Applied Reproductive Technologies Overview in Practice ... · Artificial Insemination ! Animal...
Transcript of Applied Reproductive Technologies Overview in Practice ... · Artificial Insemination ! Animal...
Larry Lanzon, Lander Veterinary Clinic August 17, 2015
2015 ARSBC, UC-‐Davis 1
Applied Reproductive Technologies in Practice You Can Do It!
Larry Lanzon D.V.M., M.P.V.M.
Overview � Artificial insemination � Embryo transfer � Ultrasonography � Ovum pickup and in vitro fertilization � Stem cell oocyte production
Responsibilities � Herd Health � Expected Outcomes ◦ AI ◦ ET
2048 recoveries mean 11.5 ova/embryos, average 6.2 viable embryos (Looney, CR, 1986.) 4440 recoveries mean 15.3 ova/embryos, average 7.7 viable embryos (LVCET 2015, unpubl.)
Artificial Insemination
Animal Selection
• Genetically and economically valuable females • Seed stock producer • Club calf producer • Commercial producer • Marketable progeny • Decreased costs
Estrus Detection Detection in donor and recipients ◦ Donor breeding ◦ Recipient estrus synchrony with transferred embryo.
Detection Methods:
• Cowboy on a horse • Paint Stick® or spray paint • Heat Patch® or Estrotect® patches
• Automatic systems (commercially available)
Heat Watch®
Afimilk®
AccuBreed®
SensOor®
Larry Lanzon, Lander Veterinary Clinic August 17, 2015
2015 ARSBC, UC-‐Davis 2
Synchronization of follicular waves
• GnRH • Prostaglandins • CIDRs
Senger, P. L. (1997). Pathways to pregnancy and parturition.
Current Conceptions, Inc., 1615 NE Eastgate Blvd.
Insemination
• Timing • Semen handling
Animal Handling • Good Facilities
• Flow
• Disposition Score • Pen Score
Range of 1-5 • Exit Score
Time/Distance
Carroll, J. Methods of assessing the stress level of cattle in order to bring about positives changes in reproduction. Society for Theriogenology. 2015. San Antonio, Texas.
Grandin, T., Review: Reducing handling stress improves both productivity and welfare. The professional Animal scientist, 1998. 14(1): p. 1-10.
Ultrasonography
Ultrasonography � Donor reproductive exam � Ovum pick up for in vitro fertilization � Superovulation response � Cow reproductive exam � Bull reproductive exam
U/S - Cow exam Complete exam � Uterus ◦ Pregnancy ◦ Fetal sex ◦ Estrus ◦ Abnormalities
� Ovaries ◦ Follicle ◦ Corpus luteum ◦ Abnormalities
Larry Lanzon, Lander Veterinary Clinic August 17, 2015
2015 ARSBC, UC-‐Davis 3
U/S - Bull exam Complete Examination � External ◦ Spermatic cord ◦ Panpiniform plexus ◦ Epidydimus - head ◦ Epidydimus - body ◦ Epidydimus - tail ◦ Testis
� Internal ◦ Prostate ◦ Ampullae ◦ Seminal vesicles
Embryo Transfer
ET - Application
• Superior females with superior males • Herd dynamics • Repopulation • Genetic reservoir • Research • Import and export
ET - Process � Selection of donor and recipients � Synchronization of donor and recipients � Superovulation of the donor female � Detection of estrus in donor and recipients � Breeding of the donor � Recovery of the embryos from the donor � Transferring embryos to the recipients � Freezing of the embryos
Selection Criteria Donor � Progeny success in ET � Calf >50 days old � No periparturient disease � BCS 2.5-3.5 � Cycling � High level of fertility � Dry
Recipient � Proven production � Virgin heifers, or ≤ 7 years � Calf >60 days of age � No periparturient disease � BCS >2.5 � Cycling
Donor superovulation
� Follicle Stimulating Hormone ◦ Reference estrus ◦ First follicular wave ◦ Second (day 9.5) and third follicular wave (day 10.5)
� CIDR® ◦ GnRH à 2 day FSH start
Larry Lanzon, Lander Veterinary Clinic August 17, 2015
2015 ARSBC, UC-‐Davis 4
Donor superovulation
• Injection Timing • FSH IM b.i.d. 10-12 hrs interval for 4 days
• Prostaglandin with last 2 FSH injections
• CIDR® removed at last prostaglandin injection
• Insemination: • 2 doses 10-12 h standing • 2 doses 10-12 h later
ET - Response Variability � Age � Breed � BCS � Season � Follicular wave size
Nutrition
Armstrong, D.T.,Theriogenology. 55(6): p. 1303-1322. Donaldson, L.E. Theriogenology, 1984. 21(6): p. 1013-1018.
Jaiswal, R., et al.,. Theriogenology, 2009. 72(1): p. 81-90. Mapletoft, R.J. IVIS Reviews in Veterinary Medicine,2006.
Singh, J., et al. Theriogenology, 2004. 62(1): p. 227-243.
ET - Embryo Recovery
• Surgical Recovery • Nonsurgical Recovery
• Syringe method • Gravity flow method
• Solutions • Commercially available • Lactated Ringers with added surfactant
• Recovery filter • Microscopic dish search
ET - Embryo Processing
Embryo Stage • Stage 1: Unfertilized • Stage 2: 2-12 cells • Stage 3: Early morula • Stage 4: Morula • Stage 5: Early blastocyst • Stage 6: Blastocyst • Stage 7: Expanded blastocyst • Stage 8: Hatched blastocyst • Stage 9: Expanding hatched blastocyst
Manual of the International Embryo Transfer Society, 4th Ed
Embryo Grading
• Grade 1: Excellent or good • Grade 2: Fair • Grade 3: Poor • Grade 4: Dead or degenerating
Larry Lanzon, Lander Veterinary Clinic August 17, 2015
2015 ARSBC, UC-‐Davis 5
ET – Transfer of Embryos
• Fresh embryo: • Recipient preparation • Embryo preparation • Embryo Transfer
• Previously frozen embryo: • Recipient preparation • Embryo preparation • Ethylene glycol cryoprotectant • Glycerol cryoprotectant
ET - Cryopreservation
• Straw labeling • Embryo preparation
• Ethylene glycol • Glycerol • Vitrification
• Embryo Freezing • Frozen embryo storage
Photos: WL Walker
Other Technologies
Ovum Pickup (OPU) In Vitro Fertilization (IVF)
� Donor selection � Valuable donor not producing with ET � Stimulated and non-stimulated � Response variability � Process � Time and Temperature
OPU and IVF Results
01/2014 -‐ 06/2015 Total DAIRY BEEF
Number of OPUs 92 41 51
Total oocytes recovered 937 546 391
per cow 10.2 13.3 7.7
Total oocytes matured 757 430 327
per cow 8.2 10.5 6.4
Total G 1 & 2 oocytes matured 512 290 222
per cow 5.6 7.1 4.4
Total Embryos produced 259 131 128
per cow 2.8 3.2 2.5
% Total 34% 30% 39% % Grade 1 and 2 Oocytes 51% 45% 58%
Stem Cell Oocyte Production
� The next ART?
� Less invasive: animal welfare & reproduction
Purified oocyte germ cells capable of proliferating in vitro when transplanted into ovaries of recipient mice generate fully functional oocytes that fertilize to produce viable embryos and offspring.
Larry Lanzon, Lander Veterinary Clinic August 17, 2015
2015 ARSBC, UC-‐Davis 6
Take Home � Responsibilities � Complete exam � Design programs � Embrace technologies
“tools for improved genetics and production”
Larry Lanzon DVM, MPVM Lander Veterinary Clinic Embryo Transfer 2930 Lander Ave. Turlock, CA 95380 209 678 3335 phone 209 634 0643 fax [email protected]