the fast-cleaving embryos indicated their maternal and zygotic origin, but transcripts of the Hsp 70 gene, represented by a single 488-bp amplicon, were not detected in slow-cleaving embryos, suggesting altered zygotic expression. Glut1 expression was prominent from the 8-16 cell stage, indicating a metabolic shift from pyruvate to glucose after the pre-compaction stage. For slow-cleaving embryos, transcripts of the Glut1 gene, represented by a single 327-bp amplicon, were absent during morula- and blastocyst-stage embryos, indicating the poor developmental competence of these embryos, which morphologically appeared normal. These transcription patterns reflect the embryonic response to the in vitro culture conditions and also correlate with the embryo quality and the speed of development of the pre-implantation buffalo (Bubalus bubalis) embryos.

scholarly journals 281REDUCTION OF POLYSPERMY IN PORCINE IN VITRO FERTILIZATION BY MODIFIED SWIM-UP METHOD

2004 ◽  
Vol 16 (2) ◽  
pp. 260
Author(s):  
C.-H. Park ◽  
B.-S. Koo ◽  
J.-I. Yun ◽  
M.-G. Kim ◽  
S.-G. Lee ◽  
...  
Keyword(s):  
Sperm Concentration ◽  
Fertilization In Vitro ◽  
In Vitro Production ◽  
Common Procedure ◽  
Developmental Potential ◽  
Vitro Fertilization ◽  
A Cell ◽  
The Difference ◽  
Vitro Production

In vitro production (IVP) of porcine embryos facilitates research related to biotechnology and biomedicine. Even though many attempts have been made to optimize the IVP of porcine embryos, the outcome is still unsatisfactory compared to other species, such as mouse and cattle. The high incidence of polyspermic fertilization is one of the major causes lowering the overall efficiency of porcine IVF. The common procedure for fertilization in vitro involves the co-culture of both gametes in the medium drop, which increases sperm concentration and incidence of polyspermy. Therefore, the present study was carried out to increase the efficiency of porcine IVF by reducing polyspermy using a modified swim-up method. This method modifies conventional swim-up washing by placing oocytes directly at the time of washing. Porcine oocytes were aspirated from ovaries and matured. Sperm pellet was prepared in the tube and mature oocytes were placed on a cell strainer with 70-μm pore size (Falcon 2350) at the top of the tube. After fertilization, the oocytes were fixed and stained for examination. Also, the developmental potential of fertilized embryos was measured to evaluate for the feasibility of this method. While penetration rates were similar in both methods (86.67±2.36% to 83.33±1.36%), there was a significant reduction of polyspermy in the modified swim-up method (17.50±1.60%) compared to the control (44.1±3.70%) (P<0.05). Subsequent culture showed higher rate of blastocyst formation in the modified swim-up method (20.44±0.99%) than in the control (15.73±3.26%) (P<0.05), even though the difference was not significant. These results suggest that, by controlling the number of spermatozoa reaching the oocytes, porcine oocytes might be protected from polyspermy in vitro. Also, the developmental potential of the fertilized embryos using this method could be improved by increasing the pool of spermatozoa with better quality. Further optimization of the procedure is required to impliment this method in routine porcine IVF.

scholarly journals Factors affecting the in vitro production of bovine embryos in a commercial program

2021 ◽  
Vol 10 (2) ◽  
pp. e16110212264
Author(s):  
Bárbara Gomes Rodrigues Nogueira ◽  
Lilian Franscico Arantes de Souza ◽  
Raquel Zaneti Puelker ◽  
Inês Cristina Giometti ◽  
Sheila Merlo Garcia Firetti ◽  
...  
Keyword(s):  
Bos Indicus ◽  
Bovine Embryos ◽  
In Vitro Production ◽  
Chi Square ◽  
Factors Affecting ◽  
Group Frequency ◽  
Sexed Semen ◽  
Using Data ◽  
Vitro Production

Variability in the production of bovine blastocysts per session of ultrasound-guided follicularaspiration(OPU) is still an obstacle in commercial production.Thus, the objective of the current work was to verify factors that influence in vitro embryo production (PIVE), using data from a commercial laboratory.Data from 2014 to 2016 were analyzed, referring to 799 OPU sessions of adult bovine females of the breeds: Nelore (n=83), Girolando (n=73), Brangus (n=49), HPB (n=20), and Senepol (n=10).The influence of five variables was analyzed: breed, genetic group, frequency of aspiration, type of semen (sexed/conventional), and seasonality (spring/summer vs. autumn/winter) on the rate of viable oocytes, cleavage, and blastocyst production (bl).The protocols for the in vitroproduction of embryos (IVP) followed the routine established by the partner laboratory of the present study. The statistical analysis of the data was performed using SAS, with the Chi-square test.The Girolando and Nelore breeds had a higher (p=0.0001) number of blastocysts/OPU session, as well as the Bos indicus blood group.Blastocyst production was higher (p=0.0059) with sexed semen compared to conventional semen (6.6 x 5.2 bl/OPU session); a higher frequency of aspirations also increased (p=0.005) the number of bl/OPU session (7.7 x 5.9).We conclude that the in vitro production of bovine embryos is influenced by the analyzed factors and knowledge of these variables could guide the commercial use of OPU-PIVE. 

264 EFFECT OF FROZEN SEMEN BATCH ON THE IN VITRO PRODUCTION OF BOVINE EMBRYOS

2010 ◽  
Vol 22 (1) ◽  
pp. 289
Author(s):  
M. B. Fernandes ◽  
T. L. G. Torregrossa ◽  
R. B. Prado ◽  
R. A. Vila ◽  
F. P. Elias ◽  
...  
Keyword(s):  
Embryo Development ◽  
Bos Indicus ◽  
Cumulus Cells ◽  
Development Rate ◽  
In Vitro Production ◽  
Vitro Fertilization ◽  
Significance Level ◽  
Frozen Semen ◽  
Vitro Production

Within an in vitro production controlled system, bulls differ with respect to their semen potential in generating embryos when the variables of maternal effect are minimized (Marquant-le-Guienne and Humblot 1992 Ann. Zootech. 41, 361-370). We have tested the hypothesis that even with this variation among bulls, there is also an intra-bull variation, according to the frozen semen batch used in the in vitro fertilization, identified with the date of ejaculate and its freezing. In an embryo commercial production system, over 12 months, 10 619 viable oocytes were obtained by ultrasound-guided follicular aspiration from 642 Nelore cows (Bos indicus). The oocytes were matured in vitro for 24 h in TCM-199 supplemented with 0.5 μg mL-1 FSH, 50 μg mL-1 LH, and 10% fetal bovine serum. They were then inseminated for 18 hours in IVF-TALP medium, using the semen from 4 bulls (A to D) subdivided into 4 frozen batches (I to IV) and selected by 45/90% Percoll gradient. Putative zygotes surrounded in cumulus cells were transferred in CR2aa medium drops (Rosenkrans and First 1994 J. Anim. Sci. 72, 434-437) for 163 h at 39°C in a humidified atmosphere of 5% CO2 in air. The oocyte distribution, the total number of blastocysts, and the embryo development rate by each bull and respective batch are described in Table 1. The chi-square test was measured with a significance level of P < 0.05 and showed that there is a difference between the used batches of each bull regarding the development rate of blastocysts 163 h after IVF Therefore, there is intra-bull variation in the ability to develop in vitro embryos according to the batch of frozen semen. Table 1.Viable oocytes (VO), total blastocysts (TB), and embryo development rate (%E) by bull and batch used in IVF

Laboratory production of buffalo (Bubalus bubalis) embryos

1998 ◽  
Vol 10 (5) ◽  
pp. 379 ◽  
Author(s):  
P. Palta ◽  
M. S. Chauhan
Keyword(s):  
In Vitro Maturation ◽  
Large Scale ◽  
Follicle Stimulating Hormone ◽  
Bubalus Bubalis ◽  
Blastocyst Stage ◽  
In Vitro Production ◽  
Nuclear Maturation ◽  
Fertilization Rates ◽  
Vitro Production

There is an increasing interest in large-scale in vitro production (IVP) of buffalo embryos through in vitro maturation (IVM), fertilization (IVF) and culture (IVC) of oocytes for faster multiplication of superior germplasm. The recovery of total and usable quality oocytes from slaughterhouse ovaries is low in this species. The nuclear maturation rates of buffalo oocytes matured in the presence of follicular fluid or serum and hormones like luteinizing hormone, follicle-stimulating hormone and oestradiol vary from 70 to 80% and are comparable to those reported for cattle oocytes. However, with fertilization rates of 40–55%, and the yield of blastocysts at around 10–15%, the efficiency of IVP is much lower than that in cattle. The in vitro sperm preparation procedures and the systems employed for performing IVF and culture of zygotes up to blastocyst stage are suboptimal and need substantial improvements. The quality and viability of blastocysts produced need to be checked by cell count, and after transfer to synchronized recipients, for development of quality control standards.

Sign in / Sign up

Export Citation Format

Share Document