Superovulation: A tool to produce disease free
embryos in wood bison (Bison bison athabascae)
Jesus Manuel Palomino DVM, MSc PhD Candidate
University of Saskatchewan
41st Annual Conference of the IETS, Versailles, FranceJanuary 10-13, 2015
Recipient of:Peter W. Farin Trainee Travel Scholarship Award
The Problem
Hartop et al., 2009
Brucellosis and tuberculosisDisease-free
areas
Population decline
Loss of genetic diversity
Reproductive Germ plasm Disease-free Technologies bio-bank offspring
Genetic diversity
Reproductive technologies to produce disease-free wood bison embryos
In vitro embryo productionSuperstimulation and oocyte collection
In-vivo-derived embryosSuperovulation and embryo collection
Progression of thesis research
Study 1. Inducing ovulation in Wood Bison
Study 2. Effect of LH vs hCG and progesterone on superovulation
Study 3. Does eCG improve the superovulatory response and embryo quality in wood bison?
Study 4. …
Study 5. …
What did we learn from Studies 1 to 3?· Ovarian superstimulation:
- 8 vs 2 vs 1 administrations of FSH· Induction of ovulation:
- hCG more effective than LH· Adjunct treatments
- eCG: Did not improve superovulation or embryo quality- Exogenous progesterone not needed: anovulatory season
Problem· Number of embryos collected: 4 embryos/cow· Number of Grade 1 & 2 (Freezable) embryos: 2 embryos/cow
Can we improve these numbers?
Study 4Effect of lengthening of FSH treatment on
superovulation and embryo quality in wood bison
Rationale: In cattle, lengthening the period of FSH treatment induced
a greater number of ovulations and CL compared with conventional treatment (Garcia Guerra et al., 2012)
Lengthening FSH treatment will rescue more subordinate follicles in the wave and they will grow to a sufficient stage of maturity to respond to LH and ovulate
Hypothesis Lengthening FSH treatment will improve the ovulation
rate and embryo quality in superovulated wood bison.
Objective: Determine the effect of lengthening of FSH treatment on
superovulatory response and embryo quality in wood bison during the ovulatory and anovulatory season
Experiment 1: Anovulatory season
Experimental design:
Experiment 2: Ovulatory season
Experimental design:
Animals and facilities Alternative Livestock Center, University of Saskatchewan 50 adult female and 4 male wood bison
Hydraulic chute to handle bison
Synchronization: Follicular ablation
Day -1 Day 0
Superovulation
Hyaluronic acid (Map 5, Bioniche)
hCG 3000 IU (Chorulon, Merck)
FSH 400 mg (Folltropin, Bioniche)
Artificial Insemination: Chilled semen
Semen collection: electroejaculation, 3 bulls, pooled Extender: Tryladil (MOFA) + 20% egg yolk Chilled semen 5º C (1-3 days)
Embryo collection: Non-surgical
Embryo classification (IETS, 2010)
cvzd
Table 1: Anovulatory seasonFSH Short FSH Long
Number of bison 16 16
Number of follicles ≥9 mm on Day 5 10.8 ± 2.0 10.8 ± 1.1
Ovulation rate (# ovulations / # follicles ≥9 mm)
52%(67/130)
54%(70/130)
Number of CL 4.5 ± 0.9 5.7 ± 1.0
Total ova-embryos 2.6 ± 0.5 3.2 ± 0.7
Grade 1 & 2 0.5 ± 0.3 0.3 ± 0.2
Grade 3 0.3 ± 0.2 0.1 ± 0.1
Grade 4 1.2 ± 0.4 1.8 ± 0.6
Recovery rate (# ova-embryos/# CL)
48 % (26/54)
51%(35/68)
Results are presented as Mean±SEM, otherwise stated.
Table 2: Ovulatory seasonFSH Short FSH Long P - value
Number of bison 14 14
Number of follicles ≥9 mm on Day 5 8.1 ± 1.1 9.8 ± 1.4 0.40
Ovulation rate (# ovulations / # follicles ≥9 mm)
60% (67/130)
74% (73/98)
0.07
Number of CL 4.6 ± 0.6 6.5 ± 0.9 0.13
Total ova-embryos 2.2 ± 0.5 4.4 ± 0.7 0.04
Grade 1 & 2 1.1 ± 0.4 2.3 ± 0.6 0.09
Grade 3 0.2 ± 0.1 0.8 ± 0.4 0.03
Grade 4 0.8 ± 0.2 1.7 ± 0.6 0.05
Recovery rate (# ova-embryos/# CL)
48 %(22/46)
68%(44/65)
0.06
Dias et al., 2013Garcia Guerra et al., 2012
Results are presented as Mean±SEM, otherwise stated.
Conclusions
Extended superstimulatory protocol improved embryo production during the ovulatory season
Embryo quality was not affected by lengthening the superstimulatory protocol
Repeat experiment with greater number of bison
Study 5: Production of disease-free embryos from
superovulated wood bison
Objective: Determine the effectiveness of washing procedures for
removing Brucella abortus from bison embryos exposed in vitro
Abstract 140
Level 3 laboratory – Intervac/VIDO
Acknowledgments Dr. Adams Dr. Mapletoft Dr. Woodbury Dr. McCorkell Dr. Muir/ Dr. Machin Dr. Anzar Dr. Mastromonaco Miriam Rodrigo, Hayder, Alisha Nick, Brad Kosala, Lyle
Department of Veterinary Biomedical Sciences