83 Production of live calves after transfer of in vitro-produced embryos in synchronised wood bison (Bison bison athabascae)

2022 ◽  
Vol 34 (2) ◽  
pp. 278
Author(s):  
M. Zwiefelhofer ◽  
G. Mastromonaco ◽  
E. Zwiefelhofer ◽  
G. Adams
Keyword(s):  
Bison Bison ◽  
Wood Bison

171 IN VIVO AND IN VITRO MATURATION OF WOOD BISON (BISON BISON ATHABASCAE) CUMULUS–OOCYTE COMPLEXES DURING THE OVULATORY SEASON

2014 ◽  
Vol 26 (1) ◽  
pp. 199
Author(s):  
M. P. Cervantes ◽  
M. Anzar ◽  
R. J. Mapletoft ◽  
J. M. Palomino ◽  
G. P. Adams
Keyword(s):  
In Vitro Maturation ◽  
Calf Serum ◽  
Germinal Vesicle ◽  
Bison Bison ◽  
Germinal Vesicle Breakdown ◽  
Cumulus Expansion ◽  
Nuclear Maturation ◽  
Wood Bison

Technologies are being developed to conserve the genetic diversity of wood bison. Knowledge of the characteristics of in vivo and in vitro maturation of the cumulus–oocyte complex (COC) are needed in wood bison to design efficient in vitro embryo production protocols. The objectives were to (1) determine the optimal interval after hCG treatment for in vivo maturation of COC in superstimulated wood bison, and (2) compare the characteristics of COC after in vitro and in vivo maturation. Ovarian synchronization was induced in 25 bison during October and November by giving a luteolytic dose of prostaglandin followed 8 days later by follicular ablation (Day –1). Ovarian superstimulation was induced with FSH (Folltropin-V) given i.m. on Day 0 (300 mg) and Day 2 (100 mg). A second luteolytic dose of prostaglandin was given on Day 3. Bison were assigned randomly to 5 groups (n = 5/group). The COC were collected by transvaginal follicle aspiration on Day 4 and were either assessed immediately (0 h, control), or matured in vitro for 24 or 30 h (in vitro maturation), or collected on Day 5 (in vivo maturation), 24 or 30 h after bison were given 2000 IU of hCG i.m. on Day 4. In vitro maturation was done in TCM-199 with 5% calf serum, 5 μg mL–1 LH, 0.5 μg mL–1 FSH, and 0.05 μg mL–1 gentamicin, at 38.5°C and in a 5% CO2 humidified atmosphere. Nuclear maturation was classified as germinal vesicle (GV), germinal vesicle breakdown (GVBD), metaphase I (MI), or metaphase II (MII) with anti-lamin AC/DAPI staining. Groups were compared by analysis of variance and Fisher's exact test (Table 1). A mean (±s.e.m.) of 7.3 ± 1.7 COC were collected per bison, with no difference among groups. The COC in the control (0 h) group were at the nonexpanded GV stage. Cumulus cells were more expanded after in vivo than in vitro maturation, and the percentage of fully expanded COC was the highest in the 30-h in vivo maturation group (87%; P < 0.05). The greatest number of oocytes reached MII stage after 24 h of in vitro maturation, and 30 h of in vivo maturation. In conclusion, nuclear maturation occurred more quickly in vitro compared with in vivo, but the degree and incidence of cumulus expansion was greater after in vivo maturation. The competence of oocytes to undergo fertilization and develop into embryos remains to be investigated. Table 1.Cumulus expansion and nuclear maturation of wood bison oocytes

133 In Vitro Embryo Production from Oocytes Collected from Non-Supertimulated Wood Bison (Bison bison athabascae) Following Maturation In Vitro Using Portable Incubators

2018 ◽  
Vol 30 (1) ◽  
pp. 206
Author(s):  
M. P. Cervantes ◽  
G. P. Adams ◽  
M. Anzar ◽  
J. M. Palomino ◽  
G. F. Mastromonaco
Keyword(s):  
Embryo Development ◽  
Transvaginal Ultrasound ◽  
Calf Serum ◽  
Cumulus Cells ◽  
Bison Bison ◽  
Embryo Production ◽  
Commercial Medium ◽  
Wood Bison ◽  
In Vitro Embryo Production

This study was done to determine the feasibility of in vitro embryo production in wood bison during the anovulatory season, without ovarian superstimulation or follicle wave synchronization, to simulate collection conditions in a wild or field setting. The experiment provided the opportunity to compare embryo development using 2 different maturation media and incubator systems. The cumulus-oocyte complexes (COC) were collected by transvaginal ultrasound-guided follicle aspiration during May from non-superstimulated bison. Compact COC were allocated to 2 groups and matured in standard maturation medium using a portable gassed incubator, or in commercial medium using a portable non-gassed incubator. In the former (Standard), the COC were placed in a round-bottomed tube containing TCM-199 medium with 5% calf serum, 5 μg mL−1 LH, 0.5 μg mL−1 FSH, and 0.05 μg mL−1 gentamicin, and the tube was placed in a portable incubator with 5% CO2. In the latter (Commercial), COC were placed in a round-bottom tube containing the commercial medium (Boviteq, Saint-Hyacinthe, QC, Canada), and placed in a portable incubator without CO2. After 24 h of maturation, oocytes were fertilized in vitro (Day 0) in Brackett-Oliphant medium at 38.5°C in a conventional incubator with 5% CO2 humidified atmosphere. Presumptive zygotes were cultured in CR1aa plus 5% calf serum, at 38.5°C and in 5% CO2, 5% O2, and 90% N2 and high humidity. Cleavage was recorded on Day 3 and embryo development was recorded on Day 7. Cleavage and transferable embryo rates (calculated from the total number of oocytes submitted to IVF) were compared between groups by chi-squared test. No difference in cleavage rates was observed between Standard and Commercial treatment groups [68.1 (32/47) v. 79.2% (57/72), respectively; P = 0.25], nor in morula plus blastocyst rates on Day 7 (36.2 v. 45.8%, respectively; P = 0.39). However, the rate of transferable embryos (grade 1 and grade 2) on Day 7 was higher in the Commercial group (38.9 v. 12.8%; P < 0.01). Of the COC in the Commercial group, a higher number of morula plus blastocyst were observed to be compact good COC (>3 layers of cumulus cells) than compact regular COC (1-3 layers of cumulus cells) (66.7 v. 31.0% respectively; P < 0.05), along with a higher number of transferable embryos on Day 7 (60.0 v. 23.8%, respectively; P < 0.05). In conclusion, wood bison oocytes collected during the anovulatory season from non-superstimulated, non-synchronized bison and matured in vitro using portable incubators were competent to develop to the morula and blastocyst stages following IVF and culture. These results are important for future plans that require transporting oocytes from remote collection sites to the IVF laboratory, particularly with respect to the effectiveness of commercial maturation media which does not require CO2 supplementation. Research was supported by the Natural Sciences and Engineering Research Council of Canada.

Effect of season and superstimulatory treatment on in vivo and in vitro embryo production in wood bison ( Bison bison athabascae )

2019 ◽  
Vol 55 (1) ◽  
pp. 54-63
Author(s):  
Jesus Manuel Palomino ◽  
Gabriela F. Mastromonaco ◽  
Miriam P. Cervantes ◽  
Reuben J. Mapletoft ◽  
Muhammad Anzar ◽  
...  
Keyword(s):  
Bison Bison ◽  
Embryo Production ◽  
Wood Bison ◽  
In Vitro Embryo Production

In vitro embryo production in wood bison ( Bison bison athabascae ) using in vivo matured cumulus-oocyte complexes

2017 ◽  
Vol 89 ◽  
pp. 122-130 ◽  
Author(s):  
Miriam P. Cervantes ◽  
J. Manuel Palomino ◽  
Muhammad Anzar ◽  
Reuben J. Mapletoft ◽  
Gabriela F. Mastromonaco ◽  
...  
Keyword(s):  
Bison Bison ◽  
Embryo Production ◽  
Wood Bison ◽  
In Vitro Embryo Production

140 EFFECTIVENESS OF WASHING PROCEDURES FOR REMOVING BRUCELLA ABORTUS FROM IN VIVO-DERIVED WOOD BISON EMBRYOS

2015 ◽  
Vol 27 (1) ◽  
pp. 162
Author(s):  
J. M. Palomino ◽  
M. P. Cervantes ◽  
G. Mastromonaco ◽  
R. J. Mapletoft ◽  
B. Allan ◽  
...  
Keyword(s):  
Zona Pellucida ◽  
Brucella Abortus ◽  
Calf Serum ◽  
Petri Dish ◽  
Bison Bison ◽  
Genetic Reserve ◽  
Wood Bison ◽  
Wood Buffalo National Park

Endemic brucellosis threatens wild herds of wood bison (Bison bison athabascae) in and around Wood Buffalo National Park, the largest genetic reserve of wood bison in the world. The overall goal of our project was to produce and preserve disease-free embryos for the purpose of conserving the genetic diversity of this species. The aim of the present experiment was to determine the effectiveness of washing procedures for removing Brucella bacteria from in vivo-derived wood bison embryos exposed in vitro to the pathogen. Wood bison cows were given 300 mg im of Folltropin diluted in 0.5% hyaluronan on the day of follicle wave emergence (Day 0) and 100 mg im of hyaluronan on Day 2, and then given 2500 IU im of hCG on Day 5 and inseminated 12 and 24 h later. Embryos were collected on Day 13. The experiment was done in 6 replicates (n = 4 bison/replicate) and an average of 9 embryos/replicate were collected. Zona pellucida-intact embryos were kept in holding medium (PBS + 2% fetal calf serum) and transported to a Biosafety Level 3 laboratory at the International Vaccine Centre, University of Saskatchewan. Embryos were transferred through 5 aliquots of holding medium to remove any contaminant before exposure to Brucella. Embryos were divided equally into 2 Petri dishes (representing later wash groups with v. without antibiotics) containing 2.7 mL of holding medium (n = 2 to 7 embryos per dish/replicate). In a Class II biosafety cabinet, Brucella abortus biovar 1 (1 × 107 to 1 × 109 CFU mL–1 in 0.3 mL) was added to each Petri dish and incubated for 2 h at 37°C in 8% CO2. A sample of holding medium was taken before exposure and after incubation for culture as negative and positive controls, respectively. After incubation, embryos in each Petri dish were subjected to a 10-step washing procedure (according to the IETS Manual, 2010) using wash medium (PBS + 0.4% BSA) without antibiotics or with antibiotics (100 IU mL–1 of penicillin + 100 μg mL–1 of streptomycin). The embryo wash medium was cultured at wash steps 1, 3, 6, and 9. After the tenth wash, the zona pellucida of each embryo was ruptured mechanically using a glass pipette and embryos were cultured individually. Culturing of samples was done on sheep blood agar and specific identification of Brucella organisms was done by PCR. Brucella abortus was detected in 3 embryos from the group washed in medium without antibiotics (3/27), whereas all embryos washed in medium with antibiotics were culture negative (0/27). Brucella abortus was not detected in wash media after the third wash in either group (with or without antibiotics). In summary, Brucella abortus was removed from 89% of in vitro-exposed wood bison embryos using the washing procedure without antibiotics, and from 100% using the washing procedure with antibiotics. Results validate the embryo washing technique for producing Brucella-free wood bison embryos. Thanks to the Canadian Food Inspection Agency for the field strain of Brucella abortus, Bioniche AH for Folltropin and embryo collection supplies, Merck AH for hCG (Chorulon), and Intervac/VIDO for technical and logistical support.

363 EXTENDING THE IN VIVO MATURATION TIME TO PERMIT FLEXIBLE TIMING OF OOCYTE COLLECTION IN SUPERSTIMULATED WOOD BISON (BISON BISON ATHABASCAE)

2015 ◽  
Vol 27 (1) ◽  
pp. 270
Author(s):  
M. W. von der Porten ◽  
M. P. Cervantes ◽  
J. M. Palomino ◽  
G. P. Adams
Keyword(s):  
Collection Efficiency ◽  
Animal Health ◽  
Reproductive Technologies ◽  
Ovulation Rate ◽  
Bison Bison ◽  
Collection Rate ◽  
Oocyte Collection ◽  
Wood Bison

Wood bison are a species threatened by endemic brucellosis and tuberculosis. Reproductive technologies are being developed in an effort to ensure the genetic diversity of wild wood bison, and to prevent disease transmission to healthy bison, livestock, and humans. For the purposes of IVF, recent results revealed that cumulus cell expansion was more extensive in in vivo- v. in vitro-matured cumulus-oocyte complexes (COC), and more oocytes reached maturity after 30 v. 24 h of in vivo maturation following hCG treatment (Cervantes et al. 2013 Reprod. Fert. Develop. 25, 283). An experiment was designed to determine the effects of an additional 4 h of in vivo maturation on follicle development, unwanted ovulation, and COC collection efficiency. Wood bison cows (n = 28) underwent transvaginal ultrasound-guided follicle ablation to induce emergence of a new follicular wave (Day 0 = day of wave emergence, 1 day after ablation) during the non-breeding season. Bison were given FSH diluted in hyaluronan IM on Days 0 (300 mg) and 2 (100 mg), and 2500 IU hCG IM on Day 4. Bison were then assigned randomly to 2 groups (n = 14 per group) in which transvaginal oocyte collection was done at either 30 or 34 h after hCG treatment. The number and size of follicles available for aspiration (i.e. = 5 mm) was compared between groups by Student's t-test. Binomial data (COC collection rate and ovulation rate) were compared by chi-square, and the proportion of cows that ovulated was compared using a Fisher's exact test. Ovulation was defined as the sudden disappearance of follicles ≥10 mm from the hCG treatment to the time of COC collection. The numbers of follicles ≥5 mm and ≥10 mm at the time of COC collection were not different between the 30 and 34 h groups (19.0 ± 1.4 v. 17.4 ± 2.4, and 9.5 ± 1.2 v. 7.7 ± 1.8), nor was the average size of follicles = 5 mm (9.9 ± 0.2 v. 9.8 ± 0.2 mm). The number of follicles aspirated was similar between the 30 and 34 h groups (16.4 ± 1.4 v. 13.4 ± 2.1), but the pre-collection ovulation rate was lower in the 30 h group (12/89 [13.5%] v. 47/147, [32.0%]; P = 0.003), as was the proportion of bison that ovulated (3/14 v. 10/14, P = 0.02). The COC collection rate was lower in the 30 v. 34 h group (64.3% v. 78.2%; P = 0.003), but the total number of COC collected per bison was similar (10.6 ± 1.7 v. 10.5 ± 1.5). Although waiting for 34 h before COC collection resulted in a larger proportion of unwanted ovulations, a greater collection efficiency in the 34 h group resulted in a similar number of COC collected per bison. We conclude that the 30 to 34 h in vivo maturation window provides flexibility for the purposes of oocyte collection and immediate in vitro fertilization in wood bison.We thank Bioniche Animal Health for providing FSH (Folltropin-V) and hyaluronan (MAP-5), and Merck Animal Health for hCG (Chorulon).

183 IN VITRO EMBRYO PRODUCTION: A TOOL TO PRESERVE THE THREATENED WOOD BISON (BISON BISON ATHABASCAE)

2016 ◽  
Vol 28 (2) ◽  
pp. 222
Author(s):  
M. P. Cervantes ◽  
J. M. Palomino ◽  
M. Anzar ◽  
R. J. Mapletoft ◽  
G. Mastromonaco ◽  
...  
Keyword(s):  
In Vitro Maturation ◽  
Animal Health ◽  
Developmental Competence ◽  
Bison Bison ◽  
High Humidity ◽  
Embryo Production ◽  
Wood Bison ◽  
In Vitro Embryo Production

In vitro embryo production is being developed as a tool to restore genetic diversity and eliminate endemic disease in wood bison. In a recent study in wood bison, we found that more oocytes reached maturity after 30 h v. 24 h of in vivo maturation following hCG treatment (Cervantes et al. 2014 Reprod. Fertil. Dev. 26, 199). An additional 4 h of in vitro maturation after an in vivo maturation period of 30 h also had a positive effect on developmental competence. The present study was designed to test the hypothesis that extending the in vivo maturation time (i.e. extending the interval between hCG treatment and cumulus-oocyte complex (COC) collection) from 30 to 34 h will improve in vitro embryo production in wood bison. Follicular wave development was synchronised among female wood bison (n = 28, 6 to 10 years old) by transvaginal follicular ablation. The study was done in 4 replicates (n = 7 bison per replicate). Bison were given FSH 1 day (300 mg) and 3 days (100 mg) after ablation for ovarian superstimulation, and hCG (2500 IU) 5 days after ablation to induce COC maturation in vivo. Bison were divided randomly into 2 groups (n = 14/group) in which COC were collected transvaginally at either 30 h or 34 h after hCG treatment. Expanded COC from the 30 h group were fertilised after 4 h of in vitro maturation, while expanded COC from the 34 h group were fertilised immediately. Oocytes and sperm were co-incubated (Day 0 = day of fertilization) for 18 h at 38.5°C in 5% CO2 in air and high humidity. Presumptive zygotes were cultured in 4-well dishes containing 500 μL well–1 of CR1aa medium at 38.5°C, 5% CO2, 5% O2, 90% N2 and high humidity, and assessed on Days 3, 7, and 8 (Day 0 = day of fertilization). Data were compared between groups by Chi-squared analysis. No effect of replicate was found. Compared to the 30 h group, the 34 h group had a greater cleavage rate [55/74 (74%) v. 49/86 (57%); (P < 0.05)], and a greater blastocyst rate on Day 7 [25/74 (34%) v. 9/86 (10%); (P < 0.05)] and Day 8 [(40/74 (54.1%) v. 32/86 (37.2%); (P < 0.05)]. We concluded that an extended period of in vivo maturation is beneficial for embryo production after in vitro fertilization in wood bison. We thank Vetoquinol Canada for providing FSH (Folltropin-V) and hyaluronan (MAP-5) and thank Merck Animal Health for hCG (Chorulon).

In vitro- production of embryos using immature oocytes collected transvaginally from superstimulated wood bison ( Bison bison athabascae )

2017 ◽  
Vol 92 ◽  
pp. 103-110 ◽  
Author(s):  
Miriam P. Cervantes ◽  
J. Manuel Palomino ◽  
Muhammad Anzar ◽  
Reuben J. Mapletoft ◽  
Gabriela F. Mastromonaco ◽  
...  
Keyword(s):  
Bison Bison ◽  
In Vitro Production ◽  
Immature Oocytes ◽  
Wood Bison ◽  
Vitro Production

In vivo and in vitro maturation of oocytes collected from superstimulated wood bison ( Bison bison athabascae ) during the anovulatory and ovulatory seasons

2016 ◽  
Vol 173 ◽  
pp. 87-96 ◽  
Author(s):  
Miriam P. Cervantes ◽  
J. Manuel Palomino ◽  
Muhammad Anzar ◽  
Reuben J. Mapletoft ◽  
Gregg P. Adams
Keyword(s):  
In Vitro Maturation ◽  
Bison Bison ◽  
Wood Bison

158 IN VITRO DEVELOPMENT OF WOOD BISON (BISON BISON ATHABASCAE) EMBRYOS BY INTERSPECIES SOMATIC CELL NUCLEAR TRANSFER

2009 ◽  
Vol 21 (1) ◽  
pp. 178 ◽  
Author(s):  
B. M. Kumar ◽  
E. St. John ◽  
P. M. Mackie ◽  
W. A. King ◽  
G. F. Mastromonaco
Keyword(s):  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Cell Number ◽  
Total Cell ◽  
Bison Bison ◽  
Blastocyst Development ◽  
Total Cell Number ◽  
Wood Bison

Wood bison (Bison bison athabascae) are currently classified as threatened in Canada. Interspecies somatic cell nuclear transfer (iSCNT) is a valuable tool for embryo production in non-domestic species in which access to gametes is limited. Unlike fertilization, SCNT allows preservation of the entire genome, thus avoiding dilution of valuable alleles, an important factor for the preservation of genetic diversity. The present study compared the developmental competence of iSCNT embryos reconstructed from adult female wood bison ear fibroblasts (bison NT) with development of embryos reconstructed from adult female cattle ear fibroblasts (cattle NT). Domestic cattle (Bos taurus) oocytes were used as recipient ooplasm for both donor cell types. In vitro fertilized (IVF) and parthenogenetic (PA) cattle embryos were used as controls. Fibroblast cultures at passages 3 to 5 confluent for 5 days were used for SCNT. Mature oocytes were enucleated, reconstructed by transfer of donor cells, and fused with an electrical stimulus of 1.5 kV cm–1 for 40 μs in 0.28 m mannitol containing 100 μm CaCl2 and MgCl2. Oocytes for parthenogenesis and following reconstruction were activated for 5 min in 5 μm ionomycin followed by 5 h in 10 μg mL–1 cycloheximide. Embryos produced by IVF, PA, and SCNT were cultured in modified synthetic oviductal fluid medium at 38.5°C in 5% CO2, 5% O2, 90% N2. Cleavage, blastocyst development to day 8, apoptosis (TUNEL assay, Roche Diagnostics, IN, USA), and total cell number were evaluated. Statistical analyses were carried out using one-way ANOVA, followed by Tukey post hoc analysis or the equivalent nonparametrical Kruskal-Wallis test. Cleavage rate was significantly (P < 0.05) higher in the IVF group than in all other groups (86.9 ± 2.9% v. 71.6 ± 4.5% to 78.1 ± 5.1%). Blastocyst rates, expressed as a percentage of cleaved embryos, were similar among all treatment groups (33.4 ± 3.3% to 39.8 ± 5.7%) except for bison NT which had significantly (P < 0.05) lower development to blastocyst (19.2 ± 5.5%). The percentages of TUNEL-positive cells among PA embryos (6.6 ± 1.5%) and bison NT embryos (6.7 ± 2.4%) were significantly (P < 0.05) higher than in IVF embryos (4.2 ± 1.0%), but similar to cattle NT embryos (5.4 ± 1.7%), which did not differ from the IVF group. Total cell number was significantly (P < 0.05) higher in the IVF group than in all other groups (133.2 ± 10.2 v. 91.2 ± 7.8 to 100.1 ± 12.9). These results confirm that in vitro-matured domestic cattle oocytes can serve as suitable recipients of wood bison somatic cells and that iSCNT may provide a possible alternative for embryo production and genetic preservation of endangered cattle species. Both the incidence of apoptotic cells and total cell number did not differ between cattle and bison NT embryos; thus other factors must play a role in the significantly decreased blastocyst development observed in bison NT embryos. This work was supported by Endangered Species Reserve Fund, Toronto Zoo, and the Canada Research Chairs program.

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