Benefits of OPU/IVF (IVP) in Dairy Cattle. M.V. Ramon Tosta Duarte Deforest WI - Reproduction Supervisor ST-Genetics

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1 Benefits of OPU/IVF (IVP) in Dairy Cattle M.V. Ramon Tosta Duarte Deforest WI - Reproduction Supervisor ST-Genetics

2 What is OPU/IVF (IVP)? ARTs used at ST Genetics Donor Selection Donors Animal Welfare Care of Juvenile Donors Equipment Environment Conditions OPU Technique Effect of age on IVP Benefits vs Challenges of IVP Advantages of SexedUltra 4M in IVP Expectations

3 Introduction ST Vienna Farms Donor center with 300 heifers In Vitro and In vivo embryo production Genomic animals Holstein (Red and BW), Jersey, Brown Swiss

4 What is OPU/IVF (IVP)? In vitro production of ruminant embryos is a three-step process involving oocyte maturation, oocyte fertilization and in vitro culture. Only 30-40% of such oocytes reach the blastocyst stage, at which they can be transferred to a recipient or frozen for future use (Lonergan, 2007). Ultrasound-guided transvaginal follicular aspiration for ovum pick-up (OPU), is a non-invasive procedure for recovering oocytes from antral follicles in live animals. Together with in vitro fertilization of oocytes, OPU has been taken as a most flexible and repeatable technique to produce embryos from any given live donor (Qi et al, 2013).

5 ARTs used at STgenetics IVP (in vitro embryo production) 70% of our Donors MOET (multiple ovulation embryo transfer) 30 % of our Donors AI (artificial insemination) 100% of our donors

6 Genetic Value Donor Selection Health status Breed (Bos Taurus vs Bos Indicus) Body condition score (BCS); Nutritional Status Reproductive evaluation (Ovaries, Follicular Population, etc) Reproductive status (Open, Pregnant) Age (Pre-Pubertal, Pubertal, Mature) Cow production status (Milking, or Dry)

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8 DELTA DAUGHTERS Donors

9 Animal Welfare

10 Care of Juvenile Donors

11 Equipment Vacuum Pump (negative pressure) Tube Warmer (temperature) Ultrasound Machine (with micro convex probe) Guide for Ovum Pick up Tubing line, silicone cork, 50 ml tube Needles for OPU

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14 Environment Conditions

15 OPU Technique Epidural prior to the procedure/animal restrained Positioning of the ovary per rectum Visualization of follicles by the transducer Ovum pick up by needle Search and evaluation of oocytes

16 Ovum Pick Up (OPU)

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18 In Vitro Production Photos by Daniela Pereira

19 Effect of Age on IVP 10 mo 11 mo Pubertals and Pre-purbertal Heifers Titolo del grafico 4,1 12,7 9 mo 10 mo 3,9 13,8 3,6 8 mo 8-9 mo 15,7 0,0 0,0 4,5 4,0 9,0 8,0 12,0 13,5 16,0 18,0 Embryos Oocytes Chart OPUs, 381 Donors, 75 bulls, 8277 Oocytes, 2604 Embryos

20 Effect of Age on IVP Mature, Pregnant Heifers and Cows mo 4,3 Titolo del grafico 13, mo mo mo 5,3 14, mo mo 4,8 13, mo mo mo 5,3 14,2 12 mo 12 mo 3,9 9,6 0,0 0 4,0 4 8, , ,0 Embryos Oocytes Chart OPUs, 381 Donors, 75 bulls, 8277 Oocytes, 2604 Embryos

21 Effect of Age on IVP 15,7 13, , ,2 13,7 14,1 13,9 9,6364 3,6 3,9 4,1 3,9 5,3 4,8 5,3 4,3 8-9 m 10 m 11 m 12 m m m Viable Oocytes Emb Prod Preg Transf Frozen Chart OPUs, 381 Donors, 75 bulls, 8277 Oocytes, 2604 Embryos

22 Effect of Age on Fertility P=0.006 N=11852

23 Effect of Age on Fertility P=0.002 N=5147

24 Benefits vs Challenges of (IVP) Benefits Non invasive technique Use of hormones or not Donors can be aspirated very often Animals can be aspirated at any time and stage (juvenile and pregnant) Can freeze or vitrify embryos Possibility to use Sexed Semen (SexedUltra 4M ) Low cost of semen (cost effective) More pregnancies per year than ET, and AI Challenges Laboratory and skilled personal is needed Pregnancy loss is slightly higher than ET and AI Overall cost is High Challenge to export to some countries

25 Advantages of SexedUltra 4M in IVP Genetic improvement Gender selection with SexedUltra 4M More replacements of the right heifers Increase in milk production, increased net merit (NM$ s), Fat, and Protein Additional income source with extra heifers Heifers calving easy Great conception rates on AI with SexedUltra 4M #1 benefit to Sexed Ultra 4M increased conception rate, close to and / or beat conventionally conception rate

26 Advantages of SexedUltra 4M in IVP Use of Sex Semen on IVF Conv semen 5,4 15,3 Sexed semen 4, Embryos Oocytes Chart 4 - Sexed semen Female/Male: 909 OPUs, 408 Donors, 67 Bulls, 3846 Embryos Conv Semen: 537 OPUs, 276 Donors, 99 Bulls, 2915 Embryos

27 Advantages of SexedUltra 4M ST Partner Herds 122,876 Insemination

28 Recipient Selection Age, Health Status, Breed, DPR Body condition score (BCS); Nutritional Status Reproductive evaluation (Ovaries, Reproductive tract, etc.) Use of CIDR sync/natural heat

29 Conception Rate According Body Score 53,1% 46,9% 48,8% 49,2% 48,1% Percentage, % 39,7% 41,5% 41,7% Conception rate d32 Conception rate d Chart 6 - Conception rate at 32 and 60 days according to body condition score (n=1,919) in dairy heifers submitted to embryo transfer.

30 Conception Rate According to DPR 53,7% 49,5% 48,3% 47,9% 49,6% 42,7% 41,8% 40,5% Percentage, % Conception rate d32 Conception rate d60 (-3.2 until -0.1) (0.1 until 2.0) (2.1 until 4.0) (4.1 until 7.2) Chart 5 - Conception rate at 32 and 60 days and according to daughter pregnancy rate (DPR - n=1,509) in dairy heifers submitted to embryo transfer.

31 Conception Rate According to the Size of Corpus luteum 48,6% 48,1% 49,9% 41,4% 41,3% 43,0% Percentage, % Conception rate at 32d Conception rate at 60d CL 17.0 mm CL mm CL 20.5 mm Chart 7 - Conception rate at 32 and 60 days according to the size of Corpus luteum on day 5 of estrous cycle in dairy heifers submitted to embryo transfer.

32 Conception Rate per Number of Breeding Percentage, % 47,5% 50,5% 48,5% 48,9% 39,3% 43,5% 41,7% 44,3% Conception rate d32 Conception rate d Chart 8 - Conception rate at 32 and 60 days according to number of breeding (IVF, n=2,027) in dairy heifers submitted to embryo transfer.

33 Conception Rate per Type of Heat 48,8% 49,5% 42,1% 41,1% Percentage, % Conception rate d32 Heat synchronization Return to Heat Conception rate d60 Chart 9 - Conception rate at 32 and 60 days according to type of heat (n=2,015) in dairy heifers submitted to embryo transfer.

34 Conception Rate Crossbreed Angus/ Holstein vs. Holstein 48,7% 41,4% 41,6% 35,1% Percentage, % Conception rate 32 days Conception rate 60 days Angus/Holstein Holstein Chart 10 - Conception rate at 32 and 60 days in Crossbreed Angus/Holstein vs. Holstein.

35 CL Side on Ovary Left Ovary Right Ovary 56,5% 43,5% 43,2% 43,4% Percentage, % Percentage of side of CL Conception rate at 32d Chart 11 - Percentage of CL and conception rate at 32 days according to the CL side on ovary (IVF, n=10,343) in dairy heifers submitted to embryo transfer.

36 Conclusion ARTs enabled ST to expedite genetic improvement Production of elite animals to improve genetics in the whole cattle industry Expedite the production of embryos from valuable donors and bulls Decrease the interval between generations Focus on the use of male semen in order to produce bulls Challenge sexed semen in young genomic donors

37 Thank you