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